Your SlideShare is downloading. ×
Metabolic syndrome   underlying mechanisms and drug therapies
Upcoming SlideShare
Loading in...5
×

Thanks for flagging this SlideShare!

Oops! An error has occurred.

×
Saving this for later? Get the SlideShare app to save on your phone or tablet. Read anywhere, anytime – even offline.
Text the download link to your phone
Standard text messaging rates apply

Metabolic syndrome underlying mechanisms and drug therapies

541
views

Published on

Published in: Health & Medicine

0 Comments
0 Likes
Statistics
Notes
  • Be the first to comment

  • Be the first to like this

No Downloads
Views
Total Views
541
On Slideshare
0
From Embeds
0
Number of Embeds
0
Actions
Shares
0
Downloads
75
Comments
0
Likes
0
Embeds 0
No embeds

Report content
Flagged as inappropriate Flag as inappropriate
Flag as inappropriate

Select your reason for flagging this presentation as inappropriate.

Cancel
No notes for slide

Transcript

  • 1. Metabolic Syndrome
  • 2. Metabolic Syndrome Underlying Mechanisms and Drug Therapies Edited by Minghan Wang Amgen, Inc., Thousand Oaks, California, USA
  • 3. Copyright Ó 2011 by John Wiley & Sons, Inc. All rights reserved Published by John Wiley & Sons, Inc., Hoboken, New Jersey Published simultaneously in Canada No part of this publication may be reproduced, stored in a retrieval system, or transmitted in any form or by any means, electronic, mechanical, photocopying, recording, scanning, or otherwise, except as permitted under Section 107 or 108 of the 1976 United States Copyright Act, without either the prior written permission of the Publisher, or authorization through payment of the appropriate per-copy fee to the Copyright Clearance Center, Inc., 222 Rosewood Drive, Danvers, MA 01923, (978) 750-8400, fax (978) 750-4470, or on the web at www.copyright.com. Requests to the Publisher for permission should be addressed to the Permissions Department, John Wiley & Sons, Inc., 111 River Street, Hoboken, NJ 07030, (201) 748-6011,fax (201) 748-6008, or online at http://www.wiley.com/go/ permission. Limit of Liability/Disclaimer of Warranty: While the publisher and author have used their best efforts in preparing this book, they make no representations or warranties with respect to the accuracy or completeness of the contents of this book and specifically disclaim any implied warranties of merchantability or fitness for a particular purpose. No warranty may be created or extended by sales representatives or written sales materials. The advice and strategies contained herein may not be suitable for your situation. You should consult with a professional where appropriate. Neither the publisher nor author shall be liable for any loss of profit or any other commercial damages, including but not limited to special, incidental, consequential, or other damages. For general information on our other products and services or for technical support, please contact our Customer Care Department within the United States at (800) 762-2974, outside the United States at (317) 572-3993 or fax (317) 572-4002. Wiley also publishes its books in a variety of electronic formats. Some content that appears in print may not be available in electronic formats. For more information about Wiley products, visit our web site at www.wiley.com. Library of Congress Cataloging-in-Publication Data: Metabolic syndrome : underlying mechanisms and drug therapies / edited by Minghan Wang. p. ; cm. Includes bibliographical references and index. ISBN 978-0-470-34342-5 (cloth) 1. Metabolic syndrome–Pathophysiology. 2. Metabolic syndrome–Chemotherapy. I. Wang, Minghan, 1966- [DNLM: 1. Metabolic Syndrome X–drug therapy. 2. Metabolic Syndrome X–physiopathology. WK 820 M58695 2011] RC662.4.M53 2011 616.3’99–dc22 2010019505 Printed in the United States of America 10 9 8 7 6 5 4 3 2 1
  • 4. Contents Introduction ix Minghan Wang Contributors xi Part One The Physiology of Metabolic Tissues Under Normal and Disease States 1. Gut as an Endocrine Organ: the Role of Nutrient Sensing in Energy Metabolism 3 Minghan Wang 2. Central Glucose Sensing and Control of Food Intake and Energy Homeostasis 29 Lourdes Mounien and Bernard Thorens 3. Abnormalities in Insulin Secretion in Type 2 Diabetes Mellitus 53 Taly Meas and Pierre-Jean Guillausseau 4. Adipokine Production by Adipose Tissue: A Novel Target for Treating Metabolic Syndrome and its Sequelae 73 Vanessa DeClercq, Danielle Stringer, Ryan Hunt, Carla G. Taylor, and Peter Zahradka 5. Hepatic Metabolic Dysfunctions in Type 2 Diabetes: Insulin Resistance and Impaired Glucose Production and Lipid Synthesis 133 Ruojing Yang v
  • 5. 6. Energy Metabolism in Skeletal Muscle and its Link to Insulin Resistance 157 Minghan Wang Part Two Metabolic Diseases and Current Therapies 7. Mechanisms and Complications of Metabolic Syndrome 179 Minghan Wang 8. Emerging Therapeutic Approaches for Dyslipidemias Associated with High LDL and Low HDL 199 Margrit Schwarz and Jae B. Kim 9. Mechanism of Action of Niacin: Implications for Atherosclerosis and Drug Discovery 235 Devan Marar, Shobha H. Ganji, Vaijinath S. Kamanna, and Moti L. Kashyap 10. Current Antidiabetic Therapies and Mechanisms 253 Minghan Wang Part Three Drug Targets for Antidiabetic Therapies 11. GLP-1 Biology, Signaling Mechanisms, Physiology, and Clinical Studies 281 Remy Burcelin, Cendrine Cabou, Christophe Magnan, and Pierre Gourdy 12. Dipeptidyl Peptidase IV Inhibitors for Treatment of Diabetes 327 C.H.S. McIntosh, S.-J. Kim, R.A. Pederson, U. Heiser, and H.-U. Demuth 13. Sodium Glucose Cotransporter 2 Inhibitors 359 Margaret Ryan and Serge A. Jabbour 14. Fibroblast Growth Factor 21 as a Novel Metabolic Regulator 377 Radmila Micanovic, James D. Dunbar, and Alexei Kharitonenkov 15. Sirtuins as Potential Drug Targets for Metabolic Diseases 391 Qiang Tong vi Contents
  • 6. 16. 11b-Hydroxysteroid Dehydrogenase Type 1 as a Therapeutic Target for Type 2 Diabetes 423 Clarence Hale and David J. St. Jean, Jr. 17. Monoclonal Antibodies for the Treatment of Type 2 Diabetes: A Case Study with Glucagon Receptor Blockade 459 Hai Yan, Wei Gu, and Murielle Veniant-Ellison Part Four Lessons Learned and Future Outlook 18. Drug Development for Metabolic Diseases: Past, Present and Future 471 Minghan Wang Index 489 Contents vii
  • 7. Introduction It has been more than 20 years since Reaven first introduced the concept of syndrome X or insulin resistance syndrome to describe the clustering of several cardiovascular risk factors. The concept has evolved over the years and is now commonly referred to as metabolic syndrome, which covers the individual metabolic abnormalities of obesity, insulin resistance, hyperglycemia, dyslipidemia (high triglycerides and low HDL), and hypertension. Patients with metabolic syndrome have increased risk of developing cardiovascular disease (CVD) and type 2 diabetes mellitus (T2DM). Despite the debates surrounding the existence and definition of metabolic syndrome, the concept has been useful in understanding the interconnections of the various risk factors that are common in a large population of patients and thereby managing the overall disease risk. From the drug discovery standpoint, all the components of metabolic syndrome are therapeutic targets for the treatment of CVD and T2DM to reduce comorbidities and overall mortality. While there is a wealth of information concerning the clinical features and mechanisms of metabolic syndrome, putting them in the physiological context relevant to the development of therapeutics is essential for drug discovery. The goal of this book is to provide comprehensive understanding of the molecular and physiological abnormalities associated with metabolic syndrome and the therapeutic strategies for drug development. Part One is devoted to gaining an integrated understanding of the metabolic abnormalities at the tissue and pathway levels that are associated with disease states. In Part Two, metabolic syndrome is discussed at the physiological level and current therapies are summarized. These sections help lay the foundation to identify pathways and molecular targets for the development of antidiabetic therapies in Part Three. Since more than 80% type 2 diabetic patients have metabolic syndrome, a large portion of this book is devoted to antidiabetic therapies. Finally, the successes and failures in developing antidiabetic and cardio- vascular drugs and lessons learned are discussed in Part Four. Although the chapters are contributed by different authors, the organization and the content of the book have been carefully designed so that the information is presented systematically. In the meantime, each chapter independently covers a subarea of metabolic or drug discovery topics, the reader has the flexibility to gain information on a specific tissue, pathway, or target in a time-efficient manner. Despite the exciting advances that have been made in developing antidiabetic and CVD therapies in the past several ix
  • 8. decades, drug discovery in these areas continues to be a challenge. I hope this book will help the reader better understand the exciting science behind metabolic drug discovery and development and develop a greater appreciation of the complexity of metabolic syndrome as well as the treatment strategies. MINGHAN WANG x Introduction
  • 9. Contributors Remy Burcelin, Rangueil Institute of Molecular Medicine, INSERM U858, Toulouse, France Cendrine Cabou, Rangueil Institute of Molecular Medicine, INSERM U858, Toulouse, France Vanessa DeClercq, Department of Human Nutritional Sciences, University of Manitoba, Winnipeg, Canada; Canadian Centre for Agri-Food Research in Health and Medicine, St. Boniface Hospital Research Centre, Winnipeg, Canada H.-U. Demuth, Probiodrug AG, Biocenter, Halle (Saale), Germany James D. Dunbar, BioTechnology Discovery Research, Lilly Research Laborato- ries, Lilly Corporate Center, Indianapolis, IN, USA Shobha H. Ganji, Department of Veterans Affairs Healthcare System, Atheroscle- rosis Research Center, Long Beach, CA, USA; Department of Medicine, University of California, Irvine, CA, USA Pierre Gourdy, Rangueil Institute of Molecular Medicine, INSERM U858, Tou- louse, France Wei Gu, Department of Metabolic Disorders, Amgen, Inc., Thousand Oaks, CA, USA Pierre-Jean Guillausseau, APHP, Department of Internal Medicine B, Hoˆpital Lariboisiere, Paris, France; Universite Paris 7, Paris, France Clarence Hale, Department of Metabolic Disorders, Amgen, Inc., Thousand Oaks, CA, USA U. Heiser, Probiodrug AG, Biocenter, Halle (Saale), Germany Ryan Hunt, Department of Human Nutritional Sciences, University of Manitoba, Winnipeg, Canada; Canadian Centre for Agri-Food Research in Health and Medicine, St. Boniface Hospital Research Centre, Winnipeg, Canada Serge A. Jabbour, Division of Endocrinology, Diabetes, and Metabolic Diseases, Department of Medicine, Jefferson Medical College, Thomas Jefferson University, Philadelphia, PA, USA xi
  • 10. Vaijinath S. Kamanna, Department of Veterans Affairs Healthcare System, Atherosclerosis Research Center, Long Beach, CA, USA; Department of Medicine, University of California, Irvine, CA, USA Moti L. Kashyap, Department of Veterans Affairs Healthcare System, Atheroscle- rosis Research Center, Long Beach, CA, USA; Department of Medicine, University of California, Irvine, CA, USA Alexei Kharitonenkov, BioTechnology Discovery Research, Lilly Research Laboratories, Lilly Corporate Center, Indianapolis, IN, USA Jae B. Kim, Global Development, Amgen, Inc., Thousand Oaks, CA, USA S.-J. Kim, Department of Cellular and Physiological Sciences, Life Sciences Institute, University of British Columbia, Vancouver, Canada; Diabetes Research Group, Life Sciences Institute, University of British Columbia, Vancouver, Canada Christophe Magnan, INSERM U858, Toulouse, France; University Paris Diderot, CNRS, Paris, France C.H.S. McIntosh, Department of Cellular and Physiological Sciences, Life Sciences Institute, University of British Columbia, Vancouver, Canada; Diabetes Research Group, Life Sciences Institute, University of British Columbia, Vancouver, Canada Devan Marar, Department of Veterans Affairs Healthcare System, Atherosclerosis Research Center, Long Beach, CA, USA; Department of Medicine, University of California, Irvine, CA, USA Radmila Micanovic, BioTechnology Discovery Research, Lilly Research Labora- tories, Lilly Corporate Center, Indianapolis, IN, USA Taly Meas, APHP, Department of Internal Medicine B, Hoˆpital Lariboisiere, Paris, France; Universite Paris 7, Paris, France Lourdes Mounien, Department of Physiology, University of Lausanne, Lausanne, Switzerland; Center for Integrative Genomics, University of Lausanne, Lausanne, Switzerland R.A. Pederson, Department of Cellular and Physiological Sciences, Life Sciences Institute, University of British Columbia, Vancouver, Canada; Diabetes Research Group, Life Sciences Institute, University of British Columbia, Vancouver, Canada Margaret Ryan, Division of Endocrinology, Diabetes, and Metabolic Diseases, Department of Medicine, Jefferson Medical College, Thomas Jefferson University, Philadelphia, PA, USA Margrit Schwarz, Department of Metabolic Disorders, Amgen, Inc., South San Francisco, CA, USA xii Contributors
  • 11. David J. St. Jean, Jr., Department of Medicinal Chemistry, Amgen, Inc., Thousand Oaks, CA, USA Danielle Stringer, Department of Human Nutritional Sciences, University of Manitoba, Winnipeg, Canada; Canadian Centre for Agri-Food Research in Health and Medicine, St. Boniface Hospital Research Centre, Winnipeg, Canada Carla G. Taylor, Department of Human Nutritional Sciences, University of Manitoba, Winnipeg, Canada; Canadian Centre for Agri-Food Research in Health and Medicine, St. Boniface Hospital Research Centre, Winnipeg, Canada Bernard Thorens, Department of Physiology, University of Lausanne, Lausanne, Switzerland; Center for Integrative Genomics, University of Lausanne, Lausanne, Switzerland Qiang Tong, USDA/ARS Children’s Nutrition Research Center, Baylor College of Medicine, Houston, TX, USA; Department of Pediatrics, Baylor College of Medicine, Houston, TX, USA; Department of Medicine, Baylor College of Medicine, Houston, TX, USA; Department of Molecular Physiology and Biophysics, Baylor College of Medicine, Houston, TX, USA Murielle Veniant-Ellison, Department of Metabolic Disorders, Amgen, Inc., Thousand Oaks, CA, USA Minghan Wang, Metabolic Disorders Research, Amgen, Inc., Thousand Oaks, CA, USA Hai Yan, Department of Protein Science, Amgen, Inc., Thousand Oaks, CA, USA Ruojing Yang, Department of Metabolic Disorders – Diabetes, Merck Research Laboratories, Rahway, NJ, USA Peter Zahradka, Department of Physiology, University of Manitoba, Winnipeg, Canada; Department of Human Nutritional Sciences, University of Manitoba, Winnipeg, Canada; Canadian Centre for Agri-Food Research in Health and Medicine, St. Boniface Hospital Research Centre, Winnipeg, Canada Contributors xiii
  • 12. Part One The Physiology of Metabolic Tissues Under Normal and Disease States
  • 13. Chapter 1 Gut as an Endocrine Organ: the Role of Nutrient Sensing in Energy Metabolism MINGHAN WANG Department of Metabolic Disorders, Amgen, Inc., Thousand Oaks, CA, USA INTRODUCTION Energy homeostasis is balanced by food intake and energy expenditure. Both events are controlled by complex sets of neuronal and hormonal actions. Food intake is driven by a central feeding drive, namely, the appetite, which is induced under the fasting state after energy consumption through physical activities. Following food digestion, the passage of nutrients through the gastrointestinal (GI) tract generates signals that produce sensations of fullness and satiation. In particular, nutrients interact with receptors in the small intestine and stimulate the release of peptide hormones, the actions of which mediate physiological adaptations in response to energy intake. The commonly known GI peptides include the incretins, glucagon-like peptide 1 (GLP-1) and glucose-dependent insulinotropic polypeptide or gastric inhibitory peptide (GIP), as well as peptide tyrosine tyrosine (PYY), cholecystokinin (CCK), and oxyntomodulin. These peptide hormones are secreted from different regions of the small intestine. GLP-1, oxyntomodulin, and PYY are secreted from endocrine L cells that are mainly distributed in the distal small intestine (1, 2), whereas GIP is secreted from endocrine K cells primarily localized in the duode- num (3, 4). CCK is secreted from I cells in the duodenum (5). Nutrients released through the digestive tract induce secretion of GI peptide hormones, which subse- quently bind to their respective receptors and trigger a cascade of physiological events. These receptors are expressed in tissues such as the central nervous system (CNS), the GI tract, and pancreas, and upon activation lead to suppression of appetite, Metabolic Syndrome: Underlying Mechanisms and Drug Therapies Edited by Minghan Wang Copyright Ó 2011 John Wiley Sons, Inc. 3
  • 14. reduced gastric emptying, and assimilation of nutrients. Nutrients can also suppress the secretion of GI peptides. For example, ghrelin, a peptide hormone released by the stomach under the fasting state that stimulates food intake (6), is suppressed after food ingestion (6). GI peptides mediate two principal physiological events: (i) the feedback response on the CNS and the stomach to reduce food intake and slow gastric emptying, and (ii) the feedforward response, mediated particularly by the incretins, to prepare tissues for nutrient integration. In this regard, the small intestine is not only an organ for nutrient absorption but also a major site for providing hormonal regulation of energy intake and storage. GLP-1 and GIP are called incretins because they act on the pancreatic b-cells to increase insulin secretion at normal or elevated glucose levels. They also regulate glucagon secretion by pancreatic a-cells. These actions represent a critical step in preparing the body to switch from the fasting state to postprandial activities. By suppressing glucagon secretion, GLP-1 shut down hepatic gluconeo- genesis and adipose lipolysis, two key biological pathways in maintaining energy homeostasis under the fasting state. In addition, GLP-1 can act directly on liver and muscle to regulate glucose metabolism independent of its incretin action (7). In the meantime, induction of insulin secretion by the incretins facilitates glucose uptake by the peripheral tissues. GLP-1 is also involved in the feedback response by acting on the CNS to suppress food intake. PYY and CCK exhibit a similar effect in the CNS underscoring the complexity of appetite regulation. The magnitude and potency of the feedback and feedforward responses depend on both the nutrient content and the length of small intestine exposed. Although both glucose and free fatty acids (FFAs) modulate the secretion of GI peptides, their actions are mediated by distinct mechanisms because they have different residence times in the small intestine and interact with different nutrient-sensing receptors. In fact, even the activity of FFAs varies with their chain length. Moreover, the intestinal length exposed to nutrients and the nutrient contact sites are important determinants in GI peptide secretion. FOOD INTAKE AND NUTRIENT-SENSING SYSTEMS IN THE GI TRACT Afteringestion,foodchimeismixedwithdigestivejuicesinthestomachandpropelled into the small intestine. The three segments of the small intestine, the duodenum, the jejunum, and the ileum, perform different digestive functions (Figure 1.1). Nutrients are generated from the digestion of carbohydrates, fat, protein, and other food components. The passage of nutrients through the small intestine not only facilitates absorption but also plays a role in regulating gastric emptying and satiety. The interactionofnutrientswiththesmallintestinesegmentsgeneratessignalsthatregulate the rate of gastric emptying and food intake. The nutrient-sensing system consists of receptors, channels, and transporters in the open-type cells on the small intestine luminalsurface.Itrespondstomacronutrientsandactivatessignalingpathwaysleading to the release of GI peptides, which subsequently act on the stomach and the CNS to 4 Chapter 1 Gut as an Endocrine Organ
  • 15. slow gastric emptying and suppress appetite, respectively (Figure 1.1). In addition, some peptides such as the incretins stimulate insulin secretion and regulate glucagon secretion to help integrate nutrients into tissues post absorption (Figure 1.1). Studies in pigs demonstrated that rapid injection of glucose into the duodenum during or immediately prior to feeding suppressed food intake (8, 9). The reduction in food intake far exceeded the energy content of the infused glucose (8, 9), suggesting that the effect of glucose on food intake is likely to be mediated by signaling events. In the meantime, hepatic portal or jugular infusion of glucose in pigs did not alter short- term food intake (10). These data suggest that the regulatory effect of glucose on food intake is a preabsorptive event and the sites of regulation are in the GI tract. To further understand the mechanisms by which dietary carbohydrates regulate energy intake, glucose was infused into the stomach or different segments of the small intestine in pigs. The infusion started 30 min prior to the meal and continued until the pigs stopped eating (11). It was found that infusion of glucose into the stomach, duodenum, jejunum, or ileum each suppressed food intake (11). But comparatively, jejunal infusion caused more reduction in food intake than elsewhere (11). These data suggest that glucose may interact with receptors or other sensing components expressed in various parts of the small intestine to control short-term energy intake. In addition to glucose, FFAs released from fat digestion also play important albeit more complex roles in controlling energy intake. Healthy human volunteers receiving ileal infusion Duodenum Jejunum Ileum Colon Stomach Food (carbohydrates, fat, protein, etc.) Enterocyte Enteroendocrine cell GI peptide Neurons Circulation in bloodstream Target tissues Figure 1.1 Localization of enteroendocrine cells in the GI tract. Enteroendocrine cells (exemplified by an L cell) are on the surface of the GI tract where their luminal sides detect nutrients passing in the lumen, leading to intracellular signals that stimulate the secretion of GI peptides. GI peptides exert biological effects by acting on their receptors in nearby neurons that transduce signals to target tissues. The peptides are carried to target tissues through the circulation and act locally on sites such as the CNS, the pancreas, and the stomach. Food Intake and Nutrient-Sensing Systems in the GI Tract 5
  • 16. of lipids consumed a smaller amount of food and energy and had delayed gastric emptying (12). Ileal lipid infusion also accelerated the sensation of fullness during a meal(12).However,intravenous (i.v.)infusionoflipids didnotaffect foodintake (12), suggesting that lipids may interact with ileal receptors to induce satiety and reduce food consumption. Further studies suggest that digestion is a prerequisite for the inhibitory effect of fat on gastric emptying and energy intake. For example, admin- istration of a lipase inhibitor increased food intake in healthy subjects or type 2 diabetic patients receiving a high-fat meal (13, 14), suggesting that FFAs, the breakdown products of fat after ingestion, rather than triglycerides, are the active nutrients that exert the regulatory effects. Likewise, sugars from carbohydrate digestion, rather than carbohydrates themselves, are the active nutrients that induce intestinal signals. Although both glucose and FFAs can stimulate a set of GI peptides that regulate appetite, gastric emptying, and insulin and glucagon release, they have differential effects. For example, glucose stimulates robustsecretion of both GLP-1 and GIP, whereas FFAs from a fat meal elicit only modest GLP-1 secretion despite equally robust GIP secretion (15). Further, not all FFAs are equally active since the stimulatory effect depends on their chain length. Although FFAs with a chain length of greater than C12 stimulate CCK release, further increase in chain length has no additional effect, and C11 or shorter FFAs are not active (16, 17). Like carbohydrate and fat meals, protein meals also activate the nutrient-sensing system but in different ways. In healthy human subjects, plasma GIP levels were elevated after both carbohydrate and fat meals but not a protein meal (15). However, intraduodenal amino acid perfusion in human subjects stimulated both GIP and insulin secretion (18, 19). Oral ingestion of mixed amino acids by healthy volunteers also increased plasma GLP-1 levels (20). These findings suggest that amino acids can function as nutrient-sensing agents, and a protein meal is likely to contribute to nutrient sensing in the GI tract. However, since mixed amino acids are not equivalent to a digested protein meal, GLP-1 secretion was studied in humans following a protein meal (15). A transient peak was observed at 30 min followed by a steady-state rise throughout the rest of the 3 h study period (15). The nutrients from the protein meal that stimulated GLP-1 secretion were a mixture of protein hydrolysates but not amino acids per se. It is important to carry out studies with protein hydrolysates that mimic the digested products in the GI tract. A protein hydrolysate (peptone) containing 31% free amino acids and 69% peptides induced the secretion of PYY and GLP-1 in the portal effluent of isolated vascularly perfused rat ileum after luminal administra- tion (21). Peptones also induced CCK secretion and transcription in STC-1 cells, an established L cell line (22, 23). Peptones made from both albumin egghydrolysate and meat hydrolysate stimulated the transcriptional expression of the proglucagon gene encoding GLP-1 in two L cell lines but not pancreatic glucagon-producing cell lines (24), suggesting that the signaling pathways mediating this effect are L cell/ small intestine specific. In STC-1 cells, the proglucagon promoter contains elements responsive to peptones (25). In contrast, the mixture of free amino acids is at best a weak stimulant (21, 24). These data suggest that free amino acids may have a limited role in protein meal-stimulated GLP-1 or PYY secretion. However, amino acids are indeed involved in nutrient sensing in the GI tract. Aromatic amino acids may play a 6 Chapter 1 Gut as an Endocrine Organ
  • 17. role in gastrin secretion because they activate the calcium-sensing receptor (CaR) on gastrin-secreting antral cells (26, 27). In addition, amino acids also stimulate CCK release (28, 29) and gastric acid secretion (30). In addition to glucose, FFAs, amino acids, and digested peptides from proteins, other nutrients are also involved in the regulation of GI peptide secretion (21). At physiological concentrations, bile acids stimulate the secretion of PYY, GLP-1, and neurotensin (NT) (21). Interestingly, the threshold concentration of taurocholate for PYY and GLP-1 stimulation is about twofold that required for stimulating NT release (21), suggesting that there is a slight difference in the sensitivity of L cells and N cells to bile acids (21). In addition to the small intestine, the stomach plays an important role in terminating a meal. When rats were implanted with an extra stomach to which a liquid diet was infused, food intake was reduced regardless of whether food was allowed to empty into the small intestine or retained in the stomach (31). This effect is not likely to be mediated by neuronal mechanisms because the implanted stomach was completely denervated (31). This result suggests that the implanted stomach may have generated hormonal signals that affect food intake, and these hormonal signals may mediate the ability of the stomach to sense nutrient quality and quantity to alter the rate of gastric emptying and amount of food ingested (32, 33). MOLECULAR MECHANISMS OF NUTRIENT SENSING It has been recognized that it is the monomeric nutrients that interact with luminal small intestinal receptors or other nutrient-sensing components and regulate the feedback and feedforward responses to food intake. What do we know about these receptors and their downstream pathways? The analogy between the intestinal nutrient sensing and taste reception by the tongue can shed new light on this question. Glucose sensing in taste buds is mediated by taste receptors expressed in the lingual epithelium (34). These receptors are G protein-coupled receptors (GPCRs) in the apical membranes of taste receptor cells (34). All the three members of the taste receptor family 1 (T1R) class of GPCRs are involved in this function by acting in combination to sense different tastes. The T1R2/T1R3 heterodimer senses sweet taste whereas the T1R1/T1R3 heterodimer senses amino acids and umami taste (35). These receptors activate a phospholipase C (PLC) b2-dependent pathway to increase intracellular Ca2 þ concentrations by coupling to the G proteins gustducin and/or transducin (34). The activated taste receptors may also stimulate the cAMP-depen- dent pathway (34). In an in vitro assay where T1R2/T1R3 were coupled to Ga15, a promiscuous G protein linked to PLC, T1R2/T1R3 responded to sweet taste stimuli, including glucose, fructose, lactose, and galactose, as well as synthetic sweet- eners (35). The activity was inhibited by the sweet taste inhibitor lactisole (35). These data indicatethat the T1R2/T1R3 complex mediates sweetsensation alongwith other components such as G proteins and PLC. Interestingly, the key components of the sweet taste transduction pathways are also expressed in the gut enteroendocrine cells (36), with the signaling events leading to GI peptide secretion by these cells (Figure 1.2). For example, the three members of Molecular Mechanisms of Nutrient Sensing 7
  • 18. ProteinFatCarbohydrates Taste receptors (i.e., T1R2/T1R3) G proteins (i.e., gustducin) PLCβ2 Glucose Glucokinase Closure of KATPchannels SGLT2 Electrogenic activity Opening of voltage- dependent Ca2+ channels ATP/ADP ratio Membrane depolarization Intracellular [Ca2+ ] Secretion of GI peptides (i.e., GLP-1) CaR, GPR93, and others GPR40, GPR119, and GPR120 FFAs Amino acids and peptides G proteins (Gs or Gq) Intracellular [Ca2+ ] or Intracellular cAMP Secretion of GI peptides G proteins Secondary messengers Secretion of GI peptides Figure 1.2 Potential signaling cascades that mediate GI nutrient sensing in response to main nutrients. Macronutrients, including sugars, FFAs, and amino acids/peptides, are derived through digestion from carbohydrates, fat, and protein. There are three potential pathways that can sense glucose: taste receptors, KATP channels, and SGLT1. FFAs and amino acids/peptides can activate GPCRs expressed in enteroendocrine cells. Activation of downstream signaling by these mechanisms triggers secretion of GI peptides. 8
  • 19. the T1R class of GPCRsare detected in brush cells, one form of solitary chemosensory cells (SCCs), in the apical membranes of rat jejunum (37). Also found in these cells are a-gustducin, transducin, and PLCb2 (37). In addition, a-gustducin is also expressed in brush cells of the stomach, the duodenum, and pancreatic ducts in rats (38, 39). Brush cells have a structure similar to lingual taste cells (39), suggesting that they may use similar nutrient-sensing pathways. Consistent with the findings in rats, T1R2, T1R3, and a-gustducin are expressed in mouse small intestine (40). Taste signaling elements, including the three subunits of gustducin (a-gustducin, Gb3, and Gg13), PLCb2, and taste receptors, were also found in human L cells (41). Taken together, these data suggest that the taste receptors and associated signaling compo- nents are present in gut cells and may be involved in nutrient sensing in a fashion similar to that by the lingual epithelium of the tongue. There are two functional consequences upon the activation of the taste receptor systems in the gut. The first is the release of GI peptides such as GLP-1, which mediates both feedback and feedforward responses to food intake as described above. Glucose induces GLP-1 secretion from enteroendocrine L cells by stimulating the taste receptors, the signal of which is mediated by the taste G protein gustducin. The role of gustducin in sugar sensing and glucose homeostasis was exemplified in a-gustducin null mice (41). In wild-type mice, ingestion of glucose induced a marked increase of GLP-1 secre- tion (41); in contrast, a-gustducin null mice exhibited defective GLP-1 secretion in response to glucose ingestion (41), suggesting that L cells of the gut sense glucose through similar mechanisms used by taste cells of the tongue. Thus, the gut cells can “taste” sugars and release mediators, such as the incretins, that in turn regulate food intake and nutrient assimilation. The second consequence of the taste receptor activation in the GI tract is elevated glucose transporter 2 (GLUT2) insertion on the apical membrane of the gut lumen to increase glucose absorption (37). The basal level of glucose absorption in the gut is mediated by sodium–glucose cotran- sporter 1 (SGLT1) and GLUT2 when glucose level is around 20 mM (37). At higher local glucose concentrations (30–100 mM), increased insertion of GLUT2 in the apical membrane occurs to facilitate additional glucose absorption (37). GLUT2 provides three to five times more capacity for glucose absorption than the SGLT1 pathway (37). Despite the above evidence that supports the role of the taste receptor system in mediating nutrient-sensing effects in the GI tract, several research groups have reported findings that dispute this notion. Although the artificial sweetener sucralose was shown to stimulate GLP-1 secretion from human L cells in vitro (41), it did not stimulate GLP-1 secretion in primary L cells (42). In addition, it did not stimulate GLP-1 or GIP release in healthy humans when delivered by intragastric infusion (43). This is in agreement with an earlier study in type 2 diabetic patients where the sweetener stevioside had no effect on GLP-1 or GIP release (44). Further, several sweeteners, including sucralose, were tested in Zucker diabetic fatty rats for their nutrient-sensing activity (45). Consistent with the previous reports, none of these sweeteners increased incretin secretion (45). Taken together, these data indicate that the role of the taste receptor system in GI nutrient sensing remains to be further clarified. Molecular Mechanisms of Nutrient Sensing 9
  • 20. Two additional signaling pathways in the GI tract have been proposed that could mediate GLP-1 secretion in response to glucose exposure. The first one is the classical glucose-sensing machinery employed by pancreatic b-cells for eliciting glucose- dependent insulin secretion (46). This machinery includes components such as ATP- sensitive potassium (KATP) channels and glucokinase (Figure 1.2). In this pathway, glucokinaseservesastherate-limitingstepinglucosemetabolismandthereforeisalso termed “glucose sensor.” Glucose metabolism increases the ATP/ADP ratio, which causes the closure of KATP channels and depolarization of the b-cell membrane. Next, membrane depolarization leads to opening of voltage-dependent Ca2 þ channels and accumulation of intracellular Ca2 þ , which triggers insulin release. Both the KATP channelsubunitsKir6.2andSUR1andglucokinaseweredetectedinGLUTagcells,an L cell line (46). In these cells, glucose concentrations between 0 and 20 mM decreased membrane conductance, caused membrane depolarization, and triggered action potentials (46). Tolbutamide also triggered action potentials in GLUTag cells (46), presumably by blocking the KATP channels. These data suggest that the classical glucose-sensing machinery involving glucokinase and KATP channels mediates glu- cose-induced GLP-1 release from L cells. However, if this notion is true, GLP-1 and GIP secretion following an oral glucose challenge should be lower in individuals with heterozygousglucokinasemutationsthatconferreducedactivity.Unfortunately,when heterozygous glucokinase mutation carriers were subjected to oral glucose tolerance test (OGTT), they did not have altered GLP-1 or GIP secretion post oral glucose challenge compared to normal controls (47). This observation suggests that the glucokinase and KATP channel pathway does not mediate incretin secretion in thegut,oritisinvolvedbutthereareotherredundantpathwaysthatcancompensatefor it. SGLT1 represents another novel glucose-sensing mechanism that triggers GLP-1 secretion (Figure 1.2). Both SGLT1 and SGLT3 are expressed in GLUTag cells (48), and GLP-1 secretion in response to glucose is inhibited by phlorizin, a SGLTinhibitor compound (48). Moreover, the EC50 value of glucose for glucose-induced GLP-1 secretionmatchestheKm ofSGLT1(49).ThesedatasuggestthatSGLT1coulddirectly mediate glucose-induced GLP-1 release. This effect could be attributed to the electrogenic activity of SGLT1 because low glucose concentrations were shown to trigger small inward currents as they enter cells (48). This current could cause membrane depolarization, which could induce GLP-1 release (Figure 1.2). Like sugars, amino acids and FFAs also regulate endocrine response to food intake through activation of their respective GPCRs in enteroendocrine cells (Fig- ure 1.2). L-Amino acids activate the T1R1/T1R3 heterodimer, which mediates umami taste in taste buds (35). These GPCRs are also expressed in the apical membranes of the gut (37) and couple to the G protein transducin to activate PLCb2 and stimulate Ca2 þ mobilization. Through this signaling system, amino acids may mediate GI peptide release and regulate food intake. In addition, the extracellular CaR may also act to sense amino acids released from protein digestion. CaR is abundantly expressed in epithelial cells and neurons of the stomach, the small intestine, and the large intestine (50). In the stomach, CaR is expressed on gastrin-releasing G cells and its activation stimulates intracellular Ca2 þ mobilization via the activation of PLC (51). 10 Chapter 1 Gut as an Endocrine Organ
  • 21. CaR can be activated by aromatic amino acids (52), suggesting that it may act as a nutrient-sensing receptor in response to a protein diet. However, in the absence of Ca2 þ , aromatic amino acids had no effect on CaR-mediated signaling (52), suggest- ing that aromatic amino acids are not CaR agonists; rather, they may act as allosteric modulators to enhance the sensitivity of CaR to its agonist Ca2 þ . The proposed role of CaR in amino acid sensing has physiological support. Analysis of human jejunal content before and 3 h after ingestion of a protein-rich meal revealed that aromatic amino acids were more preferentially released than acidic, polar, and aliphatic amino acids (53). For example, the phenylalanine concentration in jejunum could reach 2 mM (53), a level similar to the EC50 value of phenylalanine in a Ca2 þ mobi- lization assay (52). In addition, L-phenylalanine can activate CCK secretion, pre- sumably through CaR (54, 55). Further, protein hydrolysates directly activate GPR93 in enterocytes, suggesting that multiple GPCRs are involved in sensing of protein nutrients (23). The G protein species to which CaR and GPR93 are coupled are diverse; they depend on specific conditions in different cell types (56) and ligand species (23). As a result, these receptors stimulate the accumulation of a number of secondary messengers. Like glucose and amino acids, longer FFAs appear to interact directly with GPCRs in enteroendocrine cells. The FFA receptor GPR40 is a GPCR highly expressed in pancreatic b-cells mediating the FFA-stimulated glucose-depen- dent insulin secretion (57). GPR40 is activated by medium- and long-chain FFAs (57, 58). Interestingly, it is also expressed in endocrine L and K cells of the GI tract and mediates GLP-1 and GIP secretion (59). GPR120 is another GPCR expressed in the intestine especially in GLP-1 positive cells and acts as a receptor for unsaturated long- chain FFAs (60). Activation of GPR120 both in vitro and in vivo led to increased GLP- 1 secretion (60), suggesting that GPR120 is a major intestinal FFA sensing receptor that mediates incretin release. Further, a recent study indicates that GPR120 also mediates the stimulation of CCK release by FFAs (61). GPR119, a receptor for endogenous ligands oleoyl-lysophosphatidylcholine (OLPC) and oleoylethanola- mide (OEA) (62, 63), is expressed in pancreatic b-cells and upon activation enhances glucose-dependent insulin secretion (63). GPR119 is also localized in L cells and oral administration of a GPR119 agonist increased the release of both GLP-1 and GIP in normal but not GPR119 knockout mice (64), suggesting that GPR119 mediates long- chain FFA-induced incretin release. The three GPCRs trigger different intracellular signaling pathways. GPR40 is coupled to the Gq-PLC pathway and upon activation increases the intracellular Ca2 þ accumulation (65), which leads to incretin secretion. Similarly, GPR120 also induces incretin release by triggering the accumulation of intracellular Ca2 þ (60). GPR119 is coupled to Gs and stimulates intracellular cAMP accumulation (66). In addition to enteroendocrine cells, the intestinal mucosa has two other types of sensory systems, neurons and immune cells (67). The sensory neurons are involved in the control of GI motility and signaling to the CNS that controls feeding behavior (67). The immune cells protect against harmful substances that may enter the GI tract. All the three sensing systems work in concert through direct contact with the intestinal contents. Molecular Mechanisms of Nutrient Sensing 11
  • 22. REGULATION OF INCRETIN SECRETION In 1902, Bayliss and Starling discovered that acid extracts of intestinal mucosa contained a hormone that could be carried to distal tissues via blood circulation and stimulate the exocrine secretion of the pancreas, and named this factor secretin (68). To test if this factor could be used to treat diabetes, Moore et al. administered duodenal mucosa extracts orally to several type 1 diabetics but did not see clear effects (69). The term “incretin” was first proposed by La Barre in 1932 to describe a hormone extracted from the upper gut mucosa with hypoglycemic effect (70). However, the existence of incretin was not proven until 1964, when two independent research groups discovered that an oral glucose load is associated with a significantly greater insulin response than intravenous administration of the same amount of glucose in human subjects (71, 72). The incretin activity was further evaluated by conducting i.v. glucose infusion isoglycemic to the profile generated from an oral glucose challenge. Despite the identical plasma glucose profiles generated by both the oral and the i.v. routes, the oral glucose challenge stimulated greater levels of insulin and C-peptide (73, 74), suggesting that intestinal factors may be released and involved in the stimulation of insulin secretion after oral glucose ingestion. This so-called “incretin effect” describes the important communication through enteroendocrine factors from the GI tract to pancreas in response to food ingestion. This response is a key part of the feedforward mechanism that increases insulin secretion in anticipation of rising blood glucose after food ingestion. There are two incretins, GLP-1 and GIP, both of which are rapidly released to the bloodstream after meal ingestion and stimulate glucose-dependent insulin secretion (GSIS) by pancreatic b-cells. In addition, GLP-1 also suppresses glucagon release by pancreatic a-cells, food intake, and gastric emptying, and is cardiopro- tective. In contrast, GIP does not exhibit these effects. GLP-1 is secreted from intestinal L cells, which are predominantly found in the distal jejunum, ileum, colon, and rectum (1). However, the distribution of L cells throughout the GI tract is somewhat species specific. The overall L cell density in rat or pig GI tract is greater than that in human gut (1), and higher levels are located in the distal jejunum, ileum, and rectum relative to other intestinal regions in humans (1). In dogs, L cells are predominantly concentrated in the jejunum and less so in the ileum (4). Recently, GLP-1 immunoreactive cells were detected in human duodenum (75), and GLP-1 and GIP were colocalized in a subset of endocrine cells in the small intestine (76). GIP is secreted from K cells located primarily in the duodenum (3), but they can be found in other parts of the small intestine (76). For instance, in dogs, GIP-secreting K cells are equally distributed in the duodenum and the jejunum (4). Both L and K cells are open-type endocrine cells that are in immediate contact with nutrients in the intestinal lumen, allowing nutrient-dependent regulation of incretin secretion. GIP is a 42-amino acid secreted peptide initially isolated from intestinal mucosa. It was named gastric inhibitory peptide but later renamed glucose-dependent insu- linotropic peptide for its ability to stimulate insulin secretion (77). The secreted GIP from intestinal K cells is the active form GIP(1–42). GIP is rapidly cleaved at the 12 Chapter 1 Gut as an Endocrine Organ
  • 23. N-terminus by dipeptidyl peptidase-4 (DPP-4) (also known as DPP IV, DP 4, CD26, and adenosine deaminase binding protein), an amino peptidase found in almost all organs and tissues (78), producing the inactive form GIP(3–42) (79). DPP-4 also processes other peptides such as GLP-1 (79), chemokines (80–82), and neuropep- tides (83). GLP-1 is part of the proglucagon polypeptide that is expressed in both intestinal L cells and pancreatic a-cells. The proglucagon polypeptide is processed posttranslationally by prohormone convertases (PC) 1/3 and 2. PC1/3 is expressed in L cells whereas PC2 is expressed in a-cells. The tissue-specific expression of the convertase isoforms dictates which mature peptides are generated from the proglu- cagon polypeptides. In the small intestine, the posttranslational processing by PC1/3 produces GLP-1, GLP-2, glicentin, and oxyntomodulin (84, 85). In contrast, in pancreatic a-cells, PC2-mediated posttranslational processing generates glucagon, glicentin-related pancreatic peptide (GRPP), and the major proglucagon fragment (MPGF) that contains the GLP-1 and GLP-2 segments within its sequence (85). There are two equipotent active forms of GLP-1, GLP-1(7–36)amide and GLP-1(7–37). Both forms are prone to proteolytic cleavage by DPP-4 generating inactive GLP-1(9–36)amide and GLP-1(9–37), respectively (79). Carbohydrate, fat, and protein meals all stimulate GLP-1 secretion in human subjects with glucose being the strongest stimulant (15, 20). Unlike carbohydrates and fat that are also strong stimulants of GIP secretion, protein meals have no effect (15). The plasma concentrations of both hormones increase rapidly within 5–15 min after food ingestion (15, 20) but their actions are short lasting due to rapid proteolytic degradation by DPP-4 and other proteases. The plasma half-lives for intact GLP-1 and GIP are 1–2 and 7 min, respectively (86–88). DPP-4 is the main enzyme for incretin clearance as targeted disruption of the DDP-4 gene in mice led to improved stability of endogenous GLP-1 (89). The tissue distribution of DPP-4 plays an important role in GLP-1 degradation. There is a high level of DPP-4 in the endothelium of the capillaries surrounding L cells, and over 50% of newly secreted intact GLP-1 loses the N-terminal dipeptide and as a result is inactivated before entering the systemic circulation (90). The rapid rise of GLP-1 and GIP in the circulation ensures elevated GSIS in response to a meal, which is essential for the normalization of postprandial glucose. The disappearance of the incretins is in sync with the normalization of postprandial glucose. The first contributor of such a precise regulation is proteolytic degradation. In addition to DPP-4, the neutral endopeptidase 24.11 (NEP-24.11) is also involved in incretin degradation (91, 92). But DPP-4 is the main incretin degradation protease. Like DPP-4, NEP-24.11 is not selective against the incretins; it also processes other hormonal peptides (91). Its catalytic rates on vasoactive intestinal peptides (VIP) and glucagon are much faster than those on the incretins (91). The other factor that contributes to the rapid decline of plasma GLP-1 and GIP levels is a negative feedback mechanism, under which both hormones limit their own secretion by stimulating the somatostatin-mediated paracrine regulation. Somato- statin-positive D cells are located throughout the small intestine in close proximity to both L and K cells (4). In vitro, somatostatin inhibits GLP-1 secretion by L cells (93). In perfused porcine intestine, blocking somatostatin activity with a neutralizing monoclonal antibody increased GLP-1 secretion by 8–9-fold (94). Further, Regulation of Incretin Secretion 13
