8/14/2016. Diet, Gut Bacteria, and Metabolic Disease: Strategies to Promote Healthy Microbial Communities. Outline. The Human Microbiome:

Transcription

1 8/14/216 Diet, Gut Bacteria, and Metabolic Disease: Strategies to Promote Healthy Microbial Communities Kristina Martinez PhD, RD Postdoctoral Research Scholar University of Chicago Chicago, IL Outline Gut Microbiota and Microbiota Research Obesity, Diet, and Gut Microbiota Mechanisms Lipid Absorption Strategies to Promote Healthy Microbiota Prebiotics Supplementation Study Probiotics FMT Postbiotics Disclosure to Participants The Human Microbiome: Excitement in the scientific community No conflicts of interest to disclose. NIH: $115M over 5 years 1

2 8/14/216 Major Phyla of Bacteria in the Gut: Bacteroidetes Firmicutes Actinobacteria Proteobacteria Verrucomicrobia What is the gut microbiota? Characteristics of Gut Microbiota Gut microbiota Collection of microbes or the number of microbial cells harboring the gastrointestinal tract May include bacteria, yeast, archaea, and viruses Gut microbiome Collective number of genes within the gut microbiota Microbial dysbiosis Disruption of normal microbial communities or imbalance of good vs bad bacteria (i.e., pathobionts) How does our microbial community compare to us? Why do we need gut microbes? A vital organ? Pathogen protection Immune development Digestion and caloric salvage Gut development Energy balance Drug metabolism Regulatory functions 2

3 8/14/216 How do we determine the functional role of gut microbes? How do we study gut bacteria: 16S rrna Sequencing 16S ribosomal RNA gene V1 V2 V3 V4 V5 V6 V7 V8 Schematic representation of the structure of 16S rrna gene. Boxes represent constant regions and bars represent variable regions of 16S rrnagene Tom Barry, et al. A general method to generate DNA probes for microorganisms, Biotechnology (N Y). 199 Mar;8(3): Germ-free mice are used to study the effect of gut microbiota on the host. Other Technologies Metagenomics- Determine entire genome of bacterial communities Metabolomics- Determine collection of metabolites from bacterial communities Metatranscriptomics- Determine the collection of gene transcripts upregulated in bacterial communities Karlsson F et al. Diabetes 213;62: Copyright 214 American Diabetes Association, Inc. Classic Microbiology Culturing Techniques Factors Contributing to the Rise in Obesity 3

4 8/14/216 Factors Affecting Gut Microbial Structure Animal-based diet alters microbial communities within 1 week Diet Environment Host Genetics and Metabolic State David et al. 214 Nature. 55(7484): Microbial dysbiosis occurs in a genetic model of obesity Diet regardless of genotype alters gut microbial structure Firmicutes Proteobacteria Bacteroidetes Verrucomicrobia Actinobacteria Ley et al. Proc Natl Acad Sci USA (31): Carmody et al. Cell Host & Microbe : Fecal Microbiota Transplant (FMT) from lean donors resolves insulin resistance in obese subjects 4 weeks Turnbaugh et al., Sci Trans Med. (29) 6 weeks Improved symptoms of Type II Diabetes and Metabolic syndrome Lean male donors Obese male recipients BMI 3 Vrieze et al., Gastro. (212) 143:

5 8/14/216 Summary Many factors affect microbial assemblage including: Diet Genetics (and perhaps Diet > Genetics?) Metabolic State (i.e., Obesity) Do mice have impaired lipid absorption? Microbial communities from lean healthy donors may protect against symptoms associated with type 2 diabetes Microbial communities can transfer an obese phenotype if given to a lean donor Rabot et al, FASEB Journal, Vol How does this occur? D zebrafish have increased epithelial lipid vs What are the mechanisms? Pediatr Gastroenterol Hepatol Nutr. 213 March; 16(1): Semova et al. 212 Cell Host & Microbe mice are resistant to diet-induced obesity Fat Digestion and Absorption is Regulated on Multiple Levels Adequate lipid absorption relies on endocrine signaling, emulsification from bile, and absorptive capacity of small intestinal enterocytes Leone et al. Cell Host Microbe. 215 May 13;17(5): Hui DY and Howles PH. (25) Sem Cell Dev Biol. 16:183 5

