Using nutrigenomics to study ranges and plasticity in homeostasis

Michael MüllerNetherlands Nutrigenomics Centre

& Nutrition, Metabolism and Genomics GroupDivision of Human Nutrition, Wageningen University

http://twitter.com/nutrigenomics

Our scientific challenge: What's healthy?

Metabobolic homeostasis & syndrome

Metabolic health = plasticity / flexibility

• The personal genome is the starting point & we can get comprehensive information about it but we should not underestimate the challenges of “bioinformatics & databasing”

• Health is dynamic: The property to adapt to metabolic perturbations / challenges

• Feeding / fasting => autophagy => cellular homeostasis & “exercise”

• Caloric restriction => chromatin “exercise”• Food bioactives that modulate transcription (e.g. via

nuclear receptors) or chromatin activity (nutriepigenome) => cell & organ “exercise”

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Phenotype plasticityPhenotypic plasticity is the ability of an organism to change its phenotype in response to changes in the environment => nutrition, lifestyle

Duality of biological information:Epigenetic & Genetic

Modern Nutritional Science with Nutrigenomics Quantification of the nutritional phenotype

Phenotype

Metabolome

Proteome

Transcriptome

Epigenome

Genotype

Lifestyle

NutritionFoods

Microbiota

Environment

Timely relatively modest interventions in early life can have a large effect on disease

risk later

Nutrigenomics – molecular nutrition and genomics

90,000 (?)proteins

20,210 genes

100,000 (?) transcripts

5 -10,000 (?)metabolites

Müller & KerstenNature Reviews Genetics 2003

Understanding NutritionHow nutrients regulate our genes: via sensing molecular

switches

Changed organ

metabolic capacity

J Clin Invest. 2004;114:94-103J Biol Chem. 2006;28:934-44 Endocrinology. 2006;147:1508-16Physiol Genomics. 2007;30:192-204Endocrinology. 2007;148:2753-63 BMC Genomics 2007; 8:267 Arterioscler Thromb Vasc Biol. 2007;27:2420-7

Am J Clin Nutr. 2007;86(5):1515-23PLOS ONE 2008;3(2):e1681 BMC Genomics 2008; 9:231BMC Genomics 2008; 9:262J Biol Chem. 2008;283:22620-7Arterioscler Thromb Vasc Biol. 2009;29:969-74.Plos One 2009;4(8):e6796HEPATOLOGY 2010;51:511-522

Am J Clin Nutr. 2009; 90:415-24Am J Clin Nutr. 2009;90:1656-64Mol Cell Biology 2009;29:6257-67Am J Clin Nutr. 2010;91:208-17BMC Genomics 2009Physiol. Genomics 2009Circulation 2010Diabetes 2010Cell Metabolism 2010Nature 2011

Dose-dependent effects of dietary fat on development of obesity in relation to intestinal differential gene expression in C57BL/6J

mice

PLOS one 2011

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proximal middle distal

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non-Dose-dependentDose-dependent (%)

Robust & concentration dependent effects in small intestine

Differentially regulated intestinal genes by high fat diet

PLOS one 2011

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Cellular localization and specific lipid metabolism-related function of fat-dose dependently regulated

genes

PLOS one 2011

Intestinal capacity for lipid absorption

45% FAT

10% FAT

Microbiota

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Dangerous interaction - The two hitsStress from Metabolism & Inflammation

Liver, FAT & NASH/NAFLD Nonalcoholic Fatty Liver Diseases (NAFLD):

Liver component of Metabolic Syndrome

Different stages in NAFLD progression:

Molecular events involved in NASH pathogenesis: Role of PPARa (Endocrinology 2008 & Hepatology 2010) Role Kupffer cells (Hepatology 2010)

Role of macrophages in lipid metabolism (JBC 2008; Cell Metabolism 2010)

hepatic steatosis steatohepatitis (NASH) & fibrosis cirrhosis

Experimental Design

• stratificatio

n on body weight

• liver• plasma collection multiple protein assays

RNA extraction: Affx microarrays

tissue collectionrun-in diet 20 weeks diet intervention

frozen sections: histological feat.

• ep. white adipose tissue

10% low fat diet

(palm oil)

10 LFD

10 HFD

45% high fat diet

(palm oil)

20 LFD

RNA extraction: real-time PCR

paraffin sections: histological feat.

lipid content

quality control & data analysis

pipelineMouse

genome 430 2.0

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High fat diet-induced obesity

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weeks under diet intervention

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Liver TG content

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LFL LFH HFL HFH

The HFH mice develops NASH

Immunohistochemical staining confirms enhanced liver inflammation and early fibrosis in HFH mice

Macrophage CD68

Collagen

Stellate cell GFAP

Upregulation of inflammatory and fibrotic gene expression in HFH

responder mice

Adipose dysfunction in HFH mice

Change in adipose gene expression indicate adipose tissue

dysfunction

Plasma proteins as early predictive biomarker for NASH in C57Bl/6

mice

Plasma proteins as early predictive biomarker for NASH in C57Bl/6

mice

Multivariate analysis of association of protein plasma concentrations with final liver triglyceride content

Conclusions

• Our data support the existence of a tight relationship between adipose tissue dysfunction and NASH pathogenesis.

• It points to several novel potential predictive biomarkers for NASH.

Duval C, Thissen U, Keshtkar S, Accart B, Stienstra R, Boekschoten MV, Roskams T, Kersten S, Müller M.

Adipose tissue dysfunction signals progression of hepatic steatosis towards nonalcoholic steatohepatitis in C57BL/6

mice. Diabetes. 2010;59:3181-91.

Plasma Protein Profiling Reveals Protein Clusters Related to BMI and Insulin Levels in Middle-Aged Overweight Subjects

AIM• Associate plasma protein profiles with BMI• Identify potential marker profile of early

disease state

. PLoS One. 2010 Dec 23;5(12):e14422

Measurements

• Rules Based Medicine (Austin, USA)• Multiplex immunoassay• In total 124 proteins measured

– Involved in diseases, inflammation, endothelial function and metabolism

. PLoS One. 2010 Dec 23;5(12):e14422

We are different: improved phenotyping necessary to reveal phenotype clusters

. PLoS One. 2010 Dec 23;5(12):e14422

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

Very personal conclusionsHow to keep our metabolic plasticity/health

• Identify chronic (non-resolving) stress using systems “perturbation” tests & deep genomics-based phenotyping

• Solve it!– Less Inflammation– Less Metabolic Stress (sat. fat, lipogenic foods)– More Exercise (muscle & other organs) with a

“challenging” lifestyle & food pattern– Eat less from time to time

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2 Meals a day, work as long as possible & embrace challenge

Walter Breuning (1896 - 2011)

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