  • 24. intravascularinfusionofsomatostain-28stronglyinhibitedGLP-1releaseinpigs(94). This finding is consistent with other somatostatin infusion studies in rats (95), sheep (96), and human subjects (97), where somatostatin inhibited GLP-1 or GIP secretion in vivo. These data suggest that GLP-1 secretion is tonically inhibited by the local release of somatostatin-28 from epithelial paracrine D cells. Compared to somatostatin-28, the enteric neuron-derived somatostatin-14 is much weaker in influencing GLP-1 secretion (94). The suppressive effect of somatostatin on GLP- 1 secretion is mediated by somatostatin receptor subtype 5 expressed in L cells (98). Since both GLP-1 and GIP stimulate somatostatin release (99, 100), somatostatin is believed to be a key player in a negative feedback loop that controls incretin release in the gut. The existence of the negative feedback loop on GLP-1 secretion is supported by further evidence in dogs and humans. Conscious dogs were orally given a DPP-4 inhibitor, which increased meal-induced active GLP-1 levels (101). However, the total GLP-1 levels in these dogs were reduced (101), presumably due to the inhibitory effect of elevated active GLP-1 on endogenous GLP-1 secretion. A similar result was observed in healthy human volunteers who received an oral dose of a different DPP-4 inhibitor (102). These data support the notion that GLP-1 can inhibit its own secretion in vivo as part of a negative feedback loop. In addition to direct stimulation by nutrients, GLP-1 secretion is also indirectly regulated by GIP released in the proximal intestine in rodents. After a meal, nutrients are expected to reach the distal L cells and stimulate GLP-1 release via direct contact. However, this does not explain the biphasic pattern of GLP-1 secretion after a meal, including a 15–30 min rapid rise after oral ingestion followed by a second minor peak at 90–120 min (15, 103). Since all the initial findings indicate that L cells are located in the distal intestine (ileum, colon, rectum), the rapid early rise of GLP-1 after food ingestion within 5–15 min is faster than the time required for unabsorbed nutrients to reach the L cells in the distal intestine. A proposed mechanism is the existence of a neuroendocrine loop that regulates GLP-1 secretion distally once the ingested nutrients reach the proximal intestine (duodenum). This regulatory mechanism is referred to as proximal–distal neuroendocrine loop or duodeno-ileal endocrine loop. Since high GIP levels can stimulate GLP-1 secretion (104, 105), it is possible that nutrient entry into the duodenum stimulates GIP release, which in turn stimulates GLP-1 secretion in the distal intestine even before the nutrients arrive. This notion is supported by several studies. First, intraarterial infusion of GIP into perfused rat colon strongly stimulated GLP-1 secretion (106). In another study, the flow of nutrients to the distal intestine was restrained in rats to prevent direct interaction of the luminal content with the distal L cells (107). Next, when fat or glucose was placed in the duodenal lumen of these animals, GLP-1 release was induced at a level comparable to that by directly placing nutrients into the ileum (107). In the meantime, a rapid rise in GIP was also observed (107). This finding suggests that GIP released from the proximal intestine may mediate the early secretion of GLP-1 in the distal intestine. The vagus nerve appears to play an important role in this regulation because bilateral subdiaphragmatic vagotomy abolished the GLP-1 secretion by fat placed into the duodenum (108). Further, GLP-1 secretion stimulated by physiological concentra- tions of infused GIP was completely abrogated with selective hepatic branch 14 Chapter 1 Gut as an Endocrine Organ
  • 25. vagotomy (108). These data suggest that the vagus nerve mediates the GIP-stimulated GLP-1 response in the distal intestine in rats. The GIP-mediated regulation of GLP-1 release has not beenvalidated in humans. Although there is an early rise of GLP-1 after oral ingestion in humans (15, 103), GIP does not play a role in mediating this response. Intraduodenal infusion of a small amount of glucose produced a rapid and short-lasting GLP-1 response but the GIP level did not change (20), suggesting that the GLP-1 response to the duodenal glucose infusion is not mediated by GIP. In a separate study, synthetic GIP was infused into both type 2 diabetic patients and normal subjects. The exogenously administered GIP increased insulin secretion but had no effect on circulating GLP-1 level in normal subjects (109, 110). A further study was carried out in patients with upper and lower gut resections (jejunal or ileal small intestinal resections and colectomy), and it was found that a clear and early (peak at 15–30 min) GLP-1 response after food ingestion was observed in the patients with gut resection as well as controls (111). These studies demonstrate that the early GLP-1 response to food ingestion is not mediated by GIP in humans. One proposed explanation is that the early rise in GLP-1 is also a direct effect of nutrients on L cells because in contrast to previous reports that L cells are primarily located in the distally lower jejunum, ileum, colon, and rectum (1), GLP-1 positive cells were also found in human duodenum in recent studies (75, 76). These data suggest that the early rise in GLP-1 in humans after a meal could be ascribed to GLP-1 secretion from L cells in the upper gut. DISORDERS IN INCRETIN RESPONSE IN TYPE 2 DIABETES Meal- or oral glucose-induced GLP-1 response is decreased in type 2 diabetic patients as well as subjects with impaired glucose tolerance (IGT) (112, 113), but the GLP-1 response in IGT subjects trends higher than that in type 2 diabetics (112). This impairment could at least in part contributeto the disease pathogenesis. If this is one of the major causes of the defective glucose homeostasis in type 2 diabetes, adminis- tration of exogenous GLP-1 is expected to help normalize glucose control. It is encouraging that despite reduced GLP-1 secretion, type 2 diabetics still respond to GLP-1 infusion with augmented insulin release and improved glucose tolerance (109, 114). However, there are individual variations in response to exogenous GLP-1 administration among type 2 diabetics; glucose elimination is faster and lower glycemia was achieved in patients with lower baseline fasting plasma glucose (114). This finding suggests that GLP-1 treatment becomes less effective as the disease progresses. In contrast to GLP-1, GIP has diminished incretin effect in type 2 diabetic patients, suggesting that the GIP response is largely lost in the disease state (109). The underlying mechanism behind this observation is not clear. Based on these data, only GLP-1 is expected to have potential therapeutic value in treating type 2 diabetes. OTHER GI PEPTIDE HORMONES OR NEUROTRANSMITTERS In addition to the incretins, other peptide hormones and neurotransmitters are also involved in the regulation of gastric emptying, food intake, and energy metabolism. Other GI Peptide Hormones or Neurotransmitters 15
  • 26. There are many such peptides and some are yet to be assigned exact functional roles. They are secreted by different types of enteroendocrine cells distributed in different segments across the GI luminal surface. These cells sense luminal contents through direct interaction and secrete peptide hormones with regulatory effects. PYY and CCK are both involved in the regulation of food intake and gastric emptying. PYY-immunoreactive L cells are found in the distal small intestine, the colon, and the rectum (115). There are two forms of PYY, PYY(1–36) and PYY(3–36), in human blood (116), with PYY(3–36) derived from PYY(1–36) through DPP-4 proteolytic cleavage. Unlike GLP-1, both forms of PYYare bioactive. PYY(3–36) is the major form in human colonic mucosa. The plasma PYY level increases several fold after meal ingestion in humans. Compared to equivalent calories of protein and carbohydrate diets, fat is a more potent stimulus of PYY secretion (117). PYYcaninhibitgastricacidandpepsinsecretionanddelayintestinaltransittime(117), suggesting that PYY is a negative regulator of energy intake in response to food ingestion. PYY can interact with a family of Gi-coupled GPCRs, including Y1R, Y2R, Y4R, Y5R, and Y6R. Peripheral injection of PYY(3–36) was shown to inhibit food intake and reduce body weight in rats (118). PYY(3–36) also inhibited food intake in mice but not in Y2R-null mice (118), suggesting that the anorectic effects are mediated by Y2R. Consistent with the findings in animals, PYY(3–36) infusion significantly reduced appetite and food intake in human subjects of normal weight (118). Further, the circulating levels of PYY were significantly lower in obese subjects compared to lean controls, and like its effect in lean subjects, PYY infusion reduced food intake in obese individuals (119). These findings demonstrate that PYY is an anorectic agent and could be used to treat obesity. However, in contrast to peripheral administration, central administration of PYY increased food intake (120). More- over, the anorectic effects of peripheral PYY(3–36) administration could not be reproduced by some research groups (121), although others have been successful in replicating the original findings (122, 123). These discrepancies remain to be resolved with further studies. Similarly, CCK is another gut peptide involved in the regulation of food intake and related physiological activities. CCK is expressed as a 115-amino acid peptide in cells and undergoes posttranslational proteolytic processing to generate CCK- 58 (124, 125), the main circulating form. CCK is secreted by both I cells in the proximal intestine and L cells in the distal intestine (5). CCK is also found in the brain (5). Further proteolytic cleavage of CCK-58 generates smaller but still biologically active CCKs, including CCK-39, CCK-33, CCK-22, CCK-12, and CCK-8 (126). CCK is secreted and released into the blood circulation upon food ingestion and induces satiety. Two CCK receptors mediate the CCK function: CCK-1 receptor, primarily expressed in the GI tract, and CCK-2 receptor, mainly expressed in the brain. CCK-1 receptor is also expressed in the hindbrain and hypothalamus.Part of the CCK action in the brain is mediated by suppressing the expression of orexins A and B, two peptides produced in the lateral hypothalamic areas that stimulate food intake (127). The suppression of food intake by CCK was demonstrated in animal models as well as humans. Rats deficient in CCK-1 receptor had increased meal size and developed obesity (128), suggesting that the satiation signal is mediated by CCK-1 receptor. CCK administration also decreased food intake in humans by 16 Chapter 1 Gut as an Endocrine Organ
  • 27. shortening meals (129). The anorectic effects of CCK are weak because rats deficient in CCK-1 receptor developed only mild obesity (128), and CCK-1 receptor-null mice did not develop obesity (130). In addition, the anorectic effects were rapidly lost during repeated CCK administration (131), suggesting that behavioral tolerance may have developed under such a condition. These data question the suitability of CCK as an anti-obesity therapy. Other important GI peptides include oxyntomodulin, GLP-2, and ghrelin. Like GLP-1, oxyntomodulin and GLP-2 are proglucagon-derived peptides secreted from L cells (84, 85). Oxyntomodulin has been demonstrated to reduce food intake and body weight gain in rodents (132–134) and humans (135, 136). Interestingly, oxyntomo- dulin also increases energy expenditure in both animals and humans (133, 137). These effects are presumably mediated by GLP-1 receptor, although oxyntomodulin binds to it less avidly than GLP-1 (132). Oxyntomodulin also binds to glucagon receptor as its N-terminus contains the full glucagon sequence (138). However, it has a lower affinity than glucagon itself (138). The dual activation of both GLP-1 and glucagon receptors by oxyntomodulin might be a better explanation for the effects on food intake, body weight gain, and energy expenditure. Two independent studies dem- onstrated that dual activation of both GLP-1 and glucagon receptors with oxynto- modulin- or glucagon-derived peptides reduced food intake, body weight gain, body fat, hepatic steatosis, and blood glucose, and improved insulin sensitivity and lipid metabolism (138, 139). Although also derived from the proglucagon polypeptide and secreted from L cells, GLP-2 has no incretin effect. Rather, it is an intestinal growth factor. GLP-2 stimulates crypt cell proliferation and bowel growth in an ErbB- dependent manner (140, 141). GLP-2 also increases intestinal lipid absorption through activation of CD36 (142), thereby mediating a key function in response to food intake. Ghrelin is secreted from the stomach (6, 143) and is the endogenous ligand of the growth hormone (GH) secretagogues receptor (143). There are acylated and unacylated forms of ghrelin and the acylation is essential for the activity (143). Unlike the incretins or PYY, it increases food intake and is involved in meal initiation marked by a pre-meal surge (144). Ghrelin is likely involved in the long-term regulation of body weight (145). Interestingly, ghrelin improved cardiac functions in rats with heart failure (146), suggesting that there may be a role of ghrelin in regulating cardiovascular function. THE PHYSIOLOGICAL IMPORTANCE OF THE GUT: LESSONS LEARNED FROM GASTRIC BYPASS The metabolic role of the gut is further implicated in the fascinating findings from bariatric surgery, which produces dramatic and durable weight loss (147). Among many different types of bariatric surgical operations employed to treat severe obesity (147), the most commonly performed are laparoscopic adjustable gastric banding (LAGB), gastric bypass, and biliopancreatic bypass (147). Gastric bypass (or Roux-en-Y gastric bypass, RYGB) involves surgical reduction of the size of stomach and bypassing a portion of the proximal small intestine (Figure 1.3). The portion The Physiological Importance of the Gut: Lessons Learned from Gastric Bypass 17
  • 28. bypassed is connected to the distal small intestine to allow the passage of pancreatic fluids and bile into the gut (Figure 1.3). This procedure causes dramatic weight loss and has been the most effective treatment of severe obesity. In a series of 608 patients with 95% follow-up for at least 16 years, the mean weight loss was 106 lb (148). Surprisingly, more than 80% of the patients with type 2 diabetes developed complete remission of the disease after the surgery (148, 149). Weight loss does not fully explain the remission of type 2 diabetes after gastric bypass because within days after surgery the hyperglycemia and hyperinsulinemia were totally normalized (148). Although the mechanisms behind the antidiabetic effect are not entirely clear, increased insulin secretion and improved b-cell function are likely involved. Late- onset hyperinsulinemic hypoglycemia has been observed in patients after the surgery (150–152), and some may even require partial or total pancreatectomy to prevent recurrent hypoglycemia (150, 152). This phenomenon underscores the robust improvement of pancreatic function achieved by RYGB. GLP-1 and PYYare two important gut hormones that are believed to mediate the more robust beneficial effects of RYGB compared to LAGB, a procedure that restricts food intake by banding the stomach but does not involve the bypass of the proximal intestine. The metabolic effects of LAGB are therefore results of reduced food intake and weight loss. The average reduction in body weight after LAGB is 28% compared to 40% after RYGB, and the remission of type 2 diabetes occurs in 48% relative to 84% in RYGB (153, 154). One of the key differences between these two different operations is the greater GLP-1 and PYY response post meal after RYGB sur- gery (155), suggesting that these peptide hormones may play an important role in promoting weight loss and improved insulin sensitivity. As mentioned above, RYGB results in improved insulin sensitivity before weight loss in the short term. This effect RYGB Bypass Food Food Bypassed section transports bile and pancreatic fluid into the gut The upper portion is separated from the main stomach Duodenum Figure 1.3 Illustration of Roux-en-Y gastric bypass. 18 Chapter 1 Gut as an Endocrine Organ
  • 29. seems to persist even in the long run in a weight loss independent manner, although weight loss itself can lead to improved insulin sensitivity. When compared with a weight-matched group, the patients who underwent RYGB had lower fasting insulin and better insulin sensitivity (156), suggesting that in addition to weight loss something else leads to further improved insulin sensitivity in RYGB patients. In addition to suppressing appetite and weight loss after RYGB (157), the increased postprandial GLP-1 response could further improve insulin sensitivity by increasing b-cell mass and improving b-cell function. In fact, there is sustained elevation of GLP-1 secretion post meal in RYGB patients compared to normal controls (158). This may be counterintuitive because L cells are also found in human duodenum (75) and nutrient bypass of the proximal intestine is expected to cause reduction in GLP-1 release. It could be that this is a small loss relative to the robust increase in GLP-1 secretion by the distal intestine so that the total GLP-1 secretion is still elevated after RYGB. Two hypotheses have been proposed to explain the weight loss independent effect in RYGB based on the roles of the foregut and the hindgut. The hindgut hypothesis proposes that the beneficial effects result from the expedited delivery of nutrients to the distal small intestine and enhancement of physiologic signals that improve glucose homeostasis (159); the foregut hypothesis holds that the weight loss independent effect depends on the exclusion of the duodenum and proximal jejunum from nutrient passage, therefore preventing the secretion of a physiologic signal that promotes insulin resistance (159). Using nonobese diabetic Goto- Kakizaki (GK) rats, Rubino et al. demonstrated that duodenal–jejunal bypass (DJB), a stomach-preserving RYGB, improved oral glucose tolerance compared to a pair-fed sham-operated group (159). However, restoration of duodenal nutrient passage in the DJB rats reestablished impaired glucose tolerance (159), suggesting that the weight loss independent metabolic benefits in the DJB rats were likely to be driven by the nutrient bypass of the foregut. Why does the duodenal nutrient passage have a negative effect? These researchers proposed that a physiologic signal induced by duodenal nutrient passage might play a role. This negative signal could be an anti-incretin factor, which might be secreted from the proximal intestine in response to nutrient passage and stimulate insulin resistance (159). The anti-incretin factor may interfere with the incretin secretion and/or actions and ultimately inhibit insulin action (159). One of the possibilities is that the anti-incretin inhibits GLP-1 secretion and after nutrient bypass of the proximal intestine the suppression is relieved leading to elevated GLP-1 secretion. Although this hypothesis is consistent with the improved b-cell function in RYGB patients, it remains to be validated by identification of a factor with anti-incretin effect. While the anti-incretin concept helps explain the weight loss independent effects in RYGB patients, Rubino’s data do not exclude the involvement of the hindgut in the improvement of metabolic effects. In fact, a study in mouse models indicates that there is increased gluco- neogenesis in the distal intestine post DJB but not gastric banding (160), and the increased local glucose concentration is detected by a GLUT2-dependent hepato- portal sensor, which leads to reduced food intake and body weight and improved insulin sensitivity (160). Thus, it seems that different sections of the small intestine The Physiological Importance of the Gut: Lessons Learned from Gastric Bypass 19
  • 30. play important roles via distinct mechanisms to achieve beneficial metabolic effects in RYGB. SUMMARY In addition to its role in food intake and nutrient absorption, gut is also an endocrine organ for secreted GI peptides. The release of these peptides in response to food intake is mediated by the direct contact of macronutrients with enteroendocrine cells on the luminal side distributed throughout the GI tract. These GI peptides regulate a variety of physiological actions in response to food intake, including the feedback response to suppress food intake and the feedforward response for nutrient assim- ilation. The incretin GLP-1 plays important roles in both regulatory pathways. Different sets of GI peptides are stimulated in response to specific types of macronutrients. There are several potential nutrient-sensing mechanisms mediated by taste receptors, KATP channels, glucose transporters, and GPCRs. Further studies are required to clarify the relative contributions of these pathways. The robust metabolic benefits associated with RYGB suggest that changes in the secretion profiles of GI peptides may be beneficial, although the exact mechanism is still elusive. Further studies in gut biology will likely shed new light on the metabolic functions of GI peptides. REFERENCES 1. EISSELE, R., R. GOKE, S. WILLEMER, H.P. HARTHUS, H. VERMEER, R. ARNOLD, and B. GOKE. 1992. Glucagon-like peptide-1 cells in the gastrointestinal tract and pancreas of rat, pig and man. Eur J Clin Invest 22:283–291. 2. ONAGA, T., R. ZABIELSKI, and S. KATO. 2002. Multiple regulation of peptide YY secretion in the digestive tract. Peptides 23:279–290. 3. FEHMANN, H.C., R. GOKE, and B. GOKE. 1995. Cell and molecular biology of the incretin hormones glucagon-like peptide-I and glucose-dependent insulin releasing polypeptide. Endocr Rev 16:390–410. 4. DAMHOLT, A.B., H. KOFOD, and A.M. BUCHAN. 1999. Immunocytochemical evidence for a paracrine interaction between GIP and GLP-1-producing cells in canine small intestine. Cell Tissue Res 298:287–293. 5. REHFELD, J.F. 1978. Immunochemical studies on cholecystokinin. II. Distribution and molecular heterogeneity in the central nervous system and small intestine of man and hog. J Biol Chem 253:4022–4030. 6. ARIYASU, H., K. TAKAYA, T. TAGAMI, Y. OGAWA, K. HOSODA, T. AKAMIZU, M. SUDA, T. KOH, K. NATSUI, S. TOYOOKA, G. SHIRAKAMI, T. USUI, A. SHIMATSU, K. DOI, H. HOSODA, M. KOJIMA, K. KANGAWA, and K. NAKAO. 2001. Stomach is a major source of circulating ghrelin, and feeding state determines plasma ghrelin-like immunoreactivity levels in humans. J Clin Endocrinol Metab 86:4753–4758. 7. AYALA, J.E., D.P. BRACY, F.D. JAMES, B.M. JULIEN, D.H. WASSERMAN, and D.J. DRUCKER. 2009. The glucagon-like peptide-1 receptor regulates endogenous glucose production and muscle glucose uptake independent of its incretin action. Endocrinology 150:1155–1164. 8. STEPHENS, D.B. 1980. The effects of alimentary infusions of glucose, amino acids, or neutral fat on meal size in hungry pigs. J Physiol 299:453–463. 9. HOUPT, T.R., B.A. BALDWIN, and K.A. HOUPT. 1983. Effects of duodenal osmotic loads on spontaneous meals in pigs. Physiol Behav 30:787–795. 20 Chapter 1 Gut as an Endocrine Organ
  • 31. 10. STEPHENS, D.B., and B.A. BALDWIN. 1974. The lack of effect of intrajugular or intraportal injections of glucose or amino acids on food intake in pigs. Physiol Behav 12:923–929. 11. GREGORY, P.C., M. MCFADYEN, and D.V. RAYNER. 1987. The influence of gastrointestinal infusions of glucose on regulation of food intake in pigs. Q J Exp Physiol 72:525–535. 12. WELCH, I., K. SAUNDERS, and N.W. READ. 1985. Effect of ileal and intravenous infusions of fat emulsions on feeding and satiety in human volunteers. Gastroenterology 89:1293–1297. 13. O’DONOVAN, D., C. FEINLE-BISSET, J. WISHART, and M. HOROWITZ. 2003. Lipase inhibition attenuates the acute inhibitory effects of oral fat on food intake in healthy subjects. Br J Nutr 90:849–852. 14. O’DONOVAN, D., M. HOROWITZ, A. RUSSO, C. FEINLE-BISSET, N. MUROLO, D. GENTILCORE, J.M. WISHART, H.A. MORRIS, and K.L. JONES. 2004. Effects of lipase inhibition on gastric emptying of, and on the glycaemic, insulin and cardiovascular responses to, a high-fat/carbohydrate meal in type 2 diabetes. Diabetologia 47:2208–2214. 15. ELLIOTT, R.M., L.M. MORGAN, J.A. TREDGER, S. DEACON, J. WRIGHT, and V. MARKS. 1993. Glucagon- like peptide-1(7–36)amideand glucose-dependent insulinotropic polypeptide secretion in response to nutrient ingestion in man: acute post-prandial and 24-h secretion patterns. J Endocrinol 138:159–166. 16. MCLAUGHLIN, J.T., R.B. LOMAX, L. HALL, G.J. DOCKRAY, D.G. THOMPSON, and G. WARHURST. 1998.Fatty acids stimulate cholecystokinin secretion via an acyl chain length-specific, Ca2 þ -dependent mech- anism in the enteroendocrine cell line STC-1. J Physiol 513 (Pt 1): 11–18. 17. MCLAUGHLIN, J., M. GRAZIA LUCA, M.N. JONES, M. D’AMATO, G.J. DOCKRAY, and D.G. THOMPSON. 1999. Fatty acid chain length determines cholecystokinin secretion and effect on human gastric motility. Gastroenterology 116:46–53. 18. THOMAS, F.B., E.L. MAZZAFERRI, S.E. CROCKETT, H.S. MEKHJIAN, H.D. GRUEMER, and S. CATALAND. 1976. Stimulation of secretion of gastric inhibitory polypeptide and insulin by intraduodenal amino acid perfusion. Gastroenterology 70:523–527. 19. THOMAS, F.B., D. SINAR, E.L. MAZZAFERRI, S. CATALAND, H.S. MEKHJIAN, J.H. CALDWELL, and J.J. FROMKES. 1978. Selective release of gastric inhibitory polypeptide by intraduodenal amino acid perfusion in man. Gastroenterology 74:1261–1265. 20. HERRMANN, C., R. GOKE, G. RICHTER, H.C. FEHMANN, R. ARNOLD, and B. GOKE. 1995. Glucagon-like peptide-1 and glucose-dependent insulin-releasing polypeptide plasma levels in response to nutrients. Digestion 56:117–126. 21. DUMOULIN, V., F. MORO, A. BARCELO, T. DAKKA, and J.C. CUBER. 1998. Peptide YY, glucagon-like peptide-1, and neurotensin responses to luminal factors in the isolated vascularly perfused rat ileum. Endocrinology 139:3780–3786. 22. NEMOZ-GAILLARD, E., C. BERNARD, J. ABELLO, M. CORDIER-BUSSAT, J.A. CHAYVIALLE, and J.C. CUBER. 1998. Regulation of cholecystokinin secretion by peptones and peptidomimetic antibiotics in STC-1 cells. Endocrinology 139:932–938. 23. CHOI, S., M. LEE, A.L. SHIU, S.J. YO, G. HALLDEN, and G.W. APONTE. 2007. GPR93 activation by protein hydrolysate induces CCK transcription and secretion in STC-1 cells. Am J Physiol Gastro- intest Liver Physiol 292:G1366–G1375. 24. CORDIER-BUSSAT, M., C. BERNARD, F. LEVENEZ, N. KLAGES, B. LASER-RITZ, J. PHILIPPE, J.A. CHAYVIALLE, and J.C. CUBER. 1998. Peptones stimulate both the secretion of the incretin hormone glucagon-like peptide 1 and the transcription of the proglucagon gene. Diabetes 47:1038–1045. 25. GEVREY, J.C., M. MALAPEL, J. PHILIPPE, G. MITHIEUX, J.A. CHAYVIALLE, J. ABELLO, and M. CORDIER- BUSSAT. 2004. Protein hydrolysates stimulate proglucagon gene transcription in intestinal endocrine cells via two elements related to cyclic AMP response element. Diabetologia 47:926–936. 26. CONIGRAVE, A.D., A.H. FRANKS, E.M. BROWN, and S.J. QUINN. 2002. L-Amino acid sensing by the calcium-sensing receptor: a general mechanism for coupling protein and calcium metabolism? Eur J Clin Nutr 56:1072–1080. 27. RAY, J.M., P.E. SQUIRES, S.B. CURTIS, M.R. MELOCHE, and A.M. BUCHAN. 1997. Expression of the calcium-sensing receptor on human antral gastrin cells in culture. J Clin Invest 99:2328–2333. 28. STRUNZ, U.T., J.H. WALSH, and M.I. GROSSMAN. 1978. Stimulation of gastrin release in dogs by individual amino acids. Proc Soc Exp Biol Med 157:440–441. References 21
  • 32. 29. KONTUREK, S.J., T. RADECKI, P. THOR, and A. DEMBINSKI. 1973. Release of cholecystokinin by amino acids. Proc Soc Exp Biol Med 143:305–309. 30. ISENBERG, J.I., and V. MAXWELL. 1978. Intravenous infusion of amino acids stimulates gastric acid secretion in man. N Engl J Med 298:27–29. 31. KOOPMANS, H.S. 1983. A stomach hormone that inhibits food intake. J Auton Nerv Syst 9:157–171. 32. MCHUGH, P.R., T.H. MORAN, and G.N. BARTON. 1975. Satiety: a graded behavioural phenomenon regulating caloric intake. Science 190:167–169. 33. SHAFER, R.B., A.S. LEVINE, J.M. MARLETTE, and J.E. MORLEY. 1985. Do calories, osmolality, or calcium determine gastric emptying? Am J Physiol Regul Integr Comp Physiol 248:R479–R483. 34. MARGOLSKEE, R.F. 2002. Molecular mechanisms of bitter and sweet taste transduction. J Biol Chem 277:1–4. 35. LI, X., L. STASZEWSKI, H. XU, K. DURICK, M. ZOLLER, and E. ADLER. 2002. Human receptors for sweet and umami taste. Proc Natl Acad Sci USA 99:4692–4696. 36. CUMMINGS, D.E., and J. OVERDUIN. 2007. Gastrointestinal regulation of food intake. J Clin Invest 117:13–23. 37. MACE, O.J., J. AFFLECK, N. PATEL, and G.L. KELLETT. 2007. Sweet taste receptors in rat small intestine stimulate glucose absorption through apical GLUT2. J Physiol 582:379–392; erratum in J Physiol 583:411. 38. HOFER, D., and D. DRENCKHAHN. 1998. Identification of the taste cell G-protein, alpha-gustducin, in brush cells of the rat pancreatic duct system. Histochem Cell Biol 110:303–309. 39. HOFER, D., B. PUSCHEL, and D. DRENCKHAHN. 1996. Taste receptor-like cells in the rat gut identified by expression of alpha-gustducin. Proc Natl Acad Sci USA 93:6631–6634. 40. DYER, J., K.S. SALMON, L. ZIBRIK, and S.P. SHIRAZI-BEECHEY. 2005. Expression of sweet taste receptors of the T1R family in the intestinal tract and enteroendocrine cells. Biochem Soc Trans 33:302–305. 41. JANG, H.J., Z. KOKRASHVILI, M.J. THEODORAKIS, O.D. CARLSON, B.J. KIM, J. ZHOU, H.H. KIM, X. XU, S.L. CHAN, M. JUHASZOVA, M. BERNIER, B. MOSINGER, R.F. MARGOLSKEE, and J.M. EGAN. 2007. Gut- expressed gustducin and taste receptors regulate secretion of glucagon-like peptide-1. Proc Natl Acad Sci USA 104:15069–15074. 42. REIMANN, F., A.M. HABIB, G. TOLHURST, H.E. PARKER, G.J. ROGERS, and F.M. GRIBBLE. 2008. Glucose sensing in L cells: a primary cell study. Cell Metab 8:532–539. 43. MA, J., M. BELLON, J.M. WISHART, R. YOUNG, L.A. BLACKSHAW, K.L. JONES, M. HOROWITZ, and C.K. RAYNER. 2009. Effect of the artificial sweetener, sucralose, on gastric emptying and incretin hormone release in healthy subjects. Am J Physiol Gastrointest Liver Physiol 296:G735–G739. 44. GREGERSEN, S., P.B. JEPPESEN, J.J. HOLST, and K. HERMANSEN. 2004. Antihyperglycemic effects of stevioside in type 2 diabetic subjects. Metabolism 53:73–76. 45. FUJITA, Y., R.D. WIDEMAN, M. SPECK, A. ASADI, D.S. KING, T.D. WEBBER, M. HANEDA, and T.J. KIEFFER. 2009. Incretin release from gut is acutely enhanced by sugar but not by sweeteners in vivo. Am J Physiol Endocrinol Metab 296:E473–E479. 46. REIMANN, F., and F.M. GRIBBLE. 2002. Glucose-sensing in glucagon-like peptide-1-secreting cells. Diabetes 51:2757–2763. 47. MURPHY, R., A. TURA, P.M. CLARK, J.J. HOLST, A. MARI, and A.T. HATTERSLEY. 2009. Glucokinase, the pancreatic glucose sensor, is not the gut glucose sensor. Diabetologia 52:154–159. 48. GRIBBLE, F.M., L. WILLIAMS, A.K. SIMPSON, and F. REIMANN. 2003. A novel glucose-sensing mechanism contributing to glucagon-like peptide-1 secretion from the GLUTag cell line. Diabetes 52:1147–1154. 49. DIEZ-SAMPEDRO, A., M.P. LOSTAO, E.M. WRIGHT, and B.A. HIRAYAMA. 2000. Glycoside binding and translocation in Naþ -dependent glucose cotransporters: comparison of SGLT1 and SGLT3. J Membr Biol 176:111–117. 50. CONIGRAVE, A.D., and E.M. BROWN. 2006. Taste receptors in the gastrointestinal tract. II. L-Amino acid sensing by calcium-sensing receptors: implications for GI physiology. Am J Physiol Gastrointest Liver Physiol 291:G753–G761. 51. BUCHAN, A.M., P.E. SQUIRES, M. RING, and R.M. MELOCHE. 2001. Mechanism of action of the calcium- sensing receptor in human antral gastrin cells. Gastroenterology 120:1128–1139. 22 Chapter 1 Gut as an Endocrine Organ
  • 33. 52. CONIGRAVE, A.D., S.J.QUINN, and E.M. BROWN. 2000. L-Amino acid sensing by the extracellular Ca2 þ - sensing receptor. Proc Natl Acad Sci USA 97:4814–4819. 53. ADIBI, S.A., and D.W. MERCER. 1973. Protein digestion in human intestine as reflected in luminal, mucosal, and plasma amino acid concentrations after meals. J Clin Invest 52:1586–1594. 54. MANGEL, A.W., V. PRPIC, H. WONG, S. BASAVAPPA, L.J. HURST, L. SCOTT, R.L. GARMAN, J.S. HAYES, A.I. SHARARA, N.D. SNOW, et al. 1995. Phenylalanine-stimulated secretion of cholecystokinin is calcium dependent. Am J Physiol Gastrointest Liver Physiol 268:G90–G94. 55. BALLINGER, A.B., and M.L. CLARK. 1994. L-Phenylalanine releases cholecystokinin (CCK) and is associated with reduced food intake in humans: evidence for a physiological role of CCK in control of eating. Metabolism 43:735–738. 56. BRENNAN, S.C., and A.D. CONIGRAVE. 2009. Regulation of cellular signal transduction pathways by the extracellular calcium-sensing receptor. Curr Pharm Biotechnol 10:270–281. 57. ITOH, Y., Y. KAWAMATA, M. HARADA, M. KOBAYASHI, R. FUJII, S. FUKUSUMI, K. OGI, M. HOSOYA, Y. TANAKA, H. UEJIMA, H. TANAKA, M. MARUYAMA, R. SATOH, S. OKUBO, H. KIZAWA, H. KOMATSU, F. MATSUMURA, Y. NOGUCHI, T. SHINOHARA, S. HINUMA, Y. FUJISAWA, and M. FUJINO. 2003. Free fatty acids regulate insulin secretion from pancreatic beta cells through GPR40. Nature 422:173–176. 58. BRISCOE, C.P., M. TADAYYON, J.L. ANDREWS, W.G. BENSON, J.K. CHAMBERS, M.M. EILERT, C. ELLIS, N.A. ELSHOURBAGY, A.S. GOETZ, D.T. MINNICK, P.R.MURDOCK, H.R.SAULS,Jr., U.SHABON,L.D. SPINAGE,J.C. STRUM, P.G. SZEKERES, K.B. TAN, J.M. WAY, D.M. IGNAR, S. WILSON, and A.I. MUIR. 2003. The orphan G protein-coupled receptor GPR40 is activated by medium and long chain fatty acids. J Biol Chem 278:11303–11311. 59. EDFALK, S., P. STENEBERG, and H. EDLUND. 2008. Gpr40 is expressed in enteroendocrine cells and mediates free fatty acid stimulation of incretin secretion. Diabetes 57:2280–2287. 60. HIRASAWA, A., K. TSUMAYA, T. AWAJI, S. KATSUMA, T. ADACHI, M. YAMADA, Y. SUGIMOTO, S. MIYAZAKI, and G. TSUJIMOTO. 2005. Free fatty acids regulate gut incretin glucagon-like peptide-1 secretion through GPR120. Nat Med 11:90–94. 61. TANAKA, T., S. KATSUMA, T. ADACHI, T.A. KOSHIMIZU, A. HIRASAWA, and G. TSUJIMOTO. 2008. Free fatty acids induce cholecystokinin secretion through GPR120. Naunyn Schmiedebergs Arch Pharmacol 377:523–527. 62. OVERTON, H.A., A.J. BABBS, S.M. DOEL, M.C. FYFE, L.S. GARDNER, G. GRIFFIN, H.C. JACKSON, M.J. PROCTER, C.M. RASAMISON, M. TANG-CHRISTENSEN, P.S. WIDDOWSON, G.M. WILLIAMS, and C. REYNET. 2006. Deorphanization of a G protein-coupled receptor for oleoylethanolamide and its use in the discovery of small-molecule hypophagic agents. Cell Metab 3:167–175. 63. SOGA, T., T. OHISHI, T. MATSUI, T. SAITO, M. MATSUMOTO, J. TAKASAKI, S. MATSUMOTO, M. KAMOHARA, H. HIYAMA, S. YOSHIDA, K. MOMOSE, Y. UEDA, H. MATSUSHIME, M. KOBORI, and K. FURUICHI. 2005. Lysophosphatidylcholine enhances glucose-dependent insulin secretion via an orphan G-protein- coupled receptor. Biochem Biophys Res Commun 326:744–751. 64. CHU, Z.L., C. CARROLL, J. ALFONSO, V. GUTIERREZ, H. HE, A. LUCMAN, M. PEDRAZA, H. MONDALA, H. GAO, D. BAGNOL, R. CHEN, R.M. JONES, D.P. BEHAN, and J. LEONARD. 2008. A role for intestinal endocrine cell-expressed G protein-coupled receptor 119 in glycemic control by enhancing glucagon- like peptide-1 and glucose-dependent insulinotropic peptide release. Endocrinology 149:2038–2047. 65. SHAPIRO, H., S. SHACHAR, I. SEKLER, M. HERSHFINKEL, and M.D. WALKER. 2005. Role of GPR40 in fatty acid action on the beta cell line INS-1E. Biochem Biophys Res Commun 335:97–104. 66. CHU, Z.L., R.M. JONES, H. HE, C. CARROLL, V. GUTIERREZ, A. LUCMAN, M. MOLONEY, H. GAO, H. MONDALA, D. BAGNOL, D. UNETT, Y. LIANG, K. DEMAREST, G. SEMPLE, D.P. BEHAN, and J. LEONARD. 2007. A role for beta-cell-expressed G protein-coupled receptor 119 in glycemic control by enhancing glucose-dependent insulin release. Endocrinology 148:2601–2609. 67. FURNESS, J.B., W.A. KUNZE, and N. CLERC. 1999. Nutrient tasting and signaling mechanisms in the gut. II. The intestine as a sensory organ: neural, endocrine, and immune responses. Am J Physiol Gastrointest Liver Physiol 277:G922–G928. 68. BAYLISS, W.M., and E.H. STARLING. 1902. The mechanism of pancreatic secretion. J Physiol 28:325–353. References 23
  • 34. 69. MOORE, B., E.S. EDIE, and J.H. ABRAM. 1906. On the treatment of diabetes mellitus by acid extracts of duodenal mucous membrane. Biochem J 1:228–238. 70. La BARRE, J. 1932. Sur les possibilites d’un traitement du diabete par l’incretine. Bull Acad R Med Belg 12:620–634. 71. MCINTYRE, N., C.D. HOLDSWORTH, and D.S. TURNER. 1964. New interpretation of oral glucose tolerance. Lancet 2:20–21. 72. ELRICK, H., L. STIMMLER, C.J. HLAD, Jr., and Y. ARAI. 1964. Plasma insulin response to oral and intravenous glucose administration. J Clin Endocrinol Metab 24:1076–1082. 73. NAUCK, M.A., E. HOMBERGER, E.G. SIEGEL, R.C. ALLEN, R.P. EATON, R. EBERT, and W. CREUTZFELDT. 1986. Incretineffectsofincreasingglucose loads in man calculatedfrom venous insulinand C-peptide responses. J Clin Endocrinol Metab 63:492–498. 74. HAMPTON, S.M., L.M. MORGAN, J.A. TREDGER, R. CRAMB, and V. MARKS. 1986. Insulin and C-peptide levels after oral and intravenous glucose. Contribution of enteroinsular axis to insulin secretion. Diabetes 35:612–616. 75. THEODORAKIS, M.J., O. CARLSON, S. MICHOPOULOS, M.E. DOYLE, M. JUHASZOVA, K. PETRAKI, and J.M. EGAN. 2006. Human duodenal enteroendocrine cells: source of both incretin peptides, GLP-1 and GIP. Am J Physiol Endocrinol Metab 290:E550–E559. 76. MORTENSEN,K., L.L.CHRISTENSEN,J.J. HOLST,and C. ORSKOV.2003.GLP-1andGIPare colocalized in a subset of endocrine cells in the small intestine. Regul Pept 114:189–196. 77. DUPRE, J., S.A. ROSS, D. WATSON, and J.C. BROWN. 1973. Stimulation of insulin secretion by gastric inhibitory polypeptide in man. J Clin Endocrinol Metab 37:826–828. 78. MENTLEIN, R. 1999. Dipeptidyl-peptidase IV (CD26): role in the inactivation of regulatory peptides. Regul Pept 85:9–24. 79. MENTLEIN, R., B. GALLWITZ, and W.E. SCHMIDT. 1993. Dipeptidyl-peptidase IV hydrolyses gastric inhibitory polypeptide, glucagon-like peptide-1(7–36)amide, peptide histidine methionine and is responsible for their degradation in human serum. Eur J Biochem 214:829–835. 80. PROOST, P., I. De MEESTER, D. SCHOLS, S. STRUYF, A.M. LAMBEIR, A. WUYTS, G. OPDENAKKER, E. De CLERCQ, S. SCHARPE, and J. VAN DAMME. 1998. Amino-terminal truncation of chemokines by CD26/ dipeptidyl-peptidase IV. Conversion of RANTES into a potent inhibitor of monocyte chemotaxis and HIV-1-infection. J Biol Chem 273:7222–7227. 81. PROOST, P., S. STRUYF, D. SCHOLS, G. OPDENAKKER, S. SOZZANI, P. ALLAVENA, A. MANTOVANI, K. AUGUSTYNS, G. BAL, A. HAEMERS, A.M. LAMBEIR, S. SCHARPE, J. Van DAMME, and I. DE MEESTER. 1999. Truncation of macrophage-derived chemokine by CD26/dipeptidyl-peptidase IV beyond its predicted cleavage site affects chemotactic activity and CC chemokine receptor 4 interaction. J Biol Chem 274:3988–3993. 82. SHIODA, T., H. KATO, Y. OHNISHI, K. TASHIRO, M. IKEGAWA, E.E. NAKAYAMA, H. HU, A. KATO, Y. SAKAI, H. LIU, T. HONJO, A. NOMOTO, A. IWAMOTO, C. MORIMOTO, and Y. NAGAI. 1998. Anti-HIV-1 and chemotactic activities of human stromal cell-derived factor 1alpha (SDF-1alpha) and SDF-1beta are abolished by CD26/dipeptidyl peptidase IV-mediated cleavage. Proc Natl Acad Sci USA 95:6331–6336. 83. MENTLEIN, R., P. DAHMS, D. GRANDT, and R. KRUGER. 1993. Proteolytic processing of neuropeptide Y and peptide YY by dipeptidyl peptidase IV. Regul Pept 49:133–144. 84. DHANVANTARI, S., N.G. SEIDAH, and P.L. BRUBAKER. 1996. Role of prohormone convertases in the tissue-specific processing of proglucagon. Mol Endocrinol 10:342–355. 85. ROUILLE, Y., S. MARTIN, and D.F. STEINER. 1995. Differential processing of proglucagon by the subtilisin-like prohormone convertases PC2 and PC3 to generate either glucagon or glucagon-like peptide. J Biol Chem 270:26488–26496. 86. MEIER, J.J., M.A. NAUCK, D. KRANZ, J.J. HOLST, C.F. DEACON, D. GAECKLER, W.E. SCHMIDT, and B. GALLWITZ. 2004. Secretion, degradation, and elimination of glucagon-like peptide 1 and gastric inhibitory polypeptide in patients with chronic renal insufficiency and healthy control subjects. Diabetes 53:654–662. 87. VILSBOLL, T., H. AGERSO, T. KRARUP, and J.J. HOLST. 2003. Similar elimination rates of glucagon-like peptide-1 in obese type 2 diabetic patients and healthy subjects. J Clin Endocrinol Metab 88:220–224. 24 Chapter 1 Gut as an Endocrine Organ
  • 35. 88. DEACON, C.F., M.A. NAUCK, J. MEIER, K. HUCKING, and J.J. HOLST. 2000. Degradation of endogenous and exogenous gastric inhibitory polypeptide in healthy and in type 2 diabetic subjects as revealed using a new assay for the intact peptide. J Clin Endocrinol Metab 85:3575–3581. 89. MARGUET, D., L. BAGGIO, T. KOBAYASHI, A.M. BERNARD, M. PIERRES, P.F. NIELSEN, U. RIBEL, T. WATANABE, D.J. DRUCKER, and N. WAGTMANN. 2000. Enhanced insulin secretion and improved glucose tolerance in mice lacking CD26. Proc Natl Acad Sci USA 97:6874–6879. 90. HANSEN, L., C.F. DEACON, C. ORSKOV, and J.J. HOLST. 1999. Glucagon-like peptide-1-(7–36)amide is transformed to glucagon-like peptide-1-(9–36)amide by dipeptidyl peptidase IV in the capillaries supplying the L cells of the porcine intestine. Endocrinology 140:5356–5363. 91. HUPE-SODMANN, K., G.P. MCGREGOR, R. BRIDENBAUGH, R. GOKE, B. GOKE, H. THOLE, B. ZIMMERMANN, and K.VOIGT. 1995.Characterisation ofthe processingby humanneutralendopeptidase24.11 ofGLP- 1(7–36) amide and comparison of the substrate specificity of the enzyme for other glucagon-like peptides. Regul Pept 58:149–156. 92. PLAMBOECK, A., J.J. HOLST, R.D. CARR, and C.F. DEACON. 2005. Neutral endopeptidase 24.11 and dipeptidyl peptidase IV are both mediators of the degradation of glucagon-like peptide 1 in the anaesthetised pig. Diabetologia 48:1882–1890. 93. BRUBAKER, P.L. 1991. Regulation of intestinal proglucagon-derived peptide secretion by intestinal regulatory peptides. Endocrinology 128:3175–3182. 94. HANSEN, L., B. HARTMANN, T. BISGAARD, H. MINEO, P.N. JORGENSEN, and J.J. HOLST. 2000. Somatostatin restrains the secretion of glucagon-like peptide-1 and -2 from isolated perfused porcine ileum. Am J Physiol Endocrinol Metab 278:E1010–E1018. 95. HO, L.T., H.F. PU, W.J. SHEU, W.C. WANG, and P.S. WANG. 1987. Inhibition of somatostatin on glucose- induced release of gastric inhibitory polypeptide in rats. Chin J Physiol 30:45–53. 96. MARTIN, P.A., and A. FAULKNER. 1996. Effects of somatostatin-28 on circulating concentrations of insulin and gut hormones in sheep. J Endocrinol 151:107–112. 97. SALERA, M., L. PIRONI, P. GIACOMONI, S. VENTURI, M. CAPELLI, M. MIGLIOLI, and L. BARBARA. 1982. Effectofsomatostatinon fastingandglucose-stimulated gastricinhibitorypolypeptide release in man. Digestion 24:126–132. 98. CHISHOLM, C., and G.R. GREENBERG. 2002. Somatostatin-28 regulates GLP-1 secretion via somato- statin receptor subtype 5 in rat intestinal cultures. Am J Physiol Endocrinol Metab 283:E311–E317. 99. BRUBAKER, P.L., S. EFENDIC, and G.R. GREENBERG. 1997. Truncated and full-length glucagon-like peptide-1 (GLP-1) differentially stimulate intestinal somatostatin release. Endocrine 6:91–95. 100. JIA, X., J.C. BROWN, Y.N. KWOK, R.A. PEDERSON, and C.H. MCINTOSH. 1994. Gastric inhibitory polypeptide and glucagon-like peptide-1(7–36) amide exert similar effects on somatostatin secretion butoppositeeffectsongastrinsecretionfromtheratstomach.CanJPhysiolPharmacol72:1215–1219. 101. DEACON, C.F., S. WAMBERG, P. BIE, T.E. HUGHES, and J.J. HOLST. 2002. Preservation of active incretin hormones by inhibitionof dipeptidyl peptidase IV suppressesmeal-inducedincretin secretion in dogs. J Endocrinol 172:355–362. 102. EL-OUAGHLIDI, A., E. REHRING, J.J. HOLST, A. SCHWEIZER, J. FOLEY, D. HOLMES, and M.A. NAUCK. 2007. The dipeptidyl peptidase 4 inhibitor vildagliptin does not accentuate glibenclamide-induced hypo- glycemia but reduces glucose-induced glucagon-like peptide 1 and gastric inhibitory polypeptide secretion. J Clin Endocrinol Metab 92:4165–4171. 103. RASK, E., T. OLSSON, S. SODERBERG, O. JOHNSON, J. SECKL, J.J. HOLST, and B. AHREN. 2001. Impaired incretin response after a mixed meal is associated with insulin resistance in nondiabetic men. Diabetes Care 24:1640–1645. 104. GREENBERG,G.R.,B. CHAN, T.J. MCDONALD,and J. ALLEYNE. 1985.The roleof vagal integrityin gastrin releasing peptide stimulated gastroenteropancreatic hormone release and gastric acid secretion. Regul Pept 10:179–187. 105. HANSEN, L., and J.J. HOLST. 2002. The effects of duodenal peptides on glucagon-like peptide-1 secretion from the ileum. A duodeno-ileal loop? Regul Pept 110:39–45. 106. PLAISANCIE, P., C. BERNARD, J.A. CHAYVIALLE, and J.C. CUBER. 1994. Regulation of glucagon-like peptide-1-(7–36) amide secretion by intestinal neurotransmitters and hormones in the isolated vascularly perfused rat colon. Endocrinology 135:2398–2403. References 25
  • 36. 107. ROBERGE, J.N., and P.L. BRUBAKER. 1993. Regulation of intestinal proglucagon-derived peptide secretion by glucose-dependent insulinotropic peptide in a novel enteroendocrine loop. Endocrinol- ogy 133:233–240. 108. ROCCA, A.S., and P.L. BRUBAKER. 1999. Role of the vagus nerve in mediating proximal nutrient- induced glucagon-like peptide-1 secretion. Endocrinology 140:1687–1694. 109. NAUCK, M.A., M.M. HEIMESAAT, C. ORSKOV, J.J. HOLST, R. EBERT, and W. CREUTZFELDT. 1993. Preserved incretin activity of glucagon-like peptide 1 [7–36 amide] but not of synthetic human gastric inhibitory polypeptide in patients with type-2 diabetes mellitus. J Clin Invest 91:301–307. 110. NAUCK, M.A., E. BARTELS, C. ORSKOV, R. EBERT, and W. CREUTZFELDT. 1993. Additive insulinotropic effects of exogenous synthetic human gastric inhibitory polypeptide and glucagon-like peptide-1- (7–36) amideinfusedat near-physiological insulinotropichormoneand glucoseconcentrations. J Clin Endocrinol Metab 76:912–917. 111. NAUCK, M.A., J. SIEMSGLUSS, C. ORSKOV, and J.J. HOLST. 1996. Release of glucagon-like peptide 1 (GLP-1 [7–36 amide]), gastric inhibitory polypeptide (GIP) and insulin in response to oral glucose after upper and lower intestinal resections. Z Gastroenterol 34:159–166. 112. TOFT-NIELSEN, M.B., M.B. DAMHOLT, S. MADSBAD, L.M. HILSTED, T.E. HUGHES, B.K. MICHELSEN, and J. J. HOLST. 2001. Determinants of the impaired secretion of glucagon-like peptide-1 in type 2 diabetic patients. J Clin Endocrinol Metab 86:3717–3723. 113. MANNUCCI, E., A. OGNIBENE, F. CREMASCO, G. BARDINI, A. MENCUCCI, E. PIERAZZUOLI, S. CIANI, A. FANELLI, G. MESSERI, and C.M. ROTELLA. 2000. Glucagon-like peptide (GLP)-1 and leptin concentra- tions in obese patients with Type 2 diabetes mellitus. Diabet Med 17:713–719. 114. TOFT-NIELSEN, M.B., S. MADSBAD, and J.J. HOLST. 2001. Determinants of the effectiveness of glucagon- like peptide-1 in type 2 diabetes. J Clin Endocrinol Metab 86:3853–3860. 115. LUNDBERG, J.M., K. TATEMOTO, L. TERENIUS, P.M. HELLSTROM, V. MUTT, T. HOKFELT, and B. HAMBERGER. 1982. Localization of peptide YY (PYY) in gastrointestinal endocrine cells and effects on intestinal blood flow and motility. Proc Natl Acad Sci USA 79:4471–4475. 116. GRANDT, D., M. SCHIMICZEK, C. BEGLINGER, P. LAYER, H. GOEBELL, V.E. EYSSELEIN, and J.R. REEVE, Jr., 1994. Two molecular forms of peptide YY (PYY) are abundant in human blood: characterization of a radioimmunoassay recognizing PYY 1–36 and PYY 3–36. Regul Pept 51:151–159. 117. ADRIAN, T.E., A.P. SAVAGE, G.R. SAGOR, J.M. ALLEN, A.J. BACARESE-HAMILTON, K. TATEMOTO, J.M. POLAK, and S.R. BLOOM. 1985. Effect of peptide YY on gastric, pancreatic, and biliary function in humans. Gastroenterology 89:494–499. 118. BATTERHAM, R.L., M.A. COWLEY, C.J. SMALL, H. HERZOG, M.A. COHEN, C.L. DAKIN, A.M. WREN, A.E. BRYNES, M.J. LOW, M.A. GHATEI, R.D. CONE, and S.R. BLOOM. 2002. Gut hormone PYY(3–36) physiologically inhibits food intake. Nature 418:650–654. 119. BATTERHAM,R.L.,M.A.COHEN,S.M.ELLIS, C.W. LeROUX, D.J.WITHERS, G.S.FROST,M.A.GHATEI,and S.R. BLOOM. 2003. Inhibition of food intake in obese subjects by peptide YY3–36. N Engl J Med 349:941–948. 120. HAGAN, M.M. 2002. Peptide YY: a key mediator of orexigenic behavior. Peptides 23:377–382. 121. TSCHOP, M., T.R. CASTANEDA, H.G. JOOST, C. THONE-REINEKE, S. ORTMANN, S. KLAUS, M.M. HAGAN, P.C. CHANDLER, K.D. OSWALD, S.C. BENOIT, R.J. SEELEY, K.P. KINZIG, T.H. MORAN, A.G. BECK- SICKINGER, N. KOGLIN, R.J. RODGERS, J.E. BLUNDELL, Y. ISHII, A.H. BEATTIE, P. HOLCH, D.B. ALLISON, K. RAUN, K. MADSEN, B.S. WULFF, C.E. STIDSEN, M. BIRRINGER, O.J. KREUZER, M. SCHINDLER, K. ARNDT, K. RUDOLF, M. MARK, X.Y. DENG, D.C. WHITCOMB, H. HALEM, J. TAYLOR, J. DONG, R. DATTA, M. CULLER, S. CRANEY, D. FLORA, D. SMILEY, and M.L. HEIMAN. 2004. Physiology: does gut hormone PYY3–36 decrease food intake in rodents? Nature 430:1 p following 165; discussion 2 p following 165. 122. ADAMS, S.H., W.B. WON, S.E. SCHONHOFF, A.B. LEITER, and J.R. PATERNITI, Jr., 2004. Effects of peptide YY[3–36] on short-term food intake in mice are not affected by prevailing plasma ghrelin levels. Endocrinology 145:4967–4975. 123. CHELIKANI, P.K., A.C. HAVER, J.R. REEVE, Jr., D.A. KEIRE, and R.D. REIDELBERGER. 2006. Daily, intermittent intravenous infusion of peptide YY(3–36) reduces daily food intake and adiposity in rats. Am J Physiol Regul Integr Comp Physiol 290:R298–R305. 26 Chapter 1 Gut as an Endocrine Organ
