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  • 1. RESISTENSI INSULIN : DEFINISI, MEKANISME PADADIABETES DAN PEMERIKSAAN LABORATORIUMNYA A.A. WIRADEWI LESTARIBAGIAN PATOLOGI KLINIK FK UNUD / INSTALASILABORATORIUM PATOLOGI KLINIK RSUP SANGLAH DENPASAR 1
  • 2. OUTLINE INTRODUCTION INSULIN CHEMISTRY, BIOSYNTHESIS, METABOLISM GLUCOSE TRANSPORT MECHANISM AND METABOLIC EFFECT OF INSULIN INSULIN RESISTANCE AND ASSOCIATED CONDITIONS LABORATORY TESTING FOR INSULIN RESISTANCE 2
  • 3. Diabetes mellitus :a heterogeneous disorderdefined by the presence ofhyperglycemia 3
  • 4. Hyperglycemia : functional deficiency of insulin action, due to :1. Decreased in insulin secretion2. Decreased response to insulin by target tissues (insulin resistance)3. Increase counterregulatory hormones (oppose effects insulin)10% of cases : type 1 (autoimmune B cell destruction) 4
  • 5. 90% of cases : type 2 (has a stronger genetic component, associated with resistence to the effects of insulin at its sites of action and, 80% of cases associated with obesity)Primary lesi : Insulin resistance exhausted Hyperinsulinemia insulin resistance (down regulated insulin reseptors number) Impaired early secretion of insulin insulin resistance (hiperglikemia, hiperinsulinemia)Insulin resistance : hallmark of this disorder 5
  • 6. Diabetes in clinical reality – Global 2000 2030Ranking Country People with Country People with diabetes diabetes (millions) (millions)1 India 31.7 India 79.42 China 20.8 China 42.33 US 17.7 US 30.34 Indonesia 8.4 Indonesia 21.35 Japan 6.8 Pakistan 13.96 Pakistan 5.2 Brazil 11.37 Russia 4.6 Bangladesh 11.18 Brazil 4.6 Japan 8.99 Italy 4.3 Philippines 7.810 Bangladesh 3.2 Egypt 6.7 6
  • 7. Global diabetes epidemic 350 333 diabetes world wide (millions) Number of people with 300 72% 250 194 increase 200 150 100 50 0 2003 2025 Year 7 International Diabetes Federation
  • 8. Cardiometabolic Risk (CMR)Gelfand EV. et al. 2006, Vasudevan AR. et al. 2005, Després 2006 8
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  • 10. IR and β-cell dysfunction are fundamental to Type 2 diabetes Obesity IFG Diabetes Uncontrolled hyperglycaemia 350 – Postprandial 300 – glucose 250 – Glucose 200 – Fasting (mg/dl) glucose 150 – 100 – 50 – 250 – 200 – IR Relative 150 – function (%) 100 – Clinical Insulin secretion 50 – diagnosis 0– -10 -5 0 5 10 15 20 25 30 Years of diabetesAdapted from Burger HG et al. 2001. Diabetes Mellitus, Carbohydrate Metabolism, and Lipid Disorders. In Endocrinology. 4th ed. Edited by LJ DeGroot and JL Jameson. Philadelphia: W.B. Saunders Co., 2001. 10 Originally published in Type 2 Diabetes BASICS. International Diabetes Center, Minneapolis, 2000.
