Nutrition metabolism and inflammation an essential but dangerous crosstalk


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A lecture I gave in the Charite Berlin on April 16, 2013.

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  • A subpopulation of mice fed HFD develops NASH. Haematoxylin and eosin staining (D) and oil red O staining (E) of representative liver sections of the 4 subgroups
  • (Immuno)histochemical staining confirms enhanced inflammation and early fibrosis in HFH miceImmunohistochemical staining of macrophage activation in representative liver section of HFL and HFH mice using antibody against the specific macrophagemarker Cd68Collagen staining using fast green FCF/sirius red F3B. Staining of stellate cell activation using antibody against GFAP.
  • - Number of genes up- or down-regulated in the various subgroups in comparison to the LFL mice, as determined by Affymetrix GeneChip analysis. Genes with a p-value below 0.05 were considered significantly regulated. - Heat map showing changes in expression of selected genes involved in lipid metabolism, inflammation and fibrosis in liver. Changes in gene expression of selected genes as determined by real-time quantitative PCR. Mean expression in LFL mice was set at 100%. Error bars reflect standard deviation. Bars with different letters are statistically different (P<0.05 according to Student’s t-test). Number of mice per group: n=4 (LFL, HFL, HFH), n=6 (LFH).
  • Haematoxylin and eosin staining of representative adipose tissue sections. Immunohistochemical staining of macrophages using antibody against Cd68. Collagen staining using fast green FCF/sirius red F3B.
  • Adipose tissue mRNA expression of a selected group of genes was determined by quantitative real-time PCR after 21 weeks of dietary intervention. Mean expression in LFL mice was set at 100%. Error bars reflect standard deviation. * = significantly different from HFL mice according to Student’s t-test (P<0.05). Number of mice per group: n=4 (LFL, HFL, HFH), n=6 (LFH).
  • Nutrition metabolism and inflammation an essential but dangerous crosstalk

    1. 1. "Nutrition, metabolism and inflammation: an essential but dangerous crosstalk" Michael Müller Netherlands Nutrigenomics Centre & Nutrition, Metabolism and Genomics Group Division of Human Nutrition, Wageningen University
    2. 2. Our “paleolithic” genes + modern diets Paleolithic era Modern Times 1.200.000 Generations 2-3 Generations between feast en famine in energy abundance% Energy % Energy 100 Low-fat meat 100 Grain Milk/-products Chicken Isolated Carboh. Eggs Isolated Fat/Oil Fish Alcohol 50 Fruit 50 Meat Chicken Vegetables (carrots) Fish Nuts Honey Fruit Vegetables 0 0 Beans“Unsafe” foods = Many ligands of NRs “Safe” foods = Less ligands of NRs
    3. 3. Phenotype plasticityPhenotypic plasticity is the ability of an organism tochange its phenotype in response to changes in theenvironment (e.g. nutrition or exercise). CYP4A10 14 12 10 FC vs WT ctrl 8 6 4 2 0 WT KO WT KO WT KO WT KO WT KO WT KO WT KO WT KO WT KO ctrl WY feno C10:0TG C18:1TG C18:2TG C18:3TG C20:5TG C22:6TG
    4. 4. Genome plasticity
    5. 5. “We are what we eat and have eaten”Received, Recorded, Remembered & Revealed
    6. 6. Transcription-factor pathwaysmediating nutrient-gene interaction , RXRs TLR4
    7. 7. Understanding Nutrition How nutrients regulate our genes: via sensing molecular switches Changed organ metabolic capacityJ Clin Invest. 2004;114:94-103 BMC Genomics 2008; 9:262 Physiol. Genomics 2009 PLoS One. 2012;7(8):e43260.J Biol Chem. 2006;28:934-44 J Biol Chem. 2008;283:22620-7 Circulation 2010 J Hepatol. 2012 Dec;57(6):1370-3.Endocrinology. 2006;147:1508-16 Arterioscler Thromb Vasc Biol. 2009;29:969-74. Diabetes 2010 Am J Physiol Gastrointest Liver Physiol. 2012Physiol Genomics. 2007;30:192-204 Plos One 2009;4(8):e6796 Cell Metabolism 2010 Physiol Genomics. 2012 Mar 19;44(6):352-61.Endocrinology. 2007;148:2753-63 Hepatology 2010;51:511-522 Physiol Genomics. 2011;43(23):1307-18. Am J Physiol Endocrinol Metab. 2012BMC Genomics 2007; 8:267 Am J Clin Nutr. 2009; 90:415-24 PLoS One. 2011;6(4):e19145. Prog Lipid Res. 2012 Jan;51(1):63-70.Arterioscler Thromb Vasc Biol. 2007;27:2420-7 Am J Clin Nutr. 2009;90:1656-64 Nature 2011 May 22 Mol Cell Biol. 2013 Jan 22.Am J Clin Nutr. 2007;86(5):1515-23 Mol Cell Biology 2009;29:6257-67 PLoS One. 2012;7(12):e49868. Hepatology. 2013 Jan 21.PLOS ONE 2008;3(2):e1681 Am J Clin Nutr. 2010;91:208-17 PLoS One. 2012;7(11):e51066. J Nutr. 2013 Jan 16.BMC Genomics 2008; 9:231 BMC Genomics 2009 PLoS One. 2012;7(10):e47303. Carcinogenesis. 2013 March BMC Med Genomics. 2012 Aug 28
    8. 8. Nutrigenomics & molecular nutrition allows us to define the mechanistic framework Blood triglycerides
    9. 9. “2 hits” in Metabolic SyndromeToo much metabolic & inflammatory stress Nature Medicine 2012
    10. 10. de Wit NJ, Afman LA, Mensink M, Müller M Phenotyping the effect of diet on non-alcoholic fatty liver disease J Hepatol 2012.