  • 37. 124. EYSSELEIN, V.E., G.A. EBERLEIN, W.H. HESSE, M.V. SINGER, H. GOEBELL, and J.R. REEVE, Jr., 1987. Cholecystokinin-58 is the major circulating form of cholecystokinin in canine blood. J Biol Chem 262:214–217. 125. EBERLEIN, G.A., V.E. EYSSELEIN, W.H. HESSE, H. GOEBELL, M. SCHAEFER, and J.R. REEVE, Jr., 1987. Detection of cholecystokinin-58 in human blood by inhibition of degradation. Am J Physiol Gastrointest Liver Physiol 253:G477–G482. 126. CHANDRA, R., and R.A. LIDDLE. 2007. Cholecystokinin. Curr Opin Endocrinol Diabetes Obes 14:63–67. 127. GALLMANN, E., D. ARSENIJEVIC, M. SPENGLER, G. WILLIAMS, and W. LANGHANS. 2005. Effect of CCK-8 on insulin-induced hyperphagia and hypothalamic orexigenic neuropeptide expression in the rat. Peptides 26:437–445. 128. MORAN, T.H., L.F. KATZ, C.R. PLATA-SALAMAN, and G.J. SCHWARTZ. 1998. Disordered food intake and obesity in rats lacking cholecystokinin A receptors. Am J Physiol Regul Integr Comp Physiol 274: R618–R625. 129. MUURAHAINEN, N., H.R. KISSILEFF, A.J. DEROGATIS, and F.X. PI-SUNYER. 1988. Effects of cholecys- tokinin-octapeptide (CCK-8) on food intake and gastric emptying in man. Physiol Behav 44:645–649. 130. KOPIN, A.S., W.F. MATHES, E.W. MCBRIDE, M. NGUYEN, W. AL-HAIDER, F. SCHMITZ, S. BONNER-WEIR, R. KANAREK, and M. BEINBORN. 1999. The cholecystokinin-A receptor mediates inhibition of food intake yet is not essential for the maintenance of body weight. J Clin Invest 103:383–391. 131. CRAWLEY, J.N., and M.C. BEINFELD. 1983. Rapid development of tolerance to the behavioural actionsof cholecystokinin. Nature 302:703–706. 132. DAKIN, C.L., I. GUNN, C.J. SMALL, C.M. EDWARDS, D.L. HAY, D.M. SMITH, M.A. GHATEI, and S.R. BLOOM. 2001. Oxyntomodulin inhibits food intake in the rat. Endocrinology 142:4244–4250. 133. DAKIN, C.L., C.J. SMALL, A.J. PARK, A. SETH, M.A. GHATEI, and S.R. BLOOM. 2002. Repeated ICV administration of oxyntomodulin causes a greater reduction in body weight gain than in pair-fed rats. Am J Physiol Endocrinol Metab 283:E1173–E1177. 134. DAKIN, C.L., C.J. SMALL, R.L. BATTERHAM, N.M. NEARY, M.A. COHEN, M. PATTERSON, M.A. GHATEI, and S.R. BLOOM. 2004. Peripheral oxyntomodulin reduces food intake and body weight gain in rats. Endocrinology 145:2687–2695. 135. COHEN, M.A., S.M. ELLIS, C.W. Le ROUX, R.L. BATTERHAM, A. PARK, M. PATTERSON, G.S. FROST, M.A. GHATEI, and S.R. BLOOM. 2003. Oxyntomodulin suppresses appetite and reduces food intake in humans. J Clin Endocrinol Metab 88:4696–4701. 136. WYNNE,K.,A.J.PARK,C.J.SMALL,M.PATTERSON,S.M.ELLIS,K.G.MURPHY,A.M.WREN,G.S.FROST,K. MEERAN, M.A. GHATEI, and S.R. BLOOM. 2005. Subcutaneous oxyntomodulin reduces body weight in overweight and obese subjects: a double-blind, randomized, controlled trial. Diabetes 54:2390–2395. 137. WYNNE, K., A.J. PARK, C.J. SMALL, K. MEERAN, M.A. GHATEI, G.S. FROST, and S.R. BLOOM. 2006. Oxyntomodulin increases energy expenditure in addition to decreasing energy intake in overweight and obese humans: a randomised controlled trial. Int J Obes (Lond) 30:1729–1736. 138. POCAI, A., P.E. CARRINGTON, J.R. ADAMS, W. MICHAEL, G. EIERMANN, L. ZHU, X. DU, A. PETROV, M.E. LASSMAN,G. JIANG,F.LIU,C.MILLER, L.M.TOTA,G.ZHOU,X.ZHANG, M.M.SOUNTIS,A.SANTOPRETE,E. CAPITO, G.G. CHICCHI, N. THORNBERRY, E. BIANCHI, A. PESSI, D.J. MARSH, and R. SINHAROY. 2009. GLP- 1/GCGR dual agonism reverses obesity in mice. Diabetes 58:2258–2266. 139. DAY, J.W., N. OTTAWAY, J.T. PATTERSON, V. GELFANOV, D. SMILEY, J. GIDDA, H. FINDEISEN, D. BRUEMMER, D.J. DRUCKER, N. CHAUDHARY, J. HOLLAND, J. HEMBREE, W. ABPLANALP, E. GRANT, J. RUEHL, H. WILSON, H. KIRCHNER, S.H. LOCKIE, S. HOFMANN, S.C. WOODS, R. NOGUEIRAS, P.T. PFLUGER, D. PEREZ-TILVE, R. DIMARCHI, and M.H. TSCHOP. 2009. A new glucagon and GLP-1 co-agonist eliminates obesity in rodents. Nat Chem Biol 5:749–757. 140. YUSTA, B., D. HOLLAND, J.A. KOEHLER, M. MAZIARZ, J.L. ESTALL, R. HIGGINS, and D.J. DRUCKER. 2009. ErbB signaling is required for the proliferative actions of GLP-2 in the murine gut. Gastroenterology 137:986–996. 141. HADJIYANNI, I., K.K. LI, and D.J. DRUCKER. 2009. Glucagon-like peptide-2 reduces intestinal permeability but does not modify the onset of type 1 diabetes in the nonobese diabetic mouse. Endocrinology 150:592–599. References 27
  • 38. 142. HSIEH, J., C. LONGUET, A. MAIDA, J. BAHRAMI, E. XU, C.L. BAKER, P.L. BRUBAKER, D.J. DRUCKER, and K. ADELI. 2009. Glucagon-like peptide-2 increases intestinal lipid absorption and chylomicron produc- tion via CD36. Gastroenterology 137:997–1005. 143. KOJIMA, M., H. HOSODA, Y. DATE, M. NAKAZATO, H. MATSUO, and K. KANGAWA. 1999. Ghrelin is a growth-hormone-releasing acylated peptide from stomach. Nature 402:656–660. 144. CUMMINGS,D.E.,J.Q.PURNELL,R.S.FRAYO,K.SCHMIDOVA,B.E.WISSE,andD.S.WEIGLE.2001.Apreprandial rise in plasma ghrelin levels suggests a role in meal initiation in humans. Diabetes 50:1714–1719. 145. CUMMINGS, D.E., D.S. WEIGLE, R.S. FRAYO, P.A. BREEN, M.K. MA, E.P. DELLINGER, and J.Q. PURNELL. 2002. Plasma ghrelin levels after diet-induced weight loss or gastric bypass surgery. N Engl J Med 346:1623–1630. 146. NAGAYA, N., M. UEMATSU, M. KOJIMA, Y. IKEDA, F. YOSHIHARA, W. SHIMIZU, H. HOSODA, Y. HIROTA, H. ISHIDA, H. MORI, and K. KANGAWA. 2001. Chronic administration of ghrelin improves left ventricular dysfunction and attenuates development of cardiac cachexia in rats with heart failure. Circulation 104:1430–1435. 147. PORIES, W.J. 2008. Bariatric surgery: risks and rewards. J Clin Endocrinol Metab 93:S89–S96. 148. PORIES, W.J., and G.L. DOHM. 2009. Full and durable remission of type 2 diabetes? Through surgery? Surg Obes Relat Dis 5:285–288. 149. PORIES,W.J.,J.F.CARO,E.G.FLICKINGER,H.D.MEELHEIM,andM.S.SWANSON.1987.Thecontrolofdiabetes mellitus (NIDDM) in the morbidly obese with the Greenville Gastric Bypass. Ann Surg 206:316–323. 150. PATTI, M.E., G. MCMAHON, E.C. MUN, A. BITTON, J.J. HOLST, J. GOLDSMITH, D.W. HANTO, M. CALLERY, R. ARKY, V. NOSE, S. BONNER-WEIR, and A.B. GOLDFINE. 2005. Severe hypoglycaemia post-gastric bypass requiring partial pancreatectomy: evidence for inappropriate insulin secretion and pancreatic islet hyperplasia. Diabetologia 48:2236–2240. 151. SERVICE, G.J., G.B. THOMPSON, F.J. SERVICE, J.C. ANDREWS, M.L. COLLAZO-CLAVELL, and R.V. LLOYD. 2005. Hyperinsulinemic hypoglycemia with nesidioblastosis after gastric-bypass surgery. N Engl J Med 353:249–254. 152. CLANCY, T.E., F.D. MOORE, Jr., and M.J. ZINNER. 2006. Post-gastric bypass hyperinsulinism with nesidioblastosis: subtotal or total pancreatectomy may be needed to prevent recurrent hypoglycemia. J Gastrointest Surg 10:1116–1119. 153. SJOSTROM, L., A.K.LINDROOS, M. PELTONEN, J. TORGERSON,C. BOUCHARD, B. CARLSSON, S. DAHLGREN, B. LARSSON, K. NARBRO, C.D. SJOSTROM, M. SULLIVAN, and H. WEDEL. 2004. Lifestyle, diabetes, and cardiovascular risk factors 10 years after bariatric surgery. N Engl J Med 351:2683–2693. 154. BUCHWALD, H., Y. AVIDOR, E. BRAUNWALD, M.D. JENSEN, W. PORIES, K. FAHRBACH, and K. SCHOELLES. 2004. Bariatric surgery: a systematic review and meta-analysis. JAMA 292:1724–1737. 155. KORNER, J., W. INABNET, G. FEBRES, I.M. CONWELL, D.J. MCMAHON, R. SALAS, C. TAVERAS, B. SCHROPE, and M. BESSLER. 2009. Prospective study of gut hormone and metabolic changes after adjustable gastric banding and Roux-en-Y gastric bypass. Int J Obes (Lond) 33:786–795. 156. BIKMAN, B.T., D. ZHENG, W.J. PORIES, W. CHAPMAN, J.R. PENDER, R.C. BOWDEN, M.A. REED, R.N. CORTRIGHT, E.B. TAPSCOTT, J.A. HOUMARD, C.J. TANNER, J. LEE, and G.L. DOHM. 2008. Mechanism for improved insulin sensitivity after gastric bypass surgery. J Clin Endocrinol Metab 93:4656–4663. 157. le ROUX, C.W., R. WELBOURN, M. WERLING, A. OSBORNE, A. KOKKINOS, A. LAURENIUS, H. LONROTH, L. FANDRIKS, M.A. GHATEI, S.R. BLOOM, and T. OLBERS. 2007. Gut hormones as mediators of appetite and weight loss after Roux-en-Y gastric bypass. Ann Surg 246:780–785. 158. VIDAL, J., J. NICOLAU, F. ROMERO, R. CASAMITJANA, D. MOMBLAN, I. CONGET, R. MORINIGO, and A.M. LACY. 2009. Long-term effects of Roux-en-Y gastric bypass surgery on plasma glucagon-like peptide- 1 and islet function in morbidly obese subjects. J Clin Endocrinol Metab 94:884–891. 159. RUBINO, F., A. FORGIONE, D.E. CUMMINGS, M. VIX, D. GNULI, G. MINGRONE, M. CASTAGNETO, and J. MARESCAUX. 2006. The mechanism of diabetes control after gastrointestinal bypass surgery reveals a role of the proximal small intestine in the pathophysiology of type 2 diabetes. Ann Surg 244:741–749. 160. TROY, S., M. SOTY, L. RIBEIRO, L. LAVAL, S. MIGRENNE, X. FIORAMONTI, B. PILLOT, V. FAUVEAU, R. AUBERT, B. VIOLLET, M. FORETZ, J.LECLERC,A.DUCHAMPT,C.ZITOUN, B. THORENS,C.MAGNAN, G.MITHIEUX,and F. ANDREELLI. 2008. Intestinal gluconeogenesis is a key factor for early metabolic changes after gastric bypass but not after gastric lap-band in mice. Cell Metab 8:201–211. 28 Chapter 1 Gut as an Endocrine Organ
  • 39. Chapter 2 Central Glucose Sensing and Control of Food Intake and Energy Homeostasis LOURDES MOUNIEN 1,2 AND BERNARD THORENS 1,2 1 Department of Physiology, University of Lausanne, Lausanne, Switzerland 2 Center for Integrative Genomics, University of Lausanne, Lausanne, Switzerland INTRODUCTION Glucose plays an essential role in energy homeostasis by regulating the secretion of various hormones and the activation of neuronal circuits controlling feeding and energy expenditure (1, 2). Glucose-sensing systems are present at many anatomical sites including the mouth, the gastrointestinal tract, the hepatoportal vein, the liver, the endocrine pancreas, and the central nervous system (CNS). In the mouth, glucose activates taste receptors, which stimulate afferent fibers projecting to the brainstem and trigger the cephalic phase of insulin secretion (3–5). In the intestine, glucose stimulates the secretion of the gluco-incretin hormones glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP), by a mechanism that may involve the sodium-dependent glucose transporter SGLT1 and the KATP channel (6) and possibly also the activation of sweet taste receptors (7–9). A local action of GLP-1 hormones is to increase the expression of the Naþ /glucose cotransporter SGLT1 and the translocation of the glucose transporter GLUT2atthebrushborderofenterocytes,leadingtoincreasedglucoseabsorption(9). Glucose also activates autonomic and enteric neurons located in the gut mucosa (10, 11). Activation of the cholinergic neurons of the submucosal and myenteric plexus may be mediated by glucose binding to SGLT3 (12). Metabolic Syndrome: Underlying Mechanisms and Drug Therapies Edited by Minghan Wang Copyright Ó 2011 John Wiley Sons, Inc. 29
  • 40. Glucose entering the portal vein activates vagal afferents that project to the CNS (13, 14) to control several adaptive responses such as stimulation of glucose storage by liver, soleus, heart, and brown adipose tissue; inhibition of counter- regulation; termination of food intake; and stimulation of first-phase insulin secre- tion (10, 15, 16). In the liver, glucose stimulates glycogen synthesis (17) as well as the expression of genes involved in glycolysis and lipogenesis through transcriptional mechanisms mediated in large part by the glucose-sensitive transcription factor ChreBP (18, 19). In the pancreas, increase in plasma glucose triggers insulin secretion by b-cells (1, 20–22). In contrast, glucagon secretion by a-cells is triggered when glycemia falls below the euglycemic level (20, 23, 24). In the CNS, glucose modulates the activity of glucose-sensitive neurons located in the hypothalamus and brainstem. These belong to at least two classes: glucose- excited (GE) neurons, whose firing activity is increased by rises in extracellular glucose concentrations, and glucose-inhibited (GI) neurons, which are activated when glucose concentrations decrease. These glucose-sensitive neurons control glucose homeostasis, feeding behavior, and energy homeostasis. The molecular basis for glucose monitoring and regulation of firing activity is being actively investigated. Present evidence indicates that there is a large diversity in the mechanisms of glucose sensing, which may define subpopulation of either glucose-excited or glucose- inhibited neurons. Recent experimental evidence also supports a role for glial cells in glucose sensing (25, 26). The complexity of these glucose-sensing mechanisms needs to be eventually completely understood to better manage pathologies caused by deregulated glucose and energy homeostasis. Here, we mainly focus on the mechanisms of glucose sensing by neurons. BRAIN GLUCOSE SENSING Sites of Glucodetection Claude Bernardfirstimplicatedthebrainsteminglucosehomeostasiswhen he showed that puncturing the floor of the fourth cerebral ventricle of dogs rapidly induced diabetes (27). In 1953, Jean Mayer proposed that cells located in hypothalamus monitor plasma glucose levels by translating variations in glycemia into electrical or chemical signals to control feeding behavior (28). Several groups then identified GE neurons, whose electrical activity is increased by high glucose, and GI neurons activated by cellularglucoprivation (29–32). GE and GI neurons are mainly expressed in the hypothalamus, in particular in the arcuate (Arc), lateral (LHA), dorsomedial (DMH), ventromedial (VMH), and paraventricular (PVN) hypothalamic nu- clei (32–38), and in the brainstem, in the area postrema (AP), the nucleus of the solitary tract (NTS), the dorsal motor nucleus of the vagus (DMNX), and the basolateral medulla (BLM) (14, 33, 34, 36, 39–41). High glucose excited (HGE) and high glucose inhibited (HGI) neurons, whose firing activity is regulated over glucose concentration ranges(5–20 mM), havealsobeen identifiedin the Arc (35, 42). 30 Chapter 2 Central Glucose Sensing and Control of Food Intake
  • 41. Mechanisms of Glucodetection Glucose controls the electrical activity of GE neurons by a mechanism that shares similarities to glucose-induced insulin secretion by pancreatic b-cells (43, 44) (Figure 2.1a). Glucose signaling in these cells is initiated by the uptake of glucose by GLUT2 followed by its phosphorylation by glucokinase (hexokinase IV, Km for glucose $6 mM). Subsequent activation of mitochondrial metabolism and oxidative phosphorylation increases the ATP/ADP ratio. This leads to closure of ATP-sensitive potassium (KATP) channels, plasma membrane depolarization, and opening of voltage-sensitive Ca2þ channels. The influx of calcium then triggers insulin secre- tion. In GE neurons, rise in extracellular glucose also increases the cytosolic ATP/ADP ratio and induces closure of KATP channels (39, 45, 46), plasma membrane depolarization, Ca2þ entry, and neurotransmitter release (47, 48). However, recent studies suggest that some GE neurons may be activated by glucose in a KATP channel-independent manner (35, 49). Evidence also suggests that GK (50) and GLUT2 (51, 52) may not be required for activation of these neurons. The mechanisms through which GI neurons sense glucose are not well charac- terized (Figure 2.1b). However, the effect of glucose on GI neurons may be controlled by changes in Naþ /Kþ ATPase activity (30, 37) or by the opening of ATP-regulated chloride channels that leads to hyperpolarization (31, 50). In the LHA, inhibition by high glucose of the GI orexin neurons may depend on tandem-pore Kþ (TASK) or related channels (53–55). PHYSIOLOGICAL FUNCTIONS MODULATED BY CENTRAL GLUCODETECTION Food Intake and Energy Expenditure The role of glucose in the control of food intake was demonstrated in many studies (2, 28). Particularly, it has been shown that initiation of feeding is preceded by a small drop in glycemia, and preventing it by infusing glucose suppresses initiation of feeding (56). In addition, peripheral or central administration of 2-deoxyglucose (2-DG), which induces neuroglycopenia, stimulates food intake (57–59). The modulation offeeding behavior and energy expenditure by CNS is a complex process that involves hypothalamic and brainstem neuronal circuits. In the hypo- thalamus, neurons integrate nutrient (lipid and glucose), hormonal (ghrelin, insulin, PYY3–36, leptin, CCK, GLP-1, and adiponectin), and nervous signals that convey information about food absorption and the levels of stored energy (10, 60–64). These signals are detected in large part by Arc neurons expressing the anorexigenic peptides POMC/CART or the orexigenic peptides NPY/AgRP. These neurons project to melanocortin 3 and 4 receptor-expressing neurons of the PVN and LHA (20, 65). Neurons in the PVN produce the anorexigenic neuropeptides TRH and CRF whereas neurons in the LH produce the orexigenic peptides MCH and orexin (62). Together these neurons form the melanocortin pathway and regulate peripheral metabolism Physiological Functions Modulated by Central Glucodetection 31
  • 42. GLUT2 1,3 GK KATP ↑G6P KC SGLT1,3 ↑pyruvate ↑Na+ Ox. Ph. ATPase ↑H+ ↑ATP/ADP TRP VDCC ↑ROS ↑Ca2+ ↑ROS↑ATP Classical model Alternative models GLUT2 1,3 KC VDCC↑Ca2+ ? ? glucose ? (a) GLUCOSE GE neurons GLUT2 1,3 KC ↓pyruvate ↓ATP/ADP VDCC ↓ATP Classical model Alternative models GLUCOSE ↑Ca2+ (b) GLUCOSE GI neurons Na+/K+ pump Receptor TASK Ox. Ph. ATPase ClC ? ? ↓H+ VDCC GK ↓G6P GLUCOSE UCP2 UCP2 Figure 2.1 Models for the control of the electrical activity of GE and GI neurons by glucose. (a) Glucose activation offiring rate in GE neurons. In the b-cell model, glucose uptake through GLUT2, but in neurons possibly also by GLUT1 or GLUT3, is followed by its phosphorylation by GK. Pyruvate is then channeled into the mitochondria to eventually increase ATP production and induce a rise in the ATP/ADP ratio. This leads to the closure of KATP channel, membrane depolarization, and the entry of Ca2þ , which triggers the release of neurotransmitters. In an alternative mechanism, the activity of GE neurons is controlled by the electrogenic cotransport of Naþ and glucose by SGLT1, by activation of TRP channels, or by production of ROS following the activation of the oxidative phosphorylation. ROS could directly regulate the activity of Kþ channels or intracellular Ca2þ availability. UCP2 may serve as aregulatorofATPproductionthroughitsdecouplingactivityorbyreducingthe productionofROS;both mechanisms lead to reduced glucose-induced neuronal activity. (b) Activation of GI neurons by a decrease in extracellular glucose concentration. A reduction in the ATP/ADP ratio leads to closure of ClÀ channels and/or a reduction in the activity of the Naþ /Kþ ATPase, plasma depolarization, and the entry of Ca2þ that triggers the secretion of neurotransmitters. Alternatively, a new pathway involving TASKchannelmaycontroltheactivityofGIneuronsthroughamechanism thatdoesnotinvolveglucose metabolism, possibly through interaction with a putative specific receptor. KC: Kþ channel; VDCC: voltage-dependent calcium channel. 32 Chapter 2 Central Glucose Sensing and Control of Food Intake
  • 43. through activation of the autonomic nervous system and higher brain structures to control not only feeding behavior, but also arousal and reward (66–68). The hindbrain is also a site of glucodetection and may have a major role in regulating feeding in physiological conditions. Indeed, intracerebroventricular (i.c.v.) injection of 2-DG stimulates feeding only if it can have access to the brainstem (57, 69) and food uptake can be activated by injection of 5-thioglucose (5-TG) into NTS, DMNX, and BLM but not in hypothalamic nuclei (39–41, 70, 71). Glucose-sensitive neurons from the BLM are catecholaminergic and send projection to Arc and PVN (72). Destruction of these projections by immunotoxins suppresses the effect of 2-DG on food intake and on regulated expression of AgRP and NPYin the Arc (73, 74). Counterregulation When blood glucose concentrations fall below the euglycemic level, a rapid counter- regulatory response is activated to restore normoglycemia. This involves activation of the autonomic nervous system (75) that triggers glucagon secretion and the release of catecholamines from adrenal glands (76–78). The central sites of glucose detection that activate the autonomic nervous system are located in the hypothalamus and brainstem. In the hypothalamus, lesion studies as well as pharmacological and genetic approaches have provided evidence for an important role of the VMH in the control of glucagon secretion (79). For instance, glucagon release can be induced by direct injection of 2-DG in the VMH (80). In contrast, hypoglycemia-induced glucagon secretion can be suppressed by direct injection of glucose into this structure (81). Brainstem nuclei also play an important role in activating counterregulation. For instance, when the cerebral aqueduct is obstructed, 5-TG induces glucagon secretion when injected in the fourth but not in the third ventricle (69). In addition, injection of 5-TG directly in the NTS or the BLM nuclei containing the A1/C1 catecholaminergic neurons strongly stimulates glucagon secretion (70). In decere- brated rats, the hyperglycemic response to an i.p. injection of 2-DG is preserved (82) and c-fos staining revealed that the activated cells are present in the NTS, DMNX, and the catecholaminergic neurons of the BLM (72). MULTIPLICITY OF SENSING MECHANISMS One important goal of current research is to identify each type of glucose-sensing neurons and to determine which physiological functions they control. One path to reach this goal is to identify the critical proteins that allow these neurons to respond to glucose and use these proteins as markers to identify the glucose-sensing neuron subpopulations, their topographical distribution, and the neuronal circuits they form. In recent years, many ion channels, transporters, or enzymes have been described to participate in central glucose sensing. Here, we review the list of these gene products and their role in glucose sensing. Multiplicity of Sensing Mechanisms 33
  • 44. Glucose Transporters In most instances, glucose uptake and metabolism are required for glucose signaling in neurons. Glucose uptake may be catalyzed by either facilitated diffusion glucose transporters (Gluts) or Naþ -linked glucose transporters (SGLTs) (83–87), and both types of transporters have been associated with glucose sensing by hypothalamic and brainstem neurons. Glut1, Glut3, and Glut8 Glut1 is highly expressed in endothelial cells forming the blood–brain barrier and is required for glucose entry in the brain. Inactivating mutations of Glut1 reduce the concentration of glucose in the cerebrospinal fluid, which causes seizure and delayed development (88). Glut3 is a high-affinity glucose transporter expressed in neu- rons (89). Homozygous Glut3 knockout mice die during embryogenesis whereas heterozygous knockout mice have normal glucose homeostasis and feeding behav- ior (90), although they show defects in spatial learning and memory processing (91). Glut8 is a high-affinity glucose transporter expressed in neurons in different brain regions, including the hippocampus, the hypothalamus, and the brainstem (92, 93) but homozygous Glut8 knockout mice show no defect in glucose or energy homeosta- sis (94). Thus, Glut1, Glut3, and Glut8 play specific role in brain glucose metabolism unrelated to the glucose sensing and control of whole-body glucose and energy homeostasis. Glut2 Glut2 is a low-affinity glucose transporter, required for normal glucose sensing by pancreatic b-cells. In the brain, it is expressed in neurons, astrocytes, tanycytes, and epithelial cells lining the cerebral ventricles (95–99). The low level of expression of Glut2 makes its immunohistochemical detection challenging and its precise local- ization is still partly uncertain, although it is present in hypothalamic and brainstem nuclei (51, 95, 96, 100, 101). A role for central GLUT2 in glucose sensing has been suggested by i.c.v. or direct injection in the Arc of antisense oligonucleotides to reduce its expression (98, 102). This decreased feeding and body weight gain and suppressed 2-DG-induced feeding as well as the insulin response normally triggered by intracarotid glucose injection (98, 102). Studies with Glut2À/À mice, which express a transgenic glucose transporter in their b-cells to restore normal glucose- stimulated insulin secretion (ripglut1;glut2À/À mice) (103), demonstrated a role for central Glut2 in the control of glucagon secretion in response to insulin-induced hypoglycemia or 2-DG-induced neuroglucopenia (26). Transgenic complementation studies revealed that Glut2 reexpression in astrocytes but not in neurons restored the counterregulatory response to hypoglycemia (26). The same mouse model was used to study the role of Glut2 in central glucose sensing and the control of feeding. It was demonstrated that in the absence of Glut2, the mouse presented defects in both feeding initiation and termination. This was 34 Chapter 2 Central Glucose Sensing and Control of Food Intake
  • 45. correlated with suppressed regulation of the hypothalamic anorexigenic (POMC, CART) and orexigenic (NPY, AGRP) neuropeptides during the fast to refed transition or following intracerebroventricular injection of glucose (104). In contrast to the impaired counterregulatory response, the abnormal feeding behavior of GLUT2-null mice did not rely on GLUT2 expression in astrocytes (105), suggesting that regulation of counterregulation and feeding behavior depends on Glut2 expression in different cell types, astrocytes and neurons, respectively. SGLT1 and SGLT3 SGLT1 is a Naþ –glucose symporter present in the brush border of intestinal epithelial cells, GLP-1 secreting L cells, and in the proximal straight tubule of the kidney nephrons (106–108). SGLT1 transports glucose and galactose as well as the non- metabolizable analogues 3-O-methyl-D-glucose (3-O-MDG) and a-methyl-D- glucopyranoside (a-MDG) and is inhibited by phlorizin (107, 109). SGLT1 is also expressed in the hypothalamus and ependymal cells of the third and fourth ven- tricles (52, 110). A role for SGLT1 in L cell glucose sensing has been described (108, 111). As glucose uptake is electrogenic, it leads to membrane depolarization and GLP-1 secretion in a KATP-independent manner (112); GLP-1 secretion can also be triggered by the nonmetabolized substrates 3-O-MDG and a-MDG (113, 114). That SGLT1 may play a role in central glucose sensing is suggested by the finding that i.c.v. injection of phlorizin enhances food intake in rats (115) and inhibits activation of GE neurons in the VMH (44). On the other hand, hypothalamic GE neurons can be excited by a-MDG and 3-OMG (52). SGLT3 is expressed mainly in intestine, liver, kidney, and muscle (116). Pig SGLT3 transports glucose and a-MDG with relatively low affinity and, in contrast to SGLT1, does not transport galactose and 3-O-MDG (109). Human SGLT3 does not transport glucose when expressed in Xenopus laevis oocytes but may play a role as a glucose sensor in cholinergic neurons of the small intestine and at the neuromuscular junctions (12). In Xenopus oocytes expressing SGLT3, glucose produced a phlorizin- sensitive inward current that depolarizes the membrane potential by up to 50 mV (12). SGLT3 mRNA is expressed in both cultured hypothalamic neurons and adult hypothalamus, suggesting that this transporter may also be involved in central glucose sensing (52). Intracellular Mediators Glucokinase Following its uptake, glucose is phosphorylated by hexokinases. In pancreatic b-cells, glucokinase controls the flux of glucose metabolism and the dose response of glucose- stimulated insulin secretion. In the brain, glucokinase is expressed in the Arc, LHA, DMH, VMH, and PVN, as well as in the brainstem and ependymocytes of the third and fourth ventricles (99, 101, 117, 118). Pharmacological inhibition of hypothalamic Multiplicity of Sensing Mechanisms 35
  • 46. GK decreases the activity of GE neurons and increases that of GI neuronsshowingthat GK is involved in both high glucose and glucopenia detection (32, 38, 44, 51, 119). Recently, GK has also been shown to be required for glucose sensing by GI and GE neurons in the NTS and DMNX (118). A role for central GK in glucagon secretion and feeding is evidenced by i.c.v. administration of the GK inhibitor alloxan that stimulates feeding (120). Reduction of GK activity in the VMH by injection of alloxan or by adenoviral-mediated transduction of a GK-specific shRNA showed that this enzyme is essential for the counterregulatory response to insulin-induced hypoglycemia (121) and for feeding (122). AMPK AMPK is a ubiquitous enzyme formed by a catalytic and b and g regulatory subunits (123, 124). AMPK is an intracellular fuel gauge activated by increased intracellular AMP/ATP ratio and by phosphorylation by the LKB1 and Ca2þ /Calmodulin-activated kinases (124, 125). AMPK turns on catabolic pathways such as fatty acid oxidation and turns off gluconeogenesis and lipogenesis (125). In pancreatic b-cells, activation of AMPK by low glucose suppresses glucose-induced glycolysis, mitochondrial oxidative metabolism, Ca2þ influx, and insulin secretion (126). A role for hypothalamic AMPK in metabolic regulation has been initially proposed based on studies indicating that its activity is inhibited by leptin, insulin, glucose, and refeeding. AMPK activity is regulated in these conditions only in the Arc and PVN but not in the VMH, DMH, and LHA nuclei (123). Adenoviral delivery of constitutively active or dominant negative forms of AMPK in medial hypothalamic nuclei activates or inhibits feeding, respectively (123), and i.c.v. administration or direct injection in the PVN of 5-amino-4-imidazolecarboxamide riboside (AICAR) stimulates feeding (127). Similarly, neuroglucopenia induced by i.c.v. injection of 2-DG increases hypothalamic AMPK activity and feeding, an effect that can be blocked by the AMPK inhibitor compound C (128). How AMPK activity in hypothalamic neurons controls feeding is not fully understood. In neuronal cell lines and on ex vivo hypothalamic explants, low glucose concentrations and AICAR increase AMPK activity and AgRP expression (129). In accordance with these observations, the specific deletion of the a-subunit of AMPK in POMC and AgRP neurons suppressed glucose sensing by these cells but preserved normal leptin or insulin action (130). AMPK may also be involved in the counterregulatory response to hypoglycemia or to neuroglucopenia (131–134). Stimulation of AMPK by microinjection of AICAR in the VMH increases endogenous glucose production whereas compound C or expression of a dominant negative form of AMPK in ARC and VMH impaired early counterregulation as evidenced by reduced glucagon and catecholamine responses to hypoglycemia (134). In contrast, overexpression of the dominant negative AMPK in the PVN attenuated late counterregulation and corticosterone responses (134). 36 Chapter 2 Central Glucose Sensing and Control of Food Intake
  • 47. Recently, it has been shown that the counterregulatory hormone response impaired by 2-DG-induced recurrent neuroglucopenia was partially restored by i.c.v. injection of AICAR (131). At the brainstem level, AMPK activity also contributes to energy homeostasis. For instance, AMPK activity is significantly increased in the NTS of fasted compared to ad libitum fed rats (135) and injection of compound C directly in the NTS induces a decrease in food intake and body weight (135). Regulation of AMPK in the brainstem may thus mediate the anorectic action of leptin, which is reversed by AICAR injections (135). mTOR The mammalian target of rapamycin (mTOR) is a conserved serine–threonine kinase that promotes anabolic pathways such as protein synthesis in response to growth factors, nutrients (amino acids and glucose), and stress (136). These mechanisms involve the regulatory proteins 70 kDa ribosomal protein S6 kinase (S6K1) and the eukaryotic initiation factor 4E-binding protein-1 (4EBP1), which are key regulators of protein synthesis (136, 137). mTOR exists in two distinct complexes. Target of rapamycin complex 1 (TORC1) is a functional association of mTOR with the scaffolding protein raptor, whereas TORC2 is the combination of mTOR with the protein rictor. mTORC1 functions as a nutrient/energy/redox sensor controlling protein synthesis and can be inhibited by rapamycin (138, 139). mTORC2 is an important regulator of the cytoskeleton and phosphorylates the serine/threonine protein kinase Akt/PKB at a serine residue S473 (140). TSC1 and TSC2 are tumor suppressors, and their gene products form a stable complex that inhibits mTORC1 activity (141). Glucose deprivation inhibits mTOR activity, and inhibition is abol- ished in TSC mutant cells. Interestingly, AMPK inhibits mTORC1 activity by phosphorylation of TSC2 as well as raptor (136). The group of McDaniel showed that glucose elevation activates mTOR/S6K1/ 4EBP1 and protein synthesis in an amino acid-dependent manner in both rodent and human islets (142–144). In rat brain, mTOR has been located in numerous brain structures including hypothalamus, thalamus, and cortex (145). In hypothalamus, mTOR is located in PVN and Arc (145). More precisely, 90% of NPY/AgRP neurons while 45% of POMC neurons also expressed mTOR protein (145). After fasting, there is a decrease in mTOR-positive cells in Arc suggesting that mTOR activity is low after glucose privation (145). In addition, leptin also stimulates mTOR activity and inhibition of mTOR with rapamycin blunts the anorexigenic effect of leptin (145). Recently, specific ablation of TSC1 in POMC neurons induced hyperphagic obesity and alteration of the morphology of POMC neurons (146). These phenotypes are reversed by treatment with rapamycin (146) suggesting an important role of POMC neurons in the control of metabolism. Interestingly, the activation of AMPK-dependent mechanism leads to the inhibition of mTOR activity (147). Thus, AMPK and mTOR may have reciprocal functions and interact in order to control energy homeostasis. Multiplicity of Sensing Mechanisms 37
  • 48. UCP2/ROS Activation of the mitochondrial respiratory chain leads to Hþ extrusion from the mitochondria and establishment of an electrochemical gradient across the inner mitochondrial membrane. The transport of Hþ back into the mitochondrial matrix through the F0F1ATPase generates ATP. In pancreatic b-cells, the rise in intracellular ATP/ADP ratio is critical to link glucose metabolism to insulin secretion. The uncoupling protein UCP2, located in the inner mitochondrial membrane, can dissipate the electrochemical Hþ gradient, thereby decreasing the capacity of the cells to produce ATP (148). Accordingly, overexpression of UCP2 in islets or insulinoma cells blunts glucose-induced insulin secretion (149–151), and islets from UCP2À/À mice have increased ATP levels and higher secretory response to glucose (152, 153). In brain, UCP2 is widely distributed and expressed at high levels in the hypothalamus, in particular in the Arc, VMH, PVN, and LHA, as well as in the brainstem (154) where it has been proposed to play a role in glucose sensing (155). For instance, increased expression of UCP2 in POMC neurons of mice fed a high-fat diet is associated with a loss of their glucose sensitivity, which can be prevented by genepin, an inhibitor of UCP2, or by UCP2 gene inactivation (156). UCP2 was also found to be critical for activation of NPY/AgRP neurons during fasting and in response to ghrelin (157, 158). Oxidative phosphorylation is also associated with the production of reactive oxygen species (ROS) and UCP2 may act as a negative regulator of ROS produc- tion (159–161). Indeed, initial studies of UCP2À/À mice showed that their macro- phages generated 80% more ROS than those from control mice (159). Importantly, ROS are also intracellular signaling molecules (162) that can regulate the activity of voltage-gated Kþ channels (163, 164) or Ca2þ influx (165–167). In b-cells, ROS may participate in the coupling between glucose metabolism and insulin secre- tion (168–174). In the hypothalamus, there is evidence that glucose sensing may involveROS production (175). Exposure of hypothalamic slices to increase in glucose concentrations (from 5 to 20 mmol/L) stimulates ROS generation, which is reversed by addition of antioxidants. Intracarotid administration of antimycin or rotenone, which induces ROS formation, mimics the effect of glucose on Arc neurons’ activity and subsequent nervous-mediated insulin release (175). Channels KATP Channel-Dependent Mechanisms The KATP channel plays a fundamental role in coupling changes in glucose metab- olism to plasma membrane electrical activity (176). This channel is an octameric protein consisting of four copies of the pore-forming Kir6.2 channel (in pancreatic b-cells and in neurons) and four copies of the sulfonylurea receptor SUR1 (in pancreatic b-cells and brain) or SUR2B (in brain) (34, 177–180). The expression of these different subunits (Kir6.2, SUR1, and SUR2B) suggests a molecular diversity 38 Chapter 2 Central Glucose Sensing and Control of Food Intake
  • 49. of KATP in brain and especially in hypothalamus and brainstem (34, 46, 118, 156, 176, 177, 181–184). This channel is involved in central glucose sensing to control glucagon secretion and feeding. For instance, i.c.v. or intrahypothalamic injection of glibenclamide, a KATP channel inhibitor, blocks the counterregulatory response to a hypoglycemic clamp or induced by central administration of 5-TG (185). Genetic inactivation of Kir6.2 leads to impaired glucagon secretion in response to 2-DG administration and this is correlated with suppressed glucose-regulated firing of VMH neurons (46). In contrast, activation of this channel in the VMH amplifies counterregulatory hormone responses to hypoglycemia in normal and recurrently hypoglycemic rats (186). Kir6.2-null mice also have a smaller but significant feeding response than control mice to 2-DG injection (46). In addition, the acute regulation of membrane potential and firing of a subset of hypothalamic neurons by leptin and insulin is due to an action on the KATP channel that causes cell hyperpolarization (187–189). In a recent study, the role of the KATP channel in POMC neurons was addressed by transgenic expression in these neurons of a mutant Kir6.2 subunit that prevents ATP-mediated closure of the channel. This suppresses the response of these neurons to glucose without affecting feeding and induces mild glucose intolerance (156). KATP Channel-Independent Mechanisms Numerous studies suggest that the glucose-dependent firing activity of glucose- sensing neurons is also controlled by mechanisms not involving the KATP channel. Table 2.1 lists these channels and their distribution in the hypothalamus and brainstem. Electrophysiological recording of neurons from Kir6.2À/À mice showed firing activity triggered by increase in glucose concentrations from 5 to 20 mmol/L with a decrease in input resistance (35), suggesting that these HGE neurons used a KATP channel-independent mechanism to sense glucose. This response appears to depend on the opening of transient response potential (TRP) channels (35), which may play a role in maintaining intracellular Ca2þ concentration (190). The firing activity of GI neurons in response to decreased extracellular glucose may involve reduced activation of the Naþ /Kþ ATPase (30, 191), blockade of a ClÀ conductance (37, 192), or inhibition of acid-sensitive two-pore domain Kþ channels (TASK) (55). In the LHA, local application of glucose hyperpolarizes the GI orexin neurons, an effect that is prevented by ouabain (a blocker of the Naþ /Kþ ATPase) and azide (an inhibitor of energy production), suggesting that glucose exerts its inhibitory effect through Naþ /Kþ ATPase (30). The response of the orexin neurons may involve a glucose-activated Kþ conductance (55), which based on the sensitivity to pH and halothane may be the K2p Twik1-related acid-sensitive Kþ channel subunit TASK3 (55). However, glucose-induced hyperpolarization of orexin neurons is unaffected not only in TASK3 knockout mice but also in TASK1 and TASK3/ TASK1-null mice suggesting that the exact mechanisms of activation of neurons by low glucose are still incompletely understood (53). Multiplicity of Sensing Mechanisms 39
  • 50. Table 2.1 Distribution in the Hypothalamus and Brainstem of the Proteins Involved in Central Glucose Sensing Hypothalamus Brainstem Arcuate nucleus Dorsomedial nucleus Lateral hypothalamic area Paraventricular nucleus Ventromedial nucleus Ependymal layers of third ventricle Area postrema Dorsal motor nucleus of the vagus Nucleus of the solitary tract Basolateral medulla (A1/C1) Ependymal layers of fourth ventricle Transporter Glut2 ? þ þ þ þ þ ? þ þ þ þ (96–101) Glut1/3 þ þ þ þ þ þ þ þ þ þ þ (51, 97, 99) SGLT1/3 SGLT1/3 is detected in hypothalamic cultured neurons and in adult hypothalamus ND ND þ ND þ (52, 110) Enzyme Glucokinase þ þ À À þ þ À þ þ þ þ (99, 117, 118) AMPK þ þ þ þ þ ND þ þ þ þ þ (130, 135) UCP UCP2/ROS þ þ À þ þ þ þ þ þ ND þ (148, 156, 158, 175) Channels TRP þ À ND þ þ ND TRP1 is detected in brainstem tissue extract (35) CFTR/ClÀ þ ND þ þ þ ND À À þ ND ND (191, 192) Tandem- pore Kþ þ þ þ þ þ þ þ þ þ þ þ (55, 191) Na/K ATPase þ þ þ þ þ ND þ þ þ þ ND (30, 191) KATP channels þ þ þ þ þ ND þ þ þ þ ND (34, 46, 118, 156, 177, 181) þ , Positive; À, negative; ND, undetermined; ?, conflictual data. 40
  • 51. In Arc GI neurons, a role for ClÀ conductance has been evidenced for the response to low glucose concentrations (37, 192). As gemfibrozil, a cystic fibrosis transmembrane regulator (CFTR) blocker, prevents activation of GI neurons in both the Arc and VMH, the CFTR may be involved in this response (192). In the dorsal vagal complex, inhibition of Naþ /Kþ ATPase by strophanthidin or ouabain suppressed the inward currents of GI neurons and a role for ClÀ channels can be excluded (191). CONCLUSION Changes in glycemia are monitored by several systems located at different ana- tomical sites. GE and GI neurons, located in hypothalamus and brainstem, control many physiological responses such as counterregulation, feeding behavior, and energy expenditure. At the molecular level, present evidence indicates that a large number of mechanisms have evolved to tightly monitor increases or falls in glycemia. SGLT1, GLUT2, GK, and KATP channel appear to be associated with the response of GE neurons to increase in glucose concentrations. However, not all GE neurons do express GLUT2 or GK, suggesting a diversity in the function of the GE neurons. GI neurons increase their firing activity in response to fall in glucose concentrations by mechanisms that are still unclear but that do not involve the KATP channel but rather Naþ /Kþ ATPase pump, chloride channels, or TRP or TASK channels. Again, available evidence indicates that these pump and channels may be differently required by GI neurons present in different locations. AMPK, mTOR, UCP2, and the production of ROS are also involved in the response of neurons to changes in glycemia and it is so far not known whether they all contribute to the response of all GE or GI neurons or only to subpopulations of glucose-sensitive neurons. Although not fully discussed in this chapter, it has been recently suggested that some GE and GI neurons may sense glucose independently of glucose metabolism. For instance, hypothalamic GE neurons in culture are also excited by the nonme- tabolizable glucose analogue a-MDG, which is a substrate of SGLT. In addition, in GI neurons such as orexin cells, glucose-induced hyperpolarization and inhibition are unaffected by GK inhibitors and mimicked by 2-DG. Therefore, the CNS, which critically depends on glucose for its function, has evolved many glucose-sensing mechanisms to monitor all aspects of energy supply and need and modulate glucose and energy homeostasis. Deciphering the complexity of glucose sensing by the CNS and the structure of the glucose-sensing neuronal circuits that control glucose and energy homeostasis still represents a formidable challenge. REFERENCES 1. MARTY, N., M. DALLAPORTA, and B. THORENS. 2007. Brain glucose sensing, counterregulation, and energy homeostasis. Physiology (Bethesda) 22:241–251. References 41
  • 52. 2. MOBBS, C.V., F. ISODA, H. MAKIMURA, J.W. MASTAITIS, T. MIZUNO, I.W. SHU, K. YEN, and X.J. YANG. 2004. Impaired glucose signaling as a cause of obesity and the metabolic syndrome: the glucoadipo- static hypothesis. Physiol Behav 19:2–23. 3. BERTHOUD, H.R., and T.L. POWLEY. 1990. Identification of vagal preganglionics that mediate cephalic phase insulin response. Am J Physiol Regul Integr Comp Physiol 258:R523–R530. 4. BERTHOUD, H.R., D.A. BEREITER, E.R. TRIMBLE, E.G. SIEGEL, and B. JEANRENAUD. 1981. Cephalic phase, reflex insulin secretion. Neuroanatomical and physiological characterization. Diabetologia 20 (Suppl):393–401. 5. AHREN, B. 2000. Autonomic regulation ofislet hormone secretion: implicationsfor health and disease. Diabetologia 43:393–410. 6. REIMANN, F., A.M. HABIB, G. TOLHURST, H.E. PARKER, G.J. ROGERS, and F.M. GRIBBLE. 2008. Glucose sensing in L cells: a primary cell study. Cell Metab 8:532–539. 7. EGAN, J.M., and R.F. MARGOLSKEE. 2008. Taste cells of the gut and gastrointestinal chemosensation. Mol Interv 8:78–81. 8. JANG, H.J., Z. KOKRASHVILI, M.J. THEODORAKIS, O.D. CARLSON, B.J. KIM, J. ZHOU, H.H. KIM, X. XU, S.L. CHAN, M. JUHASZOVA, M. BERNIER, B. MOSINGER, R.F. MARGOLSKEE, and J.M. EGAN. 2007. Gut- expressed gustducin and taste receptors regulate secretion of glucagon-like peptide-1. Proc Natl Acad Sci USA 104:15069–15074. 9. MARGOLSKEE, R.F., J. DYER, Z. KOKRASHVILI, K.S. SALMON, E. ILEGEMS, K. DALY, E.L. MAILLET, Y. NINOMIYA, B. MOSINGER, and S.P. SHIRAZI-BEECHEY. 2007. T1R3 and gustducin in gut sense sugars to regulate expression of Naþ -glucose cotransporter 1. Proc Natl Acad Sci USA 104:15075–15080. 10. THORENS, B., and P.J. LARSEN. 2004. Gut-derived signaling molecules and vagal afferents in the control of glucose and energy homeostasis. Curr Opin Clin Nutr Metab Care 7:471–478. 11. MEI, N. 1978. Vagal glucoreceptors in the small intestine of the cat. J Physiol 282:485–506. 12. DIEZ-SAMPEDRO, A., B.A. HIRAYAMA, C. OSSWALD, V. GORBOULEV, K. BAUMGARTEN, C. VOLK, E.M. WRIGHT, and H. KOEPSELL. 2003. A glucose sensor hiding in a family of transporters. Proc Natl Acad Sci USA 100:11753–11758. 13. SHIMIZU, N., Y. OOMURA, D. NOVIN, C.V. GRIJALVA, and P.H. COOPER. 1983. Functional correlations between lateral hypothalamic glucose-sensitive neurons and hepatic portal glucose-sensitive units in rat. Brain Res 265:49–54. 14. ADACHI, A., N. SHIMIZU, Y. OOMURA, and M. KOBASHI. 1984. Convergence of hepatoportal glucose- sensitive afferent signals to glucose-sensitive units within the nucleus of the solitary tract. Neurosci Lett 46:215–218. 15. PREITNER, F., M. IBBERSON, I. FRANKLIN, C. BINNERT, M. PENDE, A. GJINOVCI, T. HANSOTIA, D.J. DRUCKER, C. WOLLHEIM, R. BURCELIN, and B. THORENS. 2004. Gluco-incretins control insulin secretion at multiple levels as revealed in mice lacking GLP-1 and GIP receptors. J Clin Invest 113:635–645. 16. BURCELIN, R., W. DOLCI, and B. THORENS. 2000. Portal glucose infusion in the mouse induces hypoglycemia: evidence that the hepatoportal glucose sensor stimulates glucose utilization. Diabetes 49:1635–1642. 17. CARABAZA, A., C.J. CIUDAD, S. BAQUE, and J.J. GUINOVART. 1992. Glucose has to be phosphorylated to activate glycogen synthase, but not to inactivate glycogen phosphorylase in hepatocytes. FEBS Lett 296:211–214. 18. DENTIN, R., P.D. DENECHAUD, F. BENHAMED, J. GIRARD, and C. POSTIC. 2006. Hepatic gene regulation by glucose and polyunsaturated fatty acids: a role for ChREBP. J Nutr 136:1145–1149. 19. KABASHIMA, T., T. KAWAGUCHI, B.E. WADZINSKI, and K. UYEDA. 2003. Xylulose 5-phosphate mediates glucose-induced lipogenesis by xylulose 5-phosphate-activated protein phosphatase in rat liver. Proc Natl Acad Sci USA 100:5107–5112. 20. THORENS, B. 2008. Glucose sensing and the pathogenesis of obesity and type 2 diabetes. Int J Obes (Lond) 32(Suppl 6):S62–S71. 21. MATSCHINSKY, F.M., B. GLASER, and M.A. MAGNUSON. 1998. Pancreatic beta-cell glucokinase: closing the gap between theoretical concepts and experimental realities. Diabetes 47:307–315. 42 Chapter 2 Central Glucose Sensing and Control of Food Intake
