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
"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
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
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
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.
Control of inflammatory responses in adipose tissue and liver by alternatively activated M2 macrophages
Massive enlargement of mesenteric lymph nodes in Angptl4-/- mice fed HFDLichtenstein et al. Cell Metab. 2010
Angptl4 is highly induced by fatty acids in macrophagesLichtenstein et al. Cell Metab. 2010
Angptl4 prevents lipid uptake andinflammation in peritoneal macrophages
Angptl4 inhibits lipolysis and subsequent foam cell formation
Angptl4 protects against lipolysis and subsequent foam cell formation
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
How inflammation is initiated and developed in obesity Annu Rev Nutr. 2012 Mar 9.
The inflammasome: guardians of the intracellular environment Annu. Rev. Cell Dev. Biol. 2012. 28:7.1–7.25
The inflammasome controls caspase-1 activation Signal 2 NLRP3 ASC Caspase-1Signal 1
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
Caspase-1 activation in adipose tissue enhances cytokine production Low Fat Diet High Fat Diet Inactive Active ActinStienstra et al. Cell Metab. 2010
Caspase-1 activation contributes to the development of adipose tissue inflammation Wild-type HFD Casp-1-/-HFD 10x 10xStienstra et al. Cell Metab. 2010
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)
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.
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?
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
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!
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
“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
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).
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