    11. 11. Proximity of metabolic and immune cells in adipose tissue and liver
    12. 12. Depletion ofKupffer Cells
    13. 13. Conclusion• The data point toward important crosstalk between Kupffer cells and hepatocytes in the regulation of hepatic triglyceride storage.• The effect of Kupffer cells on liver triglycerides is at least partially mediated by IL-1b, which potently suppresses PPARa expression and activity via NF-kB dependent inhibition of PPARa promoter activity.
    14. 14. Control of inflammatory responses in adipose tissue and liver by alternatively activated M2 macrophages
    15. 15. Energy homeostasis & obesity
    16. 16. Interaction between WAT and liver tissue essential for NASH/NAFLD in C57Bl/6 mice run-in diet 20 weeks diet intervention tissue collection • stratification • plasma collection • liver on body weight frozen sections: histological feat. multiple protein assays lipid content RNA extraction: Affx microarrays 10 LFD-3 20 LFD 0 2 4 8 12 16 20 weeks quality control & 10 HFD Mouse data analysis 10% low genome pipeline fat diet 45% high 430 2.0 (palm oil) fat diet (palm oil) • ep. white adipose tissue paraffin sections: histological feat. RNA extraction: real-time PCR
    17. 17. High fat diet-induced obesity 25 LFL HFL LFH HFH 20 BW gain (g) * * 15 ** * 10 * * * 5 * 0 0 2 4 8 12 16 20 weeks under diet intervention Liver TG content Hepatomegaly ALT plasma activity 200 10 100 Ratio LW/BW (%) *** ALT activity (UI)mg TG/g liver 160 8 ** *** 80 120 ** 6 60 * 80 4 40 * * 40 2 20 0 0 0 LFL LFH HFL HFH
    18. 18. A subpopulation of mice fed HFD develops NASH
    19. 19. Immunohistochemical staining confirms enhanced liver inflammation and early fibrosis in HFH mice HFL HFH Macrophage CD68 Collagen Stellate cell GFAP
    20. 20. Upregulation of inflammatory and fibrotic gene expression in HFH responder mice
    21. 21. Adipose dysfunction in HFH mice
    22. 22. Change in adipose gene expressionindicate adipose tissue dysfunction
    23. 23. Conclusions• The data support the existence of a tight relationship between adipose tissue dysfunction and NASH pathogenesis. Duval et al. Diabetes 2010
    24. 24. Too much saturated fat & macrophages
    25. 25. Differential regulation between saturated and unsaturated fatty acids Protective Toxic 60 5.0 50 4.0 Fold changeFold change 40 3.0 30 2.0 20 10 1.0 0 0.0
    26. 26. Differential regulation between saturated and unsaturated fatty acids Angptl4 CHOP 60 5.0 50 4.0 Fold changeFold change 40 3.0 30 2.0 20 10 1.0 0 0.0
    27. 27. Chylomicron CE /TG Angptl4 LPL CE/TG FFA Chylomicron remnant
    28. 28. Examination of the role of Angptl4 under conditions of lipid overload Low fat Angptl4 +/+ High fat Low fat Angptl4 -/- High fatLichtenstein et al. Cell Metab. 2010
    29. 29. Angptl4-- mice on HFD become very illLichtenstein et al. Cell Metab. 2010
    30. 30. Altered tissue morphology in Angptl4-/- mice fed HFDLichtenstein et al. Cell Metab. 2010
    31. 31. Systemic inflammation in Angptl4-/- mice fed HFD Angptl4+/+ Angptl4-/-
    32. 32. Inflammatory response independent of microbiotaLichtenstein et al. Cell Metab. 2010
    33. 33. No effect of medium chain or PUFA TGs
    34. 34. Massive enlargement of mesenteric lymph nodes in Angptl4-/- mice fed HFDLichtenstein et al. Cell Metab. 2010
    35. 35. Angptl4 is highly induced by fatty acids in macrophagesLichtenstein et al. Cell Metab. 2010
    36. 36. Angptl4 prevents lipid uptake andinflammation in peritoneal macrophages
    37. 37. Angptl4 inhibits lipolysis and subsequent foam cell formation
    38. 38. Angptl4 protects against lipolysis and subsequent foam cell formation
    39. 39. Conclusion• A high saturated fat diet causes massive inflammation in Angptl4-/- mice originating in mesenteric lymph nodes.• MLN-resident macrophages are protected from the pro-inflammatory effect of saturated fatty acids via expression of Angptl4, which is strongly induced by chyle and fatty acids and which via inhibition of LPL prevents lipolysis of chylomicron-TG.• In the absence of this protective mechanism, feeding a diet rich in saturated fat rapidly leads to enhanced lipid uptake into MLN-resident macrophages, triggering foam cell formation and a massive inflammatory response. Lichtenstein et al. Cell Metab. 2010
    40. 40. How inflammation is initiated and developed in obesity Annu Rev Nutr. 2012 Mar 9.