  • 53. 22. HENQUIN, J.C., M. GEMBAL, P. DETIMARY, Z.Y. GAO, C. WARNOTTE, and P. GILON. 1994. Multisite control of insulin release by glucose. Diabetes Metab 20:132–137. 23. GROMADA, J., I. FRANKLIN, and C.B. WOLLHEIM. 2007. Alpha-cells of the endocrine pancreas: 35 years of research but the enigma remains. Endocr Rev 28:84–116. 24. RORSMAN, P., S.A. SALEHI, F. ABDULKADER, M. BRAUN, and P.E. MACDONALD. 2008. K(ATP)-channels and glucose-regulated glucagon secretion. Trends Endocrinol Metab 19:277–284. 25. GUILLOD-MAXIMIN, E., A. LORSIGNOL, T. ALQUIER, and L. PENICAUD. 2004. Acute intracarotid glucose injection towards the brain induces specific c-fos activation in hypothalamic nuclei: involvement of astrocytes in cerebral glucose-sensing in rats. J Neuroendocrinol 16:464–471. 26. MARTY, N., M. DALLAPORTA, M. FORETZ, M. EMERY, D. TARUSSIO, I. BADY, C. BINNERT, F. BEERMANN, and B. THORENS. 2005. Regulation of glucagon secretion by glucose transporter type 2 (glut2) and astrocyte-dependent glucose sensors. J Clin Invest 115:3545–3553. 27. BERNARD, C. 1849. Chiens rendus diabetiques. CR Soc Biol 1:60. 28. MAYER, J. 1953. Glucostatic mechanism of regulation of food intake. N Engl J Med 249:13–16. 29. ANAND, B.K., G.S. CHHINA, K.N. SHARMA, S. DUA, and B. SINGH. 1964. Activity of single neurons in the hypothalamic feeding centers: effect of glucose. Am J Physiol 207:1146–1154. 30. OOMURA, Y., H. OOYAMA, M. SUGIMORI, T. NAKAMURA, and Y. YAMADA. 1974. Glucose inhibition of the glucose-sensitive neurone in the rat lateral hypothalamus. Nature 247:284–286. 31. ROUTH, V.H. 2002. Glucose-sensing neurons: are they physiologically relevant? Physiol Behav 76:403–413. 32. YANG, X.J., L.M. KOW, D.W. PFAFF, and C.V. MOBBS. 2004. Metabolic pathways that mediate inhibition of hypothalamic neurons by glucose. Diabetes 53:67–73. 33. BRISKI, K.P., and P.W. SYLVESTER. 2001. Co-distribution of Fos- and mu opioid receptor immuno- reactivitywithintheratseptopreopticarea andhypothalamus duringacuteglucosedeprivation:effects of the mu receptor antagonist CTOP. Neurosci Lett 306:141–144. 34. DUNN-MEYNELL, A.A., N.E. RAWSON, and B.E. LEVIN. 1998. Distribution and phenotype of neurons containing the ATP-sensitive Kþ channel in rat brain. Brain Res 814:41–54. 35. FIORAMONTI, X., A. LORSIGNOL, A. TAUPIGNON, and L. PENICAUD. 2004. A new ATP-sensitive Kþ channel-independent mechanism is involved in glucose-excited neurons of mouse arcuate nucleus. Diabetes 53:2767–2775. 36. RITTER, S., and T.T. DINH. 1994. 2-Mercaptoacetate and 2-deoxy-D-glucose induce Fos-like immu- noreactivity in rat brain. Brain Res 641:111–120. 37. SILVER, I.A., and M. ERECINSKA. 1998. Glucose-induced intracellular ion changes in sugar-sensitive hypothalamic neurons. J Neurophysiol 79:1733–1745. 38. WANG, R., X. LIU, S.T. HENTGES, A.A. DUNN-MEYNELL, B.E. LEVIN, W. WANG, and V.H. ROUTH. 2004. The regulation of glucose-excited neurons in the hypothalamic arcuate nucleus by glucose and feeding-relevant peptides. Diabetes 53:1959–1965. 39. DALLAPORTA, M., T. HIMMI, J. PERRIN, and J.C. ORSINI. 1999. Solitary tract nucleus sensitivity to moderate changes in glucose level. Neuroreport 10:2657–2660. 40. MIZUNO, Y., and Y. OOMURA. 1984. Glucose responding neurons in the nucleus tractus solitarius of the rat: in vitro study. Brain Res 307:109–116. 41. YETTEFTI, K., J.C. ORSINI, T. el OUAZZANI, T. HIMMI, A. BOYER, and J. PERRIN. 1995. Sensitivity of nucleus tractus solitarius neurons to induced moderate hyperglycemia, with special reference to catecholaminergic regions. J Auton Nerv Syst 51:191–197. 42. PENICAUD, L., C. LELOUP, X. FIORAMONTI, A. LORSIGNOL, and A. BENANI. 2006. Brain glucose sensing: a subtle mechanism. Curr Opin Clin Nutr Metab Care 9:458–462. 43. SCHUIT, F.C., P. HUYPENS, H. HEIMBERG, and D.G. PIPELEERS. 2001. Glucose sensing in pancreatic beta- cells: a model for the study of other glucose-regulated cells in gut, pancreas, and hypothalamus. Diabetes 50:1–11. 44. YANG, X.J., L.M. KOW, T. FUNABASHI, and C.V. MOBBS. 1999. Hypothalamic glucose sensor: similarities to and differences from pancreatic beta-cell mechanisms. Diabetes 48:1763–1772. 45. ASHFORD, M.L., P.R. BODEN, and J.M. TREHERNE. 1990. Glucose-induced excitation of hypothalamic neurones is mediated by ATP-sensitive Kþ channels. Pflugers Arch 415:479–483. References 43
  • 54. 46. MIKI, T., B. LISS, K. MINAMI, T. SHIUCHI, A. SARAYA, Y. KASHIMA, M. HORIUCHI, F. ASHCROFT, Y. MINOKOSHI, J. ROEPER, and S. SEINO. 2001. ATP-sensitive Kþ channels in the hypothalamus are essential for the maintenance of glucose homeostasis. Nat Neurosci 4:507–512. 47. AMOROSO, S., H. SCHMID-ANTOMARCHI, M. FOSSET, and M. LAZDUNSKI. 1990. Glucose, sulfonylureas, and neurotransmitter release: role of ATP-sensitive Kþ channels. Science 247:852–854. 48. MORIYAMA, R., H. TSUKAMURA, M. KINOSHITA, H. OKAZAKI, Y. KATO, and K. MAEDA. 2004. In vitro increase in intracellular calcium concentrations induced by low or high extracellular glucose levels in ependymocytes and serotonergic neurons of the rat lower brainstem. Endocrinology 145:2507–2515. 49. AINSCOW, E.K., S. MIRSHAMSI, T. TANG, M.L. ASHFORD, and G.A. RUTTER. 2002. Dynamic imaging of free cytosolic ATP concentration during fuel sensing by rat hypothalamic neurones: evidence for ATP-independent control of ATP-sensitive Kþ channels. J Physiol 544:429–445. 50. SONG, Z., and V.H. ROUTH. 2005. Differential effects of glucose and lactate on glucosensing neurons in the ventromedial hypothalamic nucleus. Diabetes 54:15–22. 51. KANG, L., V.H. ROUTH, E.V. KUZHIKANDATHIL, L.D. GASPERS, and B.E. LEVIN. 2004. Physiological and molecular characteristics of rat hypothalamic ventromedial nucleus glucosensing neurons. Diabetes 53:549–559. 52. O’MALLEY, D., F. REIMANN, A.K. SIMPSON, and F.M. GRIBBLE. 2006. Sodium-coupled glucose cotransporters contribute to hypothalamic glucose sensing. Diabetes 55:3381–3386. 53. GUYON, A., M.P. TARDY, C. ROVERE, J.L. NAHON, J. BARHANIN, and F. LESAGE. 2009. Glucose inhibition persists in hypothalamic neurons lacking tandem-pore Kþ channels. J Neurosci 29:2528–2533. 54. MULKEY, D.K., E.M. TALLEY, R.L. STORNETTA, A.R. SIEGEL, G.H. WEST, X. CHEN, N. SEN, A.M. MISTRY, P.G. GUYENET, and D.A. BAYLISS. 2007. TASK channels determine pH sensitivity in select respiratory neurons but do not contribute to central respiratory chemosensitivity. J Neurosci 27:14049–14058. 55. BURDAKOV, D., L.T. JENSEN, H. ALEXOPOULOS, R.H. WILLIAMS, I.M. FEARON, I. O’KELLY, O. GERASI- MENKO, L. FUGGER, and A. VERKHRATSKY. 2006. Tandem-poreKþ channels mediate inhibitionof orexin neurons by glucose. Neuron 50:711–722. 56. LOUIS-SYLVESTRE, J., and J. LE MAGNEN. 1980. Fall in blood glucose level precedes meal onset in free-feeding rats. Neurosci Biobehav Rev 4(Suppl 1):13–15. 57. BERTHOUD, H.R., and G.J. MOGENSON. 1977. Ingestive behavior after intracerebral and intracerebro- ventricular infusions of glucose and 2-deoxy-D-glucose. Am J Physiol Regul Integr Comp Physiol 233:R127–R133. 58. MISELIS, R.R.,and A.N. EPSTEIN. 1975.Feeding inducedby intracerebroventricular 2-deoxy-D-glucose in the rat. Am J Physiol 229:1438–1447. 59. NOVIN, D., D.A. VANDERWEELE, and M. REZEK. 1973. Infusion of 2-deoxy-D-glucose into the hepatic- portal system causes eating: evidence for peripheral glucoreceptors. Science 181:858–860. 60. CUMMINGS, D.E., and M.W. SCHWARTZ. 2000. Melanocortins and body weight: a tale of two receptors. Nat Genet 26:8–9. 61. GALE, S.M., V.D. CASTRACANE, and C.S. MANTZOROS. 2004. Energy homeostasis, obesity and eating disorders: recent advances in endocrinology. J Nutr 134:295–298. 62. SCHWARTZ, M.W., S.C. WOODS, D. PORTE, R.J. SEELEY, and D.G. BASKIN. 2000. Central nervous system control of food intake. Nature 404:661–671. 63. SCHWARTZ, G.J. 2000. The role of gastrointestinal vagal afferents in the control of food intake: current prospects. Nutrition 16:866–873. 64. WOODS, S.C., R.J. SEELEY, D. PORTE, JR., and M.W. SCHWARTZ. 1998. Signals that regulate food intake and energy homeostasis. Science 280:1378–1383. 65. SCHWARTZ, M.W., S.C. WOODS, D. PORTE, JR., R.J. SEELEY, and D.G. BASKIN. 2000. Central nervous system control of food intake. Nature 404:661–671. 66. BERTHOUD, H.R. 2002. Multiple neural systems controlling food intake and body weight. Neurosci Biobehav Rev 26:393–428. 67. ADAMANTIDIS, A., and L. de LECEA. 2008. Sleep and metabolism: shared circuits, new connections. Trends Endocrinol Metab 19:362–370. 68. SAKURAI, T. 2007. The neural circuit of orexin (hypocretin): maintaining sleep and wakefulness. Nat Rev Neurosci 8:171–181. 44 Chapter 2 Central Glucose Sensing and Control of Food Intake
  • 55. 69. RITTER, R.C., P.G.SLUSSER,and S. STONE. 1981.Glucoreceptors controllingfeedingand bloodglucose: location in the hindbrain. Science 213:451–452. 70. RITTER, S., T.T. DINH, and Y. ZHANG. 2000. Localization of hindbrain glucoreceptive sites controlling food intake and blood glucose. Brain Res 856:37–47. 71. YETTEFTI, K., J.C. ORSINI, and J. PERRIN. 1997. Characteristics of glycemia-sensitive neurons in the nucleus tractus solitarii: possible involvement in nutritional regulation. Physiol Behav 61:93–100. 72. RITTER, S., I. LLEWELLYN-SMITH, and T.T. DINH. 1998. Subgroups of hindbrain catecholamine neurons are selectively activated by 2-deoxy-D-glucose induced metabolic challenge. Brain Res 805:41–54. 73. FRALEY, G.S., and S. RITTER. 2003. Immunolesion of norepinephrine and epinephrine afferents to medial hypothalamus alters basal and 2-deoxy-D-glucose-induced neuropeptide Y and agouti gene- related protein messenger ribonucleic acid expression in the arcuate nucleus. Endocrinology 144:75–83. 74. RITTER, S., K. BUGARITH, and T.T. DINH. 2001. Immunotoxic destruction of distinct catecholamine subgroups produces selective impairment of glucoregulatory responses and neuronal activation. J Comp Neurol 432:197–216. 75. PERSEGHIN, G., E. REGALIA, A. BATTEZZATI, S. VERGANI, A. PULVIRENTI, I. TERRUZZI, D. BARATTI, F. BOZZETTI, V. MAZZAFERRO, and L. LUZI. 1997. Regulation of glucose homeostasis in humans with denervated livers. J Clin Invest 100:931–941. 76. CRYER, P.E. 2004. Diverse causes of hypoglycemia-associated autonomic failure in diabetes. N Engl J Med 350:2272–2279. 77. MITRAKOU, A., C. RYAN, T. VENEMAN, M. MOKAN, T. JENSSEN, I. KISS, J. DURRANT, P. CRYER, and J. GERICH. 1991. Hierarchy of glycemic thresholds for counterregulatory hormone secretion, symptoms, and cerebral dysfunction. Am J Physiol Endocrinol Metab 260:E67–E74. 78. TABORSKY, G.J., JR., B. AHREN, and P.J. HAVEL. 1998. Autonomic mediation of glucagon secretion during hypoglycemia: implications for impaired alpha-cell responses in type 1 diabetes. Diabetes 47:995–1005. 79. FRIZZELL, R.T., E.M. JONES, S.N. DAVIS, D.W. BIGGERS, S.R. MYERS, C.C. CONNOLLY, D.W. NEAL, J.B. JASPAN, and A.D. CHERRINGTON. 1993. Counterregulation during hypoglycemia is directed by widespread brain regions. Diabetes 42:1253–1261. 80. BORG, W.P., R.S. SHERWIN, M.J. DURING, M.A. BORG, and G.I. SHULMAN. 1995. Local ventromedial hypothalamus glucopenia triggers counterregulatory hormone release. Diabetes 44:180–184. 81. BORG, M.A., R.S. SHERWIN, W.P. BORG, W.V. TAMBORLANE, and G.I. SHULMAN. 1997. Local ventro- medial hypothalamus glucose perfusion blocks counterregulation during systemic hypoglycemia in awake rats. J Clin Invest 99:361–365. 82. DIROCCO, R.J., and H.J. GRILL. 1979. The forebrain is not essential for sympathoadrenal hypergly- cemic response to glucoprivation. Science 204:1112–1114. 83. SCHEEPERS, A., H.G. JOOST, and A. SCHURMANN. 2004. The glucose transporter families SGLT and GLUT: molecular basis of normal and aberrant function. JPEN J Parenter Enteral Nutr 28:364–371. 84. ASANO, T., T. OGIHARA, H. KATAGIRI, H. SAKODA, H. ONO, M. FUJISHIRO, M. ANAI, H. KURIHARA, and Y. UCHIJIMA. 2004. Glucose transporter and Naþ /glucose cotransporter as molecular targets of anti- diabetic drugs. Curr Med Chem 11:2717–2724. 85. JOOST, H.G., and B. THORENS. 2001. The extended GLUT-family of sugar/polyol transport facilitators: nomenclature, sequence characteristics, and potential function of its novel members (review). Mol Membr Biol 18:247–256. 86. ULDRY, M., M. IBBERSON, M. HOSOKAWA, and B. THORENS. 2002. GLUT2 is a high affinity glucosamine transporter. FEBS Lett 524:199–203. 87. HEDIGER, M.A., Y. KANAI, G. YOU, and S. NUSSBERGER. 1995. Mammalian ion-coupled solute transporters. J Physiol 482:7S–17S. 88. SEIDNER, G., M.G. ALVAREZ, J.I. YEH, K.R. O’DRISCOLL, J. KLEPPER, T.S. STUMP, D. WANG, N.B. SPINNER, M.J. BIRNBAUM, and D.C. DE VIVO. 1998. GLUT-1 deficiency syndrome caused by haploinsufficiency of the blood–brain barrier hexose carrier. Nat Genet 18:188–191. 89. KAYANO, T., H. FUKUMOTO, R.L. EDDY, Y.S. FAN, M.G. BYERS, T.B. SHOWS, and G.I. BELL. 1988. Evidence for a family of human glucose transporter-like proteins. Sequence and gene localization of a protein expressed in fetal skeletal muscle and other tissues. J Biol Chem 263:15245–15248. References 45
  • 56. 90. SCHMIDT, S., M. RICHTER, D. MONTAG, T. SARTORIUS, V. GAWLIK, A.M. HENNIGE, S. SCHERNECK, H. HIMMELBAUER, S.Z. LUTZ, R. AUGUSTIN, R. KLUGE, P. RUTH, H.G. JOOST, and A. SCHURMANN. 2008. Neuronal functions, feeding behavior, and energy balance in Slc2a3þ /À mice. Am J Physiol Endocrinol Metab 295:E1084–E1094. 91. ZHAO, Y., C. FUNG, D. SHIN, B.C. SHIN, S. THAMOTHARAN, R. SANKAR, D. EHNINGER, A. SILVA, and S.U. DEVASKAR. 2009. Neuronal glucose transporter isoform 3 deficient mice demonstrate features of autism spectrum disorders. Mol Psychiatry 15:286–299. 92. IBBERSON, M., B.M. RIEDERER, M. ULDRY, B. GUHL, J. ROTH, and B. THORENS. 2002. Immunolocalization of GLUTX1 in the testis and to specific brain areas and vasopressin-containing neurons. Endocri- nology 143:276–284. 93. REAGAN, L.P., D.R. ROSELL, S.E. ALVES, E.K. HOSKIN, A.L. MCCALL, M.J. CHARRON, and B.S. MCEWEN. 2002. GLUT8 glucose transporter is localized to excitatory and inhibitory neurons in the rat hippocampus. Brain Res 932:129–134. 94. MEMBREZ, M., E. HUMMLER, F. BEERMANN, J.A. HAEFLIGER, R. SAVIOZ, T. PEDRAZZINI, and B. THORENS. 2006. GLUT8 is dispensable for embryonic development but influences hippocampal neurogenesis and heart function. Mol Cell Biol 26:4268–4276. 95. ARLUISON, M., M. QUIGNON, P. NGUYEN, B. THORENS, C. LELOUP, and L. PENICAUD. 2004. Distribution and anatomical localization of the glucose transporter 2 (GLUT2) in the adult rat brain: an immunohistochemical study. J Chem Neuroanat 28:117–136. 96. ARLUISON, M., M. QUIGNON, B. THORENS, C. LELOUP, and L. PENICAUD. 2004. Immunocytochemical localization of the glucosetransporter 2 (GLUT2) in the adult rat brain.II. Electron microscopicstudy. J Chem Neuroanat 28:137–146. 97. GARCIA, M.A., C. MILLAN, C. BALMACEDA-AGUILERA, T. CASTRO, P. PASTOR, H. MONTECINOS, K. REINICKE, F. ZUNIGA, J.C. VERA, S.A. ONATE, and F. NUALART. 2003. Hypothalamic ependymal-glial cells express the glucose transporter GLUT2, a protein involved in glucose sensing. J Neurochem 86:709–724. 98. LELOUP, C., M. OROSCO, P. SERRADAS, S. NICOLAIDIS, and L. PENICAUD. 1998. Specific inhibition of GLUT2 in arcuate nucleus by antisense oligonucleotides suppresses nervous control of insulin secretion. Brain Res Mol Brain Res 57:275–280. 99. MAEKAWA, F., Y. TOYODA, N. TORII, I. MIWA, R.C. THOMPSON, D.L. FOSTER, S. TSUKAHARA, H. TSUKAMURA, and K. MAEDA. 2000. Localization of glucokinase-like immunoreactivity in the rat lower brain stem: for possible location of brain glucose-sensing mechanisms. Endocrinology 141:375–384. 100. LELOUP, C., M. ARLUISON, N. LEPETIT, N. CARTIER, P. MARFAING-JALLAT, P. FERRE, and L. PENICAUD. 1994. Glucose transporter 2 (GLUT 2): expression in specific brain nuclei. Brain Res 638:221–226. 101. LI, B., X. XI, D.S. ROANE, D.H. RYAN, and R.J. MARTIN. 2003. Distribution of glucokinase, glucose transporter GLUT2, sulfonylurea receptor-1, glucagon-like peptide-1 receptor and neuropeptide Y messenger RNAs in rat brain by quantitative real time RT-PCR. Brain Res Mol Brain Res 113:139–142. 102. WAN, H.Z., M.G. HULSEY, and R.J. MARTIN. 1998. Intracerebroventricular administration of antisense oligodeoxynucleotide against GLUT2 glucose transporter mRNA reduces food intake, body weight change and glucoprivic feeding response in rats. J Nutr 128:287–291. 103. THORENS, B., M.T. GUILLAM, F. BEERMANN, R. BURCELIN, and M. JAQUET. 2000. Transgenic reexpression of GLUT1 or GLUT2 in pancreatic beta cells rescues GLUT2-null mice from early death and restores normal glucose-stimulated insulin secretion. J Biol Chem 275:23751–23758. 104. BADY, I., N. MARTY, M. DALLAPORTA, M. EMERY, J. GYGER, D. TARUSSIO, M. FORETZ, and B. THORENS. 2006. Evidence from glut2-null mice that glucose is a critical physiological regulator of feeding. Diabetes 55:988–995. 105. MARTY, N., I. BADY, and B. THORENS. 2006. Distinct classes of central GLUT2-dependent sensors control counterregulation and feeding. Diabetes 55:S108–S113. 106. HEDIGER, M.A., M.J. COADY, T.S. IKEDA, and E.M. WRIGHT. 1987. Expression cloning and cDNA sequencing of the Naþ /glucose co-transporter. Nature 330:379–381. 107. WRIGHT, E.M. 2001. Renal Naþ -glucose cotransporters. Am J Physiol Renal Physiol 280:F10–F18. 46 Chapter 2 Central Glucose Sensing and Control of Food Intake
  • 57. 108. GRIBBLE, F.M., L. WILLIAMS, A.K. SIMPSON, and F. REIMANN. 2003. A novel glucose-sensing mechanism contributing to glucagon-like peptide-1 secretion from the GLUTag cell line. Diabetes 52:1147–1154. 109. DIEZ-SAMPEDRO, A., M.P. LOSTAO, E.M. WRIGHT, and B.A. HIRAYAMA. 2000. Glycoside binding and translocation in Naþ -dependent glucose cotransporters: comparison of SGLT1 and SGLT3. J Membr Biol 176:111–117. 110. BRISKI, K.P., and E.S. MARSHALL. 2001. Induction of ependymal, glial, and neuronal transactivation by intraventricular administration of the SGLT1 Naþ -D-glucose cotransporter inhibitor phlorizin. Neurochem Res 26:783–792. 111. DRUCKER, D.J., T. JIN, S.L. ASA, T.A. YOUNG, and P.L. BRUBAKER. 1994. Activation of proglucagon gene transcription by protein kinase-A in a novel mouse enteroendocrine cell line. Mol Endocrinol 8:1646–1655. 112. REIMANN, F., and F.M. GRIBBLE. 2002. Glucose-sensing in glucagon-like peptide-1-secreting cells. Diabetes 51:2757–2763. 113. RITZEL, U.,A.FROMME, M. OTTLEBEN,U.LEONHARDT,and G.RAMADORI. 1997.Release ofglucagon-like peptide-1 (GLP-1) by carbohydrates in the perfused rat ileum. Acta Diabetol 34:18–21. 114. SHIMA, K., T. SUDA, K. NISHIMOTO, and S. YOSHIMOTO. 1990. Relationship between molecular structures of sugars and their ability to stimulate the release of glucagon-like peptide-1 from canine ileal loops. Acta Endocrinol (Copenh) 123:464–470. 115. TSUJII, S., and G.A. BRAY. 1990. Effects of glucose, 2-deoxyglucose, phlorizin, and insulin on food intake of lean and fatty rats. Am J Physiol Endocrinol Metab 258:E476–E481. 116. KONG, C.T., S.F. YET, and J.E. LEVER. 1993. Cloning and expression of a mammalian Naþ /amino acid cotransporter with sequence similarity to Naþ /glucose cotransporters. J Biol Chem 268:1509–1512. 117. LYNCH, R.M., L.S. TOMPKINS, H.L. BROOKS, A.A. DUNN-MEYNELL, and B.E. LEVIN. 2000. Localization of glucokinase gene expression in the rat brain. Diabetes 49:693–700. 118. BALFOUR, R.H., A.M. HANSEN, and S. TRAPP. 2006. Neuronal responses to transient hypoglycaemia in the dorsal vagal complex of the rat brainstem. J Physiol 570:469–484. 119. DUNN-MEYNELL, A.A., V.H. ROUTH, L. KANG, L. GASPERS, and B.E. LEVIN. 2002. Glucokinase is the likely mediator of glucosensing in both glucose-excited and glucose-inhibited central neurons. Diabetes 51:2056–2065. 120. RITTER, S., J.M. MURNANE, and E.E. LADENHEIM. 1982. Glucoprivic feeding is impaired by lateral or fourth ventricular alloxan injection. Am J Physiol Regul Integr Comp Physiol 243:R312–R317. 121. LEVIN, B.E., T.C. BECKER, J. EIKI, B.B. ZHANG, and A.A. DUNN-MEYNELL. 2008. Ventromedial hypothalamic glucokinase is an important mediator of the counterregulatory response to insulin- induced hypoglycemia. Diabetes 57:1371–1379. 122. DUNN-MEYNELL, A.A., N.M. SANDERS, D. COMPTON, T.C. BECKER, J. EIKI, B.B. ZHANG, and B.E. LEVIN. 2009. Relationship among brain and blood glucose levels and spontaneous and glucoprivic feeding. J Neurosci 29:7015–7022. 123. MINOKOSHI, Y., T. ALQUIER, N. FURUKAWA, Y.B. KIM, A. LEE, B. XUE, J. MU, F. FOUFELLE, P. FERRE, M.J. BIRNBAUM, B.J. STUCK, and B.B. KAHN. 2004. AMP-kinase regulates food intake by responding to hormonal and nutrient signals in the hypothalamus. Nature 428:569–574. 124. HARDIE, D.G., J.W. SCOTT, D.A. PAN, and E.R. HUDSON. 2003. Management of cellular energy by the AMP-activated protein kinase system. FEBS Lett 546:113–120. 125. KAHN, B.B., T. ALQUIER, D. CARLING, and D.G. HARDIE. 2005. AMP-activated protein kinase: ancient energy gauge provides clues to modern understanding of metabolism. Cell Metab 1:15–25. 126. DA SILVA XAVIER, G., I. LECLERC, A. VARADI, T. TSUBOI, S.K. MOULE, and G.A. RUTTER. 2003. Role for AMP-activated protein kinase in glucose-stimulated insulin secretion and preproinsulin gene expression. Biochem J 371:761–774. 127. ANDERSSON, U., K. FILIPSSON, C.R. ABBOTT, A. WOODS, K. SMITH, S.R. BLOOM, D. CARLING, and C.J. SMALL. 2004. AMP-activated protein kinase plays a role in the control of food intake. J Biol Chem 279:12005–12008. 128. KIM, M.S., J.Y. PARK, C. NAMKOONG, P.G. JANG, J.W. RYU, H.S. SONG, J.Y. YUN, I.S. NAMGOONG, J. HA, I.S.PARK,I.K.LEE, B. VIOLLET,J.H. YOUN,H.K.LEE, andK.U.LEE. 2004.Anti-obesityeffectsofalpha- References 47
  • 58. lipoic acid mediated by suppression of hypothalamic AMP-activated protein kinase. Nat Med 10:727–733. 129. LEE, K., B. LI, X. XI, Y. SUH, and R.J. MARTIN. 2005. Role of neuronal energy status in the regulation of adenosine 50 -monophosphate-activated protein kinase, orexigenic neuropeptides expression, and feeding behavior. Endocrinology 146:3–10. 130. CLARET, M., M.A. SMITH, R.L. BATTERHAM, C. SELMAN, A.I. CHOUDHURY, L.G. FRYER, M. CLEMENTS, H. AL-QASSAB, H. HEFFRON, A.W. XU, J.R. SPEAKMAN, G.S. BARSH, B. VIOLLET, S. VAULONT, M.L. ASHFORD, D. CARLING, and D.J. WITHERS. 2007. AMPK is essential for energy homeostasis regulation and glucose sensing by POMC and AgRP neurons. J Clin Invest 117:2325–2336. 131. ALQUIER, T., J. KAWASHIMA, Y. TSUJI, and B.B. KAHN. 2007. Role of hypothalamic adenosine 50 -monophosphate-activated protein kinase in the impaired counterregulatory response induced by repetitive neuroglucopenia. Endocrinology 148:1367–1375. 132. MCCRIMMON, R.J., X. FAN, H. CHENG, E. MCNAY, O. CHAN, M. SHAW, Y. DING, W. ZHU, and R.S. SHERWIN. 2006. Activation of AMP-activated protein kinase within the ventromedial hypothalamus amplifies counterregulatory hormone responses in rats with defective counterregulation. Diabetes 55:1755–1760. 133. MCCRIMMON, R.J., X. FAN, Y. DING, W. ZHU, R.J. JACOB, and R.S. SHERWIN. 2004. Potential role for AMP-activated protein kinase in hypoglycemia sensing in the ventromedial hypothalamus. Diabetes 53:1953–1958. 134. HAN, S.M.,C. NAMKOONG, P.G.JANG, I.S. PARK, S.W.HONG, H. KATAKAMI,S.CHUN, S.W.KIM, J.Y. PARK, K.U. LEE, and M.S. KIM. 2005. Hypothalamic AMP-activated protein kinase mediates counter- regulatory responses to hypoglycaemia in rats. Diabetologia 48:2170–2178. 135. HAYES, M.R., K.P. SKIBICKA, K.K. BENCE, and H.J. GRILL. 2009. Dorsal hindbrain 50 -adenosine monophosphate-activated protein kinase as an intracellular mediator of energy balance. Endocri- nology 150:2175–2182. 136. WULLSCHLEGER, S., R. LOEWITH, and M.N. HALL. 2006. TOR signaling in growth and metabolism. Cell 124:471–484. 137. HARRIS, T.E., and J.C. LAWRENCE, JR., 2003. TOR signaling. Sci STKE 2003:re15. 138. HAY, N., and N. SONENBERG. 2004. Upstream and downstream of mTOR. Genes Dev 18:1926–1945. 139. KIM, D.H., D.D. SARBASSOV, S.M. ALI, J.E. KING, R.R. LATEK, H. ERDJUMENT-BROMAGE, P. TEMPST, and D.M. SABATINI. 2002. mTOR interacts with raptor to form a nutrient-sensitive complex that signals to the cell growth machinery. Cell 110:163–175. 140. SARBASSOV, D.D., S.M. ALI, D.H. KIM, D.A. GUERTIN, R.R. LATEK, H. ERDJUMENT-BROMAGE, P. TEMPST, and D.M. SABATINI. 2004. Rictor, a novel binding partner of mTOR, defines a rapamycin-insensitive and raptor-independent pathway that regulates the cytoskeleton. Curr Biol 14:1296–1302. 141. INOKI, K., M.N. CORRADETTI, and K.L. GUAN. 2005. Dysregulation of the TSC-mTOR pathway in human disease. Nat Genet 37:19–24. 142. MCDANIEL, M.L., C.A. MARSHALL, K.L. PAPPAN, and G. KWON. 2002. Metabolic and autocrine regulation of the mammalian target of rapamycin by pancreatic beta-cells. Diabetes 51:2877–2885. 143. XU, G., G. KWON, C.A. MARSHALL, T.A. LIN, J.C. LAWRENCE, JR., and M.L. MCDANIEL. 1998. Branched-chain amino acids are essential in the regulation of PHAS-I and p70 S6 kinase by pancreatic beta-cells. A possible role in protein translation and mitogenic signaling. J Biol Chem 273:28178–28184. 144. XU, G., C.A. MARSHALL, T.A. LIN, G. KWON, R.B. MUNIVENKATAPPA, J.R. HILL, J.C. LAWRENCE, JR., and M.L. MCDANIEL. 1998. Insulin mediates glucose-stimulated phosphorylation of PHAS-I by pancreatic beta cells. An insulin-receptor mechanismfor autoregulationof protein synthesisby translation. J Biol Chem 273:4485–4491. 145. COTA, D., K. PROULX, K.A. SMITH, S.C. KOZMA, G. THOMAS, S.C. WOODS, and R.J. SEELEY. 2006. Hypothalamic mTOR signaling regulates food intake. Science 312:927–930. 146. MORI, H., K. INOKI, H. MUNZBERG, D. OPLAND, M. FAOUZI, E.C. VILLANUEVA, T. IKENOUE, D. KWIATKOWSKI, O.A. MACDOUGALD, M.G. MYERS, JR., and K.L. GUAN. 2009. Critical role for hypothalamic mTOR activity in energy balance. Cell Metab 9:362–374. 147. INOKI, K., T. ZHU, and K.L. GUAN. 2003. TSC2 mediates cellular energy response to control cell growth and survival. Cell 115:577–590. 48 Chapter 2 Central Glucose Sensing and Control of Food Intake
  • 59. 148. RICHARD, D., S. CLAVEL, Q. HUANG, D. SANCHIS, and D. RICQUIER. 2001. Uncoupling protein 2 in the brain: distribution and function. Biochem Soc Trans 29:812–817. 149. CHAN, C.B., D. De LEO, J.W. JOSEPH, T.S. MCQUAID, X.F. HA, F. XU, R.G. TSUSHIMA, P.S. PENNEFATHER, A.M. SALAPATEK, and M.B. WHEELER. 2001. Increased uncoupling protein-2 levels in beta-cells are associated with impaired glucose-stimulated insulin secretion: mechanism of action. Diabetes 50:1302–1310. 150. CHAN, C.B., P.E. MACDONALD, M.C. SALEH, D.C. JOHNS, E. MARBAN, and M.B. WHEELER. 1999. Overexpression of uncoupling protein 2 inhibits glucose-stimulated insulin secretion from rat islets. Diabetes 48:1482–1486. 151. HONG, Y., B.D. FINK, J.S. DILLON, and W.I. SIVITZ. 2001. Effects of adenoviral overexpression of uncoupling protein-2 and -3 on mitochondrial respiration in insulinoma cells. Endocrinology 142:249–256. 152. ZHANG, C.Y., G. BAFFY, P. PERRET, S. KRAUSS, O. PERONI, D. GRUJIC, T. HAGEN, A.J. VIDAL-PUIG, O. BOSS, Y.B. KIM, X.X. ZHENG, M.B. WHEELER, G.I. SHULMAN, C.B. CHAN, and B.B. LOWELL. 2001. Uncoupling protein-2 negatively regulates insulin secretion and is a major link between obesity, beta cell dysfunction, and type 2 diabetes. Cell 105:745–755. 153. LOWELL, B.B., and G.I. SHULMAN. 2005. Mitochondrial dysfunction and type 2 diabetes. Science 307:384–387. 154. RICHARD, D., R. RIVEST, Q. HUANG, F. BOUILLAUD, D. SANCHIS, O. CHAMPIGNY, and D. RICQUIER. 1998. Distribution of the uncoupling protein 2 mRNA in the mouse brain. J Comp Neurol 397:549–560. 155. HORVATH, T.L., Z.B. ANDREWS, and S. DIANO. 2009. Fuel utilization by hypothalamic neurons: roles for ROS. Trends Endocrinol Metab 20:78–87. 156. PARTON, L.E., C.P. YE, R. COPPARI, P.J. ENRIORI, B. CHOI, C.Y. ZHANG, C. XU, C.R. VIANNA, N. BALTHASAR, C.E. LEE, J.K. ELMQUIST, M.A. COWLEY, and B.B. LOWELL. 2007. Glucose sensing by POMC neurons regulates glucose homeostasis and is impaired in obesity. Nature 449:228–232. 157. ANDREWS, Z.B., Z.W. LIU, N. WALLLINGFORD, D.M. ERION, E. BOROK, J.M. FRIEDMAN, M.H. TSCHOP, M. SHANABROUGH, G. CLINE, G.I. SHULMAN, A. COPPOLA, X.B. GAO, T.L. HORVATH, and S. DIANO. 2008. UCP2 mediates ghrelin’s action on NPY/AgRP neurons by lowering free radicals. Nature 454:846–851. 158. COPPOLA, A., Z.W. LIU, Z.B. ANDREWS, E. PARADIS, M.C. ROY, J.M. FRIEDMAN, D. RICQUIER, D. RICHARD, T.L. HORVATH, X.B. GAO, and S. DIANO. 2007. A central thermogenic-like mechanism in feeding regulation: an interplay between arcuate nucleus T3 and UCP2. Cell Metab 5:21–33. 159. ARSENIJEVIC, D., H. ONUMA, C. PECQUEUR, S. RAIMBAULT, B.S. MANNING, B. MIROUX, E. COUPLAN, M.C. ALVES-GUERRA, M. GOUBERN, R. SURWIT, F. BOUILLAUD, D. RICHARD, S. COLLINS, and D. RICQUIER. 2000. Disruption of the uncoupling protein-2 gene in mice reveals a role in immunity and reactive oxygen species production. Nat Genet 26:435–439. 160. BAI, Y., H. ONUMA, X. BAI, A.V. MEDVEDEV, M. MISUKONIS, J.B. WEINBERG, W. CAO, J. ROBIDOUX, L.M. FLOERING, K.W. DANIEL, and S. COLLINS. 2005. Persistent nuclear factor-kappa B activation in Ucp2À/À mice leads to enhanced nitric oxide and inflammatory cytokine production. J Biol Chem 280:19062–19069. 161. MOUKDAR, F., J. ROBIDOUX, O. LYGHT, J. PI, K.W. DANIEL, and S. COLLINS. 2009. Reduced antioxidant capacity and diet-induced atherosclerosis in uncoupling protein-2-deficient mice. J Lipid Res 50:59–70. 162. RHEE, S.G. 2006. Cell signaling. H2O2, a necessary evil for cell signaling. Science 312:1882–1883. 163. ARCHER, S.L., X.C. WU, B. THEBAUD, R. MOUDGIL, K. HASHIMOTO, and E.D. MICHELAKIS. 2004. O2 sensing in the human ductus arteriosus: redox-sensitive Kþ channels are regulated by mitochondria- derived hydrogen peroxide. Biol Chem 385:205–216. 164. PAN, Y., J. WENG, Y. CAO, R.C. BHOSLE, and M. ZHOU. 2008. Functional coupling between the Kv1.1 channel and aldoketoreductase Kvbeta1. J Biol Chem 283:8634–8642. 165. KRAFT, R., C. GRIMM, K. GROSSE, A. HOFFMANN, S. SAUERBRUCH, H. KETTENMANN, G. SCHULTZ, and C. HARTENECK. 2004. Hydrogen peroxide and ADP-ribose induce TRPM2-mediated calcium influx and cation currents in microglia. Am J Physiol Cell Physiol 286:C129–C137. References 49
  • 60. 166. TABET, F., C. SAVOIA, E.L. SCHIFFRIN, and R.M. TOUYZ. 2004. Differential calcium regulation by hydrogen peroxide and superoxide in vascular smooth muscle cells from spontaneously hypertensive rats. J Cardiovasc Pharmacol 44:200–208. 167. TODT, I., A. NGEZAHAYO, A. ERNST, and H.A. KOLB. 2001. Hydrogen peroxide inhibits gap junctional coupling and modulates intracellular free calcium in cochlear Hensen cells. J Membr Biol 181:107–114. 168. ARMANN, B., M.S. HANSON, E. HATCH, A. STEFFEN, and L.A. FERNANDEZ. 2007. Quantification of basal and stimulated ROS levels as predictors of islet potency and function. Am J Transplant 7:38–47. 169. BINDOKAS, V.P., A. KUZNETSOV, S. SREENAN, K.S. POLONSKY, M.W. ROE, and L.H. PHILIPSON. 2003. Visualizing superoxide production in normal and diabetic rat islets of Langerhans. J Biol Chem 278:9796–9801. 170. LELOUP, C., C. TOURREL-CUZIN, C. MAGNAN, M. KARACA, J. CASTEL, L. CARNEIRO, A.L. COLOMBANI, A. KTORZA, L. CASTEILLA, and L. PENICAUD. 2009. Mitochondrial reactive oxygen species are obligatory signals for glucose-induced insulin secretion. Diabetes 58:673–681. 171. MORGAN, D., E. REBELATO, F. ABDULKADER, M.F. GRACIANO, H.R. OLIVEIRA-EMILIO, A.E. HIRATA, M.S. ROCHA, S. BORDIN, R. CURI, and A.R. CARPINELLI. 2009. Association of NAD(P)H oxidase with glucose-induced insulin secretion by pancreatic beta-cells. Endocrinology 150:2197–2201. 172. PI, J., Y. BAI, K.W. DANIEL, D. LIU, O. LYGHT, D.EDELSTEIN, M. BROWNLEE, B.E. CORKEY, and S. COLLINS. 2009. Persistent oxidative stress due to absence of uncoupling protein 2 associated with impaired pancreatic beta-cell function. Endocrinology 150:3040–3048. 173. PI, J., Y. BAI, Q. ZHANG, V. WONG, L.M. FLOERING, K. DANIEL, J.M. REECE, J.T. DEENEY, M.E. ANDERSEN, B.E. CORKEY, and S. COLLINS. 2007. Reactive oxygen species as a signal in glucose-stimulated insulin secretion. Diabetes 56:1783–1791. 174. PI, J., Q. ZHANG, J. FU, C.G. WOODS, Y. HOU, B.E. CORKEY, S. COLLINS, and M.E. ANDERSEN. 2010. ROS signaling, oxidative stress and Nrf2 in pancreatic beta-cell function. Toxicol Appl Pharmacol 244:77–83. 175. LELOUP, C., C. MAGNAN, A. BENANI, E. BONNET, T. ALQUIER, G. OFFER, A. CARRIERE, A. PERIQUET, Y. FERNANDEZ, A. KTORZA, L. CASTEILLA, and L. PENICAUD. 2006. Mitochondrial reactive oxygen species are required for hypothalamic glucose sensing. Diabetes 55:2084–2090. 176. ASHCROFT, F.M., and F.M. GRIBBLE. 1999. ATP-sensitive Kþ channels and insulin secretion: their role in health and disease. Diabetologia 42:903–919. 177. KARSCHIN, C., C. ECKE, F.M. ASHCROFT, and A. KARSCHIN. 1997. Overlapping distribution of K(ATP) channel-forming Kir6.2 subunit and the sulfonylurea receptor SUR1 in rodent brain. FEBS Lett 401:59–64. 178. SHI, N.Q., B. YE, and J.C. MAKIELSKI. 2005. Function and distribution of the SUR isoforms and splice variants. J Mol Cell Cardiol 39:51–60. 179. THOMZIG, A., G. LAUBE, H. PRUSS, and R.W. VEH. 2005. Pore-forming subunits of K-ATP channels, Kir6.1 and Kir6.2, display prominent differences in regional and cellular distribution in the rat brain. J Comp Neurol 484:313–330. 180. INAGAKI, N., T. GONOI, J.P. T. CLEMENT, N. NAMBA, J. INAZAWA, G. GONZALEZ, L. AGUILAR-BRYAN, S. SEINO, and J. BRYAN. 1995. Reconstitution of IKATP: an inward rectifier subunit plus the sulfonylurea receptor. Science 270:1166–1170. 181. KARSCHIN, A., J. BROCKHAUS, and K. BALLANYI. 1998. KATP channel formation by the sulphonylurea receptors SUR1 with Kir6.2 subunits in rat dorsal vagal neurons in situ. J Physiol 509(Pt 2): 339–346. 182. DALLAPORTA,M., J.PERRIN,andJ.C.ORSINI.2000.Involvement ofadenosinetriphosphate-sensitiveKþ channels in glucose-sensing in the rat solitary tract nucleus. Neurosci Lett 278:77–80. 183. AGUILAR-BRYAN, L., and J. BRYAN. 1999. Molecular biology of adenosine triphosphate-sensitive potassium channels. Endocr Rev 20:101–135. 184. SEINO, S. 1999. ATP-sensitive potassium channels: a model of heteromultimeric potassium channel/ receptor assemblies. Annu Rev Physiol 61:337–362. 185. EVANS, M.L., R.J. MCCRIMMON, D.E. FLANAGAN, T. KESHAVARZ, X. FAN, E.C. MCNAY, R.J. JACOB, and R.S. SHERWIN. 2004. Hypothalamic ATP-sensitive Kþ channels play a key role in sensing 50 Chapter 2 Central Glucose Sensing and Control of Food Intake
  • 61. hypoglycemia and triggering counterregulatory epinephrine and glucagon responses. Diabetes 53:2542–2551. 186. MCCRIMMON, R.J., M.L. EVANS, X. FAN, E.C. MCNAY, O. CHAN, Y. DING, W. ZHU, D.X. GRAM, and R.S. SHERWIN. 2005. Activation of ATP-sensitive Kþ channels in the ventromedial hypothalamus amplifies counterregulatory hormone responses to hypoglycemia in normal and recurrently hypoglycemic rats. Diabetes 54:3169–3174. 187. MIRSHAMSI, S., H.A. LAIDLAW, K. NING, E. ANDERSON, L.A. BURGESS, A. GRAY, C. SUTHERLAND, and M.L. ASHFORD. 2004. Leptin and insulin stimulation of signalling pathways in arcuate nucleus neurones: PI3K dependent actin reorganization and KATP channel activation. BMC Neurosci 5:54. 188. SPANSWICK, D., M.A. SMITH, V.E. GROPPI, S.D. LOGAN, and M.L. ASHFORD. 1997. Leptin inhibits hypothalamic neurons by activation of ATP-sensitive potassium channels. Nature 390:521–525. 189. SPANSWICK, D., M.A. SMITH, S. MIRSHAMSI, V.H. ROUTH, and M.L. ASHFORD. 2000. Insulin activates ATP-sensitive Kþ channels in hypothalamic neurons of lean, but not obese rats. Nat Neurosci 3:757–758. 190. PUTNEY, J.W. 2005. Physiological mechanisms of TRPC activation. Pflugers Arch 451:29–34. 191. BALFOUR, R.H., and S. TRAPP 2007. Ionic currents underlying the response of rat dorsal vagal neurones to hypoglycaemia and chemical anoxia. J Physiol 579:691–702. 192. FIORAMONTI, X., S. CONTIE, Z. SONG, V.H. ROUTH, A. LORSIGNOL, and L. PENICAUD. 2007. Character- ization of glucosensing neuron subpopulations in the arcuate nucleus: integration in neuropeptide Y and pro-opio melanocortin networks? Diabetes 56:1219–1227. References 51
  • 62. Chapter 3 Abnormalities in Insulin Secretion in Type 2 Diabetes Mellitus TALY MEAS 1,2 AND PIERRE-JEAN GUILLAUSSEAU 1,2 1 APHP, Department of Internal Medicine B, Hoˆpital Lariboisiere, APHP Paris 2 Universite Paris 7, Paris, France INTRODUCTION According to the World Health Organization (WHO) and the American Diabetes Association (ADA), type 2 diabetes mellitus (T2DM) is defined as resulting from defects in both insulin secretion and insulin sensitivity. Since the discovery of plasma insulin radioimmunoassay by Salomon Berson and Rosalyn Yalow (1), evidence has been obtained that insulin secretion is severely impaired in T2DM. Numerous functional defects such as b-cell dysfunction and other pathological abnormalities have been described in T2DM patients. Functional alterations that lead to b-cell dysfunction, including abnormalities in the kinetics of insulin secretion and quanti- tative and qualitative defects, all progress with time. Pathological abnormalities then can at least in part explain functional alterations, which include b-cell loss and its progression and reduced b-cell mass. NORMAL GLUCOSE HOMEOSTASIS Normal glucose homeostasis represents the balance between glucose appearance in systemic circulation (endogenous or meal-derived) and tissue glucose uptake and Metabolic Syndrome: Underlying Mechanisms and Drug Therapies Edited by Minghan Wang Copyright Ó 2011 John Wiley Sons, Inc. 53
  • 63. utilization. This balance is tightly regulated and plasma glucose concentrations are maintained within a narrow range. Normal fasting and 2 h post-glucose load plasma glucose levels are defined as 60–110 mg/dL and 140 mg/dL, respectively. Glucose homeostasis is maintained by the highly coordinated interaction of three physiolog- ical processes: insulin secretion, tissue glucose uptake, and hepatic glucose produc- tion (HGP). In the fasting state, plasma glucose is almost exclusively provided by hepatic production via glycogenolysis and gluconeogenesis. Hepatic glucose appear- ance rate in the circulation is matched with tissue glucose uptake rate, which occurs mostly in tissues that require glucose such as the central nervous system. Fasting plasma glucose (FPG) concentration is maintained at constant levels by the matched regulation of glucose production and glucose uptake. Following a meal, rising plasma glucose levels promote hepatic glucose uptake and insulin-independent glucose disposal and stimulate the release and the production of insulin by pancreatic b-cells. Increased plasma insulin concentrations suppress HGP primarily by decreasing glycogenolysis and gluconeogenesis and increasing glucose disposal through stim- ulation of peripheral glucose uptake (mostly in the muscle). These responses minimize hyperglycemia and ensure the return of mealtime glycemic levels to pre-meal values (2–4). INSULIN SECRETION AND EFFECTS ON TARGET TISSUES Glucose is the primary regulator of pancreatic b-cells by direct stimulation of insulin secretion and by modulating the insulin response to gut hormones and neural factors released during nutrient consumption. Insulin, like many hormones, displays rapid variations in plasma concentrations with frequent secretory peaks (periodicity 5–10 min), and less frequent larger oscillations (periodicity 60–120 min) (5). Normal insulin secretion in response to intravenous glucose follows a two-phase pattern. The first phase of insulin secretion (early or acute secretion) is rapid and sharp, reaching a maximum at 3–5 min and lasting for approximately 10 min. It represents mainly the release of stored granules. Second phase (late secretion) is gradual and persists for as long as glucose levels remain elevated. It stems from both stored secretory granules and de novo insulin synthesis. Early phase of insulin secretion is pivotal in the transition from the fasting state to the fed state with several different functions: to suppress HGP (6, 7), to suppress lipolysis (7), and to cross the endothelial barrier to prepare target cells for insulin action (8). Liver is a pivotal site in glucose metabolism regulation, and is responsive to minute changes in portal plasma insulin and glucagon concentrations. Early in the absorptivestate duringor after a meal, first-phaseinsulin secretion exertsan inhibitory effect on HGP by suppressing glycogenolysis and gluconeogenesis rate. In the fasting state, liver is almost the exclusive source of plasma glucose and therefore the most important site of insulin-mediated basal glucose release. Insulin’s first effect on skeletal muscle occurs during the absorptive state. Muscle is the major site of insulin-mediated dietary glucose uptake through stimulation of the insulin-sensitiveglucose transport system.Insulinpromotes conversionof glucose 54 Chapter 3 Abnormalities in Insulin Secretion in Type 2 Diabetes Mellitus
  • 64. into glycogen by enhancing glycogen synthase activity through regulation of a cyclic adenosine monophosphate (cAMP)-mediated cascade. Free fatty acid (FFA) metabolism plays an important role in maintaining glucose homeostasis both in the postabsorptive and absorptive states. Adipose tissue is highly sensitive to insulin. The meal-stimulated increase in plasma insulin inhibits lipolysis and FFA release from adipose tissue. Physiologic elevation of FFAs enhances hepatic gluconeogenesis and inhibits glucose uptake and utilization in insulin-sensitive tissues. Therefore, insulin action on adipose tissue affects glucose metabolism in liver and muscle, and in the mealtime suppression of FFA release contributes to the increase in peripheral glucose uptake and utilization (9–11). MEASURING INSULIN SECRETION AND b-CELL FUNCTION Measurement of Insulin Secretion Insulin secretion is markedly influenced by the route of glucose administration. When glucose is administered via the gastrointestinal tract, a much greater stimulation of insulin secretion is observed compared with similar hyperglycemia created with intravenous glucose. The difference in insulin secretion between intravenous versus oral glucose administration is referred to as the incretin effect and is mediated by glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic peptide (GIP) (12). Insulin secretion in response to intravenous glucose also differs from oral glucose in its temporal pattern. Following glucose ingestion, there is a gradual rise in plasma glucose concentration reflecting the slow rate of glucose absorption, and this is accompanied by a gradual increase in plasma insulin. The abrupt rise in plasma glucose following intravenous administration causes a rapid and transient increase in plasma insulin concentration (first-phase insulin secretion), which lasts for 10 min. This is followed by a slower, sustained rise in plasma insulin (second-phase insulin secretion), which persists as long as plasma glucose remains elevated (13). The hyperglycemic clamp is considered thegold standard for measuring first- and second-phase insulin secretion. Intravenous glucose tolerance test (IVGTT) has been widely used to assess insulin secretion. The acute insulin response (AIR) (0–10 min) correlates with first-phase insulin response during the hyperglycemic clamp (14). A disadvantage of IVGTT is that the plasma glucose concentration declines rapidly following glucose injection, precluding any second-phase insulin secretory response measurement. Indexes of insulin secretion derived from oral glucose tolerance test (OGTT) provide an estimate of insulin secretion during the more physiological route of glucose administration. The insulinogenic index (increment in plasma insulin  increment in plasma glucose) during the first 30 min of the OGTT has been used widely in epidemiological studies as a surrogate measure of first-phase insulin secretion, although not extensively validated (15). In clinical studies, insulin secretion is evaluated by measuring plasma insulin or C-peptide response to oral or intravenous glucose. The amount of insulin secreted must be related to the increment in plasma glucose concentration, which provides the Measuring Insulin Secretion and b-Cell Function 55
  • 65. stimulus to b-cells. In normogluco-tolerant subjects, the amount of insulin secreted in response to glucose correlates inversely with peripheral insulin sensitivity (16). Reduced insulin sensitivity, through as-yet unidentified mechanisms, enhances plasma insulin response to any given glucose stimulus. Therefore, if one aims at comparing b-cell function between subjects with different insulin sensitivity, an insulin secretion/insulin resistance index (disposition index) should be used (17). The Hyperbolic Sensitivity–Secretion Relationship To understand the role of b-cells, it has been useful to elucidate the quantitative relationship between insulin sensitivity and insulin action as it exists in nondiabetic individuals. Some years ago, R. Bergman postulated that, if b-cell function was normal, the sensitivity-secretion relationship could be expressed more efficiently as a rectangular hyperbola (18). The product of insulin sensitivity and insulin secretory response (insulin sensitivity index  first-phase insulin response to glucose stimu- lation) would equal a constant, which was named the “disposition index (DI).” Based on a limited data set obtained in human volunteers, this author postulated that shifts in insulin sensitivity would be accompanied by compensatory alterations in b-cell sensitivity to glucose. The single parameter, DI, can thus be envisioned to predict the normal b-cell response adequate for any degree of insulin resistance. The DI is thus a measure of the ability of b-cells to compensate for insulin resistance. It can be considered a measure of pancreatic functionality in nondiabetic individuals (17). S. Kahn et al. were the first to confirm the hyperbolic relationship in a cohort of 96 nondiabetic subjects. They assessed the relationship between insulin response to intravenous stimuli and insulin sensitivity by quantifying these two variables in a large cohort of healthy subjects of less than 45 years of age (16). The nature of this relationship implies that the product of insulin sensitivity and insulin response is a constant for a given degree of glucose tolerance. This hyperbolic relationship exists whether the insulin response is examined following intravenous administration of glucose or nonglucose insulin secretagogues. Based on these analyses, it is apparent that the variations in insulin release in response to differences in insulin sensitivity are due to changes in the secretory capacity of b-cells rather than their sensitivity to glucose (19). Additional confirmations have emerged from studies with large cohorts (20). It appears that the proposed relationship provides a quantitative and convenient approach to expressing normal metabolic functionality in vivo. ALTERATIONS IN INSULIN SECRETION KINETICS IN T2DM Alterations in Pulsatile Insulin Release In nondiabetic subjects, when endogenous insulin secretion is experimentally abol- ished by somatostatin infusion, pulsatile insulin administration is more effective in controlling glycemia than its continuous administration (21). Moreover, in T1DM patients, pulsatile insulin administration compared to continuous subcutaneous 56 Chapter 3 Abnormalities in Insulin Secretion in Type 2 Diabetes Mellitus
