How gut microbiota might influence fat storage and metabolism

ILSI SEAR Australasia Symposium

Future Directions in Saturated Fats: composition and metabolism

School of Molecular Bioscience Andrew Holmes

Microbes that colonize you as an infant give rise to a stable

community that influences you for the rest of your life. �

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1. Overview of our gut microbiota?�"How many cells? What kinds of species?�

"How we interact with it �

2. Why microbes matter. �"Microbes influence our gene expression patterns�

"Microbial activity influences our metabolism�

3. Detecting links with our microbiota�"Diversity measurement and scale "�

"An obesity-associated microbiota �

"High-fat diet microbiota �

Flint H et al. (2008) Nature Reviews Microbiology 6, 121-131 Polysaccharide utilization by gut bacteria: potential for new insights from genomic analysis

Total Energy content �

< 104 g-1

103 - 105 108

1011 - 1012

Stomach

Duodenum

Colon Jejunum

Ileum

The available energy is in polysaccharides not degraded by host enzymes or secretions

Absorption of host-accessible sugars, lipids

and amino acids is

‘complete’ by the ileum

The dominant gut-associated bacteria are members of the

Bacteroidetes and Firmicutes.

Specialize in degrading digestion-resistant

carbohydrate.

Host-microbe interaction is greatest in intestines and reflects: �

What microbes are made of (surface components, breakdown products)

What they do (metabolites produced and substrates consumed)

How our tissues sense the gut compartment (dendritic cell projections, PRRs)

What we export to the gut lumen (mucin, sIgA, bile, peptides)

What crosses in from the gut lumen (sugars, lipids, fatty acids, amino acids, microbes)

Over 1000 species, but >90% in just 2 phyla and share many features�Does diversity matter?�

Figure from Flint H et al. (2008)

Main energy source for colonocytes. Net production influences epithelial health

and gut barrier function

Absorbed and transported to liver. Net production has

potential to influence host energy balance.

Different bacteria have different strategies to degrade polysaccharide and use different pathways to extract energy from the solubilized

sugars – giving different end products�

See effect of microbes by comparing germ free with experimentally- or naturally-colonized animals�

Foetus�

Sterile�

Neonate� Child (mini adult)�

Sterile�

Birthday �Inoculation day �

Diverse community�(>1000 “species”)�

Sterile�

Defined community�(1 or more species)�

Natural�

Microbiota �

Germ-free�

Gnotobiont �

(known microbes)�

No community�

Gnotobiont models: Some features of GF animals do not need complex community�

L.V. Hooper and J.I. Gordon (2001) Glycobiology 11: 1R–10R.

Germ free�

Conv colonized�

B. theta. colonized�

Stappenbeck et al., (2002) Proc. Natl. Acad. Sci. U. S. A. 99: 15451–15455.

Conv. colonized Germ free

Epithelial fucosylation �

Intestinal vascularization�

Gnotobionts don’t do everything: microbe community required for normal phenotype�

System � GF phenotype �Nutrition ""vitamins K & B required�

""lower body fat, but eat more! �Metabolism "increased bilirubin �

""increased fecal nitrogen �

"decreased fatty acids�Intestinal "reduced mass�

"increased goblet cells�

"altered lectin composition �

Immune "decreased sIgA �

"low systemic Ig �"GALT undeveloped�

"Altered cytokine profile "�

www.narl.org.tw/en/topic/topic.php?topic_id=1

From Backhed et al. (2004) PNAS 101: 15718

From: Turnbaugh et al. (2006) An obesity-associated gut microbiome with increased capacity for energy harvest. Nature 444, 1027-131

Caloric content of feces is lower in obese mice - more energy has been extracted.�

Higher fatty acid concentrations in cecum of obese mice - more food converted to absorbable forms�

Association studies: Some microbiota-influenced properties linked to health issues (obesity, IBD).�

Microbiota: �

association �or�

cause?�

Transplantation experiments show composition of microbiota affects outcome�

Germ free ob/ob mouse

Obesity-associated microbiota (normal

ob/ob mouse)

Lean-associated microbiota (normal

mouse)

ob/ob mice conventionalized with

a LAM have less increase in body fat

From: Turnbaugh et al. (2006) Nature 444, 1027-131

ob/ob mice conventionalized with an OAM have greater increase in body fat

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Regulatory poise of immune system is influenced by microbiota composition. Inflammatory response contributes pathology

Immune system may influence microbiota composition

Host Factors (Genetic background, immunological state)

Interventions (Diet, pharmaceuticals, exercise)

Gut Microbiota (composition, activity)

Human Health

Microbiota composition influences metabolism and absorption: Diets may not deliver the anticipated calorie deficit

Oral drug delivery may not give the anticipated dose Drug metabolism may modulate the anticipated biological effect

Diet differences may influence microbiota composition Antibiotic may alter microbiota composition

The problem: Interventions give variable and/or transient

outcomes that are not explained by host-intervention

interaction axis

Working hypotheses

What are the implications of gut microbial community structure for human health?�

The emergent properties of the GUT MICROBIOME are such that therapeutic management of many diseases and disorders requires consideration of gut

microbiota - and could also involve management of gut microbiota�

Systemic inflammation,

dyslipidemia and insulin resistance

Systemic inflammation,

dyslipidemia and insulin resistance

Liver

Cytokine - & fat-laden

PortalCirculation

Inflamed & steatotic liver

Diet (caloric overload & pro -inflammatory profile)