6 8/14/216 Pathways for Intestinal lipid Absorption mice have decreased FA transport across brush borders Jejunum Ileum 3 H Oleic Acid Uptake nmol/mg Protein H Oleic Acid Uptake nmol/mg Protein Iqbal J, Hussain M M Am J Physiol Endocrinol Metab 29;296:E1183-E1194 Overall Goal: To determine if 1) gut microbes regulate lipid absorption in the small intestine of mice and 2) the mechanisms underlying this interaction. Hypothesis: mice have impaired lipid absorption on a high fat diet due to reduced hormonal signaling from the gut. mice have altered intestinal expression of lipid transporters cd36 Gene Expression (Relative to gapdh) Duodenum cd36 Jejunum Ileum mgat2 Gene Expression (Relative to gapdh).4.2. Duodenum mgat2 Jejunum Ileum Unpublished data, Chang Lab mice have a decreased rate of triglyceride absorption Experimental Design LF PUFA MF LF PUFA MF 3H Triolein (dpms/ml) Plasma 3H Triolein Hours 14C Cholesterol (dpms/ml) 1 5 Plasma 14C Cholesterol Hours Week BW BW BW FI FI BW FI BW FI Unpublished data, Chang Lab 6

7 8/14/216 mice do not gain weight on high milkfat (MF) diet A Body Weight (% Baseline) B Epididymal Fat Weight (% Body Weight) Body Weight - Mice Week Epidydimal Fat Weight Body Weight (% Baseline) Mesenteric Fat Weight (% Body Weight) Body Weight - Mice Week Mesenteric Fat Weight Unpublished data, Chang Lab MF SO LF MF SO LF mice have impaired pancreatic secretion of lipase and amylase Lipase Activity Pancreas Lipase Activity Jejunum Unpublished data, Chang Lab H2 CO + mice have larger gallbladders and elevated stool TG vs mice Gallbladder Length entionalizing mice with MF microbiota restores CCK expression cck secretin cck Gene Expression (Relative to gapdh) D secretin Gene Expression (Relative to gapdh) D Unpublished data, Chang Lab Corn Oil (CO) Challenge Experimental Design Fast Fast CO or H2O Gavage Harvest Hours CO or H2O Gavage Harvest Hours Pancreas and intestinal Luminal contents collected Pancreas and intestinal Luminal contents collected entionalization of mice with MF microbes restores TG and Chol absorption to a greater extent than LF microbes 3H Triolein (dpms/ul) Plasma 3H Triolein 1h 3h 5h 7h D MF-MF D MF-LF D LF-LF 14C Cholesterol (dpms/ul) Plasma 14C Cholesterol 1h 3h 5h 7h D MF-MF D MF-LF D LF-LF 7