  • 66. administration is associated with a 40% reduction in insulin doses needed to maintain normal glycemic control (22). The lower efficacy of continuous administration is related to the downregulation of insulin membrane receptors. Pulsatile insulin release is related to oscillations in intracellular Ca2 þ concentrations, which regulate exo- cytosis of insulin granules (5). Lack of oscillatory secretion can be caused by excessive intracellular Ca2 þ concentrations and may alter islet pattern (23). Prolonged exposure of islets to high Ca2 þ concentrations has been shown to be associated with apoptotic signals in b-cells (5). b-Cell “pace-maker” is severely altered in T2DM patients where reduction or absence of rapid secretory peaks is observed,andtheseabnormalitiesarepresentintheearlyphasesofthedisease(24–27). First-Phase Insulin Secretion in Initial Stages of T2DM or First- Degree Relatives At the time of diagnosis of T2DM, first-phase insulin secretion is abolished (9, 28–30), and the late phase is reduced and delayed. Reduction in first-phase insulinsecretion takes place early in thecourseof the disease,as ithas been reportedin subjects with impaired glucose tolerance (IGT) and IFG (31) as well as normogly- cemic first-degree relatives of patients with T2DM (32). The abolition of first-phase insulin secretion has been found not only in patients with overt T2DM but also at the initial stage of the disease such as IGT and impaired fasting glucose (IFG). The abolition of first-phase insulin secretion predicts further conversion of IGT or IFG to overt diabetes. Therefore, use offirst-phase insulin secretion as a marker of T2DM has been proposed by some researchers. The decrease in first-phase insulin secretion after intravenous glucose in patients with mild abnormalities of glucose tolerance has been reported well before IGT received its definition by the World Health Organiza- tion (33). Long-term follow-up studies of patients with IGT have demonstrated conversion from IGT to T2DM in more than 50% of the cases. Thus, IGT should be considered as an at-high-risk state for development of T2DM. Most of the studies performed in patients with IGT revealed abolition or decrease in first-phase insulin secretion (32–35). QUANTITATIVE AND QUALITATIVE ALTERATIONS IN INSULIN SECRETION In T2DM, a marked decrease in basal and glucose-stimulated plasma insulin concentration has been reported (36, 37) regardless of whether body mass index (BMI) is increased. Specific measurement of insulin and prohormones in T2DM patients by an immunoradiometric assay according to Hales coworkers (38) revealed true insulin deficiency. This defect is masked in T2DM patients by elevated circulating insulin propeptides equally represented by proinsulin 32–33 and 64–65. These peptides account for more than 40% of the circulating peptides compared to 5% in nondiabetic subjects (38, 39). Excess in proinsulin in T2DM is not a consequence of hyperstimulation of b-cells as it is devoid of the states of Quantitative and Qualitative Alterations in Insulin Secretion 57
  • 67. secondary hyperinsulinemia such as obesity (40) and liver diseases (41). It seems to indicate a diseased b-cell state rather than an altered functional state. PROGRESSION OF ABNORMALITIES OF INSULIN SECRETION Progression of Abnormalities of Insulin Secretion When Progressing from Initial Steps to Overt T2DM Longitudinal studies evaluating both early phase insulin secretion and insulin sensi- tivity have shown that defects in both functions can predict the development of overt diabetes (42, 43). Longitudinal studies have shown that the transition from normal glucose tolerance (NGT) to diabetes is associated with a progressive deterioration in earlyinsulinsecretion.PimaIndians,progressingtoIGTduringamean5.1yearfollow- up, were compared to subjects who remained NGT. Progression to IGT was accom- paniedbya27%reductioninAIR.Afurther51%reductioninAIRwasobservedduring progression from IGT to diabetes. Increases in body weight and a 31% decrease in insulinsensitivitywerealsoobservedinpatientsprogressingtodiabetes.Incontrast,in patients who remained NGT, while a similar decrease in insulin sensitivity was observed, AIR increased by 30%. This compensatory effect of insulin resistance by the b-cells explains the absence of progression of these subjects to diabetes. Similar conclusionscanbedrawnfromalong-term(7–9years)longitudinalstudyperformedin normoglycemic relatives of patients with T2DM (44). b-Cell function, evaluated by determiningDIvalues,decreasedby38%insubjectswhoprogressedfromNGTtoIGT but by only 20% in subjects who remained NGT. Progressionof Abnormalities of Insulin SecretioninOvertT2DM Over Time Worsening of insulin secretion deficiency with time is a characteristic of overt T2DM. This gradual reduction is evidenced by longitudinal studies of large cohorts (45, 46). Studies in the control group of the UKPDS indicated that residual insulin secretory capacity was decreased by 50% at the time of diagnosis of diabetes with a further decrease of 15% 6 years later (45). This decrease was linear at least during the 6 year follow-up period. If one extends the line toward the left as a way to hypothesize disease progression in the past, the actual beginning of the disease may have happened 10 years ago. This extrapolation is consistent with the results drawn from the retinal status at the time of T2DM diagnosis according to Harris et al. (47). If one extends the line toward the right as a way to predict potential progression in the future, the line crosses the abscissas axis 10–12 years after the date of T2DM diagnosis. Thus, these data suggest that the natural history of progressive b-cell death has a length of 20–25 years. Different mechanisms have been proposed to explain the progressive reduction in insulin secretion, including glucotoxicity (48), lipotoxicity (49), and the effects of advanced glycation end products (AGEs) (50, 51). The deposition of an islet amyloid substance, also known as amylin (52), may also play a role. 58 Chapter 3 Abnormalities in Insulin Secretion in Type 2 Diabetes Mellitus
  • 68. MECHANISMS OF b-CELL FAILURE Glucose-Stimulated Insulin Secretion and Glucotoxicity In pancreatic b-cells,glucose is transportedacross cytoplasmic membranevia specific transporters, glucose transporter 1 (GLUT1) and 2 (GLUT2), and is rapidly phos- phorylated by a specific glucokinase with a high Km for glucose. The combination of transport and phosphorylation determines metabolic flux through glycolysis in b-cells. Increased glycolytic flux in b-cells results in a rapid increase in the production of reducing equivalents and increased electron transfer to the mitochondrial matrix, leading to increased ATP production in mitochondria and increased ATP/ADP ratio in the cytoplasm. This in turn results in several sequential events: the closure of the ATP-sensitive Kþ (KATP) channels, depolarization of the cytoplasmic membrane, influx of extracellular Ca2 þ , a rapid increase in intracellular Ca2 þ , and activation of protein kinases, which then mediate exocytosis of insulin (53, 54). As glucose is the key physiological regulator of insulin secretion, it appears possible that it also regulates the long-term adaptation of insulin production by regulating b-cell turnover. However, it is important to stress that in human b-cells in vitro, graded increase in glucose from a physiological concentration of 5.6 to 11.2 mmol/L and above induces apoptosis. However, in rat islets, the same graded glucose increment decreases apoptosis, indicating that glucose affects the survival of islets differently in these species. This difference has created some confusion in the field (55). It also highlights the importance of genetic backgrounds in glucose sensitivity of the islets. Glucotoxicity of the islets can be defined as nonphysiological and potentially irreversible b-cell damage caused by chronic exposure to supraphy- siological glucose concentrations along with the characteristic decreases in insulin synthesis and secretion caused by decreased insulin gene expression (56). Gluco- toxicity, on the other hand, implies the gradual, time-dependent establishment of irreversible damage to cellular components of insulin production and consequently to insulin content and secretion (56). Glucose-induced apoptosis in b-cells is probably linked to the relative specificity of this toxicity toward b-cells but not to other islet or most nonislet cell types. The b-cell is extremely sensitiveto small changesin ambient glucose. When these changes are of short duration and lie within the physiological range, such as after a meal, they lead to insulin secretion. When changes are of longer duration and more pronounced in magnitude, they could be translated by the b-cell glucose-sensing pathways into proapoptotic signals (57, 58). Reactive Oxygen Species The toxic role of oxygen species, which are produced in excess in uncontrolled diabetes, is a pertinent explanation of the b-cell apoptosis (59). Long-term hyper- glycemia also induces the generation of reactive oxygen species (ROS), leading to chronic oxidative stress because the islets express very low levels of antioxidant enzymes and activity. In b-cells, hyperglycemia induces mitochondrial production of Mechanisms of b-Cell Failure 59
  • 69. superoxides that activates uncoupling protein 2 (UCP2), resulting in a decrease in intracellular ATP/ADP ratio and reduced glucose-stimulated insulin secretion (60). Diabetic islets are characterized by reduction in glucose-evoked insulin secretion, decreased cytosolic ATP and ATP/ADP ratio, abnormal hyperpolarization of the mitochondrial membrane, hyperexpression of UCP2 of complexes I and V of the respiratory chain, and high levels of a marker of oxidative stress, nitrotyrosine (61). These observations support the role of ROS in reduced b-cell function in T2DM. ROS,particularlyhydroxylradicals,interferewithnormalprocessingofthemRNA of pancreas duodenum homeobox-1 (PDX-1), a transcription factor required for insulin gene expression and glucose-induced insulin secretion as well a critical regulator of b-cell survival (55, 56). The generation of ROS and reactive nitrogen species ultimately activatesstress-inducedpathwayssuchasnuclearfactorkB(NF-kB),stresskinases,and hexosamines. Del Guerra et al., using islets isolated from the pancreas of patients with T2DM and matched nondiabetic controls, demonstrated that several functional and moleculardefectsarepresentinT2DMislets(62).HeconfirmedthatT2DMisletsrelease lessinsulinthancontrolislets.Thisperturbationisaccompaniedbyalteredexpressionof glucose transporters and glucokinase, reduced activation of AMP-activated protein kinase (AMPK) and alterations in some transcription factors regulating b-cell differ- entiationandfunction.Thelevelsofoxidativestressmarkers,suchasnitrotyrosineand8- hydroxy-20 -deoxyguanosine, were significantly higher in T2DM than in control islets, and correlated with the degree of impairment in glucose-stimulated insulin release. The additionofglutathioneintheincubationmediumcausedreductionofoxidativestress(as suggested by diminished levels of nitrotyrosine), improved glucose-stimulated insulin secretionandincreasedinsulinmRNAexpression(62).Thus,DelGuerraetal.concluded that the functional impairment of T2DM islets could be, at least in part, reversible by reducingisletcell oxidative stress (62). Itisimportant to emphasize thatin this study the percentage of b-cells was only slightly ($10%), although significantly, reduced in diabetic islets compared withcontrol islets (62). As proposed byRobertson et al. (56), if thesteadydeclineinb-cellfunctioninT2DMisattributableinanysignificantmannerto chronic oxidative stress-induced apoptosis but not deterioration in b-cell replication, interference with apoptosis by antioxidants or any other therapy might provide a much needed new treatment approach to stabilize b-cell. Excessive ROS not only damage cells directly by oxidizing DNA, protein, and lipids, but also indirectly by activating stress-sensitive intracellular signaling path- ways such as NF-kB, p38 MAPK, JNK/SAPK, hexosamine, and others. Activation of these pathways results in the increased expression of numerous gene products that may cause cellular damage and play a major role in the etiology of the late complications of diabetes. In addition, recent in vitro and in vivo data suggest that activation of the same or similar stress pathways results in insulin resistance and impaired insulin secretion (63). Lipotoxicity T2DM is associated with dyslipidemia characterized by an increase in circulating FFAs and changes in lipoprotein profile. Acute elevation of FFAs in healthy humans 60 Chapter 3 Abnormalities in Insulin Secretion in Type 2 Diabetes Mellitus
  • 70. induced hyperinsulinemia; there is also an increase in glucose-stimulated insulin secretion after prolonged FFA infusion (48 and 96 h) (64, 65), but not in nondiabetic individuals genetically predisposed to developing T2DM (65). In healthy control subjects, the FFA-induced insulin resistance was compensated by the enhanced insulin secretion, whereas persistently elevated FFAs may contribute to progressive b-cell failure (b-cell lipotoxicity) in individuals genetically predisposed to T2DM. Santomauro et al. (66) demonstrated that overnight administration of the nicotinic acid analogue acipimox lowered plasma FFAs as well as fasting insulin and glucose levels, reduced insulin resistance, and improved OGTT in lean and obese nondiabetic subjects and in subjects with IGTand T2DM. The significant decrease in insulin levels suggested that plasma FFAs support between 30 and 50% of basal insulin levels. A sustained (7 day) reduction in plasma FFA concentrations in T2DM with acipimox was also associated with enhanced insulin-stimulated glucose disposal (reduced insulin resistance), decreased content of intramyocellular long-chain fatty acyl metabolites, improvement in OGTT with a slight decrease in mean plasma insulin levels (67). These data suggest that physiological increases in plasma FFA concen- trations in humans enhance glucose-stimulated insulin secretion and are unlikely to be “lipotoxic” to b-cells (11) but may contribute to progressive b-cell failure in at least some individuals who are genetically predisposed to developing T2DM (65). For all studies reported in relation to the FFA–b-cell interaction, it is important to emphasize that the stimulatory effects on glucose-stimulated insulin secretion are physiological in nature, particularly during the fasted-to-fed transition. Circulating FFAs help maintain a basal rate of insulin secretion, keeping adipose tissue lipolysis in check. In rodent islets, increased FFAs have been shown to be proapoptotic in b-cells (68). Exposure of cultured human islets to saturated FFAs such as palmitate is highly toxic to b-cells, inducing b-cell apoptosis, decreased b-cell proliferation, and impaired b-cell function. In contrast, monounsaturated FFAs such as oleate are protective against both palmitate and glucose-induced apoptosis and induce b-cell proliferation. The deleterious effect of palmitic acid is mediated by ceramide-mitochondrial apoptotic pathways, whereas induction of the mitochondrial protein Bcl-2 by oleic acid may contribute to the protective effect of monounsaturated FFAs such as palmitoleic or oleic acids (69). The physiological or pathological significance of the effects of fatty acids and glucose on pancreatic b-cell function is a matter of debate. Although one can reasonably assert that fatty acid-induced b-cell death is clearly a toxic manifestation, their effects on functional parameters such as insulin secretion or gene expression are more difficult to categorize as either beneficial or deleterious responses in a short time frame, although they are clearly deleterious in the long run. Islet Cell Amyloid The relevance of amyloid deposition in the deterioration of b-cell function has been the subject of debate for many years. Deposits composed mainly of islet amyloid polypeptide (IAPP), also known as amylin, have been reported in up to 90% of T2DM Mechanisms of b-Cell Failure 61
  • 71. individuals compared with 10–13% of nondiabetic counterparts (70). IAPP is a 37-amino acid b-cell peptide that is costored and coreleased with insulin from b-cells in response to insulin secretagogues. Transgenic mice expressing human IAPP (hIAPP) in b-cells were obese and spontaneously developed diabetes characterized by islet amyloid deposition and decreased b-cell mass (71). Prospective studies in these mice support the hypothesis that the mechanism of the decreased b-cell mass is increased apoptosis (70). Alternatively, it is possible that IAPP formation is secondary to the onset of hyperglycemia and not of primary importance in the pathophysiology of T2DM (72). In a recently published review of islet amyloid (73), the authors concluded that in human T2DM, islet amyloidosis largely results from diabetes-related pathologies such as diabetes-associated abnormal proinsulin processing, which could contribute to the destabilization of granular IAPP, and therefore, it is not an etiological factor for hyperglycemia. REDUCTION IN b-CELL MASS IN T2DM The normal pancreas contains approximately 1 million islets of Langerhans, and each islet includes b-cells (60–80%), a-cells (20–30%), somatostatin secreting d-cells (5–15%), and pancreatic polypeptide secreting cells (PP-cells). As mentioned above, b-cell mass is regulated by apoptosis, hypo- and hyperplasia, replication and neogenesis (74, 75). In other words, regulation of b-cell mass is a dynamic process where the actual mass represents the net balance between replication, growth, and neogenesis on one side and necrosis/apoptosis on the other. The phenomenon is also known as b-cell plasticity and allows adaptation to changes in demand of b-cell function (76). Such process is disrupted in T2DM where functional defects and decreased b-cell mass coexist. Both impaired proliferation and increased apoptosis may contribute to the loss of b-cell mass. Increased apoptosis has been observed in Zucker diabetic fatty (ZDF) rats, an animal model of T2DM (77). In these animals, expansion of b-cell mass in response to insulin resistancewas shown to be inadequate. However, no defects in proliferation or neogenesis could be identified, suggesting that excessive rate of cell death by apoptosis could play a major role. It is difficult to distinguish the two mechanisms, cell formation and cell death, from each other in human tissue sections because dead cells are rapidly removed from the islet by macrophages and neighboring cells, making it hard to quantify cell death. Nonethe- less, apoptosis is currently believed to represent the main cause for loss of b-cell mass in T2DM. This view is supported by necropsy data where pancreatic tissues from T2DM patients were compared to those from nondiabetic subjects (72). Moreover, elevated activities of apoptotic mediators caspase-3 and -8 have been found in b-cells from islets of T2DM patients (78). Most studies addressing the issue of b-cell loss have concluded that there is a marked reduction in the number of b-cells in postmortem specimens of pancreas obtained at necropsy of T2DM patients. In contrast to the adaptive increases in b-cell mass observed in rodent models of obesity (79) and obese human subjects (80), 62 Chapter 3 Abnormalities in Insulin Secretion in Type 2 Diabetes Mellitus
  • 72. a marked reduction in b-cell mass in patients with T2DM has been reported by numerous groups. Recent data have provided new insights into islet pathology of T2DM and the mechanisms responsible for decreased b-cell mass (72). Pancreatic tissue samples from 124 autopsies have been examined with analysis stratified according to BMI (less or above 27 kg/m2 ). In this study, 91 obese (41 patients with T2DM, 15 subjects with IGT, and 35 nondiabetic subjects) and 33 lean subjects (16 patients with T2DM and 15 nondiabetic subjects) were included. Relative b-cell volume, frequency of b-cell apoptosis, b-cell replication, and neogenesis (new islet formation from exocrine ducts) were assessed. Compared to weight-matched con- trols, pancreas from overweight and lean T2DM patients presented with 63 and 41% deficits in relative b-cell volume, respectively. A similar decrease (41%) was observed in subjects with IGT. No difference was seen regardless of what previous T2DM treatments the patients received (diet, sulfonylureas, or insulin). Relative b-cell volume was increased in overweight patients compared to lean ones due to increased neogenesis. b-Cell replication was found to be low in all groups. Neogenesis, while increased in overweight patients, was not different between overweight T2DM patients and nondiabetic subjects, or between the lean T2DM patients and nondiabetic subjects. The most remarkable abnormality observed in islet samples from T2DM patients was increased b-cell apoptosis. Frequency of b-cell apoptosis was increased tenfold in normal weight patients and threefold in overweight patients compared to respective control groups. In this study, islet amyloid was present only in a minority of cases, around 10%, of patients with T2DM or IGT. There are two potential explanations for these results either small islet amyloid pancreatic peptide oligomers (nondetectable by light microscopy) are present and responsible for b-cell loss, or islet amyloid is not crucial in the pathogenesis of T2DM. The authors concluded that b-cell mass is decreased in T2DM due to increased b-cell apoptosis. A confirmation has been provided by recent in vitro data, indicating an increased rate of apoptosis in islets exposed to high glucose concentrations (81). Another recent study (82) quantified the b-cell mass in pancreas obtained at autopsy of 57 T2DM and 52 nondiabetic subjects of European origin. Sections from the body and tail of pancreas were immunostained for insulin. The b-cell mass was calculated from the volume density of b-cells (measured by point-counting meth- ods) and the weight of the pancreas. The pancreatic insulin concentration was measured in some of the subjects. The main findings of this study were that b-cell mass slightly increases with BMI in both nondiabetic and T2DM subjects. On average, b-cell mass is 35–39% lower in T2DM than in nondiabetic subjects and so is the concentration of pancreatic insulin; but the variations of individual values are large. b-Cell mass does not correlate with the age of T2DM subjects at diagnosis of the disease, but decreases throughout the duration of clinical diabetes. Whether the loss of b-cell mass precedes and precipitates the clinical onset of the disease remains uncertain. It is also unclear if it accounts alone for the defects in insulin secretion. Prospective noninvasive studies measuring b-cell mass, insulin secretion, and insulin action in the same individuals are necessary to unequivocally address these questions (83). Reduction in b-Cell Mass in T2DM 63
  • 73. LINKAGE OF REDUCED b-CELL MASS AND DYSFUNCTION The role of reduced b-cell mass in the alterations of insulin secretion that char- acterizes T2DM has not yet fully elucidated. Assessing the possible contribution of a low b-cell mass to the development of T2DM is not easy. Does one become diabetic when the number of properly functioning b-cells has decreased, or when b-cell function has deteriorated beyond a threshold level needed to maintain normal glucose homeostasis, or both? Longitudinal in vivo measurements of b-cell mass, b-cell function, and insulin action in the same individuals would be needed to unequivocally address the issue. Still, clinical observations and experimental data support a close interrelation- ship between the two parameters. A large proportion of liver-related pancreatic donors who underwent a 50% pancreatectomy developed diabetes (84). Pharmaco- logical or surgical reduction of b-cell mass in rodents results all in impaired insulin secretion (85). More recently, Matveyenko and Butler (86) carefully analyzed the effect of 50% pancreatectomy in normal dogs and showed that partial pancreatectomy resulted in IFG and IGT. Partial pancreas resection was associated with reduction of both basal and glucose-stimulated insulin secretion. Altogether, these data support a mechanistic role of reduced b-cell mass in the development of alterations in glucose homeostasis and progression toward T2DM. In conclusion, it seems that the major defect leading to decreased b-cell mass in T2DM is inappropriate apoptosis, while new islet formation and b-cell replication are normal. Therefore, therapeutic approaches designed to arrest apoptosis could have quite an impact on prevention and treatment of the disease. THE COMPENSATION OF INSULIN RESISTANCE BY b-CELLS In nondiabetic controls, b-cell adapts its secretion rate to the level required by insulin sensitivity so that plasma glucose concentrations remain normal. A hyperbolic relation has thus been observed between insulin secretion and sensitivity in nondi- abetic subjects (42). In uncomplicated obesity, insulin resistance is compensated by increased b-cell mass and insulin hypersecretion (19, 87). If compensation is absent or even incomplete, plasma glucose concentrations rise gradually defining the incipient stages IFG or IGTand then overt diabetes. Inability of the b-cell to adjust its secretion rate to increased insulin demand explains why glucose intolerance appears in the physiological setting of aging (88) and gestational diabetes. ORIGIN OF b-CELL DYSFUNCTION As indicated above, b-cell dysfunction is present at the early stages of the disease, that is, IFG or IGT, and in normoglycemic first-degree relatives of patients with T2DM (32, 89, 90). These results rule out the hypothesis of a hyperinsulinemic state preceding T2DM, which was evoked from findings using nonspecific insulin assays (over-estimating “true” insulin concentrations), or from pseudo-longitudinal 64 Chapter 3 Abnormalities in Insulin Secretion in Type 2 Diabetes Mellitus
  • 74. studies describing the “Starling curve of the pancreas.” But plasma glucose levels and insulin-sensitivity status were not taken into account in these studies. Impacts of Genetics and/or Environment Studies of genetic susceptibility for T2DM in families revealed high concordance for T2DM in monozygotic twins compared with a lower occurrence in heterozygotic ones (80–90% compared to 40–50%). There is also a high frequency of T2DM in subjects with family history of diabetes (50% if both parents affected; 25–30% if only one first- degree relative is affected). In monogenic subtypes of diabetes (MODY or MIDD), insulin deficiency is predominant. However, these subtypes represent only a minority of T2DM. In the recent years, multiple loci associated with T2DM have been discovered by gene candidate and/or genome scan strategies in large cohorts of patients and relatives (91). Presently, 18 susceptibility variants for T2DM have been described (92, 93), but their individual weight in the development of the disease is weak. Relative risk for developing T2DM in association with these variants ranges from 1.06 for ADAMTS9 to 1.37 for TCF7L2 (91). In a study aimed at evaluating the risk associated with the 18 loci in a large cohort of Scandinavian subjects with a mean follow-up period of 23.5 years, variants in 11 genes were associated with T2DM. These genes include TCF7L2 (Transcription factor 7-like 2), PPARG (peroxisome proliferator-activated receptor g), FTO (fat mass and obesity), KCNJ11 (ATP- sensitive Kþ channel), NOTCH2 [Notch homolog 2 (Drosophila, WFS1 for Wolfram syndrome 1; wolframin)], CDKAL1 (CDK5 regulatory subunit associated protein 1- like 1), IGF2BP2 (insulin-like growth factor 2 mRNA binding protein 2), SLC30A8 [solute carrier family 30 (zinc transporter) member 8], JAZF1 (JAZF zinc finger 1), and HHEX (hematopoietically expressed homeobox) (94). Among these 11 variants, 8 were associated with alterations in insulin secretion. Recently, a major type 2 diabetes susceptibility gene, TCF7L2, which accounts for 20% of all cases, was identified by Grant et al. in Icelanders (95). Studies conducted in European Caucasian, Asian Indian, and Afro-Caribbean populations of both sexes have confirmed the ubiquitous distribution of the association (96). TCF7L2 is associated with alterations in insulin secretion. Genotype–phenotype relationship studies disclosed severely impaired insulin secretion in carriers of T2DM suscep- tibility variants (97). Nongenetic factors, particularly insufficient supply of nutrients during fetal development and the first years of life, may also be involved in a defective development of the islets. This defect may result in a reduced b-cell mass, and/or a reduction in the ability to compensate when insulin resistance is present in cases of pregnancy, overweight or obesity, low physical exercise levels, and aging. In this respect, Hales et al. have shown that subjects with birth weight in the lowest quintiles are more prone to IGTand T2DM in adulthood (98). Barker coworkers proposed that T2DM associated with a low birth weight could be the consequence of impaired b-cell function. This may result from in utero undernutrition during a critical period of fetal life and lead to abnormal development of the endocrine pancreas. This hypothesis has Origin of b-Cell Dysfunction 65
  • 75. been supported by studies using animal models (99). If rodents are subjected to an overall reduction in maternal food intake (50% of the normal daily ration) during the last week of pregnancy and throughout the lactation, the offspring showed intra utero growth restriction. They were born with a reduced b-cell proliferation rate. Moreover, these alterations have consequences during adulthood. Inadequate pancreatic func- tions can develop in situations of increased insulin demand such as ageing or pregnancy. Further, fetal (or in utero) programming is associated with deterioration in glucose tolerance, insulinopenia, and b-cell mass reduction (100, 101). In humans, Barker reported that low birth weight was associated with defective insulin secretion in 21 year old adults during OGTT (102). But these data are not confirmed by other groups. A pathological study has shown that small gestational age does not alter fetal pancreas development and morphology in comparison to appropriate growth for gestational age (103). In a case study comparing young adults born SGA or appropriate for gestational age, subjects born SGA did not demonstrate any evidence of impairment of either the first- or the second-phase insulin secretion (104). Using another model, Flanagan et al. reached the same conclusion in a different adult cohort (103). In 8 year old Indian children, low birth weight is associated with insulin resistance without abnormality of insulin secretion (105). CONCLUSIONS Substantial evidence supports the view that T2DM is a heterogeneous disorder. There is a progressive deterioration in b-cell function over time in T2DM. The UKPDS indicated that pancreatic islet function has been found to be at about 50% of normal capacity at the time of T2DM diagnosis regardless of the degree of insulin resistance. The decline of b-cell function is the limited capacity to compensate for insulin resistance. Both insulin resistance and b-cell dysfunction are usually present in classical T2DM as well as most individuals with IGT. The defect of insulin secretion in T2DM is related to two confounding components, insulin deficiency and b-cell secretory defect. On the other hand, there is an impaired glucose sensing in the b-cells. The reduction of b-cell mass is attributable to accelerated b-cell apoptosis. Identi- fication of the factors conferring susceptibility (mainly genetic) and those that may accelerate such process (mainly environmental and metabolic) represents a major imperative, which could lead to new therapeutic strategies of slowing if not arresting b-cell loss, thus contributing to more robust glycemic control and reduction of the risk of developing long-term diabetic complications. Although difficult, with the new understanding of b-cell biology, it seems that we may be a little bit closer to a solution. REFERENCES 1. YALOW, R.S., and S.A. BERSON. 1960. Immunoassay of endogenous plasma insulin in man. J Clin Invest 39:1157–1175. 2. DINNEEN, S., J. GERICH, and R. RIZZA. 1992. Carbohydrate metabolism in non-insulin-dependent diabetes mellitus. N Engl J Med 327:707–713. 66 Chapter 3 Abnormalities in Insulin Secretion in Type 2 Diabetes Mellitus
  • 76. 3. PORTE,D.,Jr.,2001.Clinicalimportanceofinsulinsecretionanditsinteractionwithinsulinresistancein the treatment of type 2 diabetes mellitus and its complications. Diabetes Metab Res Rev 17:181–188. 4. DEFRONZO, R.A. 1997. Insulin resistance: a multifaceted syndrome responsible for NIDDM, obesity, hypertension, dyslipidaemia and atherosclerosis. Neth J Med 50:191–197. 5. BERGSTEN, P. 2000. Pathophysiology of impaired pulsatile insulin release. Diabetes Metab Res Rev 16:179–191. 6. LUZI, L., and R.A. DEFRONZO. 1989. Effect of loss of first-phase insulin secretion on hepatic glucose production and tissue glucose disposal in humans. Am J Physiol 257:E241–E246. 7. REBRIN, K., G.M. STEIL, S.D. MITTELMAN, and R.N. BERGMAN. 1996. Causal linkage between insulin suppression of lipolysis and suppression of liver glucose output in dogs. J Clin Invest 98:741–749. 8. GETTY, L., M. HAMILTON-WESSLER, M. ADER, M.K. DEA, and R.N. BERGMAN. 1998. Biphasic insulin secretion during intravenous glucose tolerance test promotes optimal interstitial insulin profile. Diabetes 47:1941–1947. 9. PFEIFER, M.A., J.B. HALTER, and D. PORTE, Jr., 1981. Insulin secretion in diabetes mellitus. Am J Med 70:579–588. 10. PORTE, D., Jr., and A.A. PUPO. 1969. Insulin responses to glucose: evidence for a two pool system in man. J Clin Invest 48:2309–2319. 11. BODEN, G., and G.I. SHULMAN. 2002. Free fatty acids in obesity and type 2 diabetes: defining their role in the development of insulin resistance and beta-cell dysfunction. Eur J Clin Invest 32(Suppl 3):14–23. 12. RASK, E., T. OLSSON, S. SODERBERG, J. HOLST JJ, A. TURA, G. PACINI, and B. AHREN. 2004. Insulin secretion and incretin hormones after oral glucose in non-obese subjects with impaired glucose tolerance. Metabolism 53:624–631. 13. DEFRONZO, R.A.,J.D.TOBIN, andR. ANDRES. 1979.Glucoseclamptechnique: a methodforquantifying insulin secretion and resistance. Am J Physiol 237:E214–E223. 14. KORYTKOWSKI, M.T., S.L. BERGA, and M.J. HORWITZ. 1995. Comparison of the minimal model and the hyperglycemic clamp for measuring insulin sensitivity and acute insulin response to glucose. Metabolism 44:1121–1125. 15. PHILLIPS, D.I., P.M. CLARK, C.N. HALES, and C. OSMOND. 1994. Understanding oral glucose tolerance: comparison of glucose or insulin measurements during the oral glucose tolerance test with specific measurements of insulin resistance and insulin secretion. Diabet Med 11:286–292. 16. KAHN, S.E., R.L. PRIGEON, D.K. MCCULLOCH, E.J. BOYKO, R.N. BERGMAN, M.W. SCHWARTZ, J.L. NEIFING, W.K. WARD, J.C. BEARD, J.P. PALMER, et al. 1993. Quantification of the relationship between insulin sensitivity and beta-cell function in human subjects. Evidence for a hyperbolic function. Diabetes 42:1663–1672. 17. BERGMAN, R.N., M. ADER, K. HUECKING, and G. Van CITTERS. 2002. Accurate assessment of beta-cell function: the hyperbolic correction. Diabetes 51(Suppl 1):S212–S220. 18. BERGMAN, R.N., L.S. PHILLIPS, and C. COBELLI. 1981. Physiologic evaluation of factors controlling glucosetolerance in man: measurement ofinsulinsensitivity and beta-cellglucose sensitivity from the response to intravenous glucose. J Clin Invest 68:1456–1467. 19. KAHN, S.E. 2003. The relative contributions of insulin resistance and beta-cell dysfunction to the pathophysiology of Type 2 diabetes. Diabetologia 46:3–19. 20. WAGENKNECHT, L.E., E.J. MAYER, M. REWERS, S. HAFFNER, J. SELBY, G.M. BOROK, L. HENKIN, G. HOWARD, P.J. SAVAGE, M.F. SAAD, et al. 1995. The insulin resistance atherosclerosis study (IRAS) objectives, design, and recruitment results. Ann Epidemiol 5:464–472. 21. PAOLISSO, G., A.J. SCHEEN, D. GIUGLIANO, S. SGAMBATO, A. ALBERT, M. VARRICCHIO, F. D’ONOFRIO, and P.J. LEFEBVRE. 1991. Pulsatile insulin delivery has greater metabolic effects than continuous hormone administration in man: importance of pulse frequency. J Clin Endocrinol Metab 72:607–615. 22. BRATUSCH-MARRAIN, P.R., M. KOMJATI, and W.K. WALDHAUSL. 1986. Efficacy of pulsatile versus continuous insulin administration on hepatic glucose production and glucose utilization in type I diabetic humans. Diabetes 35:922–926. 23. TRUMP, B.F., and I.K. BEREZESKY. 1995. Calcium-mediated cell injury and cell death. FASEB J 9:219–228. References 67
  • 77. 24. POLONSKY, K.S., B.D. GIVEN, L.J. HIRSCH, H. TILLIL, E.T. SHAPIRO, C. BEEBE, B.H. FRANK, J.A. GALLOWAY, and E. Van CAUTER. 1988. Abnormal patterns of insulin secretion in non-insulin- dependent diabetes mellitus. N Engl J Med 318:1231–1239. 25. O’MEARA, N.M., J. STURIS, E. Van CAUTER, and K.S. POLONSKY. 1993. Lack of control by glucose of ultradian insulin secretory oscillations in impaired glucose tolerance and in non-insulin-dependent diabetes mellitus. J Clin Invest 92:262–271. 26. O’RAHILLY, S.P., Z. NUGENT, A.S. RUDENSKI, J.P. HOSKER, M.A. BURNETT, P. DARLING, and R.C. TURNER. 1986. Beta-cell dysfunction, rather than insulin insensitivity, is the primary defect in familial type 2 diabetes. Lancet 2:360–364. 27. O’RAHILLY, S., R.C. TURNER, and D.R. MATTHEWS. 1988. Impaired pulsatile secretion of insulin in relatives of patients with non-insulin-dependent diabetes. N Engl J Med 318:1225–1230. 28. CERASI, E., and R. LUFT. 1967. The plasma insulin response to glucose infusion in healthy subjects and in diabetes mellitus. Acta Endocrinol (Copenh) 55:278–304. 29. FUJITA, Y., A.R. CHACRA, A.L. HERRON, Jr., and H.S. SELTZER. 1975. Influence of antecedent carbohydrate intake on the biphasic insulin response to intravenous glucose. Diabetes 24:1072–1080. 30. METZ, S.A., J.B. HALTER, and R.P. ROBERTSON. 1979. Paradoxical inhibition of insulin secretion by glucose in human diabetes mellitus. J Clin Endocrinol Metab 48:827–835. 31. RATZMANN,K.P., B. SCHULZ, P.HEINKE,and D.MICHAELIS. 1981.Quantitativeandqualitativechanges in the early insulin response to glucose in subjects with impaired carbohydrate tolerance. Diabetes Care 4:85–91. 32. ERIKSSON, J., A. FRANSSILA-KALLUNKI, A. EKSTRAND, C. SALORANTA, E. WIDEN, C. SCHALIN, and L. GROOP. 1989. Early metabolic defects in persons at increased risk for non-insulin-dependent diabetes mellitus. N Engl J Med 321:337–343. 33. CERASI, E., R. LUFT, and S. EFENDIC. 1972. Decreased sensitivity of the pancreatic beta cells to glucose in prediabetic and diabetic subjects. A glucose dose-response study. Diabetes 21:224–234. 34. DAVIES, M.J., G. RAYMAN, A. GRENFELL, I.P. GRAY, J.L. DAY, and C.N. HALES. 1994. Loss of the first phase insulin response to intravenous glucose in subjects with persistent impaired glucose tolerance. Diabet Med 11:432–436. 35. YONEDA, H., H. IKEGAMI, Y. YAMAMOTO, E. YAMATO, T. CHA, Y. KAWAGUCHI, Y. TAHARA, and T. OGIHARA. 1992. Analysis of early-phase insulin responses in nonobese subjects with mild glucose intolerance. Diabetes Care 15:1517–1521. 36. TEMPLE, R.C., C.A. CARRINGTON, S.D. LUZIO, D.R. OWENS, A.E. SCHNEIDER, W.J. SOBEY, and C.N. HALES. 1989. Insulin deficiency in non-insulin-dependent diabetes. Lancet 1:293–295. 37. DAVIES, M.J., G. RAYMAN, I.P. GRAY, J.L. DAY, and C.N. HALES. 1993. Insulin deficiency and increased plasma concentration of intact and 32/33 split proinsulin in subjects with impaired glucose tolerance. Diabet Med 10:313–320. 38. SOBEY, W.J., S.F. BEER, C.A. CARRINGTON, P.M. CLARK, B.H. FRANK, I.P. GRAY, S.D. LUZIO, D.R. OWENS, A.E. SCHNEIDER, K. SIDDLE, et al. 1989. Sensitive and specific two-site immunoradio- metric assays for human insulin, proinsulin, 65–66 split and 32–33 split proinsulins. Biochem J 260:535–541. 39. TEMPLE, R.C., P.M. CLARK, D.K. NAGI, A.E. SCHNEIDER, J.S. YUDKIN, and C.N. HALES. 1990. Radioimmunoassay may overestimate insulin in non-insulin-dependent diabetics. Clin Endocrinol (Oxf) 32:689–693. 40. WANG, P.W., F. ABBASI, M. CARANTONI, Y.D. CHEN, S. AZHAR, and G.M. REAVEN. 1997. Insulin resistance does not change the ratio of proinsulin to insulin in normal volunteers. J Clin Endocrinol Metab 82:3221–3224. 41. KRUSZYNSKA, Y.T., D.S. HARRY, V. MOHAMED-ALI, P.D. HOME, J.S. YUDKIN, andN. MCINTYRE. 1995.The contribution of proinsulin and des-31,32 proinsulin to the hyperinsulinemia of diabetic and nondi- abetic cirrhotic patients. Metabolism 44:254–260. 42. WEYER, C., C. BOGARDUS, D.M. MOTT, and R.E. PRATLEY. 1999. The natural history of insulin secretory dysfunction and insulin resistance in the pathogenesis of type 2 diabetes mellitus. J Clin Invest 104:787–794. 68 Chapter 3 Abnormalities in Insulin Secretion in Type 2 Diabetes Mellitus
  • 78. 43. LUNDGREN, H., C. BENGTSSON, G. BLOHME, L. LAPIDUS, and J. WALDENSTROM. 1990. Fasting serum insulin concentration and early insulin response as risk determinants for developing diabetes. Diabet Med 7:407–413. 44. CNOP, M., J. VIDAL, R.L. HULL, K.M. UTZSCHNEIDER, D.B. CARR, T. SCHRAW, P.E. SCHERER, E.J. BOYKO, W.Y. FUJIMOTO, and S.E. KAHN. 2007. Progressive loss of beta-cell function leads to worsening glucose tolerance in first-degree relatives of subjects with type 2 diabetes. Diabetes Care 30:677–682. 45. U.K., prospective diabetes study, 16. 1995. Overview of 6 years’ therapy of type II diabetes: a progressive disease. U.K. Prospective Diabetes Study Group. Diabetes 44:1249–1258. 46. LEVY, J., A.B. ATKINSON, P.M. BELL, D.R. MCCANCE, and D.R. HADDEN. 1998. Beta-cell deterioration determines the onset and rate of progression of secondary dietary failure in type 2 diabetes mellitus: the 10-year follow-up of the Belfast Diet Study. Diabet Med 15:290–296. 47. HARRIS, M.I., R. KLEIN, T.A. WELBORN, and M.W. KNUIMAN. 1992. Onset of NIDDM occurs at least 4–7 yr before clinical diagnosis. Diabetes Care 15:815–819. 48. ROSSETTI, L., A. GIACCARI, and R.A. DEFRONZO. 1990. Glucose toxicity. Diabetes Care 13:610–630. 49. UNGER, R.H. 1995. Lipotoxicity in the pathogenesis of obesity-dependent NIDDM. Genetic and clinical implications. Diabetes 44:863–870. 50. OLSON, L.K., J.B. REDMON, H.C. TOWLE, and R.P. ROBERTSON. 1993. Chronic exposure of HIT cells to high glucose concentrations paradoxically decreases insulin gene transcription and alters binding of insulin gene regulatory protein. J Clin Invest 92:514–519. 51. MATSUOKA, T., Y. KAJIMOTO, H. WATADA, H. KANETO, M. KISHIMOTO, Y. UMAYAHARA, Y. FUJITANI, T. KAMADA, R. KAWAMORI, and Y. YAMASAKI. 1997. Glycation-dependent, reactive oxygen species- mediated suppression of the insulin gene promoter activity in HIT cells. J Clin Invest 99:144–150. 52. HOPPENER, J.W., B. AHREN, and C.J. LIPS. 2000. Islet amyloid and type 2 diabetes mellitus. N Engl J Med 343:411–419. 53. HABER, E.P., J. PROCOPIO, C.R. CARVALHO, A.R. CARPINELLI, P. NEWSHOLME, and R. CURI. 2006. New insights into fatty acid modulation of pancreatic beta-cell function. Int Rev Cytol 248:1–41. 54. NEWSHOLME, P., D. KEANE, H.J. WELTERS, and N.G. MORGAN. 2007. Life and death decisions of the pancreatic beta-cell: the role of fatty acids. Clin Sci (Lond) 112:27–42. 55. DONATH,M.Y.,J.A.EHSES,K.MAEDLER,D.M.SCHUMANN,H.ELLINGSGAARD,E.EPPLER,andM.REINECKE. 2005. Mechanisms of beta-cell death in type 2 diabetes. Diabetes 54(Suppl 2):S108–S113. 56. ROBERTSON, R.P., J. HARMON, P.O. TRAN, Y. TANAKA, and H. TAKAHASHI. 2003. Glucose toxicity in beta- cells: type 2 diabetes, good radicals gone bad, and the glutathione connection. Diabetes 52:581–587. 57. DONATH, M.Y., and P.A. HALBAN. 2004. Decreased beta-cell mass in diabetes: significance, mechan- isms and therapeutic implications. Diabetologia 47:581–589. 58. WAJCHENBERG, B.L. 2007. Beta-cell failure in diabetes and preservation by clinical treatment. Endocr Rev 28:187–218. 59. KRAUSS, S., C.Y. ZHANG, L. SCORRANO, L.T. DALGAARD, J. ST-PIERRE, S.T. GREY, and B.B. LOWELL. 2003. Superoxide-mediated activation of uncoupling protein 2 causes pancreatic beta cell dysfunction. J Clin Invest 112:1831–1842. 60. CHAN, C.B., M.C. SALEH, V. KOSHKIN, and M.B. WHEELER. 2004. Uncoupling protein 2 and islet function. Diabetes 53(Suppl 1):S136–S142. 61. ANELLO, M., R. LUPI, D. SPAMPINATO, S. PIRO, M. MASINI, U. BOGGI, S. Del PRATO, A.M. RABUAZZO, F. PURRELLO, and P. MARCHETTI. 2005. Functional and morphological alterations of mitochondria in pancreatic beta cells from type 2 diabetic patients. Diabetologia 48:282–289. 62. Del GUERRA, S., R. LUPI, L. MARSELLI, M. MASINI, M. BUGLIANI, S. SBRANA, S. TORRI, M. POLLERA, U. BOGGI, F. MOSCA, S. Del PRATO, and P. MARCHETTI. 2005. Functional and molecular defects of pancreatic islets in human type 2 diabetes. Diabetes 54:727–735. 63. VINCENT, A.M., M.J. STEVENS, C. BACKUS, L.L. MCLEAN, and E.L. FELDMAN. 2005. Cell culture modeling to test therapies against hyperglycemia-mediated oxidative stress and injury. Antioxid Redox Signal 7:1494–1506. 64. BODEN, G. 1998. Free fatty acids (FFA), a link between obesity and insulin resistance. Front Biosci 3: d169–d175. References 69
  • 79. 65. KASHYAP, S., R. BELFORT, A. GASTALDELLI, T. PRATIPANAWATR, R. BERRIA, W. PRATIPANAWATR, M. BAJAJ, L. MANDARINO, R. DEFRONZO, and K. CUSI. 2003. A sustained increase in plasma free fatty acids impairs insulin secretion in nondiabetic subjects genetically predisposed to develop type 2 diabetes. Diabetes 52:2461–2474. 66. SANTOMAURO, A.T., G. BODEN, M.E. SILVA, D.M. ROCHA, R.F. SANTOS, M.J. URSICH, P.G. STRASSMANN, and B.L. WAJCHENBERG. 1999. Overnight lowering of free fatty acids with Acipimox improves insulin resistance and glucose tolerance in obese diabetic and nondiabetic subjects. Diabetes 48:1836–1841. 67. BAJAJ, M., S. SURAAMORNKUL, A. ROMANELLI, G.W. CLINE, L.J. MANDARINO, G.I. SHULMAN, and R.A. DEFRONZO. 2005. Effect of a sustained reduction in plasma free fatty acid concentration on intramuscular long-chain fatty Acyl-CoAs and insulin action in type 2 diabetic patients. Diabetes 54:3148–3153. 68. SHIMABUKURO, M., Y.T. ZHOU, M. LEVI, and R.H.UNGER. 1998. Fatty acid-induced beta cell apoptosis:a link between obesity and diabetes. Proc Natl Acad Sci USA 95:2498–2502. 69. MAEDLER, K., J. OBERHOLZER, P. BUCHER, G.A. SPINAS, and M.Y. DONATH. 2003. Monounsaturated fatty acids prevent the deleterious effects of palmitate and high glucose on human pancreatic beta-cell turnover and function. Diabetes 52:726–733. 70. BUTLER, A.E., J. JANSON, W.C. SOELLER, and P.C. BUTLER. 2003. Increased beta-cell apoptosis prevents adaptive increase in beta-cell mass in mouse model of type 2 diabetes: evidence for role of islet amyloid formation rather than direct action of amyloid. Diabetes 52:2304–2314. 71. SOELLER, W.C., J. JANSON, S.E. HART, J.C. PARKER, M.D. CARTY, R.W. STEVENSON, D.K. KREUTTER, and P.C. BUTLER. 1998. Islet amyloid-associated diabetes in obese A(vy)/a mice expressing human islet amyloid polypeptide. Diabetes 47:743–750. 72. BUTLER, A.E., J. JANSON, S. BONNER-WEIR, R. RITZEL, R.A. RIZZA, and P.C. BUTLER. 2003. Beta-cell deficit and increased beta-cell apoptosis in humans with type 2 diabetes. Diabetes 52:102–110. 73. CLARK, A., and M.R. NILSSON. 2004. Islet amyloid: a complication of islet dysfunction or an aetiological factor in Type 2 diabetes? Diabetologia 47:157–169. 74. BONNER-WEIR, S., and G.C. WEIR. 2005. New sources of pancreatic beta-cells. Nat Biotechnol 23:857–861. 75. DOR, Y., J. BROWN, O.I. MARTINEZ, and D.A. MELTON. 2004. Adult pancreatic beta-cells are formed by self-duplication rather than stem-cell differentiation. Nature 429:41–46. 76. Del PRATO, S., and P. MARCHETTI. 2004. Beta- and alpha-cell dysfunction in type 2 diabetes. Horm Metab Res 36:775–781. 77. PICK, A., J. CLARK, C. KUBSTRUP, M. LEVISETTI, W. PUGH, S. BONNER-WEIR, and K.S. POLONSKY. 1998. Role of apoptosis in failure of beta-cell mass compensation for insulin resistance and beta-cell defects in the male Zucker diabetic fatty rat. Diabetes 47:358–364. 78. MARCHETTI, P., S. Del GUERRA, L. MARSELLI, R. LUPI, M. MASINI, M. POLLERA, M. BUGLIANI, U. BOGGI, F. VISTOLI, F. MOSCA, and S. Del PRATO. 2004. Pancreatic islets from type 2 diabetic patients have functional defects and increased apoptosis that are ameliorated by metformin. J Clin Endocrinol Metab 89:5535–5541. 79. FLIER, S.N., R.N.KULKARNI, and C.R. KAHN. 2001.Evidence for a circulatingislet cell growth factor in insulin-resistant states. Proc Natl Acad Sci USA 98:7475–7480. 80. KLOPPEL, G., M. LOHR, K. HABICH, M. OBERHOLZER, and P.U. HEITZ. 1985. Islet pathology and the pathogenesis of type 1 and type 2 diabetes mellitus revisited. Surv Synth Pathol Res 4:110–125. 81. BIARNES, M., M. MONTOLIO, V. NACHER, M. RAURELL, J. SOLER, and E. MONTANYA. 2002. Beta-cell death and mass in syngeneically transplanted islets exposed to short- and long-term hyperglycemia. Diabetes 51:66–72. 82. RAHIER, J., Y. GUIOT, R.M. GOEBBELS, C. SEMPOUX, and J.C. HENQUIN. 2008. Pancreatic beta-cell mass in European subjects with type 2 diabetes. Diabetes Obes Metab 10(Suppl 4):32–42. 83. HENQUIN, J.C., E. CERASI, S. EFENDIC, D.F. STEINER, and C. BOITARD. 2008. Pancreatic beta-cell mass or beta-cell function? That is the question! Diabetes Obes Metab 10(Suppl 4):1–4. 84. KENDALL, D.M., D.E. SUTHERLAND, J.S. NAJARIAN, F.C. GOETZ, and R.P. ROBERTSON. 1990. Effects of hemipancreatectomy on insulin secretion and glucose tolerance in healthy humans. N Engl J Med 322:898–903. 70 Chapter 3 Abnormalities in Insulin Secretion in Type 2 Diabetes Mellitus