Genetics Obesity

Gut Dysfunction

Microb

iota

profile

Impaired barrier

function

Inflamm

ation

Mesenteric fat

Endotoxins

Liver

Leaky gut

Lipid & macrophage accumulation

Systemic inflammation,

dyslipidemia and insulin resistance

Systemic inflammation,

dyslipidemia and insulin resistance

Liver

Cytokine - & fat-laden

PortalCirculation

Inflamed & steatotic liver

Diet (caloric overload & pro -inflammatory profile)

Genetics Obesity

Gut Dysfunction

Microb

iota

profile

Impaired barrier

function

Inflamm

ation

Mesenteric fat

Endotoxins

Liver

Leaky gut

Lipid & macrophage accumulation

Lam et al (2011) Role of the gut in visceral fat inflammation and metabolic disorders Obesity (online early)

Increased fat storage - obesity�

Genetic factors�

Poor diet (excess energy; inappropriate macronutrient ratio)�

Selective pressure on microbial community�

Sampling intensity: At any one point in time

there are an estimated 1014 bacterial cells in the GIT.

Sampling pattern 2: Different anatomical regions

of the gut have different functions – and different

microbiota

Sampling pattern 1: There is continual turnover

of the microbiota by ingestion, growth and

defecation

Monitoring and measuring associations between gut microbiota and animal phenotype�

1013  g-­‐1  

The 16S rRNA gene is sampled by PCR - we count PCR products

 No. different sequence types ≈ No. biological types

 Abundance of sequence ≈ abundance of cells

 Sequence relationships correspond to evolutionary

history.

Cells with different properties have different genomes�

We can isolate cells and measure properties or isolate DNA and

measure sequences�

Cells ‘Isolated’ genes

http

://bi

olog

y.cl

c.uc

.edu

/fank

haus

er/L

abs/

Isolated colonies

Measuring microbial community structure: evol distance and discrete state methods�

‘Populations’: fingerprinting by DGGE�

Zoetendal et al. (1998) Appl. Env. Microbiol. 64:3854-3859 �

‘Clades’: phylogenetic trees�

Ley et al. (2006) Cell 124:837-848 �

Taxonomic resolution, biological relevance and sampling power�

Microbial community varies between

individuals�Sub-Species correlated

with particular metabolites �

From: Li et al. (2008) Proc. Natl. Acad. Sci. USA 105, 2117-2122

Different microbiome properties will show patterns at different

levels of resolution �

Published studies have implicated low ratio of Bacteroidetes:Firmicutes with obesity�

Fig

1. Le

y et

al.

(200

6) N

atur

e 44

4:10

22�

Obesity and community structure outcomes after dietary intervention �

12 individuals, 4 timepoints, ca 400 sequences per sample, (18,348 in total)�

6 individuals on low Carb diet �

6 individuals on low fat diet

The Korean diet study: do gut microbiota influence diet success for weight loss?�

The study� A clinical trial to examine the acceptability and efficacy of a traditional Korean diet for Westerners (PIs Caterson, Kang).�

Project’s microbiology aims� Measure the correlation between gut community differences and weight loss and metabolic health outcomes�

 Identify diet-induced changes in key gut microbes �

70 patients

35 Western Diet

35 Korean Diet

12 weeks

Diet-induced changes KD

vs WD cohorts

Microbiota-influenced outcomes low wt loss vs high

wt loss

No significant difference in weight loss outcome or metabolic improvement after 12 weeks between diets�

Calorie-restricted diet (Western) shows trend to shifting Bacteroides:Firmicutes populations but High fibre diet (Korean) did not �

Pre-diet community

Diet exposed community

(n= 9)

(n= 9)

Which aspects of community change reflect diet-selection for ‘better’ microbes�

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Three closely related

members of the Firmicutes

respond differently to high fat diet

Zhang et al. (2010) The ISME Journal 4:232-241

High fat diet induces obesity and metabolic disease in mice

The change in gut microbiota over time is proportional to weight loss�Te

chni

cal R

eplic

ates

Te

mpo

ral R

eplic

ates

3.9 % change

6.7 % change

-16.2% -11.6%

% change over diet period

222 201

18.8% 21.4%

Subject

Weight loss

▲ Western diet

▲ Korean diet

0 12wk 0 12wk Time

Community change was important – is it: �

What the organisms do?�or�

How the host reacts to them?�

T5KO mice develop obesity and metabolic disease: Mice on the same diet, but differing in ability to perceive microbe-associated molecular patterns.

M Vijay-Kumar et al. Science 2010;328:228-231

Points to consider�

Host Factors (Genetic)

Environment Gut Microbiota

Metabolic Health

QTL mapping in mouse intercross line shows host genetic factors influence

microbiota relative abundance.�

We acquire our microbes from our immediate environment – some times of our life are more

critical than others.�

Ingesta impacts community structure and vice versa�

We require our immune system for stable microbial

community structure�

Ingesta (food and bioactive molecules) impact many

aspects of our health and metabolism�

Acknowledgements �

School of Molecular Bioscience, University of Sydney�Yi Vee Chew – (KD study, mouse nutritional geometry)�Feyza Colakoglu – (mouse nutritional geometry)�Connie Ha – (KD study, HFD and barrier function)�

Korean Diet Study (Funded by Korean Govt) �

Ian Caterson, Nick Fuller (and many more) – Institute for Obesity, Nutrition and Exercise, University of Sydney�

Jae-Heon Kang, Institute for Clinical Nutrition, Inje University, Seoul�

High Fat Diet and Gut Barrier Function �

Len Storlien, Yan Lam - IONE, University of Sydney�

Nutritional Geometry�

Steve Simpson – School of Biological Sciences, University of Sydney �Aisling McMahon, Sam Solon - ANZAC Institute