  • 11. INSULIN CHEMISTRY, BIOSYNTHESIS, AND METABOLISM 11
  • 12. CHEMISTRY Is protein hormon, Store in beta-cell of pancreas as crystaline form (+Zn) Consist of 2 chains α and β chain connected by disulfide bridge. In alpha chain there is intra disulfide bridge between two cystein aa (6 and 11) Digestion of insulin by proteolytic enzyme inactivate the hormon, and for this reason it can’t be given orally 12
  • 13. BIOSYNTHESIS Human insulin gene located on the short arm of chromosome 11. A precursor molecule, preproinsulin, a peptide of MW 11.500 is translated from preproinsuline mRNA in Endoplasmic Reticulum pancreatic beta cell. Microsomal enzyme cleave preproinsulin to proinsulin (MW 9000) immediately after synthesis 13
  • 14.  Proinsulin is transported to Golgi apparatus, where packaging into secretory granule. Maturation of the secretory granule is associated to conversion of proinsulin to insulin and C- peptide Normal mature secretory granule contain insulin and C-peptide in equimolar amounts. 14
  • 15.  Insuline and C-peptide are then release from cells. C-peptide is released in an amount equimolar to insulin. Half life in plasma 3 - 5 minutes. Its metabolize fastly Liver is the main organ for insuline metabolism (50%). The others kidney and placenta. 15
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  • 18. Insuline secretion stimulation Glucose stimulate insulin secretion and suppress glucagon secretion. Initiate by ATP/ADP ratio within the cell  close the membrane ATP-sensitive K channel. depolarisation of cell  voltage change  open the Ca channel. entry Ca-ion  stimulate first short phase of insulin secretion. Increase concentration of long chain Acetyl-CoA mol : The second, more prolonged phase 18
  • 19. Biphasic Insuline secretion Fig 20.2 19
  • 20. Homeostaticregulation ofblood glucoseBlood glucose levels arenot fixed. For example,they rise after a meal,and then return to “pre-meal” values later.A meal includeschemical energy(including sugar)beyond immediate need.That excess is notdiscarded; its saved forlater use. There’s aregulated process to dothis. We’ll be focusingon glucose exclusively. 20
  • 21. GLUCOSE TRANSPORT MECHANISM ANDMETABOLIC EFFECT OF INSULIN 21
  • 22. Insulin receptor & action Insulin receptor is the members of the growth factor family, are membrane glycoprotein composed of two protein sub unit encoded by a single gene. The larger sub unit (alpha) MW 135.000 D resides entirely extracellularly, where its bind insulin mol. The alpha sub unit connected by disulfide linkage to the smaller beta subunit MW 95.000 D. This sub unit cross the membrane and its cytoplasmic domain contain a tyrosine kinase activity that initiates intra cellular signaling pathways. 22
  • 23. Tissue-specific glucose transportersName Tissue Km Properties Facilitated transport: glucose moves down concentration gradientGLUT1 RBC, most others Medium (5mM) Basal glucose uptakeGLUT2 Liver, pancreatic High (7-20mM) Allows equilibration with b-cells blood glucoseGLUT3 Brain Low (1mM) Constant uptakeGLUT4 Muscle, fat Medium (5 mM) Insulin-regulated uptake 23
  • 24. Glucose transport mechanism by insulin signaling, Depres 1999 24
  • 25. Insulin effects1. PARACRINE Effect First target cells reached by insulin is alpha cells of Langerhans islet  inhibit glucagon secretion.2. ENDOCRINE Effect Liver, Muscle, adipose tissue 25
  • 26. Metabolic effect of insulinIn the liver : promote anabolism promote glicogenesis and glicogen storage suppress lipolysis and promote synt of protein and lipogenesis Inhibit gluconeogenesis and promote glycolisis. Inhibited catabolism Inhibiting hepatic glicogenolysis, ketogenesis and gluconeogenesis. 26
  • 27.  In adipose tissue insulin stimulate triglyceride synthesis and storage, by induce production of lipoprotein lipase, increase glucose transport, and inhibit intracellular lipolysis In muscle stimulate protein synthesis, by increasing asam amino transport, promote glikogenesis 27
  • 28. Biological Action of Insulin Vasodilatation Cardio-protective  NO release Animals, human  eNOS expression Platelet inhibition Anti-apoptotic  NO release in platelets Heart, other tissues  cAMP Anti-oxidant Anti-atheroscelrotic Vascular (other) actions ApoE null mouse  ROS generation IRS-1 null mouse IRS-2 null mouse Anti-inflammatory Profibrinolytic  NFkB,  IkB Anti-thrombotic  PAI-1  TF  MCP  ICAM-1  CRP Dandona et al. Circulation 2005; 111: 1448-1454 28