  • 80. 85. BONNER-WEIR, S., D.F. TRENT, and G.C. WEIR. 1983. Partial pancreatectomy in the rat and subsequent defect in glucose-induced insulin release. J Clin Invest 71:1544–1553. 86. MATVEYENKO, A.V., and P.C. BUTLER. 2008. Relationship between beta-cell mass and diabetes onset. Diabetes Obes Metab 10(Suppl 4):23–31. 87. GERICH, J.E. 2000. Insulin resistance is not necessarily an essential component of type 2 diabetes. J Clin Endocrinol Metab 85:2113–2115. 88. RODER, M.E., R.S. SCHWARTZ, R.L. PRIGEON, and S.E. KAHN. 2000. Reduced pancreatic B cell compensation to the insulin resistance of aging: impact on proinsulin and insulin levels. J Clin Endocrinol Metab 85:2275–2280. 89. MITRAKOU, A., D. KELLEY, M. MOKAN, T. VENEMAN, T. PANGBURN, J. REILLY, and J. GERICH. 1992. Role of reduced suppression of glucose production and diminished early insulin release in impaired glucose tolerance. N Engl J Med 326:22–29. 90. PIMENTA, W., M. KORYTKOWSKI, A. MITRAKOU, T. JENSSEN, H. YKI-JARVINEN, W. EVRON, G. DAILEY, and J. GERICH. 1995. Pancreatic beta-cell dysfunction as the primary genetic lesion in NIDDM. Evidence from studies in normal glucose-tolerant individuals with a first-degree NIDDM relative. JAMA 273:1855–1861. 91. MCCARTHY, M.I., and A.T. HATTERSLEY. 2008. Learningfrom moleculargenetics: novel insights arising from the definition of genes for monogenic and type 2 diabetes. Diabetes 57:2889–2898. 92. SAXENA, R., B.F. VOIGHT, V. LYSSENKO, N.P. BURTT, P.I. de BAKKER, H. CHEN, J.J. ROIX, S. KATHIRESAN, J.N. HIRSCHHORN, M.J. DALY, T.E. HUGHES, L. GROOP, D. ALTSHULER, P. ALMGREN, J.C. FLOREZ, J. MEYER, K. ARDLIE, K. BENGTSSON BOSTROM, B. ISOMAA, G. LETTRE, U. LINDBLAD, H.N. LYON, O. MELANDER, C. NEWTON-CHEH, P. NILSSON, M. ORHO-MELANDER, L. RASTAM, E.K. SPELIOTES, M.R. TASKINEN, T. TUOMI, C. GUIDUCCI, A. BERGLUND, J. CARLSON, L. GIANNINY, R. HACKETT, L. HALL, J. HOLMKVIST, E. LAURILA, M. SJOGREN, M. STERNER,A. SURTI, M. SVENSSON, M. SVENSSON, R. TEWHEY, B. BLUMENSTIEL, M. PARKIN, M. DEFELICE, R. BARRY, W. BRODEUR, J. CAMARATA, N. CHIA, M. FAVA, J. GIBBONS, B. HANDSAKER, C. HEALY, K. NGUYEN, C. GATES, C. SOUGNEZ, D. GAGE, M. NIZZARI, S.B. GABRIEL, G.W. CHIRN, Q. MA, H. PARIKH, D. RICHARDSON, D. RICKE, and S. PURCELL. 2007. Genome-wide association analysis identifies loci for type 2 diabetes and triglyceride levels. Science 316:1331–1336. 93. MEIGS, J.B., J. DUPUIS, A.G. HERBERT, C. LIU, P.W. WILSON, and L.A. CUPPLES. 2005. The insulin gene variable number tandem repeat and risk of type 2 diabetes in a population-based sample of families and unrelated men and women. J Clin Endocrinol Metab 90:1137–1143. 94. LYSSENKO, V., A. JONSSON, P. ALMGREN, N. PULIZZI, B. ISOMAA, T. TUOMI, G. BERGLUND, D. ALTSHULER, P. NILSSON, and L. GROOP. 2008. Clinical risk factors, DNA variants, and the development of type 2 diabetes. N Engl J Med 359:2220–2232. 95. GRANT, S.F., G. THORLEIFSSON, I. REYNISDOTTIR, R. BENEDIKTSSON, A. MANOLESCU, J. SAINZ, A. HELGASON, H. STEFANSSON, V. EMILSSON, A. HELGADOTTIR, U. STYRKARSDOTTIR, K.P. MAGNUSSON, G.B. WALTERS, E. PALSDOTTIR, T. JONSDOTTIR, T. GUDMUNDSDOTTIR, A. GYLFASON, J. SAEMUNDSDOTTIR, R.L. WILENSKY, M.P. REILLY, D.J. RADER, Y. BAGGER, C. CHRISTIANSEN, V. GUDNASON, G. SIGURDSSON, U. THORSTEINSDOTTIR, J.R. GULCHER, A. KONG, and K. STEFANSSON. 2006. Variant of transcription factor 7-like 2 (TCF7L2) gene confers risk of type 2 diabetes. Nat Genet 38:320–323. 96. ZEGGINI, E., and M.I. MCCARTHY. 2007. TCF7L2: the biggest story in diabetes genetics since HLA? Diabetologia 50:1–4. 97. SAXENA, R., L. GIANNINY, N.P. BURTT, V. LYSSENKO, C. GIUDUCCI, M. SJOGREN, J.C. FLOREZ, P. ALMGREN, B. ISOMAA, M. ORHO-MELANDER, U. LINDBLAD, M.J. DALY, T. TUOMI, J.N. HIRSCHHORN, K.G. ARDLIE, L.C. GROOP, and D. ALTSHULER. 2006. Common single nucleotide polymorphisms in TCF7L2 are reproducibly associated with type 2 diabetes and reduce the insulin response to glucose in nondiabetic individuals. Diabetes 55:2890–2895. 98. HALES, C.N., D.J. BARKER, P.M. CLARK, L.J. COX, C. FALL, C. OSMOND, and P.D. WINTER. 1991. Fetal and infant growth and impaired glucose tolerance at age 64. BMJ 303:1019–1022. 99. GAROFANO, A., P. CZERNICHOW, and B. BREANT. 1997. In utero undernutrition impairs rat beta-cell development. Diabetologia 40:1231–1234. 100. GAROFANO, A., P. CZERNICHOW, and B. BREANT. 1999. Effect of ageing on beta-cell mass and function in rats malnourished during the perinatal period. Diabetologia 42:711–718. References 71
  • 81. 101. BLONDEAU, B., A. GAROFANO, P. CZERNICHOW, and B. BREANT. 1999. Age-dependent inability of the endocrine pancreas to adapt to pregnancy: a long-term consequence of perinatal malnutrition in the rat. Endocrinology 140:4208–4213. 102. PHILLIPS, D.I., D.J. BARKER, C.N. HALES, S. HIRST, and C. OSMOND. 1994. Thinness at birth and insulin resistance in adult life. Diabetologia 37:150–154. 103. FLANAGAN, D.E., V.M. MOORE, I.F. GODSLAND, R.A. COCKINGTON, J.S. ROBINSON, and D.I. PHILLIPS. 2000. Fetal growth and the physiological control of glucose tolerance in adults: a minimal model analysis. Am J Physiol Endocrinol Metab 278:E700–E706. 104. JAQUET, D., D. CHEVENNE, P. CZERNICHOW, and C. LEVY-MARCHAL. 2000. No evidence for a major beta- cell dysfunction in young adults born with intra-uterine growth retardation. Pediatr Diabetes 1:181–185. 105. BAVDEKAR,A.,C.S.YAJNIK,C.H.FALL,S.BAPAT,A.N.PANDIT,V.DESHPANDE,S.BHAVE,S.D.KELLINGRAY, and C. JOGLEKAR. 1999. Insulin resistance syndrome in 8-year-old Indian children:small at birth, big at 8 years, or both? Diabetes 48:2422–2429. 72 Chapter 3 Abnormalities in Insulin Secretion in Type 2 Diabetes Mellitus
  • 82. Chapter 4 Adipokine Production by Adipose Tissue: A Novel Target for Treating Metabolic Syndrome and its Sequelae VANESSA DECLERCQ 2,3 , DANIELLE STRINGER 2,3 , RYAN HUNT 2,3 , CARLA G. TAYLOR 2,3 , AND PETER ZAHRADKA 1,2,3 1 Department of Physiology, University of Manitoba, Winnipeg, Canada 2 Department of Human Nutritional Sciences, University of Manitoba, Winnipeg, Canada 3 Canadian Centre for Agri-Food Research in Health and Medicine, St. Boniface Hospital Research Centre, Winnipeg, Canada INTRODUCTION The topic of this chapter is adipokines, hormones produced and secreted by adipose tissue that have multiple effects on metabolism. In normal healthy adults, adipokines regulate the utilization and storage of lipids and help to coordinate their distribution throughout the body. Glucose metabolism is also modulated by various adipokines. Additionally, certain adipokines can influence the functions of specific target tissues, of which the heart, vasculature, brain, pancreas, and liver are included in this chapter. For instance, adipokines have a major role in maintaining the normal function of vascular tissues, independent of its metabolic state. Thus, when adipokine production is altered by obesity, a variety of changes can ensue, involving one or more of these tissues. For this reason, therapeutic strategies to correct adipokine imbalance will not only be useful for treating metabolic disorders, they will also help to minimize the end organ damage. Metabolic Syndrome: Underlying Mechanisms and Drug Therapies Edited by Minghan Wang Copyright Ó 2011 John Wiley Sons, Inc. 73
  • 83. We have attempted to provide an overview of the role of adipokines and described the changes that occur in obesity as they relate to the metabolic syndrome (MetS). As well, we have outlined the state-of-the-art surgical, lifestyle, dietary, genetic, and pharmacological strategies available for manipulating adipokine levels. This provides a platform for understanding the impact that may be achieved by interventions designed to influence adipokine production, such as lifestyle, diet, surgery, and pharmaceuticals. MetS AND OBESITY MetS refers to acluster of metabolic abnormalities characterized by the coexistence of abdominal obesity, dyslipidemia, hypertension, insulin resistance, and glucose intolerance (1). It has been recently proposed that inflammation and nonalcoholic fatty liver disease (NAFLD) also be considered as elements of MetS (2). Although obesity and insulin resistance are generally recognized by different health organiza- tions as central characteristics of MetS, the diagnostic criteria arevariable (3). Several health organizations, including the World Health Organization, the National Cho- lesterol Education Program, the European Group for the Study of Insulin Resistance, and the American Association of Clinical Endocrinology have proposed their own diagnostic criteria for MetS; consequently, it has been difficult to estimate its true prevalence. Based on the criteria of the National Cholesterol Education Program and the results of the Third National Health and Nutrition Examination Survey, it was estimated that 47 million people in the United States have characteristics of MetS (4). In Canada, estimates reach as high as 26% of the population, or roughly 8 million people; however, the prevalence among ethnic subsets of the population range from 11% in the Inuit to as high as 45% in First Nations people (5). Furthermore, the prevalence of MetS among adolescents is increasing. Based on data collected from the Fourth National Health and Nutrition Examination Survey (1999–2000), the overall prevalence of MetS was 6.4%, or approximately 2 million adolescents, representing an increase of 2.2% from the period of 1988–1994. The rising prevalence of MetS is likely attributable to increasing obesity rates. Regardless of definition or diagnostic criteria, older age, reduced physical fitness, and higher percentage of body fat are associated with increased risk of MetS (5). In 1979, 13.8% of Canadian adults were obese. Results from the recent Canadian Community Health Survey reveal that, presently, 36.1% of adult Canadians have a body mass index (BMI) between 25 and 30, while 23.1% have a BMI greater than 30 (6). Globally, approximately 1.6 billion people are overweight, and 400 million people are obese (World Health Organization, 2005), and these individuals are at increased risk of developing MetS. In addition to obesity, MetS is considered a risk factor for diabetes, insulin resistance, and cardiovascular disease. Although all three conditions are interrelated, they are linked to distinct organ systems. The liver, pancreas, heart, and blood vessels, in particular, are affected by MetS and their failure is ultimately responsible for death (7). While it has long been recognized that obesity has a significant role in MetS, 74 Chapter 4 Adipokine Production by Adipose Tissue
  • 84. only in 1994 with the discovery of leptin, a hormone that is produced by adipose tissue (8), was it possible to begin to understand how adipose tissue exerts its effects on the metabolic state of other tissues. MetS AND ADIPOSE TISSUE Adipose tissue contributes to both the onset and progression of MetS. Among the factors that influence the emergence of MetS are the total amount of adipose tissue and adipose tissue distribution, both of which affect the endocrine, inflammatory, and metabolic functions of adipose tissue. These changes in adipose properties coincide with adipocyte enlargement, which implicates cellular hypertrophy as the underlying cause. This state has been termed adipocyte dysfunction, and several excellent reviews on this topic have been written recently (9–12). White adipose tissue is the main type of adipose associated with obesity and related pathologies such as insulin resistance, hyperlipidemia, hypertension, coronary heart disease (CHD), and type 2 diabetes (13). Traditionally, adipose tissue was thought to provide cushioning, heat insulation, and energy storage. We now recognize that adipose tissue is an important endocrine organ, secreting adipokines that operate as hormones and paracrine factors that have key roles in modulating glucose and lipid metabolism (14, 15). White adipose tissue consists of adipocytes and adipocyte precursors, vascular tissue, and immune cells, primarily macrophages (13). Adipocytes, which represent the specialized cells of adipose tissue that store lipid, only account for approximately 50% of the cellular content of adipose tissue (9). Mature adipocytes are spherical in shape and vary greatly in size. During the first year of life, proliferation and differentiation of adipocytes is the highest, and these processes slow down consid- erably during adolescence (16). In adults that maintain energy balance, cell prolif- eration remains rather stable. However, when energy intake exceeds expenditure, expansion of adipose mass occurs (17, 18), initially by adipocyte hypertrophy and subsequently by hyperplasia (17, 18). Alterations in adipokine production in MetS occur primarily as a result of adipocyte hypertrophy. The metabolic effects of adipocyte dysfunction are mani- fested primarily through increased leptin secretion and decreased adiponectin secretion, however, secretion of adipokines such as monocyte chemoattractant protein-1 (MCP-1), which attracts macrophages to the adipose tissue (19, 20), is also elevated. Consequently, the increase in adipose tissue mass promotes macro- phage infiltration of the adipose tissue, leading to twice as many macrophages in visceral adipose tissue compared to subcutaneous adipose tissue (21). The accumu- lation of macrophages in white adipose tissue in response to MCP-1 occurs through the CC chemokine receptor-2 (22), however, the extracellular matrix may contribute to macrophage-independent adipose inflammation due to the stress it places on adipocytes as they begin to enlarge (23). The increase in macrophages leads to chronic inflammation of the adipose tissue, since they are the source of many proinflammatory cytokines (20, 24). In fact, it now appears that the interaction between adipocytes MetS and Adipose Tissue 75
  • 85. and macrophages is one of the factors that leads to the hypersecretion of proathero- genic, proinflammatory, and prodiabetic adipokines, and the reduced secretion of protective, anti-inflammatory adipokines from adipose tissue that are characteristic of MetS (20, 24, 25). Thebloodvesselsthatpermeateadiposetissuealsosecreteanumberofcytokines, but their contribution to MetS has received limited attention. It is well established that vascular function can be altered in response to proinflammatory cytokines that originate from macrophages (26). Thus, adipose inflammation in MetS may be a factor in vascular dysfunction. However, at least one molecule released from vascular tissue, nitric oxide (NO), may also contribute to adipose tissue dysfunction in MetS. Adipose Tissue: Visceral Versus Subcutaneous Dysregulation of adipokine production and secretion from site-specific adipose depots (subcutaneous and visceral) plays an important role in mediating the insulin resistance, diabetes and cardiovascular disease that accompany MetS (27). Subcu- taneous adipose tissue mainly accumulates around the gluteal and femoral regions, whereas visceral adipose tissue is composed of omental and mesenteric adipose (28). However, intra-abdominal adipose appears to have the strongest association with MetS (29–31). The association between visceral adipose and MetS may be linked to an increase in free fatty acids (FFAs) entering the portal circulation (32, 33), and the resultant overabundance of circulating FFAs can in turn contribute to the development of insulin resistance (34, 35). Alternatively, the fact that subcutaneous and visceral adipose tissues demonstrate differences in the production of adipokines (36–40) suggests that they have different roles in modulating metabolism. For example, while leptin mRNA levels are higher in subcutaneous adipose, angiotensinogen mRNA levels are higher in visceral adipose, whereas tumor necrosis factor (TNF)-a mRNA levels are similar in both depots (38). These differences underscore how preferential expression of certain adipokines by different adipose depots can affect metabolism. Leptin Leptin regulates body weight and energy expenditure (41) as well as glucose and lipid metabolism, angiogenesis, immunity, and blood pressure homeostasis (27). Leptin has also been shown to cause vasodilatation in coronary arteries (42), but in obese individuals leptin-induced NO production is impaired due to leptin resistance (42, 43). Circulating levels of leptin are directly related to obesity, with increasing adipose mass associated with increases in serum leptin (44). Subcutaneous adipose produces approximately 80% of the circulating leptin (45), however, expression and secretion of leptin are correlated with cell size in both depots (46, 47). Adiponectin Adiponectin is expressed in adipose tissue. In the circulation, it exists in two forms full-length (primarily as trimers, hexamers, and multimeric complexes) and globular 76 Chapter 4 Adipokine Production by Adipose Tissue
  • 86. adiponectin (48, 49). Adiponectin has been shown to have antiatherogenic (50, 51) and anti-inflammatory (52) properties and is significantly reduced in patients with cardiovascular disease (53) and CHD (54, 55). Hypoadiponectinemia is also associated with the development of obesity-related hypertension (56) and type 2 diabetes mellitus (57). Unlike most other adipokines, circulating adiponectin concentrations are reduced in obesity (especially with increased visceral adipose tissue), type 2 diabetes mellitus, CHD, and MetS (58– 61). Likewise, secretion of adiponectin from adipose tissue is decreased with increased adipose mass (62), with omental adipose secreting more than subcutaneous adipose (63–65). Resistin Resistin is expressed by human adipocytes, but the majority is expressed by the macrophages embedded in adipose tissue (66). Data on levels of circulating resistin in obese humans are very inconsistent (67–69), however, there is good evidence that expression of resistin in adipose tissue is differentially regulated depending on the disease model (obesity, diabetes, and insulin resistance) (70–72). The visceral depot in mice appears to have the highest resistin expression (73). Likewise, Zucker diabetic fatty (ZDF) rats have higher resistin mRNA levels in visceral compared to subcu- taneous adipose tissue (39). In humans, similar results have been observed, with abdominal adipose expressing and secreting more resistin than subcutaneous adipose tissue (74, 75). Inflammatory Cytokines Proinflammatory molecules such as TNF-a, C-reactive protein (CRP), and interleu- kin-6 (IL-6) are increased in the plasma of obese individuals (76–80). Adipose TNF-a levels are increased in obesity (81–83) and TNF-a appears to be produced equally by both subcutaneous and visceral adipose depots in humans (84). In contrast, about 30% of circulating IL-6 in obese individuals originates from adipose tissue, primarily from the visceral adipose (27, 37, 84). Renin-Angiotensin System Angiotensinogen is a major component of the renin-angiotensin system (RAS), and a fundamental regulator of systemic blood pressure. Angiotensinogen is the precursor to the vasoconstrictor angiotensin II (AngII), and thus plays an important role in hypertension (85) and vascular inflammation (86). Angiotensinogen is expressed in adipocytes (14), with higher levels in visceral adipose compared to subcutaneous adipose (38). Adipose tissue also has the ability to produce AngII due to the presence of a tissue-localized RAS (87). Interestingly, an increase in the activity of adipose tissue RAS is observed in individuals with obesity-related hypertension (88). MetS and Adipose Tissue 77
  • 87. Perivascular Adipose Perivascular adipose is the term that is applied to the adipose tissue located around vascular structures, including those present on the heart. A correlation has been shown between epicardial adipose tissue and some of the components of MetS such as waist circumference, diastolic blood pressure, and fasting insulin, but not circulating triglycerides or HDL (89). Rat aortic rings surrounded by perivascular adipose tissue display a lower contractile response compared to aortic rings without perivascular adipose (90). Similarly, vessels that have perivascular adipose such as the mesenteric arteries also show a reduced contractile response (91). Perivascular adipose has recently been shown to produce a variety of adipokines andbeinvolvedinvascularinflammation.Forexample,inepicardialadipose(includes surface oftheheartespeciallyaroundthecoronaryarteries),adiponectinmRNAlevels arelower in patients withCHD (92, 93).In humans, thethickness ofepicardial adipose tissue correlates with abdominal visceral adipose tissue and fasting insulin, and it is thought to behave like visceral adipose tissue (89). IL-6 and plasminogen activator inhibitor-1 (PAI-1) are higher in abdominal omental adipose compared to epicardial adipose, whereas leptin levels in subcutaneous abdominal adipose are higher than in epicardial adipose (93). More recently, Cheng et al. (94) found that adiponectin levels were lower in abdominal adipose compared to epicardial adipose, whereas TNF-a, IL-6, leptin, and visfatin were higher in abdominal compared to epicardial fat. In a recent study of subcutaneous, visceral, and perivascular adipose, Chatterjee et al. (95) reported that mice had higher levels of adiponectin and leptin in abdominal (epididymal)adiposecomparedtosubcutaneousadipose,whereaslevelsofadiponectin andleptinwerelowerinperivascularadiposecomparedtosubcutaneousadipose.When mice were fed a high fat (42% energy) diet for 2 weeks, leptin levels increased in all tissue depots but adiponectin levels were significantly reduced only in the perivascular adipose compared to chow fed animals (95). In primary human adipocytes, release of inflammatory mediators such as IL-8, IL-6, and MCP-1 was highest, and leptin and adiponectin secretion was lowest in cells from perivascular adipose relative to subcutaneous and visceral (peri-renal) adipose (95). Interestingly, perivascular adipo- cytes were smaller in size than adipocytes from subcutaneous or visceral adipose, a finding that correlated with a reduction in lipid droplet accumulation (95). While adipose tissue has a significant role in MetS, the morbidity associated with MetS is the result of changes in the properties of tissues that are targets for the actions of adipokines. Thus, while alterations in adipokine production may be a prime factor in the onset of MetS, the response of these target tissues is likely responsible for MetS progression. MetS AND THE LIVER: NONALCOHOLIC FATTY LIVER DISEASE The term nonalcoholic fatty liver disease encompasses a spectrum of hepatic disorders, beginning with simple hepatic steatosis characterized by intracytoplasmic 78 Chapter 4 Adipokine Production by Adipose Tissue
  • 88. lipid droplets within hepatocytes (96). Inflammation and necrosis of hepatocytes marks the progression to the second stage, nonalcoholic steatohepatitis (NASH). Further inflammatory damage leads to fibrosis, with half of NASH patients progres- sing to this stage (97). Fifteen percent of patients with fibrosis advance to cirrho- sis (97). If not detected and treated, cirrhosis can cause portal hypertension, hepatocellular carcinoma, and even liver failure (98, 99). It is estimated that 3% of patients with NAFLD develop liver failure or require liver transplantation (97). Insulin resistance is a precursor for the development of NAFLD. It is estimated that up to 75% of patients with type 2 diabetes mellitus have some form of NAFLD, and past history of type 2 diabetes mellitus is associated with a 26-fold increase in the risk of steatohepatitis (100, 101). Ninety-eight percent of patients with NASH are insulin resistant, and 87% exhibit attributes of MetS (102). Persons with type 2 diabetes mellitus and fatty liver have substantially higher insulin resistance than those with diabetes but without fatty liver (103). Studies have also shown that insulin resistance, elevated serum triacylglycerol (TAG) levels, and hyperinsulinemia are associated with NAFLD, regardless of body weight and BMI (104). Although there is strong evidence for an association among obesity, insulin resistance and NAFLD, nondiabetic and/or normal weight patients with NASH can also exhibit markers of insulin resistance (105). Obesity is also closely correlated with NAFLD, as the risk and severity of hepatic steatosis and steatohepatitis in obese patients is proportional to the degree of obesity (101). Although BMI and hepatic fat content are positively correlated, the relationship between waist circumference and hepatic fat content is stronger, highlighting a role for visceral adiposity in the development of hepatic steatosis (106– 108). It has been suggested that 30–40% of the variation in hepatic fat content can be explained by the variability in visceral adipose tissue (103). Hypertrophy of visceral adipose tissue and the resulting inflammatory response is a potential explanation for the strong relationship between visceral adipose tissue and fatty liver (109). The inflammatory response initiated by expanding visceral adipose tissue recruits macrophages which then secrete various proinflammatory cytokines. Prolonged exposure of adipocytes to these proinflammatory cytokines induces insulin resistance and leads to impaired insulin-mediated suppression of lipolysis. Conse- quently, there is an increased flux of FFAs from the visceral adipose tissue into the portal vein, resulting in direct delivery offatty acids to the liver (110). Once they reach the liver, these FFAs can then be taken up by hepatocytes and bound to coenzyme A (CoA). The fatty acyl-CoAs can form hepatic TAG, but they can also interfere with insulin signaling and cause hepatocyte insulin resistance (111). In addition to increased release of FFAs, altered production and release of adipokines by hypertrophic adipose tissue represents another possible link between obesity and hepatic steatosis (112). In particular, increased production of IL-6 and TNF-a (20, 24, 113) can suppress the production of adiponectin, an anti-inflamma- tory adipokine that appears to have an important role in the development of hepatic steatosis (114). The relationship between adiponectin and hepatic steatosis is highlighted by the strong inverse relationship between circulating adiponectin and hepatic fat content (115, 116) as well as hepatic insulin resistance (116). In addition, MetS and the Liver: Nonalcoholic Fatty Liver Disease 79
  • 89. genetic variability in the adiponectin receptor gene affects hepatic fat accumulation, supporting the important role of adiponectin signaling in the pathophysiology of hepatic steatosis in humans (117). However, the mechanism has yet to be elucidated. Adipokines and NAFLD Much of our knowledge of the contribution of adipokines to the development of hepatic steatosis is derived from studies in obese and/or insulin resistant animal models, where it is evident that certain adipokines are prosteatotic, while others are antisteatotic. Expression of hepatic TNF-a is elevated in the ob/ob mouse, a model which develops obesity and insulin resistance due to a mutation in the leptin gene (118). Fatty liver, which is characteristic of this mouse model, is reversed upon treatment with a neutralizing TNF-a antibody (119), suggesting a likely role for TNF- a in the development of hepatic steatosis. However, the actual therapeutic potential for modulating TNF-a expression may be limited, as circulating levels of this adipokine do not reflect expression in tissue (120). Conversely, ob/ob mice given adiponectin display reduced hepatomegaly, he- patic lipid content, serum alanine transaminase (ALT), TAG, and FFA after only 2 weeks of treatment (121). In humans (both adults and children), plasma concentra- tions of adiponectin are significantly lower in patients with NAFLD compared to both obese and healthy people (122–125). Furthermore, plasma adiponectin concentration is inversely associated with hepatic insulin sensitivity and hepatic lipid content (116). The amelioration of hepatic steatosis by adiponectin is hypothesized to occur via three possible mechanisms:stimulation of lipid oxidation via activation of AMP-activated protein kinase (AMPK), suppression of lipogenesis by decreasing activity of key lipogenic enzymes such as acetyl-CoA carboxylase (ACC) and fatty acid synthase (FAS), and activation of peroxisome proliferator activated receptor-a (PPARa), a transcription factor involved in lipolytic gene expression (114, 121, 126). Leptin is considered another important regulator of hepatic fat, although the mechanisms responsible for the protective effect of this adipokine are not fully understood. The observation that leptin deficiency leads to hepatic steatosis, which is reversible upon leptin treatment, suggests a protective role for this adipokine against hepatic fat accumulation (127). Even more convincing evidence for a direct effect of leptin signaling in the process of hepatic lipid deposition comes from studies in the ZDF rat (128), which lacks a functional leptin receptor. When infused with a recombinant adenovirus containing the gene for a functional leptin receptor, hepatic steatosis is markedly decreased and, since almost all of the infused functional leptin receptor construct is taken up by the liver, the liverbecomes the only leptin-responsive tissue (128). Therefore, any reduction in hepatic lipid content resulting from infection with the normal leptin receptor is due to the direct action of endogenous hyperlepti- nemia on the now leptin-responsive liver. Potential mechanisms responsible for this protective effect include stimulation of lipid oxidation by activating AMPK (129), reduced lipid synthesis, and increased lipid export (130). Development of leptin resistance due to chronic exposure of hepatocytes to high levels of circulating leptin is 80 Chapter 4 Adipokine Production by Adipose Tissue
  • 90. the likely explanation for hepatic steatosis despite elevated leptin in obesity. An interaction between resistin and leptin could also be responsible for the development of hepatic steatosis, since a cross between resistin-null and ob/ob mice does not exhibit fatty liver even though the degree of obesity is equivalent to that of ob/ob mice (131). Over time, hepatic steatosis will progress to steatohepatitis and finally fibrosis. The latter represents the point at which cirrhosis becomes evident. Hepatic stellate cells are normally found in a quiescent state in the healthy liver and become activated in response to liver injury. When this occurs, they begin to secrete collagen, which, if not stopped, is ultimately the cause of fibrosis. Evidence that adipokines influence fibrosis is now accumulating (132). Adiponectin, in particular, may have a protective role because it can suppress stellate cell activation by platelet-derived growth factor (PDGF) and connective tissue growth factor (CTGF) (133). In contrast, the proin- flammatory effects of resistin may promote disease progression (134). MetS AND THE PANCREAS The metabolic consequences of MetS on the pancreas are not as well defined as those of other organs such as liver and skeletal muscle. In one study of 104 adults with MetS, fatty pancreas, as detected by sonography, was present in 77% of the participants (135). Interestingly, coexistence of fatty pancreas and fatty liver were observed in 68% of the participants. After adjustment for age, BMI and lipid profile, fatty pancreas was independently related to insulin resistance (HOMA-IR), visceral adipose and ALT; furthermore, the number of MetS characteristics was significantly higher in the fatty pancreas group compared to the nonfatty pancreas group. Although increased pancreatic fat content has been negatively correlated with b-cell function (136), the relationship between pancreatic fat content and reduced b-cell function remains controversial (137), as other research has observed hyper- secretion of insulin and no detrimental effects on functional characteristics of b-cells in the presence of pancreatic fat deposition (138). In humans, histological exami- nation of fatty pancreas has revealed that the majority of fat is present in adipocytes within the exocrine tissue or in adipose tissue within the interlobular space, not within the islets themselves (139). This is contrary to what is observed in animal models such as the ZDF rat and high fat/high sucrose-fed swine, where higher pancreatic fat is associated with fibrotic, irregular, atrophied, and vacuolar islets containing lipid droplets and reduced insulin content (140). Adipokines likely influence pancreatic function, but few studies have examined this link. For instance, it was recently shown that the adiponectin type 1 receptor (AdipoR1) is reduced in the pancreas of obese mice (141). Leptin can inhibit glucagon release by a-cells in culture (142). Circulating leptin, but not adiponectin, levels also appear to be altered in acute pancreatitis, but a causal relationship has not been identified (143). Proinflammatory adipokines such as IL-6 and MCP-1 have also been reported to participate in the pathogenesis of pancreatitis (144). MetS and the Pancreas 81
  • 91. MetS AND THE BRAIN The effects of MetS on the brain result in changes in appetite, both in the fed and fasted states. The brain responds to short-term updates on food intake from gut-derived hormones such as cholecystokinin, peptide YY, and ghrelin while signals of long-term adiposity come from adipose tissue via the adipokine leptin (145, 146). Abdominal obesity, elevated serum leptin levels, and hypothalamic leptin resistance are common features of the MetS (147). Instead of the normal reduction in food intake and increase in energy expenditure that should accompany increased leptin secretion from adipose tissue, a leptin resistant state inhibits the amplified satiety message from reaching the hypothalamus. The leptin signal is only as strong as what is able to cross the blood–brain barrier, a process mediated by the short-form leptin receptor (ObRa). The leptin resistant state results in hypothalamic leptin insufficiency despite elevated blood levels (148). A decrease in central leptin levels affects pancreatic insulin secretion, energy expenditure and glucose metabolism as a result of impaired hypothalamic signaling (148, 149). These changes, in turn, can lead to obesity, hyperglycemia, hyperinsulinemia, and hypertriglyceridemia. This phenotype is mirrored in the db/db mouse model of type 2 diabetes mellitus, which lacks a functional leptin receptor. Impaired peripheral leptin signalling does not significantly alter insulin levels, energyexpenditure, or adiposity when comparedto impairedcentralleptinsignaling or resistance (150). Hypothalamic neurons in the arcuate nucleus produce the appetite stimulating peptides neuropeptide Y (NPY) and agouti-regulated peptide (AGRP), both of which are inhibited by leptin. A third peptide that is stimulated by leptin, pre- pro-opionmelanocortin (POMC), is a precursor to a-melanocyte stimulating hormone (a-MHS), which suppresses appetite and leads to increased energy expenditure (151, 152). The disruption in signaling between leptin and NPY may be central to leptin resistance induced by energy rich diets (148). The interaction of the signal transduction and activator of transcription-3 (STAT-3) and the leptin receptor is important in the regulation of energy homeostasis in NPYand AGRP expressing neurons in the arcuate nucleus (153). Restoring the leptin-induced restraint on NPY release may be an important therapeutic target in future treatment of obesity and MetS. Dysregulation of the endocannabinoid system (ECS) along with leptin resistance is linked to abdominal obesity and may exacerbate key risk factors that lead to the development of cardiovascular disease and type 2 diabetes mellitus. Inflammation, an altered blood lipid profile, hepatic steatosis, and insulin and leptin resistance have all been associated with chronic endocannabinoid receptor stimulation. Endocannabi- noids are lipid mediators, produced endogenously from membrane phospholipid precursors and triglycerides in response to elevated intracellular calcium (154). The majority of research has focused on two main endocannabinoids, arachidonoyl ethanolamide (or anandamide, AEA), and 2-arachidonoylglycerol (2-AG), which are derived from arachidonic acid (155). AEA and 2-AG are able to mimic the pharmacological effects of D9 -tetrahydrocannabinol, the active compound of mar- ijuana that stimulates appetite (156). Both AEA and 2-AG concentrations are elevated in the plasma and adipose tissue of obese humans (156). Diabetes also plays a role, as 82 Chapter 4 Adipokine Production by Adipose Tissue
  • 92. obese type 2 diabetic patients have greater 2-AG levels than nondiabetic weight matched controls (157). Two G protein-coupled ECS receptors are currently known. Cannabinoid receptor 1 (CB1) is highly abundant in the central nervous system, in particular the hypothalamus. CB1 receptors are also found peripherally in liver, gut, skeletal muscle, pancreas, and white adipose tissues. This receptor is linked to increased fat mass, decreased adipocyte proliferation, accelerated adipogenesis, elevated expres- sion of hepatic sterol regulatory element binding protein-1 (SREBP-1), ACC and FAS, and decreased glucose uptake by various peripheral tissues (158). The CB2 receptor is expressed by immune and hematopoietic cells as well as in the pancreas and white adipose tissue. The effect of the CB1 receptor on metabolism, however, is not limited to appetite. Activation of CB1 can affect glucose, insulin, cholesterol, TAG, and leptin levels, all independent of food intake. Upregulation of PPARg mRNA mediated by CB1 activation leads to TAG accumulation in adipocytes (159). Cross talk exists between the leptin and ECS signaling systems, however, it seems to occur downstream of the leptin receptor. Both ob/ob (leptin deficient) and db/db mice, which have elevated levels of AEA and 2-AG in their hypothalamus, show reduced appetite following blockade of CB1 (160, 161). CB1-null mice remain sensitive to intracerebroven- tricular leptin injection while having relatively low fasting leptin levels when challenged with a high fat diet (162). CB1-null mice are also resistant to diet-induced obesity and weigh approximately 30% less than regular adult mice due to decreased food intake (163). MetS AND THE CARDIOVASCULAR SYSTEM Numerous epidemiological studies have established a relationship between MetS and cardiovascular disease risk (164). Although the mechanisms responsible for promot- ing the progression of cardiovascular disease in MetS remain to be identified, a strong case can be made that the onset of insulin resistant and hyperglycemic states, which areexacerbated byobesity,underliesthepathophysiological changesthat ensue (165). This concept is supported by the fact that accelerated atherosclerosis and restenosis are highly prevalent in diabetes, even in the absence of obesity (166). On the other hand, evidence is mounting that adipose tissue can influence vascular and cardiac tissues directly via adipokines (10). A clear understanding of the positive and negative actions of these hormones may assist in the development of interventional strategies that can be directed at the adipose tissue and/or the relevant target organs. Although thisapproach may seem paradoxical, altering adipokine production is now recognized as a plausible means of combating cardiovascular disease. Diseases of the Heart and Vasculature The majority of cardiovascular disorders stem from either a reduction in the pumping efficiency of the heart or a diminution of blood flow through the vessels. The most MetS and the Cardiovascular System 83
  • 93. common cardiovascular disorder associated with MetS is atherosclerosis, a progres- sive decrease in the diameter of the vessel lumen due to presence of plaque (167). Although the hyperlipidemia component of MetS may feature prominently in these circumstances, there is also evidence that MetS-induced alterations in the levels of various circulating factors may independently influence atherosclerosis. Regardless of the underlying cause, the resultant inability to provide adequate oxygen and nutrients eventually leads to organ failure. Whether this produces an acute (heart attack, stroke) or a chronic (heart failure, renal failure, neuropathy) response ultimately determines the degree of morbidity associated with the condition. Arterial narrowing and stiffening due to atherosclerosis can also affect cardiovascular hemodynamics, which in turn can cause hypertension and an increase in cardiac workload. These conditions promote cardiac hypertrophy, which in time develops into heart failure even in the absence of a heart attack. Certain features of hemo- dynamic disorders may also be attributed to MetS independent of atherosclerotic disease (167). On the other hand, genetic abnormalities and alterations in electrical conductance are forms of cardiovascular disease that typically are not a consequence of MetS (168), although dietary intake of long-chain omega-3 fatty acids has been shown to reduce the mortality attributable to arrhythmia (169). Vascular Actions of Adipokines The vascular system serves as a conduit for blood, and thus provides the oxygen and nutrients needed for cells to function. A constant flow rate must therefore be maintained, and this requires tight control of blood pressure within a certain range. Likewise, vessels must be able to repair themselves if they are injured. The vascular response to these stimuli is mediated by various hormones, including adipokines. The incidence of cardiovascular disease in MetS, which is typically associated with higher circulating leptin and lower levels of circulating adiponectin, begs the question whether adipokines are causal factors. Certain studies have linked adipo- kines to endothelial dysfunction (170), whereas recent prospective studies by Sattar et al. (171, 172) have revealed that neither leptin nor adiponectin are strongly correlated with CHD risk. In fact, the association between leptin and CHD may be a consequence of the close correlation of leptin with BMI (172). These results are supported by an independent assessment of adipokine levels in elderly indivi- duals (173). Indeed, leptin may more accurately predict diabetes than cardiovascular disease (174). The latter observation may clarify the association between leptin and cardiovascular disease, since accelerated atherosclerosis is a hallmark of the diabetic state. Regardless of these results with CHD, it remains plausible that adipokines contribute to other forms of cardiovascular disease. Adipokines have been reported to have multiple vascular effects. The contribu- tion of specific adipokines is described in relation to normal and pathological blood vessel function. Various reviews have been written on this topic (12, 175–177) and we have therefore tried to emphasize the more recent publications and concepts. 84 Chapter 4 Adipokine Production by Adipose Tissue
  • 94. Leptin This hormone was originally identified by its ability to regulate food intake (178). However, evidence that leptin stimulated the sympathetic nervous system led to experiments that showed it was capable of increasing blood pressure (179). More recently,itwasshownthatleptinreceptorsarepresentoncellsofthevasculature(180), which suggests that leptin may be able to directly influence their function. This has become an area of intense research, given that leptin levels are elevated in MetS. (a) Vascular Expression. Leptin is primarily secreted by adipose tissue. Other cells that have been shown to express leptin, albeit in small amounts, include fibroblasts and osteoblasts (181, 182). There is no evidence of leptin pro- duction by cells of the healthy vasculature, however, Reyes et al. (183) have shown it is produced by sinusoidal endothelial cells. The importance of this localized production has not been determined. Nevertheless, its presence may be sufficient to promote vascular disease progression. The leptin receptor (ObR) has been detected on coronary endothelial cells in culture (184) and on the endothelial and smooth muscle cells (SMCs) of normal vascular tissue (180). These data are consistent with leptin’s ability to directly affect these cells in culture (185, 186). Changes in ObR expression apparently occur during atherogenesis. Schroeter et al. (180) detected a decrease in ObR staining of SMCs of atherosclerotic lesions, whereas strong staining was associated with macrophages. There was no apparent change in endothelial ObR levels. A subsequent study by the same group confirmed that ObR levels in atherosclerotic plaque were associated with macrophage infiltration (187). (b) Vascular Tone. Blood pressure is maintained by a balance of vasoconstric- tory and vasodilatory factors. Blood pressure is affected by MetS, as indicated by the prevalence of hypertension in this condition (188). A close association with obesity is evidenced by reports of improvements in blood pressure in conjunction with weight loss (189). The latter may be linked to leptin, which also declines with weight (190), since it has been shown that leptin can act both directly and indirectly to modulate vascular tone. The sympathetic nervous system is a key regulatory system for vascular tone. Leptin is known to act centrally via the hypothalamus to suppress appetite (191). As part of this process, leptin activates sympathetic neurons, and their activation may increase blood pressure (192, 193). The most direct experiment to demonstrate this relationship involved injection of leptin into the ventromedial hypothalamus of healthy rats, which resulted in an increase in blood pressure (194). At the same time, it has been difficult to clearly establish the link between these systems in the hypertensive state (195), and therefore it is not clear whether leptin contributes to hypertension via this mechanism. It is quite plausible that sympathetic activation of the kidney may be the mechanism by which leptin operates. Alternatively, leptin may function by affecting the vascular wall directly. MetS and the Cardiovascular System 85
  • 95. Hypertension is closely associated with stiffening of the arterial wall and hypertrophy/hyperplasia of the medial smooth muscle layer. Leptin has been shown to stimulate SMC hypertrophy (196) and proliferation (197), and activation of either process is a prerequisite for medial enlargement. On the other hand, leptin has also been reported to block SMC proliferation (198). Inhibition of cell proliferation may be the result of a leptin-dependent increase in the production of the vasodilator NO via stimulation of endo- thelial nitric oxide synthase (eNOS) (199). As well, leptin has been shown to interfere with AngII-dependent vasoconstriction (200), a major factor in hypertension. Based on these data, it may therefore be presumed that leptin resistance could result in hypertension due to the lack of NO and loss of vasoconstrictor inhibition, although both processes may be connected (201). Alternatively, leptin may alter the balance between NO and peroxynitrite production (202), thus inducing endothelial dysfunction by increasing the levels of molecules associated with oxidative stress (203). Although more direct links between hypertension and leptin are lacking, there are never- theless a number of studies that show a correlative relationship between circulating leptin levels and hypertension (190, 204, 205). It is also possible that these actions of leptin are indirect, and are the result of sympathetic activation by leptin (206, 207), which is a feature of MetS (208). Evidence that the vagal afferent nerves are targeted by leptin, thus interfering with baroreflex function and increasing blood pressure, has recently been reported (209). (c) Vascular Injury. The onset and progression of vascular disease is closely linkedtothedevelopmentofendothelialdysfunction,aconsequenceofinjury tothevesselwall.Hyperleptinemiaisassociatedwithendothelialdysfunction and arterial stiffening (210). Leptin resistance, however, may serve as an adaptive mechanism to prevent this outcome (210). A major consequence of endothelial dysfunction is activation of the underlying SMCs. This process requires the modulation of SMC phenotype, which switches from the con- tractile state present in the healthy vessel to the synthetic state that is characterized by migration, proliferation, and secretion of extracellular matrix proteins (211). Similar events occur when the vascular wall is injured, eithermechanically(e.g.,bypassgraftsurgery)orbychemicalmediators(e.g., hyperlipidemia). The resultant attempt to repair the vessel wall often leads to the formation of a lesion that can interfere with blood flow. Although both atherosclerotic(chronicinjury)andrestenotic(acuteinjury)lesionsmayhave different constituents, it is multiplication of SMCs in the intimal space (neointimal hyperplasia) that underlies lesion formation. Leptin promotes neointimal hyperplasia (212), a fact which may explain thehigheratherosclerosisratesindiabeteswithhyperleptinemia.Leptinlikely operates by stimulating SMC proliferation (197). Leptin could promote proliferation by increasing the responsiveness of cells to mitogenic agents. Juan et al. (71) reported that leptin increased endothelin-1 type A receptor 86 Chapter 4 Adipokine Production by Adipose Tissue