    41. 41. The inflammasome: guardians of the intracellular environment Annu. Rev. Cell Dev. Biol. 2012. 28:7.1–7.25
    42. 42. The inflammasome controls caspase-1 activation Signal 2 NLRP3 ASC Caspase-1Signal 1
    43. 43. Caspase-1 is activated in adipose tissue during the development of obesity Obesity Relative gene expression Diet-induced obesity Genetically-induced obesityStienstra et al. Cell Metab. 2010
    44. 44. Caspase-1 activation in adipose tissue enhances cytokine production Low Fat Diet High Fat Diet Inactive Active ActinStienstra et al. Cell Metab. 2010
    45. 45. Caspase-1 activation contributes to the development of adipose tissue inflammation Wild-type HFD Casp-1-/-HFD 10x 10xStienstra et al. Cell Metab. 2010
    46. 46. Does inflammasome activation contribute to the development of insulin resistance? Yes Plasma Insulin levels Hyperinsulinemic euglycemic clamp study in HFD-fed Wild-type and caspase-1-/- animalsPlasma concentration (pg/ml) LFD HFD 90 8000 80 GIR (μl min-1 kg-1) 70 6000 ** * * 60 ** 50 4000 ** 40 30 2000 20 Casp1 -/- 10 WT 0 0 10 20 40 60 80 Time (minutes)
    47. 47. Why do immune cells infiltrate the adipose tissue during the development of obesity ?• Obesity promotes the presence of harmed (due to hypoxia/lipotoxicity) adipocytes that need to be removed• Cleaning up of dying/old cells: a highly regulated mechanism• Clearance of dying cells is an important fundamental process serving multiple functions in the regulation of normal tissue turnover and homeostasis• The turnover rate in human adipose tissue has been estimated to be ∼1 million cells per second each day.
    48. 48. Human adipose tissue gene expression levels after weight lossWeight loss is accompanied by a reduction in NLRP3 gene expression and downstream target genes Nat Med. 2011, 17(2):179-88 Mol Med. 2011, 17(7-8): 840–845 What drives inflammasome activation?
    49. 49. Visceral adipose tissue of mildly obese individuals is characterizedby enhanced caspase-1 activity levels and higher production of IL- 1 as compared to subcutaneous adipose tissue Stienstra et al Cell Metabolism 2012
    50. 50. Summary Summary Inflammasome-mediated caspase-1 is activated in adipose tissue during obesity Inhibition of caspase-1 in obese animals improves adipose tissue inflammation and insulin sensitivity Inhibition of the downstream target IL-1 improves glycaemic control and insulin sensitivity in patients with Type 2 Diabetes Potential novel treatment options: CRIDs, IL-37, novel targets of caspase-1 & loosing weight!
    51. 51. Human Nutrition & (anti)inflammory response
    52. 52. Fish-oil supplementation induces anti-inflammatory gene expression profiles in human blood mononuclear cells Less inflammation & decreased pro-arteriosclerosis markers = Anti-immuno-senescence Bouwens et al. Am J Clin Nutr. 2009
    53. 53. “Obese-linked” pro-inflammatorygene expression profile by saturated fatSFA diet MUFA diet • The SFA-rich diet: • Induces a pro- inflammatory obese-linked gene expression profile • Decreases expression and plasma level of the anti- inflammatory cytokine adiponectin • “Personal Transcriptomes” Van Dijk et al. AJCN 2009
    54. 54. General conclusions• (Over)nutrition and inflammation are intimately linked => non-resolving metabolic and pro-inflammatory stress (the two hits).• It will be essential to get a better understanding of the very early events that lead to non-resolving organ inflammation and the precise role of nutrition (causal or preventive) in this pathophysiological development.• We need biomarkers for organ function (“2 hit state”) to be able to specifically target and modulate.• The challenge will be the translation of the findings from mice studies to the human situation (“individual” health).
    55. 55. Sander KerstenLydia AfmanGuido HooiveldWilma SteegengaPhilip de GrootMark BoekschotenNicole de WitRinke Stienstra& many PhDsChristian TrautweinFolkert KuipersBen van OmmenHannelore DanielBart StaelsEdith FeskensLeif SanderDirk HallerEline SlagboomDaniel ThomeMihai Nitea& many more