  • 29. INSULIN RESISTANCE (IR) AND ASSOCIATED CONDITIONS 29
  • 30.  IR : exist any time a normal amount of insulin produce a less than normal biologic response. Inadequate tissue response to insuline hormon (decreased insulin sensitivity and or decreased responsiveness) 30
  • 31. Decreased insulin sensitivity and responsiveness, Joslin 2005 31
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  • 33.  The mechanisms responsible for insulin resistance syndromes include : * Genetic or primary target cell defects (Defect in insulin receptor expression or sequence. Rare, but represents severe form) *Autoantibodies to insulin *Accelerated insulin reseptor degradation/ Insulin reseptor downregulation 33
  • 34. In type 2 Diabetes, defects at multiple level : Decreases in receptor concentration Decrease in receptor kinase activity Decrease in concentration and phosphorylation of IRS-1 and IRS-2 Decrease in PI 3-kinase activity Decrease in glucose-transporter translocation Defects in activity of intracellular enzymesChronic hyperinsulinemia on vasculature 34
  • 35. Site of IR========================================Site of rest. Poss. defect Role in DM-------------------------------------------------------------Prereseptor Ins.receptor.antibody rare.Receptor Decreased number not important or affinity of ins reseptor in DMPost receptor Defect in signal most probable Reduce IRS-1 source of IR Decrease pyruvat kinase or glycogen syntaseGLUT Defective translocation Important of GLUT to cell membrane 35
  • 36. Insulin resistance mechanism in obesity, Depres 1999 36
  • 37. Central role of FFA in insulin resistance 37
  • 38. Tissue insulin resistance 38
  • 39. A unifying hypotesis of type 2diabetes 39
  • 40. Insulin Resistance in :1. Muscle : increased acumulation of fat and secondary insulin resistance, hypertriglyceridemia, and increased levels of free fatty acid2. In liver : increased hepatic glucose output3. In Brain : Increase in appetite, more obesity and further defects in hepatic glucose output4. In β-cells : defects in glucose sensing and thereby to relative insulin deficiency.All defects associated with type 2 diabetes 40
  • 41. LABORATORY TESTINGFOR INSULIN RESISTANCE 41
  • 42. Gold standard :hyperinsulinemic -euglycemic clamp glucose clamp techniqueEstimate of insulin action based on theability of insulin to limit the increase inplasma glucose concentration inresponse to continuous glucose infusionBut : impractical, expensive, time-consuming and labor intensive 42
  • 43. Variety methods of varying complexity inestimate IR (own advantages anddisadvantages).Homeostasis Model Assessment (HOMA-IR model)First described in 1985 by Matthews,strong correlation with glucose clamptechnique 43
  • 44. HOMA-IR Fasting insulin (μU/mL) x Fasting glucose (mmol/L) 22,5 HOMA-IR normal person without metabolic disorders and without obesity : 2,77 Low HOMA-IR = high insulin sensitivity = low insulin resistance High HOMA-IR = high insulin resistance = prediabetic state 44
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  • 49. Insulin TestingMetode : Chemiluminescent (sandwich)Insulin , a biotinylated monoclonal insulin-specificantibody, and a monoclonal insulin-specific antibodylabeled with a ruthenium complexTotal duration assay : 18 minutesSample : serum , Li-heparin, K3-EDTA, and sodiumcitrate plasma, avoid hemolysisSample stability : room temperature 8 h; 2-80C 24 h ;<= -200C 6 M 49
  • 50. Reference range : 6 – 26 U/mL or 43 – 186 pmol/L newborn 3 – 20 U/mL critical values > 30 U/mL  Drugs that may cause increased insulin levels : corticosteroids. Levodopa, oral contraceptives  In the second to third trimester of pregnancy, there is a relative insulin resistance with a progressive decrease of plasma glucose and immunoreactive insulin 50
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  • 54. CONCLUSION Insulin resistance : hallmark of type 2 Diabetes melitus Insulin is protein hormon; its secretion stimulate by glucose IR : decreased insulin sensitivity and or decreased responsiveness In type 2 Diabetes : defects at multiple level (receptor and post receptor) 54
  • 55. • HOMA-IR : laboratory testing for insulin resistance• HOMA-IR : formula fasting insulin and fasting glucose• Standardize insulin immunoassay 55
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