  • 96. expression, leading to enhanced proliferation in response to endothelin-1. Although indirect stimulation of SMC proliferation by leptin is supported by the lack of atherosclerosis in mice and rats that do not express leptin or lack a functional leptin receptor, a recent study by Lloyd et al. (213) has shown that atherosclerosisstilloccursinatripleknockoutmousethatdoesnotexpressthe LDL receptor, ApoE and leptin. This study indicates that a lack of leptin is insufficient topreventatherosclerosis under conditions where extremehyper- lipidemic conditions exist. An alternate view has been proposed by Bohlen etal.(198)whofoundthatleptinpreventsvascularSMCproliferationinvitro. A similar finding was made by Nair et al. (214) with airway smooth muscle. Procopioet al.(199)recently reportedthat leptin stimulates eNOS expression by endothelial cells via AMPK, which could explain how leptin inhibits cell proliferation. These latter observations do not agree with the findings of Bodary et al. (215). These researchers compared neointimal formation in wild type, leptin deficient (ob/ob), and leptin receptor defective (db/db) mice and found that their inability to respond to leptin protection against formation of a neointimal lesion. Interestingly, lesion formation was equivalent to wild type in mice with a leptin receptor deficient in STAT-3 signalling (leptrs/s ), although these animals were as obese as the db/db mice (215). These data suggest that exacerbation of vascular lesion formation by leptin is not a function of STAT-3-dependent signaling, but STAT-3 does mediate the effects of leptin on obesity. (d) Inflammation and Thrombosis. Vascular injury triggers an inflammatory response that results in attachment and infiltration of leukocytesas well as the differentiation of monocytes into macrophages. This process is driven by the release of chemoattractants from the injured endothelial cells and SMCs, and results in the release of inflammatory cytokines that further disturb endo- thelial function (216). Additionally, the altered surface properties of dys- functional endothelial cells lead to greater adhesion of leukocytes. This in turn can precipitate formation of a thrombus or blood clot. Leptin can indirectly influence inflammation and thrombosis through an increase in the production of CRP (217). CRP is an acute phase protein that originates primarily from the liver, but sites of extrahepatic production include vascular SMCs and macrophages (218). Elevated levels of CRP in the circulation have been linked to increased thrombosis, possibly as a result of its ability to stimulate the expression of adhesion molecules by endothelial cells. Thus, the resultant increase in leukocyte attachment promotes both progression of atherosclerotic lesion formation and elevation of the risk of thrombosis. In parallel, innate production of CRP by the vascular and inflammatory cells may exacerbate the inflammatory state of adipose tissue and thus intensify the resultant dysfunction caused by inflammation. Leptin also enhances thrombosis by increasing platelet aggregation (219). The leptin receptor (ObRb) is present on platelets (220), and leptin binding MetS and the Cardiovascular System 87
  • 97. results in release ofintracellular calcium (221). Additionally, leptin enhances aggregation in response to ADP (221). Although leptin has no additional effect on platelet characteristics, and obesity does not trigger leptin resis- tance in platelets, the increased circulating levels of leptin may be sufficient to explain the increased platelet aggregation observed in obesity (222). (e) Angiogenesis. Tissue hypoxia results in the release of paracrine factors that promote the formation of new blood vessels to perfuse the region that is oxygen deficient. This process can thus enhance recovery after a heart attack by providing new blood vessels to the damaged region of the heart. At the same time, angiogenesis allows the enlargement of atherosclerotic plaques by providing oxygen to the cells that form the core of the lesion. Leptin appears to be a potent proangiogenic factor (223, 224) that operates by promoting endothelial dysfunction and cell proliferation (225). Leptin enhances the rate of angiogenic tube formation through the release of matrix metalloproteinases, enzymes that degrade the extracellular matrix and thus provide channels for elongation of the nascent capillar- ies (225). Thus, leptin supports the progression of MetS by assisting in the formation of vessels to carry nutrients during the expansion phase of obesity (226). Adiponectin Adiponectin is currently regarded as a potent vasoprotective hormone based on its ability to prevent atherosclerosis (227). Adiponectin likely operates through the endothelial cells since an inverse association exists between circulating adiponectin levels and endothelial dysfunction (228). As such, it is expected that adiponectin will affect a variety of vascular functions. But whether adiponectin functions directly on the vascular tissues or indirectly through induction of other cytokines remains unclear in many circumstances. (a) Vascular Expression. The abundant production of adiponectin by normal adipose tissue greatly exceeds that of other tissues. For this reason, produc- tion by other cell types has only recently been recognized. It was shown by Wolf et al. (229) that endothelial cells are capable of secreting adiponectin, at least in certain vascular beds. Interestingly, adiponectin is highly expressed in fetal SMCs (230), but is apparently not found in SMCs of the adult vasculature. Regardless, adiponectin secreted from periadventitial adipose tissue, the adipocytes found around blood vessels, may be more pertinent to its role in vascular function than production by cells of the vasculature itself, and possibly even with respect to circulating adiponectin (231). On the other hand, the primary adiponectin receptors, AdipoR1 and AdipoR2, are expressed by both vascular SMCs and endothelial cells (232). Their impor- tance in vascular disease onset has been established by Zhang et al. (233), who showed that increased expression of the receptors increases the anti- inflammatory actions of adiponectin. 88 Chapter 4 Adipokine Production by Adipose Tissue
  • 98. (b) Vascular Tone. An inverse correlation between hypertension severity and adiponectin levels has been identified(190), butacausal relationship remains to be proven. Cao et al. (234) have reported that blood pressure decreases when adiponectin levels increase. Fesus et al. (235) suggest that adiponectin functions as a vasodilator. This effect can certainly be linked to the fact that vascular tone is controlled in part by periadventitial adipose, a major source of local adiponectin (236), and the fact that changes in the properties of periadventitial adipose tissue have been linked to the onset of hyperten- sion (237). The most compelling evidence of a link to blood pressure regulation is the fact that adiponectin induces expression of eNOS and can stimulate production of NO (56). Only one study, however, has examined directly the effect of introducing adiponectin into a hypertensive ani- mal (238). These data suggest adiponectin can influence vascular tone, however, this may be mediated through the central nervous system rather than systemically. (c) Vascular Injury. The primary cause of most vascular disease is failure of the endothelial cell barrier, and this is especially true when blood vessels are injured. Several recent reviews of the effects of adiponectin on the vascu- lature as it relates to the development of atherosclerotic disease have been published (239–242). However, an interesting question has recently emerged: is a decrease in circulating adiponectin levels a cause of endothelial dysfunction? If this is the case, as suggested by Cao et al. (228), then changes in adiponectin production by adipose tissue may be the causal factor for the onset of cardiovascular disease in obesity. The consequence of the loss of a protective agent such as adiponectin may thus be disease progression. For instance, glucose-induced formation of reactive oxygen species is sup- pressed by adiponectin in endothelial cells (243). Likewise, secretion of adiponectin is linked to paraoxonase-1 (PON1) (244), a peroxidase that protects LDL from oxidation and is associated both with a reduction in atherosclerotic disease and increased longevity (245). Adiponectin also improves endothelial dysfunction, characterized as adecrease in the response of vessels to factors that trigger vasodilation (246), by activating the AMPK- NOS pathway (247), and NO is a potent anti-proliferative agent (248). Although adiponectin may affect vascular remodeling in response to injury via this mechanism, there is also evidence that adiponectin can influence SMCs directly. Both SMC proliferation and migration are restricted in the presence of adiponectin (249, 250). These actions would likewise explain the inhibition of restenosis observed with adiponectin (251), which is supported by the negative association of adiponectin and restenosis (252). Interestingly, adiponectin also protects against arterial calcification (253). (d) Inflammation and Thrombosis. Production of adiponectin by macrophages may provide some positive benefits (229), particularly if accompanied by the release of anti-inflammatory cytokines. Increased NO release in response to adiponectin would also reduce inflammation (239). Additionally, MetS and the Cardiovascular System 89
  • 99. adiponectin may suppress reactive oxygen species formation in endothelial cells, thereby reducing both oxidative and nitrative stress (254). The prevention of endothelial dysfunction by adiponectin reduces the expression of adhesion molecules, which reduces the risk of thrombosis by decreasing leukocyte adherence (61). The anti-thrombotic properties of adiponectin may also result from a decrease in platelet aggregation (255), although adiponectin was unable to block propylgallate-induced platelet aggregation in vitro when added to the blood of healthy humans (256). Interestingly, the CD40 ligand (CD40L) is elevated in MetS and has been shown to exacerbate inflammation (257). Since CD40L is a target of adiponectin, it has been proposed that the anti-inflammatory actions of adiponectin result from its ability to lower circulating levels of CD40L (257). The relationship between adiponectin and its truncated globular adiponectin version is not a topic of this review, but it has been shown that globular adiponectin can cause platelet activation through an interaction with the collagen receptor (258). (e) Angiogenesis. Low levels of circulating adiponectin are correlated with a decrease in collateral vessel formation in persons with occluded coronary arteries (259), while the converse is true when high levels are present (260). Although other factors affected by MetS may also be responsible, and addressing this point will only be possible by intervention studies, there is other experimental evidence that supports an inhibitory role for adipo- nectin in angiogenesis. Adiponectin blocks endothelial cell migration in response to vascular endothelial growth factor (261). Cyclooxygenase-2 (Cox-2) may mediate this process, since angiogenesis in response to adi- ponectin does not occur in Cox-2 deficient mice (262). Caloric restriction also promotes revascularization, and involves an adiponectin-dependent mechanism that relies on AMPK and eNOS (263). Interestingly, adiponectin promotes migration of endothelial progenitor cells (264), which may provide an additional explanation for its ability to block both atherosclerotic disease and restenosis. Resistin Resistin is an adipokine that is primarily produced by adipocytes in rodents, but macrophagesare theprimary source of theresistinexpressed byhuman adipose tissue. The infiltration of macrophages into adipose tissue likely explains the increase in circulatingresistinseen in MetS (265).Onthe other hand, ithas also beenreportedthat circulating resistin levels are not correlated with MetS in humans (266). Interestingly, SMCs subjected to cyclical stretch or hypoxic conditions also produce resistin (267, 268), although the physiological relevance of this process has not been investigated. Resistin induces fatty acid binding protein in endothelial cells, possibly promoting hypertension via this mechanism (269). Alternatively, resistin blocks the effect of vasodilatorysubstances (270).Resistin is associatedwith inflammation (271),and can promote the release of proinflammatory cytokines from endothelial cells (272). 90 Chapter 4 Adipokine Production by Adipose Tissue
  • 100. Interestingly, resistin secretion is elevated in hyperhomocysteinemia and stimulation of SMC migration may lead to neointimal hyperplasia under these conditions (273). Visfatin Visfatin, also known as pre-B-cell colony-enhancing factor (PBEF) and nicotinamide phosphoribosyltransferase (Nampt), is specifically released by adipocytes (274). Visfatin is the secreted form of Nampt and likely has a different function than the intracellular protein, as indicated by the fact that these forms have different molecular masses (274). Visfatin is elevated in the proinflammatory state, and circulating levels increase in parallel with waist circumference (275). However, the available evidence suggests visfatin does not correlate with the presence of MetS (276). Rather, visfatin levels appear indicative solely of visceral fat accumulation (277). The lack of an association with MetS may be puzzling given that intracellular Nampt regulates Sirt1 activity, and this protein is closely linked with cell metabolic state and the positive actions of caloric restriction (278). Other Adipokines Vaspin (visceral adipose tissue-derived serine proteinase inhibitor) is secreted primarily by visceral adipose tissue and circulating levels vary with nutritional state (279). Furthermore, vaspin is associated both with endothelial dysfunction (280) and proatherogenic inflammation of smooth muscle (281). At this time, however, there is no evidence to link vaspin with atherosclerosis (282). Although apelin is secreted by white adipose tissue, it is also produced by many other cell types. Nevertheless, circulating apelin levels are increased in obesity, suggesting it may function as a hormone to influence other tissues (283). Apelin correlates with CHD, but not diabetes (284). Although it is claimed that there is a link between apelin and vascular injury, this view is not supported by the fact injury is also prevalent in diabetes (285). Apelin expression is induced by hypoxia, and subse- quently promotes endothelial proliferation (286). A consequence of this interaction is the upregulation of angiogenesis (287), which would enable an increase in adipose mass and therefore obesity through the formation of blood vessels to oxygenate the new tissue (287). The localized production of apelin by other tissues may have a similar function and thereby ensure tissue perfusion under conditions when blood flow is reduced (288). This would be beneficial in the case of cardiac ischemia (289). However, whether secretion of apelin by adipose can influence these other tissues has not been determined. Adipokines and the Heart The heart serves primarily as a pump to move nutrients, oxygen, and waste products to and from our tissues via the bloodstream. To accomplish this task, the heart has an efficient system for deriving energy from fatty acids. For this reason, alterations in MetS and the Cardiovascular System 91
  • 101. metabolic state such as those that occur in MetS can significantly affect cardiac function. This altered metabolic state can have multiple effects on the cardiovascular system, from increasing the resting heart rate (290) to increasing the mortality of patients with heart failure (291, 292). While most heart disease can be attributed to changes in vascular function, MetS can also affect the heart muscle directly. Heart failure is the most serious consequence of reduced blood flow as well as increased workload due to hypertension or a valve disorder. Each of these conditions is associated with ischemia, a reduction in oxygen and nutrient access. As a result, the heart modifies its metabolism as a means of adapting to the lower oxygen levels. In parallel, the heart may begin a physical transformation depending upon the degree of damage inflicted on the muscle, especially if there is an acute loss of blood flow such as occurs during a heart attack. Ischemia also results in the secretion of cytokines from the heart that can promote the growth of new blood vessels and thus improve oxygen transport. What has become clear recently is that adipokines can influence progres- sion of heart failure independent of their effects on blood vessels (293). And, as was seen with vascular disease, leptin and adiponectin have received the most attention with respect to heart failure. Leptin The LIPID (Long-Term Intervention with Pravastatin in Ischaemic Disease) study recently released results that indicate circulating leptin levels are predictive of recurrent cardiovascular events such as death, stroke, and heart attack in males who have experienced a heart attack or been hospitalized for angina (294). Interestingly, adiponectin was not associated with recurrences, which suggests leptin secretion is only elevated when the heart muscle itself is compromised. Currently, the source of this leptin is unknown, but it can be speculated that it is secreted from the pericardial adipose tissue. The leptin receptor is expressed by cardiomyocytes, which explains their ability to respond to leptin. Three distinct responses to leptin have been identified: (i) stimulation of fatty acid oxidation (295), (ii) decreased cardiac contractility (296), and (iii) cardiac hypertrophy (297). It is the latter response that likely explains the positive correlation between circulating leptin levels and poor prognosis for those with heart failure (298–300). Alternatively, leptin levels become elevated as a means of suppressing cardiac hypertrophy, and leptin resistance in MetS counters the benefits expected from this response (173, 293). Adiponectin Adiponectin is synthesized by cardiomyocytes (301). Furthermore, Skurk et al. (302) have suggested adiponectin production by cardiac tissue is controlled via a mech- anism that is distinct from the adipose tissue. Their study also revealed that cardiac adiponectin production is suppressed in heart failure, while AdipoR1 and AdipoR2 remain unchanged. Interestingly, high levelsof adiponectin are associated with poorer prognosis for persons with heart failure (303), although this is converse to the findings of Soderberg et al. (294). In part, these different conclusions may reflect different 92 Chapter 4 Adipokine Production by Adipose Tissue
  • 102. responses between healthy and obese individuals (304). As well, adiponectin might induce the expression of proinflammatory cytokines that negatively affect the heart tissue (305). Adiponectin may also directly stimulate enlargement of the heart and thus promote heart failure (306). Natriuretic peptides released by the heart stimulate secretion of adiponectin from adipocytes, even in patients with congestive heart failure (307), but whether this has positive or negative consequences on heart failure has yet to be determined. In contrast to its actions on heart failure, adiponectin provides protection against ischemia-reperfusion injury. Endothelial cells appear to have a significant role in this process. Reduction of oxidative and nitrative stress may represent the mechanism of action (254, 308). The AMPK-NOS pathway also contributes to the protective effects of adiponectin on this process (309, 310). Cardiac-specific production of adiponectin in response to ischemia is elevated through activation of hypoxia-inducible factor-1 (hif-1) (311). Adiponectin also is activated in the ischemic brain, and likely represents the underlying mechanism for its cerebroprotective actions (312). Other Adipokines There is evidence that high serum resistin levels are associated with a risk for heart failure (313, 314), possibly due to its relationship with cardiac injury (315). Both ischemic injury and heart failurewill affect the heart’s pumping action, so the increase in resistin levels may be explained by the fact that mechanical stretch induces resistin expression by cardiomyocytes (316). A recent report indicates that resistin protects against myocardial infarction (317). It is therefore possible that resistin has a cardioprotective role in acute injury, but that it is ineffective with respect to long- term cardiac dysfunction such as heart failure. Apelin may protect against cardiac ischemia-reperfusion injury (289, 318), but whether adipose tissue, in particular epicardial adipose, plays a role in this process is uncertain. Visfatin also appears to have cardioprotective actions. In a recent study by Lim et al. (319), it was shown that administration of visfatin was capable of reducing cell death during an episode of ischemia-reperfusion. In contrast, fatty acid binding protein-4 (FABP4), an adipokine released in higher levels in MetS, has been shown to suppress cardiac contractility (320). This activity would have negativeconsequences for recovery after aheart attack and likely promote the development of heart failure in persons with MetS. THERAPEUTIC INTERVENTIONS The ability to alter adipokine levels is expected to provide a means of improving health given the strong links between adipokines and certain disease states. On the other hand, implementing approaches that can successfully manipulate adipokine levels is fraught with difficulty, since little is yet known about the mechanisms that regulate adipokine synthesis and secretion. On the other hand, it is possible to propose Therapeutic Interventions 93
  • 103. that small molecule agonists or antagonists of adipokine receptors may be equally effective. At the same time, will the expected positive effects be realized? It is generally accepted that raising adiponectin levels will have numerous health benefits. Even so, there is the chance that negative outcomes may result, as was suggested by Wannamethee (321) who reported that elevated levels of adiponectin may increase the incidence of CHD. The following section will describe research studies designed to address these issues, and provide an overview of behavioral, nutritional, and pharmacological intervention strategies that have been tested in either animal models or humans. Surgery Altered adipokine expression is tightly linked to adipocyte dysfunction, which is primarily due to adipocyte hypertrophy. Both animal and human studies have shown that weight loss restores the adipokine balance to one with fewer proinflammatory cytokines and more adiponectin (322, 323). These improvements are clearly linked to a reduction in adipocyte size (324). Bariatric surgery likewise improves adipocyte function according to the observed drop in circulating leptin levels and increase in circulating adiponectin (325). However, short- and long-term changes in glucose metabolism and insulin resistance due to caloric restriction and fat mass reduction that transiently affect adipokine production could influence interpretation of the results (326). Also noteworthy is the fact that removal of subcutaneous adipose exacerbates the inflammatory response, but this is followed by a reduction in proinflammatory adipokines (327, 328). Thus, surgical removal of adipose tissue appears generally beneficial to the subject, but whether this is the best treatment approach remains to be determined. Lifestyle Greater caloric intake than utilization has long been recognized as the major cause of obesity. Consequently, increased exercise or decreased food consumption have been touted as the best means for managing this disease. This approach alone will not be successful for those individuals whose obesity is caused by genetic or endocrine abnormalities. Behavioral strategies (hypocaloric diets and exercise programs) that target weight loss have resulted in increased plasma adiponectin levels in adults with MetS (329, 330), diabetes (60, 323), and obesity (331), however, this does not seem to be the case in obese adolescent girls (332). Although few studies reported to date have addressed the question of adipocyte dysfunction directly, weight reduction has been confirmed to alter biomarkers of obesity. Choi et al. (333) have shown that levels of circulating adipocyte fatty acid binding protein (A-FABP) correlate with BMI. Furthermore, it was shown that A-FABP, which is higher in obese individuals, decreases when weight is lost. In dogs, weight loss is also associated with a reduction in proinflammatory adipokines such as TNF-a (322). Interestingly, some adipokines such as leptin and TNF-a are 94 Chapter 4 Adipokine Production by Adipose Tissue
  • 104. consistently improved with weight loss regimens, while others such as adiponectin are not (323). At the same time, adiponectin has been the focus of considerable study due to its association with metabolic and cardiovascular disease (334), as well as its status as a surrogate for adipocyte dysfunction (335). The concept that weight loss leads to improved adipose tissue function is strongly supported by the results of Pasarica et al. (336). In their study, the authors showed correlations among adipocyte size, adipokine production, and weight loss induced by lifestyle modification. Specifically, a 13% reduction in weight led to a decline in the number of large adipocytes, and this resulted in a 36% increase in circulating adiponectin levels. Interestingly, major proinflammatory adipokines (IL-6, TNF-a) did not change. Specific cardiovascular risk factors were not examined in this cohort, but significant improvements in glucose utilization were obtained. Similar data have been reported by Varady et al. (324), who found the level of improvement was dependent upon the amount of weight loss. Thus, it is reasonably clear that weight loss regimens can have a positive effect on the production of adipokines. Lifestyle interventions such as diet or physical activity that induce weight loss improve all factors of the MetS (337–339). Dietary Components In relation to changes in dietary patterns, Bradley et al. (340) have recently shown that no significant changes in circulating adipokine levels occur when successful weight loss programs emphasize the reduction of specific macronutrients (e.g., carbohydrate or fat) from the diet. However, there may be other dietary constituents in our food that are capable of influencing adipokine levels. Some specific dietary components have been shown to increase plasma adiponectin levels. For instance, Decorde et al. (341) have shown that a melon extract provided to obese hamsters increased adiponectin levels by 61%. With the same animal model, Decorde et al. (341) showed that a diet enriched in grape phenolics could also positively modify adipokine levels. Addi- tionally, resveratrol, a compound found in the skin of red grapes, has shown beneficial effects for reducing epididymal adipocyte size in mice (342) as well as increasing the circulating concentration of adiponectin, reducing TNF-a production and enhancing eNOS expression in visceral adipose tissue of obese Zucker rats (343). This study in rats also showed that resveratrol improved several parameters of MetS including dyslipidemia, hypertension, hyperinsulinemia, and inflammatory markers (343). Likewise, millet can increase adiponectin levels in mice with type 2 diabetes mellitus (344). Recently, Jobgen et al. (345) showed that dietary supplementation with L-arginine for 12 weeks reduces adipocyte size in diet-induced obese rats, and lowers serum concentrations of glucose, TAG, and leptin while increasing levels of NO metabolites. On the other hand, serum insulin and adiponectin levels were not affected by L-arginine supplementation (345). Oolong tea consumption for 1 month increased plasma adiponectin levels in patients with previous myocardial infarction and stable angina pectoris (346). Interestingly, omega-3 fatty acids from fish, specifically docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA), also elevate circulating adiponectin in mice, and this occurs regardless of food intake and Therapeutic Interventions 95
  • 105. adiposity (347). Similar results have been reported with Rhizoma Dioscoreae Tokoronis extracts, Pollack (fish) protein, and mushroom-derived chitosan (348– 350), but none of these studies examined the relationship with adipocyte dysfunction. A definitive link between adipocyte size and the secretion of cardioprotective and proinflammatory cytokines has been identified (351, 352). While this association is being increasingly recognized, few studies have utilized this parameter to determine whether specific dietary components can improve adipocyte function. Pilvi et al. (353) showed recently that incorporation of lactalbumin into low-fat rodent chow resulted in smaller adipocytes after weight loss than the low-fat chow alone. Likewise, both a dietary herb and fiber combination (354) and persimmon leaf (355) have been shown to reduce adipocyte size in conjunction with improving adipocyte functional parameters. While the increase in smaller adipocytes and improved function are typically a product of the weight loss properties of these supplements, it has also been shown that improved function can be induced without a corresponding loss of weight. Noto et al. (356) conducted a study in which a mixture of conjugated linoleic acid (CLA) isomers was fed to obese (fa/fa) Zucker rats for 8 weeks. CLA has been proposed as a weight loss agent, but evidence of efficacy remains equivocal (357–360). In this case, the CLA regimen failed to reduce adipose mass, but there were significant improve- ments in the function of various organs, specifically the liver, kidneys, and pancre- as (356, 361, 362). These changes in physiological parameters were associated with a decrease in adipocyte size, with a concomitant increase in cell number, and elevated circulating levels of adiponectin (363). As well, there were decreases in a number of proinflammatory mediators (361). These data and similar results from Nagao et al. (364) suggest it is possible to dissociate obesity from the end organ damage that typically occurs with increased weight. Interestingly, Lasa et al. (365) examined the effects of a single CLA isomer in the Syrian Golden hamster, a model of diet- induced obesity. Although many reports suggest trans-10,cis-12 CLA is the most biologically active isomer, no changes in adipose mass or adipocyte size were obtained. No other organs were examined, so it is not possible to ascertain whether this treatment had additional effects on the physiology of these animals. Likewise, there have been no beneficial effects observed with CLA treatment in most human studies on obesity management (366). However, the combination of CLAwith omega- 3 fatty acids over a 12-week period increased plasma adiponectin levels in young obese (BMI 30–36 kg/m2 ) men (367). Additional studies have been directed solely at adipocytes in culture based on the rationale that compounds that inhibit the differ- entiation of preadipocytes would interfere with weight gain (368). Whether these strategies would be effective in vivo has not been stringently tested. An argument can be made that maintaining cells in the preadipocyte state would not promote the production of protective adipokines such as adiponectin, although a reduction in proinflammatory cytokines might be expected. A number of phytochemicals have been tested for their ability to interfere with adipogenesis. These include xanthohumol, a prenylflavonoid present in hops and therefore found in small quantities in beer (369); genistein, quercetin, and resveratrol, polyphenols found in many plant species (370); epigallocatechin-3-gallate from 96 Chapter 4 Adipokine Production by Adipose Tissue
  • 106. tea (371); and guggulsterone, a compound present in tree gum that has been reported to exhibit cholesterol lowering properties (372). Of this brief list, resveratrol has also been reported to inhibit leptin release from adipocytes (373). An interesting alternative to utilizing a food product directly is to modify the item prior to ingestion. An example that has shown some success is biotransformation with bacteria. Vuong et al. (374) used this approach to modify blueberry juice. The resultant material was not only able to prevent adipocyte differentiation in vitro, its incorporation into the diet of obese mice also raised adiponectin levels, reduced weight by decreasing appetite, and was able to prevent onset of diabetes and obesity when provided to young animals. In all cases, however, none of these positive results have been translated to human studies, and concerns still exist regarding their implementation in humans (375). The data that have accumulated suggest there are a number of strategies that can be used to restore, at least in part, the normal functioning of adipose tissue in the absence of pharmacological intervention. However, it has to be noted that most studies have utilized animal models, and for this reason it is difficult to extrapolate to humans. Furthermore, few studies have examined the pharmacological aspects of using food- based therapies for modulating the morbidities associated with obesity. In particular, the issue of concentration has received little consideration. While isolated compounds may be effective when used in vitro, delivering a similar dose to an animal or human may be very difficult, especially if it is at low concentrations in a product intended for oral consumption. As well, the form of the compound may be distinct from the pharmacological version. For instance, quercetin is typically found in a glucosidic form (rutin) in plants, and the activity of the conjugate may be different than the aglycoside. Within this context, passage through the gastrointestinal system or subsequently through the liver might also affect activity through the metabolic actions of these organs. Nevertheless, the indication that it is possible to improve overall health without a requirement to lose adipose mass, as was suggested by Noto et al. (363), does present a novel concept for the development of new therapeutic agents that can operate successfully without the need to induce weight loss. Adipokine Therapy Pharmacological approaches to alter serum adipokine levels represent the state of the art in this field (242). However, before drugs with these adipokine modulatory properties were identified, direct delivery of adipokines was used in the first attempts to modulate obesity. The discovery of leptin was hailed as a breakthrough because it provided the first indication that obesity and appetite were controlled by biological factors. Numerous attempts were subsequently made to test the assumption that elevating leptin levels by direct infusion would lead to reduced weight as a consequence of a decrease in food intake. Leptin infusion into the mediobasal hypothalamus of rats both suppressed white adipose tissue lipogenesis and decreased vascular tone (376). Inhibition of lipogenesis occurs by reducing the expression of SREBP-1c and PPARg and their target genes FAS and ACC (163). Sympathetic Therapeutic Interventions 97
  • 107. innervations stimulate lipolysis through the activation of hormone-sensitive lipase along with increased expression of phosphoenol-pyruvate carboxykinase (PEPCK) and uncoupling protein-2 (UCP2). Independently of feeding behavior, leptin is able to simultaneously reduce de novo lipogenesis and FFA uptake in white adipose while preserving lean body mass through promotion of protein synthesis (376–378). While these data were encouraging, leptin infusion yielded few positive results in humans, and the best results were obtained in cases of leptin deficiency. The consequence of leptin replacement, therefore, is the amelioration of lipodystrophy, which is charac- terized as a marked loss of adipose tissue (379). TheuseofexogenousleptinasamethodfortreatingMetSorobesityisnotpractical unless it is injected directly in to the hypothalamic circulation because most MetS patients already have highly elevated endogenous leptin production as well as leptin resistance.Treatmentwithrecombinantleptinhasminimaleffectonadiposityinobese patients,evenatsupraphysiologicaldoses(380).However,infusionofrecombinantrat leptinintoFisherBrownNorwayandSprague-Dawleyratsforsevendayssignificantly reducedtotalintra-abdominalfatandcausedaNO-dependentdecreaseinmeanarterial pressure(381).Likewise,infusionofleptinandamylinincombinationhasbeenshown to restore leptin responsiveness in diet-induced obese rats (382). Increased respon- siveness to leptin was determined by the increased phosphorylation of STAT-3 in the ventromedial hypothalamus (146). The improved sensitivity to leptin resulted in decreased caloric consumption and weight loss. The increased leptin sensitivity was not explained by the reduction in caloric intake alone (146). The same effect was demonstrated in humans, with combination therapy of pramlintide (amylin-analogue) and metreleptin (recombinant human leptin) resulting in a mean weight loss of 12.7 Æ 0.9% after 24 weeks, with the weight loss being significantly different from the control group after 4 weeks. In contrast, monotherapy of pramlintide and metre- leptin yielded weight loss of 8.4 Æ 0.9% and 8.2 Æ 1.3%, respectively (146). Like leptin, adiponectin has been shown to have a wide range of physiological effects. In particular, increasing serum adiponectin levelsis expected to influence both insulin sensitivity and vascular function. At this point, however, no direct infusion studies have been performed in humans, although numerous animal experiments have shown the validity of the basic premise (383). In contrast to employing the full-length protein, Lyzogubov et al. (384) describe a novel approach that uses a peptide containing sequence present in the globular domain of adiponectin that is responsible for AdipoR1 binding. Interperitoneal injection of this peptide prevented ocular neovascularization in an animal model of macular degeneration by blocking endo- thelial cell proliferation. Infusion of recombinant resistin in Sprague-Dawley rats worsened glucose homeostasis (385) and, in a similar way, intraperitoneal injections of resistin into mice impaired glucose homeostasis and insulin action (66). These results suggest that resistin may contribute to insulin resistance. Even so, longer studies using different animal models are needed. Clinical therapy with HIV protease inhibitors reduces plasma adiponectin concentrations and causes metabolic disorders such hyperlipidemia and atheroscle- rosis. HIV protease inhibitors reduce adiponectin secretion from 3T3-L1 adipocytes, 98 Chapter 4 Adipokine Production by Adipose Tissue
  • 108. however, infusion of adiponectin in mice ameliorates protease inhibitor-induced elevations of TAG and FFAs (386). Infusion of recombinant adiponectin significantly reduces insulin resistance by decreasing TAG content in muscle and liver in obese mice, thus ameliorating hyperglycemia and hyperinsulinemia (387). Infusion of peptides can also affect pancreatic function. Kapica et al. (388) have shown that leptin, apelin, and obestatin promote output of fluid and protein from the pancreas. As well, administration of exogenous resistin improved blood flow to the pancreas (389). Given these roles in pancreatic function, it is not surprising that circulating adipokine levels may also be linked to pancreatitis (143). Gene Therapy Treatment of MetS may also be possible through the application of gene therapy. This field of research is in the early stages; however, some promising progress has been made. One type of gene therapy involves the use of adenoviruses, double-stranded DNA molecules carrying the genetic code for a particular gene. Some studies have used adenoviruses to transfer adipokine genes and study disease progression or treatment. For example, adenoviral transfer of the leptin gene into nonobese rats reverses streptozotocin-induced diabetes (390). Leptin insufficiency may be over- come by an intravenous injection of recombinant adeno-associated viral vectors that encode the leptin gene (rAAV-lep). A single injection leads to increased circulating leptin levels and normalizes body weight in obese rodents such as the ob/ob mouse. Experiments in genetically obese, diet-induced obese, or wild-type rodents given leptin either intracerebroventricularly or in specific hypothalamic sites results in leptin-induced downregulation of NPY signaling for the rodent’s lifetime (149). This restraint on NPY signaling leads to decreased levels of circulating triglycerides and FFAs and prevents insulin hypersecretion, a process that would normally precede weight gain (149). A single rAAV-lep intracerebroventricular injection normalizes blood glucose levels and prolongs life span of streptozotocin-induced diabetic mice and rats as well NOD (nonobese diabetic) mice by inhibiting the normal catabolic effects of a total lack of insulin (390). Apolipoprotein E-deficient mice treated with recombinant adenovirus expressing full-length adiponectin have a 30% reduction in aortic lesions (50). Furthermore, adiponectin-knockout (KO) mice developed hypertension; however, adenovirus- delivered adiponectin lowered elevated blood pressure in KO mice (56) and signi- ficantly decreased plasma TAG levels in normal mice (391). Based on these results, it would appear that viral delivery can successfully be used to elevate adipokine levels. Whether this approach will be feasible in the long term will depend upon development of vectors capable of constitutive, tissue-specific expression of these proteins. Pharmacological Interventions The identification of drugs for weight control has received considerable interest. Drugs such as orlistat or sibutramine have been shown to reduce visceral obesity and Therapeutic Interventions 99
  • 109. improve parameters of MetS (392, 393). In general, it is assumed that weight reduction through pharmacological management will result in the same improve- ments in health as would be expected from lifestyle changes (394, 395). In particular, the changes would involve reductions in leptin and proinflammatory cytokines (e.g., IL-6, TNF-a) in conjunction with elevated adiponectin levels. As well, it is becoming evident that some of these pharmacological agents operate by modifying adipocyte dysfunction. The sections below describe the effect of both established and recently introduced pharmacological agents on adipokine production. It should be noted that compounds not currently employed may be more effective than those already available, but their utility as drugs has never been examined. For instance, it has been shown that induction of hypoxia inducible factor-1 (hif-1) or heme oxidase-1 (HO-1) under conditions of low oxygen levels will increase adiponectin expres- sion (311). While activation of HO-1 can also be achieved with SnCl2 (234), it is unlikely that this compound will ever be used clinically since it is a strong irritant of mucosal membranes. However, the knowledge that HO-1 is a potential therapeutic target may lead to interventions that are feasible (396). Statins Statins are drugs developed to inhibit a key regulatory enzyme in the cholesterol synthesis pathway, HMG-CoA reductase. By reducing the ability to synthesize cholesterol in the liver, circulating cholesterol levels decrease. While this relationship is well recognized, statins also have pleiotropic effects that are independent of their cholesterol lowering actions. Targets of statins that have received considerable attention include RhoA and Rac1 (397). RhoA and Rac1 are key intracellular signaling proteins that regulate numerous cellular functions, among them prolifer- ation and differentiation. It has recently been established that statins can prevent the differentiation of 3T3-L1 adipocytes in culture (398). This would explain their ability to modulate expression of leptin, resistin, and adiponectin in animals and humans (399–401). These data would also explain the weight loss attained with statin treatment in obese individuals with type 2 diabetes (402), although weight loss has not been reported in other statin trials. Given the relative safety profile of statins, there appears to be considerable potential for these drugs as weight-loss agents. On the other hand, if weight loss is not a major characteristic of statins, or it is limited to specific conditions such as type 2 diabetes mellitus, there is still substantial evidence that statins improve adipose function. In the latter case, increases in adiponectin levels could explain why statins ameliorate the vascular effects of obesity, and this improvement could explain the well-known cardioprotective actions of statins. For instance, in patients with CHD, 6 months of pravastatin treatment significantly increased plasma adiponectin levels along with improving other factors of MetS such as lowering CRP levels, total cholesterol, LDL-cholesterol, and improving hyperinsulinemia and hyperglyce- mia (403). Similarly, a recent study by Nomura et al. (404) demonstrated significant increases in serum adiponectin levels and reductions in total and LDL-cholesterol after 6 months of treatment with pravastatin. Likewise, hyperlipidemic patients with 100 Chapter 4 Adipokine Production by Adipose Tissue
  • 110. NASH show increased plasma adiponectin and reduced TNF-a levels after 24 months of atorvastatin treatment (405). However, 6 months of treatment with atorvasta- tin (406) or 3 months of rosuvastatin (407) in patients with type 2 diabetes mellitus did not alter plasma adiponectin levels but did improve, as expected, total and LDL- cholesterol levels. The conflicting data may be due to the fact that different statins were used in these studies, as was indicated by Koh et al. (408). These investigators showed that a 2-month treatment with simvastatin significantly decreased plasma adiponectin levels and reduced insulin sensitivity while pravastatin significantly increased adiponectin levels and improved insulin sensitivity. Atorvastatin in patients with steatohepatitis and hyperlipidemia improved steatosis and steatohepatitis, and this was paralleled with a 25% increase in serum adiponectin; however, it is unknown if the improvement in steatohepatitis was related to the increase in circulating adiponectin (405). PPAR Agonists Both fibrates and thiazolidinediones (TZDs), like statins, were developed as agents for cholesterol lowering and increasing insulin sensitivity, respectively. These com- pounds operate by activating PPARs, key regulators of body metabolism. Fibrates are PPARa ligands, and therefore stimulate b-oxidation of fatty acids (409). Pathological weight loss is a side effect noted for the most commonly prescribed fibrate, fenofibrate, and these results are supported by evidence that this compound also prevents weight gain in animals (410). The increase in b-oxidation induced by fenofibrate may also explain its ability to reduce adipocyte size, and concomitantly decrease circulating levels of proinflammatory adipokines (411). In agreement with these data, it has been shown that fibrates are capable of increasing adiponectin levels in humans with hypertriglyceridemia (412). These results therefore support the concept that fibrates positively affect adipocyte function, and likely provide benefits that extend beyond their ability to lower serum lipids. The TZDs, in contrast to the fibrates, are ligands for PPARg and work as insulin sensitizers (413). PPARg is also an essential mediator of adipogenesis, and therefore agents such as rosiglitazone and pioglitazone are capable of affecting adipocyte function (414, 415). The latter likely explains why rosiglitazone increases plasma adiponectin levels and decreases resistin in obese persons with type 2 diabetes mellitus (416). However, the data showing effects of TZDs on resistin in these studies is not consistent (417–419). Even in animal studies, the data on TZDs and resistin are not as clear as that for adiponectin. Some studies have shown that TZDs upregulate resistin mRNA levels in adipose tissue of ob/ob mice and ZDF rats (420) whereas others report downregulated mRNA levels in adipose tissue of db/db mice and in 3T3- L1 adipocytes (72, 421, 422). Both human and animal studies have shown that PPARg agonists increase plasma adiponectin levels (423–425). Many studies have shown the benefits of TZDs in increasing adiponectin levels and reducing proinflammatory mediators such as IL-6 and CRP in patients with MetS (328, 417–419). Work by Krzyzanowska et al. (426) suggests there is a relationship between adiponectin and FFA levels. The fact that rosiglitazone treatment promotes an increase in plasma Therapeutic Interventions 101