8 8/14/216 HF diet-induced dysbiosis increases lipid absorption through impacting digestion and lipid uptake How do prebiotics promote metabolic benefits? Increase expression of antimicrobial peptides against deleterious bacteria Short chain fatty acid (SCFA) production and SCFAmediated stimulation of intestinal gluconeogenesis and increased epithelial integrity Increase satiety and insulin sensitivity via release of gut peptide hormones including polypeptide YY (PYY) and glucagon-like peptide (GLP)-1, respectively Decrease relative abundance of pathogenic bacteria and increase abundance of commensal bacteria Targeting the Gut Microbiota Prebiotics Probiotics Synbiotics Fecal Microbiota Transplant (FMT) Postbiotics Study Objective Determine if a dietary prebiotic (e.g., inulin) can remodel the gut microbiota (e.g., composition and function) in a well-defined human sample population (e.g., metabolic syndrome), while demonstrating possible effects on host metabolism (e.g., improved glucose tolerance). Hypothesis Daily supplementation with 8 g of a FOS-rich prebiotic for 3 weeks will improve glucose tolerance in pre-diabetic borderline obese males. Prebiotics Definition: Prebiotics are foods that promote growth of beneficial bacteria Resistant to gastric acidity Non-digestible; fermented by bacteria Promote healthy microbial communities (Viladomiua et al. 213) Non-digestible carbohydrates Inulin-type fructans Oligofructose Galactans Examples Prebiotin Oligofructose Inulin Study Design Population: n = 8 males Overweight/obese (BMI 25-35) Prediabetic (FBG > 1) Intervention: 3 week single arm intervention Study product: Prebiotin FOSenriched 1% inulin Comparisons: Baseline vs End of Study Experimental Outcomes: Host Fasting blood glucose, insulin, GTT including glucose and insulin measurements Dietary intake (3 day diet record) Lipid Panel Microbe 16s rrna sequencing Biolog Targeted metabolites: SCFAs, H2S, bile acids 8

9 8/14/216 Stool Collection 2 collection kits provided at baseline and end of study 2 samples to be collected within a 3 day period Time and date noted Provided 3 day bowel habits diary prior to and at end of 3 week supplementation Demographics/Baseline Measures 4 non-hispanic white, 3 African American, 1 Asian/Pacific Islander Mean Age: /- 2.9 BMI: 3.2 +/- 1.2 Body Fat % (DEXA) /- 1.4% FBG /- 2.7 mg/dl at screening Matsuda Insulin Sensitivity index Change in MISI Relative to Baseline Matsuda Insulin Sensitivity Index Responders NonResponders Change in MISI Relative to Baseline Matsuda Insulin Sensitivity Index Responders NonResponders Prebiotic Supplementation A commercially-available oligofructose-rich fiber supplement Used in at least two NIH-funded studies. 8 g FOS-inulin per day for 3 weeks. Maximum tolerable amount recommended by the company and found to be an effective dose in promoting bifidogenic activity in previous studies (Glen Gibson 27). All Fiber product purchased for the study came from the same lot number. Microbial communities differs b/t responders and non-responders Responder Non-Responder P=.3 Responders displayed improved glucose and insulin tolerance Modest phylum level shifts are in responders but not non-responders Glucose (mg/dl) Responders GTT Glucose Glucose (mg/dl) Non-Responders GTT Glucose Day Day Time Time Insulin (miu/ml) Responders GTT Insulin Insulin (miu/ml) Non-Responders GTT Insulin Day Day Time Time 9

10 8/14/216 Can the microbiota confer improved glucose tolerance in naïve mice fed an obesogenic diet? Acknowledgements UChicago: Chang Lab Department of Medicine Eugene Chang - PI Vanessa Leone Anu Nadimpalli Mark Musch Daina Ringus Joe Pierre Alex Bobe Noelle Patno Patty Ojeda Kali Deans Biofortis Merieux NutriSciences: Chad Cook Deanne Liska Argonne National Laboratory NIH - Funding Source T32DK774 and DK97268 Conclusions 5% of subjects displayed improved glucose tolerance and/or insulin sensitivity ( Responders ) Responders appeared to have alterations in microbial structure whereas Non-Responders did not Inulin significantly increased butyrate and acetate levels from baseline to endpoint but no differences in levels between responders and non-responders Inulin may promote mild improvement in glucose tolerance in some subjects after only 3 weeks of supplementation Thank You & Embrace Your Microbiota! Overall Conclusions Intimate link between gut microbes and host physiology including energy extraction and metabolism Several mechanisms are being investigated to understand this relationship Many therapeutic options to restructure gut microbiota are available and on the rise Much research is needed to solidify the best approach in using these therapies 1