  • 111. adiponectin while decreasing FFA levels suggests this effect is mediated by adipo- cytes. The effect on adiponectin would also explain why rosiglitazone protects against endothelial dysfunction induced by FFAs (427). Randomized, placebo-controlled trials of pioglitazone treatment in patients with nonalcoholic steatohepatitis have shown that as little as 6 months of treatment can reduce hepatic lipid content, normalize liver function, and improve hepatic insulin sensitivity (428, 429). The reduction in liver lipid content was inversely associated with an increase in plasma adiponectin. Paradoxically, although patients given pioglitazone experienced an increase in percentage of body fat, plasma adiponectin increased. Adiponectin- stimulated activation of AMPK is thought to be an important factor in mediating the metabolic effects of TZDs in the liver (429). Renin-Angiotensin System Inhibition AngII is a vasoactive molecule that is not only essential for normal cardiovascular function, but can also have detrimental effects ifit is present at chronically high levels. The pathological effects of AngII include atherosclerosis and hypertension.Typically, angiotensinogen is secreted by the liver. It is then cleaved to angiotensin I by renin, which is produced by the kidney. AngII is then produced by cleavage with angiotensin converting enzyme (ACE), which is present on the luminal surface of endothelial cells. An important discovery was the identification of angiotensinogen, a precursor to AngII, as an adipokine (27). Furthermore, adipose tissue contains all of the compo- nents of the renin-angiotensin system necessary to convert angiotensinogen to AngII. Local production of AngII therefore does not require the circulating enzymes, but adipose-derived AngII can influence systemic levels. In this way, AngII production by adipocytes can promote the development of hypertension. Another way that AngII can impact health is through suppression of adiponectin production (430), which in turn can exacerbate endothelial dysfunction. The negative actions of AngII suggest that inhibitors of AngII production will have beneficial health effects. This has been seen with ACE inhibitors, a popular class of antihypertensive agents. These com- pounds function by lowering systemic AngII levels, and thus reduces blood pressure. In addition, they have been shown to lower body weight and increase plasma adiponectin levels in rats (431), presumably by decreasing adipocyte size (432). In humans, AngII receptor blockers (ARBs), as well as ACE inhibitors, increase plasma adiponectin concentrations (433–435). Specifically, ACE inhibitors such as ramipril and valsartan have been shown to increase adiponectin concentrations in patients with MetS (436). These results may explain the broad protective effects against renal, cardiovascular, and neural disease ascribed to ACE inhibitors (437). Interestingly, ARBs are not as effective as ACE inhibitors in raising adiponectin. The sole exception is telmisartan, an ARB that stimulates PPARg as well as blocks the AngII AT1 receptor (target of ARBs) (438, 439). The fact that these observations were made in human studies suggests this approach may have a great value in the treatment of various diseases linked to vascular dysfunction. While it is clear that elevating adiponectin may be a useful therapeutic approach for treating cardiovascular disease (440), some caution may be advised before 102 Chapter 4 Adipokine Production by Adipose Tissue
  • 112. implementing these therapies. This statement is based on the fact that some treatments designed to reduce blood pressure may negatively influence adiponectin. For in- stance, ACE inhibitors have been shown to increase circulating adiponectin, but in combination with a diuretic, adiponectin levels decrease (441). This may be due to the fact that diuretic monotherapy is also associated with a reduction of adiponectin (442). AMPK Activation AMPK plays an important role in regulating glucose and lipid metabolism. Activation of AMPK results in reduced deposition of lipids and enhances oxidation of stored fat, thus it may be a possible target for treatment of MetS (443). In humans, metformin, a member of the biguanide family of antidiabetic drugs that activate AMPK, has been shown to reduce serum leptin, insulin and glucose concentrations despite having no affect on body weight or adipose tissue mass (444). Interestingly, adiponectin also activates AMPK resulting in free fatty acid oxidation and glucose uptake by skeletal muscle (126, 445) and suppression of glucose production (126). Metformin also increases resistin protein levels in abdominal (epididymal) adipose tissue in db/db mice (446). Interestingly, metformin can also affect the expression of resistin in hepatic tissues and downregulation of resistin levels by metformin may lead to improved insulin sensitivity (447). Endocannabinoid Receptor Antagonists The ECS has been identified as an important modulator of metabolism (448), and it is now clear that the ECS plays arolein food intake and adipose accumulation in humans and animals (448–450). The discovery of two G protein-coupled receptors (CB1 and CB2) for D9 -tetrahydrocannabinol has led to the identification of numerous endog- enous cannabinoid receptor ligands and the development of various receptor agonists and antagonists. Activation of CB1 increases food intake, while blocking the CB1 receptor suppresses food intake. The CB2 receptor, in contrast, appears to modulate insulin secretion by the pancreas (451) and has been recently reported to influence hepatic steatosis, inflammation, and insulin resistance in obesity (452). Treatment with a CB1 receptor antagonist (rimonabant) increases plasma adiponectin in both obese humans and rats (453–455). Recently, Despres et al. (456) have shown that one year treatment with rimonabant, significantly reduces the ratio of intra-abdominal (visceral) adipose to subcutaneous adipose tissue while increasing serum adiponectin. While these studies show rimonabant is linked to an elevation of adiponectin levels, it appears this effect may be a consequence of weight loss (457). Five placebo-controlled randomized clinical trials have been completed using rimonabant, two of which included participants with type 2 diabetes melli- tus (458). Rimonabant is recommended for patients with a BMI greater than 30 kg/ m2 and/or abnormal blood lipids. Pooled, one year data from the Rimonabant in Obesity (RIO) program showed that 20 mg/day of rimonabant resulted in significant weight loss as well as improvements in other end points such as HDL cholesterol, Therapeutic Interventions 103
  • 113. TAG, fasting glucose levels, as well as a 0.6% reduction in hemoglobin A1c (HbA1c) levels in diabetes (458, 459). With respect to other adipokines, a CB1 knockout mouse shows increased leptin resistance in association with hepatic steatosis, but little is known about the relationship between the ECS and leptin. It is possible that they operate as antag- onizing systems in the hypothalamus in terms of satiety (460, 461), which agrees with evidence that leptin may regulate CB1 expression in the hypothalamus (160). Interestingly, resveratrol has been recently shown to bind to the CB1 receptor (462), which may explain its ability to affect leptin release. Further investigation will be necessary to fully understand the effects of the ECS on other adipokines. At the same time, the withdrawal of rimonabant from the market due to psychiatric side effects associated with depression (155) and an increase in death rate as a result of intensive treatment intended to lower HbA1c levels to 6.5% (463) suggests this class of drugs may not provide the panacea from obesity as it first appeared. Cyclooxygenase Inhibition Cox-2 inhibition has been recently examined as a mechanism of reducing inflam- matory disease. Obesity is considered an inflammatory disease due to the high numbers of macrophages that infiltrate the adipose tissue (21). Lijnen et al. (464) recently examined the effect of the Cox-2 inhibitor rofecoxib (Vioxx) in an animal model of diet-induced obesity. These investigators found that the treatment attenuated weight gain and reduced adipocyte size. Furthermore, there were fewer macrophages present in the adipose tissue and 30% less leptin was produced. No differences in adiponectin or TNF-a were detected. Similar results were obtained with wild-type mice (465). Although some Cox-2 inhibitors have been withdrawn from the market, these compounds might represent an alternative route for modulating adipose function. The efficacy of these drugs may be due to the ability of adipocyte-derived prostaglandin E2 (PGE2) to stimulate the secretion of leptin by adipocytes (466). In retrospect, these results indicate PGE2 could also be classed as an adipokine. Inhibition of Monocyte Infiltration Weisberg et al. (467) showed that antagonists of the CC chemokine receptor-2, which is present on monocytes and binds MCP-1, increased adiponectin expression in adipose tissue of obese mice without altering weight or adipose mass. Although gross changes in body composition did not occur, there was a reduction in hepatic steatosis and improved insulin sensitivity. It is presumed that a reduction in monocyte infiltration into the adipose tissue is the primary cause of these positive changes. Interestingly, deletion of the MCP-1 gene did not produce similar effects (468), but this may be due to the fact that CC chemokine receptor-2 binds other members of the MCP family and thus there is sufficient redundancy to ensure that macrophages will still be drawn to adipose tissue as it enlarges due to excess caloric intake. This novel approach warrants further investigation given the significant contribution of macro- phage infiltration to inflammation-mediated adipose dysfunction. 104 Chapter 4 Adipokine Production by Adipose Tissue
  • 114. Vitamin A All-trans-retinoic acid (RA) is the active metabolite of vitamin A and is a ligand of the retinoid X receptor, a cofactor of numerous nuclear hormone receptors including the PPARs. Several studies have investigated the contribution of RA to the actions of nuclear hormone receptors, and as a matter of course have identified RA-specific changes in cellular activity. One relevant example is a decrease in leptin expression in 3T3-L1 adipocytes and human adipose tissue explants with RA treatment in vitro (469). In agreement with this observation, both RA and vitamin A have been shown to lower plasma leptin levels in rats when given in pharmacological doses, but there were no changes in body weight or adipose mass (470). Interestingly, there was no effect of RA on adipokine levels in a human population except for a lowering of resistin (471). Of note, 13-cis retinoic acid, a compound considered to have anti- inflammatory properties and is used for the treatment of acne, was shown to increase insulin resistance in adults. Paradoxically, it also increases plasma adiponectin levels (472, 473). Effects on TNF-a have also been reported (474). Ciliary Neurotrophic Factor Ciliary neurotrophic factor (CNTF) is a neurocytokine that suppresses AMPK in the hypothalamus (475) and thus exhibits similarities to leptin (476). At the same time, it can act peripherally to modulate metabolism by stimulating AMPK in skeletal muscle (477), and has been shown to induce weight loss in both animals and humans (478, 479). This weight reduction has been linked to changes in adiponectin production by the adipose tissue (480). It appears that remodeling of adipocytes by CNTF may represent the mechanism by which it achieves these results (481). These impressiveresults indicate there is strong potential for future development in this area. SUMMARY The identification of leptin in 1994 indicated for the first time that adipose tissue plays a significant role in metabolic homeostasis, and its importance in this process is now well established. Adipokine production by adipose tissue has a regulatory role in the fasting/feeding cycle, with leptin contributing to satiation. Adipokines also link adipose tissue to various organs (liver, skeletal muscle) that are important for lipid and carbohydrate metabolism, but what has now become more accepted is the role of adipokines in modulating the function of other tissues, such as the pancreas, brain, heart, and blood vessels. At this point in time, exploration of the therapeutic potential of adipokines has revealed that it is possible to manipulate their production. The underlying mechanisms that influence adipokine levels in pathological conditions, however, have not been delineated to the point that specific drug targeting is possible. Until this transpires, careful investigation of the physiological effects of any inter- vention will require close attention, particularly from the standpoint that their manipulation has the potential to negatively affect a variety of metabolic pathways. But on a positive note, this approach appears to have great potential for treating Summary 105
  • 115. metabolic diseases, even if weight loss is not achieved, by improving the function of adipose tissue. REFERENCES 1. POTENZA, M.V., and J.I. MECHANICK. 2009. The metabolic syndrome: definition, global impact, and pathophysiology. Nutr Clin Pract 24:560–577. 2. RECTOR, R.S., J.P. THYFAULT, Y. WEI, and J.A. IBDAH. 2008. Non-alcoholic fatty liver disease and the metabolic syndrome: an update. World J Gastroenterol 14:185–192. 3. MAGLIANO, D.J., J.E. SHAW, and P.Z. ZIMMET. 2006. How to best define the metabolic syndrome. Ann Med 38:34–41. 4. FORD, E.S., W.H. GILES, and A.H. MOKDAD. 2004. Increasing prevalence of the metabolic syndrome among U.S. Adults. Diabetes Care 27:2444–2449. 5. LIU, J., A.J. HANLEY, T.K. YOUNG, S.B. HARRIS, and B. ZINMAN. 2006. Characteristics and prevalence of the metabolic syndrome among three ethnic groups in Canada. Int J Obes (Lond) 30:669–676. 6. TJEPKEMA, M. 2005. Insomnia. Health Rep 17:9–25. 7. MEIGS, J.B. 2002. Epidemiology of the metabolic syndrome. 2002. Am J Manag Care 8:S283–S292. 8. ZHANG, Y., R. PROENCA, M. MAFFEI, M. BARONE, L. LEOPOLD, and J.M. FRIEDMAN. 1994. Positional cloning of the mouse obese gene and its human homologue. Nature 372:425–432. 9. BAYS, H.E., J.M. GONZALEZ-CAMPOY, G.A. BRAY, A.E. KITABCHI, D.A. BERGMAN, A.B. SCHORR, H.W. RODBARD, and R.R. HENRY. 2008. Pathogenic potential of adipose tissue and metabolic conse- quences of adipocyte hypertrophy and increased visceral adiposity. Expert Rev Cardiovasc Ther 6:343–368. 10. BAYS, H.E. 2009. “Sick fat,” metabolic disease, and atherosclerosis. Am J Med 122:S26–S37. 11. BLUHER, M. 2009. Adipose tissue dysfunction in obesity. Exp Clin Endocrinol Diabetes 117:241–250. 12. HAJER, G.R., T.W. VAN HAEFTEN, and F.L. VISSEREN. 2008. Adipose tissue dysfunction in obesity, diabetes, and vascular diseases. Eur Heart J 29:2959–2971. 13. SETHI, J.K. 2004. A burst of energy in metabolic disease research. Genome Biol 5:327. 14. AHIMA, R.S., and J.S. FLIER. 2000. Adipose tissue as an endocrine organ. Trends Endocrinol Metab 11:327–332. 15. AHIMA, R.S. 2006. Adipose tissue as an endocrine organ. Obesity (Silver Spring) 14 (Suppl 5): 242S–249S. 16. KIESS, W., S. PETZOLD, M. TOPFER, A. GARTEN, S. BLUHER, T. KAPELLEN, A. KORNER, and J. KRATZSCH. 2008. Adipocytes and adipose tissue. Best Pract Res Clin Endocrinol Metab 22:135–153. 17. FAUST, I.M., P.R. JOHNSON, J.S. STERN, and J. HIRSCH. 1978. Diet-induced adipocyte number increase in adult rats: a new model of obesity. Am J Physiol 235:E279–E286. 18. HIRSCH, J., and B. BATCHELOR. 1976. Adipose tissue cellularity in human obesity. Clin Endocrinol Metab 5:299–311. 19. BRUUN, J.M., A.S. LIHN, S.B. PEDERSEN, and B. RICHELSEN. 2005. Monocyte chemoattractant protein-1 release is higher in visceral than subcutaneous human adipose tissue (AT): implication of macro- phages resident in the AT. J Clin Endocrinol Metab 90:2282–2289. 20. WEISBERG, S.P., D. MCCANN, M. DESAI, M. ROSENBAUM, R.L. LEIBEL, and A.W. FERRANTE, JR. 2003. OBESITY IS ASSOCIATED WITH MACROPHAGE ACCUMULATION IN ADIPOSE TISSUE. J CLIN INVEST 112:1796–1808. 21. CANCELLO, R., and K. CLEMENT. 2006. Is obesity an inflammatory illness? Role of low-grade inflammation and macrophage infiltration in human white adipose tissue. BJOG 113:1141–1147. 22. BOULOUMIE, A., C.A. CURAT, C. SENGENES, K. LOLMEDE, A. MIRANVILLE, and R. BUSSE. 2005. Role of macrophage tissue infiltration in metabolic diseases. Curr Opin Clin Nutr Metab Care 8:347–354. 23. KHAN, T., E.S. MUISE, P. IYENGAR, Z.V. WANG, M. CHANDALIA, N. ABATE, B.B. ZHANG, P. BONALDO, S. CHUA, and P.E. SCHERER. 2009. Metabolic dysregulation and adipose tissue fibrosis: role of collagen VI. Mol Cell Biol 29:1575–1591. 106 Chapter 4 Adipokine Production by Adipose Tissue
  • 116. 24. XU, H., G.T. BARNES, Q. YANG, G. TAN, D. YANG, C.J. CHOU, J. SOLE, A. NICHOLS, J.S. ROSS, L. A. TARTAGLIA, and H. CHEN. 2003. Chronic inflammation in fat plays a crucial role in the development of obesity-related insulin resistance. J Clin Invest 112:1821–1830. 25. MATSUZAWA, Y. 2005. Whiteadiposetissue and cardiovascular disease. Best PractRes ClinEndocrinol Metab 19:637–647. 26. LOPPNOW, H., K. WERDAN, and M. BUERKE. 2008. Vascular cells contribute to atherosclerosis by cytokine- and innate-immunity-related inflammatory mechanisms. Innate Immun 14:63–87. 27. FRUHBECK, G. 2004. The adipose tissue as a source of vasoactive factors. Curr Med Chem Cardiovasc Hematol Agents 2:197–208. 28. LAFONTAN, M., and M. BERLAN. 2003. Do regional differences in adipocyte biology provide new pathophysiological insights? Trends Pharmacol Sci 24:276–283. 29. PHILLIPS, L.K., and J.B. PRINS. 2008. The link between abdominal obesityand the metabolic syndrome. Curr Hypertens Rep 10:156–164. 30. DESPRES, J.P., B.J. ARSENAULT, M. COTE, A. CARTIER, and I. LEMIEUX. 2008. Abdominal obesity: the cholesterol of the 21st century? Can J Cardiol 24 (Suppl D): 7D–12D. 31. DESPRES, J.P., A. CARTIER, M. COTE, and B.J. ARSENAULT. 2008. The concept of cardiometabolic risk: bridging the fields of diabetology and cardiology. Ann Med 40:514–523. 32. BERGMAN, R.N. 2000. Non-esterified fatty acids and the liver: why is insulin secreted into the portal vein? Diabetologia 43:946–952. 33. RODRIGUEZ, A., V. CATALAN, J. GOMEZ-AMBROSI, and G. FRUHBECK. 2007. Visceral and subcutaneous adiposity:are bothpotentialtherapeutictargets fortacklingthe metabolicsyndrome?CurrPharmDes 13:2169–2175. 34. WANG, H., and R.H. ECKEL. 2009. Lipoprotein lipase: from gene to obesity. Am J Physiol Endocrinol Metab 297:E271–E288. 35. DELARUE, J., and C. MAGNAN. 2007. Free fatty acids and insulin resistance. Curr Opin Clin Nutr Metab Care 10:142–148. 36. FAIN, J.N., A.K. MADAN, M.L. HILER, P. CHEEMA, and S.W. BAHOUTH. 2004. Comparison of the release of adipokines by adipose tissue, adipose tissue matrix, and adipocytes from visceral and subcutaneous abdominal adipose tissues of obese humans. Endocrinology 145:2273–2282. 37. FRIED, S.K., D.A. BUNKIN, and A.S. GREENBERG. 1998. Omental and subcutaneous adipose tissues of obese subjects release interleukin-6: depot difference and regulation by glucocorticoid. J Clin Endocrinol Metab 83:847–850. 38. DUSSERRE, E.,P.MOULIN,andH.VIDAL.2000.DifferencesinmRNAexpressionofthe proteinssecreted by the adipocytes in human subcutaneous and visceral adipose tissues. Biochim Biophys Acta 1500:88–96. 39. ATZMON,G.,X.M. YANG, R.MUZUMDAR,X.H.MA,I.GABRIELY, andN.BARZILAI.2002.Differentialgene expression between visceral and subcutaneous fat depots. Horm Metab Res 34:622–628. 40. EINSTEIN,F.H.,G.ATZMON,X.M. YANG, X.H.MA, M. RINCON, E. RUDIN,R.MUZUMDAR, andN.BARZILAI. 2005. Differential responses of visceral and subcutaneous fat depots to nutrients. Diabetes 54:672–678. 41. FRIEDMAN, J.M., and J.L. HALAAS. 1998. Leptin and the regulation of body weight in mammals. Nature 395:763–770. 42. BELTOWSKI, J., G. WOJCICKA, and A. JAMROZ. 2003. Stimulatory effect of leptin on nitric oxide production is impaired in dietary-induced obesity. Obes Res 11:1571–1580. 43. RAHMOUNI, K., D.A. MORGAN, G.M. MORGAN, A.L. MARK, and W.G. HAYNES. 2005. Role of selective leptin resistance in diet-induced obesity hypertension. Diabetes 54:2012–2018. 44. CORREIA, M.L., and K. RAHMOUNI. 2006. Role of leptin in the cardiovascular and endocrine complications of metabolic syndrome. Diabetes Obes Metab 8:603–610. 45. ANTUNA-PUENTE, B., B. FEVE, S. FELLAHI, and J.P. BASTARD. 2008. Adipokines: the missing link between insulin resistance and obesity. Diabetes Metab 34:2–11. 46. MONTAGUE, C.T., J.B. PRINS, L. SANDERS, J. ZHANG, C.P. SEWTER, J. DIGBY, C.D. BYRNE, and S. O’RAHILLY. 1998. Depot-related gene expression in human subcutaneous and omental adipocytes. Diabetes 47:1384–1391. References 107
  • 117. 47. VAN HARMELEN, V., S. REYNISDOTTIR, P. ERIKSSON, A. THORNE, J. HOFFSTEDT, F. LONNQVIST, and P. ARNER. 1998. Leptin secretion from subcutaneous and visceral adipose tissue in women. Diabetes 47:913–917. 48. TSAO, T.S., H.E. MURREY, C. HUG, D.H. LEE, and H.F. LODISH. 2002. Oligomerization state-dependent activation of NF-kappa B signaling pathway by adipocyte complement-related protein of 30 kDa (Acrp30). J Biol Chem 277:29359–29362. 49. XU, A., K.W. CHAN, R.L. HOO, Y. WANG, K.C. TAN, J. ZHANG, B. CHEN, M.C. LAM, C. TSE, G.J. COOPER, and K.S. LAM. 2005. Testosterone selectively reduces the high molecular weight form of adiponectin by inhibiting its secretion from adipocytes. J Biol Chem 280:18073–18080. 50. OKAMOTO, Y., S. KIHARA, N. OUCHI, M. NISHIDA, Y. ARITA, M. KUMADA, K. OHASHI, N. SAKAI, I. SHIMOMURA, H. KOBAYASHI, N. TERASAKA, T. INABA, T. FUNAHASHI, and Y. MATSUZAWA. 2002. Adiponectin reduces atherosclerosis in apolipoprotein E-deficient mice. Circulation 106:2767–2770. 51. OUCHI, N.,S. KIHARA, Y. ARITA, M. NISHIDA, A.MATSUYAMA, Y. OKAMOTO, M. ISHIGAMI, H.KURIYAMA,K. KISHIDA, H. NISHIZAWA, K. HOTTA, M. MURAGUCHI, Y. OHMOTO, S. YAMASHITA, T. FUNAHASHI, and Y. MATSUZAWA. 2001. Adipocyte-derived plasma protein, adiponectin, suppresses lipid accumulation and class A scavenger receptor expression in human monocyte-derived macrophages. Circulation 103:1057–1063. 52. GOLDSTEIN, B.J., and R. SCALIA. 2004. Adiponectin: a novel adipokine linking adipocytes and vascular function. J Clin Endocrinol Metab 89:2563–2568. 53. National Cholesterol Education Program (NCEP) Expert Panel on Detection, Evaluation, and Treatment of High Blood Cholesterol in Adults (Adult Treatment Panel III). 2002. Third report of the National Cholesterol Education Program (NCEP) Expert Panel on detection, evaluation, and treatment of high blood cholesterol in adults (adult treatment panel III) final report. Circulation 106:3143–3421. 54. KUMADA, M., S. KIHARA, S. SUMITSUJI, T. KAWAMOTO, S. MATSUMOTO, N. OUCHI, Y. ARITA, Y. OKAMOTO, I. SHIMOMURA, H. HIRAOKA, T. NAKAMURA, T. FUNAHASHI, Y. MATSUZAWA, and Osaka CAD Study Group. Coronary Artery Disease. 2003. Association of hypoadiponectinemia with coronary artery disease in men. Arterioscler Thromb Vasc Biol 23:85–89. 55. ZOCCALI, C., F. MALLAMACI, G. TRIPEPI, F.A. BENEDETTO, S. CUTRUPI, S. PARLONGO, L.S. MALATINO, G. BONANNO, G. SEMINARA, F. RAPISARDA, P. FATUZZO, M. BUEMI, G. NICOCIA, S. TANAKA, N. OUCHI, S. KIHARA, T. FUNAHASHI, and Y. MATSUZAWA. 2002. Adiponectin, metabolic risk factors, and cardiovascular events among patients with end-stage renal disease. J Am Soc Nephrol 13:134–141. 56. OHASHI, K., S. KIHARA, N. OUCHI, M. KUMADA, K. FUJITA, A. HIUGE, T. HIBUSE, M. RYO, H. NISHIZAWA, N. MAEDA, K. MAEDA, R. SHIBATA, K. WALSH, T. FUNAHASHI, and I. SHIMOMURA. 2006. Adiponectin replenishment ameliorates obesity-related hypertension. Hypertension 47:1108–1116. 57. LINDSAY, R.S., T. FUNAHASHI, R.L. HANSON, Y. MATSUZAWA, S. TANAKA, P.A. TATARANNI, W.C. KNOWLER, and J. KRAKOFF. 2002. Adiponectin and development of type 2 diabetes in the Pima Indian population. Lancet 360:57–58. 58. SALAS-SALVADO, J., M. GRANADA, M. BULLO, A. COROMINAS, P. CASAS, and M. FOZ. 2007. Plasma adiponectin distribution in a Mediterranean population and its association with cardiovascular risk factors and metabolic syndrome. Metabolism 56:1486–1492. 59. ARITA, Y., S. KIHARA, N. OUCHI, M. TAKAHASHI, K. MAEDA, J. MIYAGAWA, K. HOTTA, I. SHIMOMURA, T. NAKAMURA, K. MIYAOKA, H. KURIYAMA, M. NISHIDA, S. YAMASHITA, K. OKUBO, K. MATSUBARA, M. MURAGUCHI, Y. OHMOTO, T. FUNAHASHI, and Y. MATSUZAWA. 1999. Paradoxical decrease of an adipose-specific protein, adiponectin, in obesity. Biochem Biophys Res Commun 257:79–83. 60. HOTTA, K., T. FUNAHASHI, Y. ARITA, M. TAKAHASHI, M. MATSUDA, Y. OKAMOTO, H. IWAHASHI, H. KURIYAMA, N. OUCHI, K. MAEDA, M. NISHIDA, S. KIHARA, N. SAKAI, T. NAKAJIMA, K. HASEGAWA, M. MURAGUCHI, Y. OHMOTO, T. NAKAMURA, S. YAMASHITA, T. HANAFUSA, and Y. MATSUZAWA. 2000. Plasma concentrations of a novel, adipose-specific protein, adiponectin, in type 2 diabetic patients. Arterioscler Thromb Vasc Biol 20:1595–1599. 61. OUCHI, N., S. KIHARA, Y. ARITA, K. MAEDA, H. KURIYAMA, Y. OKAMOTO, K. HOTTA, M. NISHIDA, M. TAKAHASHI, T. NAKAMURA, S. YAMASHITA, T. FUNAHASHI, and Y. MATSUZAWA. 1999. Novel modulator 108 Chapter 4 Adipokine Production by Adipose Tissue
  • 118. for endothelial adhesion molecules: adipocyte-derived plasma protein adiponectin. Circulation 100:2473–2476. 62. HOFFSTEDT, J., E. ARVIDSSON, E. SJOLIN, K. WAHLEN, and P. ARNER. 2004. Adipose tissue adiponectin production and adiponectin serum concentration in human obesity and insulin resistance. J Clin Endocrinol Metab 89:1391–1396. 63. MOTOSHIMA, H., X. WU, M.K. SINHA, V.E. HARDY, E.L. ROSATO, D.J. BARBOT, F.E. ROSATO, and B.J. GOLDSTEIN. 2002. Differential regulation of adiponectin secretion from cultured human omental and subcutaneous adipocytes: effects of insulin and rosiglitazone. J Clin Endocrinol Metab 87:5662–5667. 64. FISHER, F.M., P.G. MCTERNAN, G. VALSAMAKIS, R. CHETTY, A.L. HARTE, A.J. ANWAR, J. STARCYNSKI, J. CROCKER, A.H. BARNETT, C.L. MCTERNAN, and S. KUMAR. 2002. Differences in adiponectin protein expression: effect of fat depots and type 2 diabetic status. Horm Metab Res 34:650–654. 65. DEGAWA-YAMAUCHI, M., K.A. MOSS, J.E. BOVENKERK, S.S. SHANKAR, C.L. MORRISON, C.J. LELLIOTT, A. VIDAL-PUIG, R. JONES, and R.V. CONSIDINE. 2005. Regulation of adiponectin expression in human adipocytes: effects of adiposity, glucocorticoids, and tumor necrosis factor alpha. Obes Res 13:662–669. 66. STEPPAN,C.M.,S.T.BAILEY,S.BHAT,E.J.BROWN,R.R.BANERJEE,C.M.WRIGHT,H.R.PATEL,R.S.AHIMA, and M.A. LAZAR. 2001. The hormone resistin links obesity to diabetes. Nature 409:307–312. 67. YANNAKOULIA,M., N. YIANNAKOURIS, S. BLUHER,A.L. MATALAS, D.KLIMIS-ZACAS, andC.S. MANTZOROS. 2003. Body fat mass and macronutrient intake in relation to circulating soluble leptin receptor, free leptin index, adiponectin, and resistin concentrations in healthy humans. J Clin Endocrinol Metab 88:1730–1736. 68. DEGAWA-YAMAUCHI, M., J.E. BOVENKERK, B.E. JULIAR, W. WATSON, K. KERR, R. JONES, Q. ZHU, and R.V. CONSIDINE. 2003. Serum resistin (FIZZ3) protein is increased in obese humans. J Clin Endocrinol Metab 88:5452–5455. 69. AZUMA, K., F. KATSUKAWA, S. OGUCHI, M. MURATA, H. YAMAZAKI, A. SHIMADA, and T. SARUTA. 2003. Correlation between serum resistin level and adiposity in obese individuals. Obes Res 11:997–1001. 70. STEPPAN, C.M., and M.A. LAZAR. 2002. Resistin and obesity-associated insulin resistance. Trends Endocrinol Metab 13:18–23. 71. JUAN, C.C., L.C. AU, V.S. FANG, S.F. KANG, Y.H. KO, S.F. KUO, Y.P. HSU, C.F. KWOK, and L.T. HO. 2001. Suppressed gene expression of adipocyte resistin in an insulin-resistant rat model probably by elevated free fatty acids. Biochem Biophys Res Commun 289:1328–1333. 72. HAUGEN, F., A. JORGENSEN, C.A. DREVON, and P. TRAYHURN. 2001. Inhibition by insulin of resistin gene expression in 3T3-L1 adipocytes. FEBS Lett 507:105–108. 73. STEPPAN, C.M., and M.A. LAZAR. 2004. The current biology of resistin. J Intern Med 255:439–447. 74. MCTERNAN, P.G., C.L. MCTERNAN, R. CHETTY, K. JENNER, F.M. FISHER, M.N. LAUER, J. CROCKER, A.H. BARNETT, and S. KUMAR. 2002. Increased resistin gene and protein expression in human abdominal adipose tissue. J Clin Endocrinol Metab 87:2407. 75. FAIN, J.N., P.S. CHEEMA, S.W. BAHOUTH, and M. LLOYD HILER. 2003. Resistin release by human adipose tissue explants in primary culture. Biochem Biophys Res Commun 300:674–678. 76. PRADHAN, A.D., J.E. MANSON, N. RIFAI, J.E. BURING, and P.M. RIDKER. 2001. C-reactive protein, interleukin 6, and risk of developing type 2 diabetes mellitus. JAMA 286:327–334. 77. KERN, P.A., S. RANGANATHAN, C. LI, L. WOOD, and G. RANGANATHAN. 2001. Adipose tissue tumor necrosis factor and interleukin-6 expression in human obesity and insulin resistance. Am J Physiol Endocrinol Metab 280:E745–E751. 78. DANDONA, P., R. WEINSTOCK, K. THUSU, E. ABDEL-RAHMAN, A. ALJADA, and T. WADDEN. 1998. Tumor necrosis factor-alpha in sera of obese patients: fall with weight loss. J Clin Endocrinol Metab 83:2907–2910. 79. DANDONA, P., A. ALJADA, and A. BANDYOPADHYAY. 2004. Inflammation: the link between insulin resistance, obesity and diabetes. Trends Immunol 25:4–7. 80. VOZAROVA, B., C. WEYER, K. HANSON, P.A. TATARANNI, C. BOGARDUS, and R.E. PRATLEY. 2001. Circulating interleukin-6 in relation to adiposity, insulin action, and insulin secretion. Obes Res 9:414–417. References 109
  • 119. 81. RUDIN, E., and N. BARZILAI. 2005. Inflammatory peptides derived from adipose tissue. Immun Ageing 2:1. 82. HO, R.C., K.P. DAVY, M.S. HICKEY, and C.L. MELBY. 2005. Circulating tumor necrosis factor alpha is higher in non-obese, non-diabetic Mexican Americans compared to non-Hispanic white adults. Cytokine 30:14–21. 83. MOON, Y.S., D.H. KIM, and D.K. SONG. 2004. Serum tumor necrosis factor-alpha levels and components of the metabolic syndrome in obese adolescents. Metabolism 53:863–867. 84. FONTANA, L., J.C. EAGON, M.E. TRUJILLO, P.E. SCHERER, and S. KLEIN. 2007. Visceral fat adipokine secretion is associated with systemic inflammation in obese humans. Diabetes 56:1010–1013. 85. DAS, U.N. 2005. Is angiotensin-II an endogenous pro-inflammatory molecule? Med Sci Monit 11: RA155–RA162. 86. THAM, D.M., B. MARTIN-MCNULTY, Y.X. WANG, D.W. WILSON, R. VERGONA, M.E. SULLIVAN, W. DOLE, and J.C. RUTLEDGE. 2002. Angiotensin II is associated with activation of NF-kappaB-mediated genes and downregulation of PPARs. Physiol Genomics 11:21–30. 87. MASSIERA, F., J. SEYDOUX, A. GELOEN, A. QUIGNARD-BOULANGE, S. TURBAN, P. SAINT-MARC, A. FUKAMIZU, R. NEGREL, G. AILHAUD, and M. TEBOUL. 2001. Angiotensinogen-deficient mice exhibit impairment of diet-induced weight gain with alteration in adipose tissue development and increased locomotor activity. Endocrinology 142:5220–5225. 88. BOUSTANY, C.M., K. BHARADWAJ, A. DAUGHERTY, D.R. BROWN, D.C. RANDALL, and L.A. CASSIS. 2004. Activation of the systemic and adipose renin-angiotensin system in rats with diet-induced obesity and hypertension. Am J Physiol Regul Integr Comp Physiol 287:R943–R949. 89. IACOBELLIS, G., M.C. RIBAUDO, F. ASSAEL, E. VECCI, C. TIBERTI, A. ZAPPATERRENO, U. DI MARIO, and F. LEONETTI. 2003. Echocardiographic epicardial adipose tissue is related to anthropometric and clinical parameters of metabolic syndrome: a new indicator of cardiovascular risk. J Clin Endocrinol Metab 88:5163–5168. 90. LOHN, M., G. DUBROVSKA, B. LAUTERBACH, F.C. LUFT, M. GOLLASCH, and A.M. SHARMA. 2002. Periadventitial fat releases a vascular relaxing factor. FASEB J 16:1057–1063. 91. VERLOHREN, S., G. DUBROVSKA, S.Y. TSANG, K. ESSIN, F.C. LUFT, Y. HUANG, and M. GOLLASCH. 2004. Visceral periadventitial adipose tissue regulates arterial tone of mesenteric arteries. Hypertension 44:271–276. 92. IACOBELLIS, G., D. PISTILLI, M. GUCCIARDO, F. LEONETTI, F. MIRALDI, G. BRANCACCIO, P. GALLO, and C.R. DI GIOIA. 2005. Adiponectin expression in human epicardial adipose tissue in vivo is lower in patients with coronary artery disease. Cytokine 29:251–255. 93. BAKER, A.R., N.F. SILVA, D.W. QUINN, A.L. HARTE, D. PAGANO, R.S. BONSER, S. KUMAR, and P.G. MCTERNAN. 2006. Human epicardial adipose tissue expresses a pathogenic profile of adipocy- tokines in patients with cardiovascular disease. Cardiovasc Diabetol 5:1. 94. CHENG, K.H., C.S. CHU, K.T. LEE, T.H. LIN, C.C. HSIEH, C.C. CHIU, W.C. VOON, S.H. SHEU, and W.T. LAI. 2008. Adipocytokines and proinflammatory mediators from abdominal and epicardial adipose tissue in patients with coronary artery disease. Int J Obes (Lond) 32:268–274. 95. CHATTERJEE, T.K., L.L. STOLL, G.M. DENNING, A. HARRELSON, A.L. BLOMKALNS, G. IDELMAN, F.G. ROTHENBERG, B. NELTNER, S.A. ROMIG-MARTIN, E.W. DICKSON, S. RUDICH, and N.L. WEINTRAUB. 2009. Proinflammatory phenotype of perivascular adipocytes: influence of high-fat feeding. Circ Res 104:541–549. 96. NEUSCHWANDER-TETRI, B.A., and S.H. CALDWELL. 2003. Nonalcoholic steatohepatitis: summary of an AASLD Single Topic Conference. Hepatology 37:1202–1219. 97. SHETH, S.G., F.D. GORDON, and S. CHOPRA. 1997. Nonalcoholic steatohepatitis. Ann Intern Med 126:137–145. 98. ZAFRANI, E.S. 2004. Non-alcoholic fatty liver disease: an emerging pathological spectrum. Virchows Arch 444:3–12. 99. LUYCKX, F.H., P.J. LEFEBVRE, and A.J. SCHEEN. 2000. Non-alcoholic steatohepatitis: association with obesity and insulin resistance, and influence of weight loss. Diabetes Metab 26:98–106. 100. MALNICK, S.D., M. BEERGABEL, and H. KNOBLER. 2003. Non-alcoholic fatty liver: a common manifestation of a metabolic disorder. QJM 96:699–709. 110 Chapter 4 Adipokine Production by Adipose Tissue
  • 120. 101. WANLESS, I.R., and J.S. LENTZ. 1990. Fatty liver hepatitis (steatohepatitis) and obesity: an autopsy study with analysis of risk factors. Hepatology 12:1106–1110. 102. ANGELICO, F., M. DEL BEN, R. CONTI, S. FRANCIOSO, K. FEOLE, D. MACCIONI, T.M. ANTONINI, and C. ALESSANDRI. 2003. Non-alcoholic fatty liver syndrome: a hepatic consequence of common metabolic diseases. J Gastroenterol Hepatol 18:588–594. 103. KELLEY, D.E., T.M. MCKOLANIS, R.A. HEGAZI, L.H. KULLER, and S.C. KALHAN. 2003. Fatty liver in type 2 diabetes mellitus: relation to regional adiposity, fatty acids, and insulin resistance. Am J Physiol Endocrinol Metab 285:E906–E916. 104. MARCHESINI, G., M. BRIZI, A.M. MORSELLI-LABATE, G. BIANCHI, E. BUGIANESI, A.J. MCCULLOUGH, G. FORLANI, and N. MELCHIONDA. 1999. Association of nonalcoholic fatty liver disease with insulin resistance. Am J Med 107:450–455. 105. COMERT, B., M.R. MAS, H. ERDEM, A. DINC, U. SAGLAMKAYA, M. CIGERIM, O. KUZHAN, T. UNAL, and F. KOCABALKAN. 2001. Insulin resistance in non-alcoholic steatohepatitis. Dig Liver Dis 33:353–358. 106. HSIAO, T.J., J.C. CHEN, and J.D. WANG. 2004. Insulin resistance and ferritin as major determinants of nonalcoholic fatty liver disease in apparently healthy obese patients. Int J Obes Relat Metab Disord 28:167–172. 107. CHURCH, T.S., J.L. KUK, R. ROSS, E.L. PRIEST, E. BILTOFT, and S.N. BLAIR. 2006. Association of cardiorespiratory fitness, body mass index, and waist circumference to nonalcoholic fatty liver disease. Gastroenterology 130:2023–2030. 108. KOTRONEN, A., L. JUURINEN, A. HAKKARAINEN, J. WESTERBACKA, A. CORNER, R. BERGHOLM, and H. YKI-JARVINEN. 2008. Liver fat is increased in type 2 diabetic patients and underestimated by serum alanine aminotransferase compared with equally obese nondiabetic subjects. Diabetes Care 31:165–169. 109. MESHKANI, R., and K. ADELI. 2009. Hepatic insulin resistance, metabolic syndrome and cardiovascular disease. Clin Biochem 42:1331–1346. 110. JENSEN, M.D. 2006. Is visceral fat involved in the pathogenesis of the metabolic syndrome? Human model. Obesity (Silver Spring) 14 (Suppl 1): 20S–24S. 111. BERK, P.D. 2008. Regulatable fatty acid transport mechanisms are central to the pathophysiology of obesity, fatty liver, and metabolic syndrome. Hepatology 48:1362–1376. 112. RODEN, M. 2006. Mechanisms of disease: hepatic steatosis in type 2 diabetes—pathogenesis and clinical relevance. Nat Clin Pract Endocrinol Metab 2:335–348. 113. HOTAMISLIGIL, G.S. 2006. Inflammation and metabolic disorders. Nature 444:860–867. 114. KADOWAKI, T., and T. YAMAUCHI. 2005. Adiponectin and adiponectin receptors. Endocr Rev 26:439–451. 115. KANTARTZIS, K., K. RITTIG, B. BALLETSHOFER, J. MACHANN, F. SCHICK, K. PORUBSKA, A. FRITSCHE, H.U. HARING, and N. STEFAN. 2006. The relationships of plasma adiponectin with a favorable lipid profile, decreased inflammation, and less ectopic fat accumulation depend on adiposity. Clin Chem 52:1934–1942. 116. BUGIANESI, E., U. PAGOTTO, R. MANINI, E. VANNI, A. GASTALDELLI, R. DE IASIO, E. GENTILCORE, S. NATALE, M. CASSADER, M. RIZZETTO, R. PASQUALI, and G. MARCHESINI. 2005. Plasma adiponectin in nonalco- holic fatty liver is related to hepatic insulin resistance and hepatic fat content, not to liver disease severity. J Clin Endocrinol Metab 90:3498–3504. 117. STEFAN, N., F. MACHICAO, H. STAIGER, J. MACHANN, F. SCHICK, O. TSCHRITTER, C. SPIETH, C. WEIGERT, A. FRITSCHE, M. STUMVOLL, and H.U. HARING. 2005. Polymorphisms in the gene encoding adiponectin receptor 1 are associated with insulin resistance and high liver fat. Diabetologia 48:2282–2291. 118. TILG, H.,and A.M.DIEHL.2000.Cytokinesin alcoholicandnonalcoholicsteatohepatitis. N EnglJ Med 343:1467–1476. 119. LI, Z., S. YANG, H. LIN, J. HUANG, P.A. WATKINS, A.B. MOSER, C. DESIMONE, X.Y. SONG, and A.M. DIEHL. 2003. Probiotics and antibodies to TNF inhibit inflammatory activity and improve nonalcoholic fatty liver disease. Hepatology 37:343–350. 120. FERNANDEZ-REAL, J.M., and W. RICART. 2003. Insulin resistance and chronic cardiovascular inflam- matory syndrome. Endocr Rev 24:278–301. References 111
  • 121. 121. XU, A., Y. WANG, H. KESHAW, L.Y. XU, K.S. LAM, and G.J. COOPER. 2003. The fat-derived hormone adiponectin alleviates alcoholic and nonalcoholic fatty liver diseases in mice. J Clin Invest 112:91–100. 122. AYGUN, C., O. SENTURK, S. HULAGU, S. URAZ, A. CELEBI, T. KONDUK, B. MUTLU, and Z. CANTURK. 2006. Serum levels of hepatoprotective peptide adiponectin in non-alcoholic fatty liver disease. Eur J Gastroenterol Hepatol 18:175–180. 123. PAGANO, C., G. SOARDO, W. ESPOSITO, F. FALLO, L. BASAN, D. DONNINI, G. FEDERSPIL, L.A. SECHI, and R. VETTOR. 2005. Plasma adiponectin is decreased in nonalcoholic fatty liver disease. Eur J Endocrinol 152:113–118. 124. LOUTHAN, M.V., S. BARVE, C.J. MCCLAIN, and S. JOSHI-BARVE. 2005. Decreased serum adiponectin: an early event in pediatric nonalcoholic fatty liver disease. J Pediatr 147:835–838. 125. ZOU, C.C., L. LIANG, F. HONG, J.F. FU, and Z.Y. ZHAO. 2005. Serum adiponectin, resistin levels and non- alcoholic fatty liver disease in obese children. Endocr J 52:519–524. 126. YAMAUCHI, T., J. KAMON, Y. MINOKOSHI, Y. ITO, H. WAKI, S. UCHIDA, S. YAMASHITA, M. NODA, S. KITA, K. UEKI, K. ETO, Y. AKANUMA, P. FROGUEL, F. FOUFELLE, P. FERRE, D. CARLING, S. KIMURA, R. NAGAI, B.B. KAHN, and T. KADOWAKI. 2002. Adiponectin stimulates glucose utilization and fatty-acid oxidation by activating AMP-activated protein kinase. Nat Med 8:1288–1295. 127. JAVOR,E.D., M.G.GHANY, E.K. COCHRAN, E.A.ORAL, A.M.DEPAOLI, A.PREMKUMAR, D.E.KLEINER,and P. GORDEN. 2005. Leptin reverses nonalcoholic steatohepatitis in patients with severe lipodystrophy. Hepatology 41:753–760. 128. LEE, Y., M.Y. WANG, T. KAKUMA, Z.W. WANG, E. BABCOCK, K. MCCORKLE, M. HIGA, Y.T. ZHOU, and R.H. UNGER. 2001. Liporegulation in diet-induced obesity. The antisteatotic role of hyperleptinemia. J Biol Chem 276:5629–5635. 129. ANDREELLI, F., M. FORETZ, C. KNAUF, P.D. CANI, C. PERRIN, M.A. IGLESIAS, B. PILLOT, A. BADO, F. TRONCHE, G. MITHIEUX, S. VAULONT, R. BURCELIN, and B. VIOLLET. 2006. Liver adenosine monophosphate-activated kinase-alpha2 catalytic subunit is a key target for the control of hepatic glucose production by adiponectin and leptin but not insulin. Endocrinology 147:2432–2441. 130. SINGH, A., M. WIRTZ, N. PARKER, M. HOGAN, J. STRAHLER, G. MICHAILIDIS, S. SCHMIDT, A. VIDAL-PUIG, S. DIANO, P. ANDREWS, M.D. BRAND, and J. FRIEDMAN. 2009. Leptin-mediated changes in hepatic mitochondrial metabolism, structure, and protein levels. Proc Natl Acad Sci USA 106:13100–13105. 131. SINGHAL, N.S., R.T. PATEL, Y. QI, Y.S. LEE, and R.S. AHIMA. 2008. Loss of resistin ameliorates hyperlipidemia and hepatic steatosis in leptin-deficient mice. Am J Physiol Endocrinol Metab 295: E331–E338. 132. BERTOLANI, C., and F. MARRA. 2008. The role of adipokines in liver fibrosis. Pathophysiology 15:91–101. 133. KAMADA, Y., S. TAMURA, S. KISO, H. MATSUMOTO, Y. SAJI, Y. YOSHIDA, K. FUKUI, N. MAEDA, H. NISHIZAWA, H. NAGARETANI, Y. OKAMOTO, S. KIHARA, J. MIYAGAWA, Y. SHINOMURA, T. FUNAHASHI, and Y. MATSUZAWA. 2003. Enhanced carbon tetrachloride-induced liver fibrosis in mice lacking adiponectin. Gastroenterology 125:1796–1807. 134. PAGANO, C., G. SOARDO, C. PILON, C. MILOCCO, L. BASAN, G. MILAN, D. DONNINI, D. FAGGIAN, M. MUSSAP, M. PLEBANI, C. AVELLINI, G. FEDERSPIL, L.A. SECHI, and R. VETTOR. 2006. Increased serum resistin in nonalcoholic fatty liver disease is related to liver disease severity and not to insulin resistance. J Clin Endocrinol Metab 91:1081–1086. 135. LEE, J.S., S.H. KIM, D.W. JUN, J.H. HAN, E.C. JANG, J.Y. PARK, B.K. SON, S.H. KIM, Y.J. JO, Y.S. PARK, and Y.S. KIM. 2009. Clinical implications of fatty pancreas: correlations between fatty pancreas and metabolic syndrome. World J Gastroenterol 15:1869–1875. 136. TUSHUIZEN, M.E., M.C. BUNCK, P.J. POUWELS, S. BONTEMPS, J.H. VAN WAESBERGHE, R.K. SCHINDHELM, A. MARI, R.J. HEINE, and M. DIAMANT. 2007. Pancreatic fat content and beta-cell function in men with and without type 2 diabetes. Diabetes Care 30:2916–2921. 137. SAISHO, Y., A.E. BUTLER, and P.C. BUTLER. 2008. Pancreatic fat content and beta-cell function in men with and without type 2 diabetes: response to Tushuizen et al. Diabetes Care 31:e38; author reply e39. 112 Chapter 4 Adipokine Production by Adipose Tissue
  • 122. 138. CAMASTRA, S., M. MANCO, A. MARI, S. BALDI, A. GASTALDELLI, A.V. GRECO, G. MINGRONE, and E. FERRANNINI. 2005. Beta-cell function in morbidly obese subjects during free living: long-term effects of weight loss. Diabetes 54:2382–2389. 139. SAISHO, Y., A.E. BUTLER, J.J. MEIER, T. MONCHAMP, M. ALLEN-AUERBACH, R.A. RIZZA, and P.C. BUTLER. 2007. Pancreas volumes in humans from birth to age one hundred taking into account sex, obesity, and presence of type-2 diabetes. Clin Anat 20:933–942. 140. YIN, W., D. LIAO, M. KUSUNOKI, S. XI, K. TSUTSUMI, Z. WANG, X. LIAN, T. KOIKE, J. FAN, Y. YANG, and C. TANG. 2004. NO-1886 decreases ectopic lipid deposition and protects pancreatic beta cells in diet- induced diabetic swine. J Endocrinol 180:399–408. 141. WADE, T.E., A. MATHUR, D. LU, D.A. SWARTZ-BASILE, H.A. PITT, and N.J. ZYROMSKI. 2009. Adiponectin receptor-1 expression is decreased in the pancreas of obese mice. J Surg Res 154:78–84. 142. TUDURI, E., L. MARROQUI, S. SORIANO, A.B. ROPERO, T.M. BATISTA, S. PIQUER, M.A. LOPEZ-BOADO, E.M. CARNEIRO, R. GOMIS, A. NADAL, and I. QUESADA. 2009. Inhibitory effects of leptin on pancreatic alpha-cell function. Diabetes 58:1616–1624. 143. ADRYCH, K., M. SMOCZYNSKI, E. STELMANSKA, J. KORCZYNSKA, E. GOYKE, and J. SWIERCZYNSKI. 2008. Serum adiponectin and leptin concentrations in patients with chronic pancreatitis of alcoholic and nonalcoholic origin. Pancreas 36:120–124. 144. ZYROMSKI, N.J., A. MATHUR, H.A. PITT, D. LU, J.T. GRIPE, J.J. WALKER, K. YANCEY, T.E. WADE, and D.A. SWARTZ-BASILE. 2008. A murine model of obesity implicates the adipokine milieu in the pathogenesis of severe acute pancreatitis. Am J Physiol Gastrointest Liver Physiol 295:G552–G558. 145. CHEN, H.C., J.D. ROTH, B.E. SCHROEDER, and C. WEYER. 2008. Role of islet-, gut-, and adipocyte- derived hormones in the central control of food intake and body weight: implications for an integrated neurohormonal approach to obesity pharmacotherapy. Curr Diabetes Rev 4:79–91. 146. ROTH, J.D., B.L. ROLAND, R.L. COLE, J.L. TREVASKIS, C. WEYER, J.E. KODA, C.M. ANDERSON, D.G. PARKES, and A.D. BARON. 2008. Leptin responsiveness restored by amylin agonism in diet-induced obesity: evidence from nonclinical and clinical studies. Proc Natl Acad Sci USA 105:7257–7262. 147. DESPRES, J.P., I. LEMIEUX, J. BERGERON, P. PIBAROT, P. MATHIEU, E. LAROSE, J. RODES-CABAU, O.F. BERTRAND, and P. POIRIER. 2008. Abdominal obesity and the metabolic syndrome: contribution to global cardiometabolic risk. Arterioscler Thromb Vasc Biol 28:1039–1049. 148. KALRA, S.P. 2008. Disruption in the leptin-NPY link underlies the pandemic of diabetes and metabolic syndrome: new therapeutic approaches. Nutrition 24:820–826. 149. KALRA, S.P. 2007. Global life-long health benefits of repression of hypothalamic NPY system by central leptin gene therapy. Curr Top Med Chem 7:1675–1681. 150. GUO, K., J.E. MCMINN, T. LUDWIG, Y.H. YU, G. YANG, L. CHEN, D. LOH, C. LI, S. CHUA, JR., AND Y. ZHANG. 2007. DISRUPTION OF PERIPHERAL LEPTIN SIGNALING IN MICE RESULTS IN HYPERLEPTINEMIA WITHOUT ASSOCIATED METABOLIC ABNORMALITIES. ENDOCRINOLOGY 148:3987–3997. 151. MARATOS-FLIER, E. 2008. The long reach of leptin. Nat Med 14:604–606. 152. CONE, R.D., M.A. COWLEY, A.A. BUTLER, W. FAN, D.L. MARKS, and M.J. LOW. 2001. The arcuate nucleus as a conduit for diverse signals relevant to energy homeostasis. Int J Obes Relat Metab Disord 25 (Suppl 5): S63–S67. 153. GONG, L., F. YAO, K. HOCKMAN, H.H. HENG, G.J. MORTON, K. TAKEDA, S. AKIRA, M.J. LOW, M. RUBINSTEIN, and R.G. MACKENZIE. 2008. Signal transducer and activator of transcription-3 is required in hypothalamic agouti-related protein/neuropeptide Y neurons for normal energy homeo- stasis. Endocrinology 149:3346–3354. 154. PERRINI, S., A. LEONARDINI, L. LAVIOLA, and F. GIORGINO. 2008. Biological specificity of visceral adipose tissue and therapeutic intervention. Arch Physiol Biochem 114:277–286. 155. CARR, T.P., E.D. JESCH, and A.W. BROWN. 2008. Endocannabinoids, metabolic regulation, and the role of diet. Nutr Res 28:641–650. 156. DI MARZO, V. 2008. The endocannabinoid system in obesity and type 2 diabetes. Diabetologia 51:1356–1367. 157. GASPERI, V., F. FEZZA, N. PASQUARIELLO, M. BARI, S. ODDI, A.F. AGRO, and M. MACCARRONE. 2007. Endocannabinoids in adipocytes during differentiation and their role in glucose uptake. Cell Mol Life Sci 64:219–229. References 113
  • 123. 158. GARY-BOBO, M., G. ELACHOURI, B. SCATTON, G. LE FUR, F. OURY-DONAT, and M. BENSAID. 2006. The cannabinoid CB1 receptor antagonist rimonabant (SR141716) inhibits cell proliferation and increases markers of adipocyte maturation in cultured mouse 3T3 F442A preadipocytes. Mol Pharmacol 69:471–478. 159. MATIAS, I., M.P. GONTHIER, P. ORLANDO, V. MARTIADIS, L. DE PETROCELLIS, C. CERVINO, S. PETROSINO, L. HOAREAU, F. FESTY, R. PASQUALI, R. ROCHE, M. MAJ, U. PAGOTTO, P. MONTELEONE, and V. DI MARZO. 2006. Regulation, function, and dysregulation of endocannabinoids in models of adipose and beta- pancreatic cells and in obesity and hyperglycemia. J Clin Endocrinol Metab 91:3171–3180. 160. DI MARZO,V.,S.K. GOPARAJU, L. WANG, J.LIU, S. BATKAI, Z.JARAI, F. FEZZA, G.I.MIURA,R.D.PALMITER, T. SUGIURA, and G. KUNOS. 2001. Leptin-regulated endocannabinoids are involved in maintaining food intake. Nature 410:822–825. 161. SCHERER, T., and C. BUETTNER. 2009. The dysregulation of the endocannabinoid system in diabesity— a tricky problem. J Mol Med 87:663–668. 162. RAVINET TRILLOU, C., C. DELGORGE, C. MENET, M. ARNONE, and P. SOUBRIE. 2004. CB1 cannabinoid receptor knockout in mice leads to leanness, resistance to diet-induced obesity and enhanced leptin sensitivity. Int J Obes Relat Metab Disord 28:640–648. 163. WIERZBICKI, A.S. 2006. Rimonabant: endocannabinoid inhibition for the metabolic syndrome. Int J Clin Pract 60:1697–1706. 164. ALEXANDER, C.M., P.B. LANDSMAN, S.M. TEUTSCH, S.M. HAFFNER, Third National Health and Nutrition Examination Survey (NHANES III), and National Cholesterol Education Program (NCEP). 2003. NCEP-defined metabolic syndrome, diabetes, and prevalence of coronary heart disease among NHANES III participants age 50 years and older. Diabetes 52:1210–1214. 165. CHAIT, A., and K.E. BORNFELDT. 2009. Diabetes and atherosclerosis: is there a role for hyperglycemia? J Lipid Res 50 (Suppl): S335–S339. 166. HAMMOUD, T., J.F. TANGUAY, and M.G. BOURASSA. 2000. Management of coronary artery disease: therapeutic options in patients with diabetes. J Am Coll Cardiol 36:355–365. 167. QIAO, Q., W. GAO, L. ZHANG, R. NYAMDORJ, and J. TUOMILEHTO. 2007. Metabolic syndrome and cardiovascular disease. Ann Clin Biochem 44:232–263. 168. POIRIER, P. 2