Role of metagenomic determinants in childhood obesity and body fat partitioning

Transcription

1 The American Association of Clinical Endocrinologists AACE 9th Annual Meeting on August 10-12, 2018 Role of metagenomic determinants in childhood obesity and body fat partitioning Nicola Santoro MD, PhD Yale University

2 Outline of this presentation Clinical characteristics and complications of pediatric obesity Role of the gut microbiome in the pathogenesis of obesity Evidence of the effect of gut microbiome on obesity complications: cardiovascular diseases. Future Perspectives

3 Outline of this presentation Clinical characteristics and complications of pediatric obesity Role of the gut microbiome in the pathogenesis of obesity Evidence of the effect of gut microbiome on obesity complications: cardiovascular diseases. Future Perspectives

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5 From: Trends in Obesity Prevalence Among Children and Adolescents in the United States, Through JAMA. 2016;315(21): doi: /jama

6 Patterns of Overweight at 7 Years of Age, 13 Years of Age, and Early Adulthood (EA) and the Risk of Type 2 Diabetes at 30 to 60 Years of Age Bjerregaard LG NEJM 2018

7 Are all obese children equal? Complication Prevalence Metabolic Syndrome 35%-50% Dyslipidemia 30%-45% Prediabetes 12%-15% Type 2 diabetes 2%-5% NAFLD 30%-45% Weiss R et al. NEJM 2004 Sinha R et al. NEJM 2002 Trico D et al. Hepatology 2018

8 Cellular and molecular differences in the SAT in the obese adolescent with a low VAT/SAT ratio and a high VAT/SAT ratio. Caprio S et al. Gastroenterology 2017

9 Increased inflammatory gene expression and macrophage infiltration in abdominal SAT of the high VAT/VAT+SAT group Caprio S et al. Gastroenterology 2017

10 Insulin-resistant obese adolescents have increased lipolysis and impaired suppression of HGP associated with increased WAT IL6 concentrations. Caprio S et al. Gastroenterology 2017

11 Ectopic fat distribution is a major risk factor for developing metabolic complications Normal Liver NAFLD

12 Model of lipid flux through the liver Glycogen Glycolysis DNL TG synthesis DNL 26.1% De novo lipogenesis is the synthesis of fatty acids from carbs Large VLDL Modified from Donnely KL et al., Journal of Clinical Investigation 2005

13 NAFLD Non alcoholic fatty liver disease (NAFLD) is characterized by macrovesicular steatosis in more than 5% of hepatocytes in an individual without significant history of alcohol intake. NAFLD encompasses a range of disease severity spanning from simple steatosis to non alcoholic steatohepatitis (NASH), which in turn can progress to cirrhosis. Non alcoholic fatty liver disease (NAFLD) has emerged as one of the most common complications of childhood obesity affecting almost 30% of obese children. Schwimmer JB et al Pediatrics :

14 NAFLD PROGRESSION Steatosis Steatohepatitis Cirrhosis FFA

15 Kaplan Meier survival curve of children with non-alcoholic fatty liver disease (NAFLD) as compared to the general United States population of same age and sex. Feldstein AE et al Gut 2009;58:

16 Why a Pediatric Endocrinologist should study NAFLD?

17 Fatty Liver as and Insulin Resistance We studied 23 obese adolescents with high HFF (HFF >5.5%) and 20 obese adolescents with low HFF (HFF <5.5%), matched for age, Tanner stage, BMI z score, and percentages of body fat, visceral fat, and IMCL Percent suppression of hepatic glucose production and lipolysis and muscle insulin sensitivity in low ( ) and high ( ) liver fat content groups, during the low- and high-dose insulin infusion. D'Adamo E et al. Diabetes Care 2010;33:

18 Fatty Liver is a Metabolic Disease 1. NAFLD is associated with deterioration of insulin sensitivity and glucose tolerance independent of visceral and intra-myocellular lipid. 2. Subjects with NAFLD show a higher prevalence of metabolic syndrome, pre-diabetes and type 2 diabetes than age, gender, ethnicity and obesity matched individuals.

19 Childhood Obesity 1. Childhood obesity is a complex disease. 2. Childhood obesity is characterized by a variety of complications that can be explained by environmental and genetic factors.

20 Genetics and Metagenomics determinants of early onset obesity In 1986, Albert Stunkard offered the most compelling evidence yet that one's weight could be largely determined by one's parentage (1986). Stunkard and colleagues used a Danish adoption registry of 540 adults, the majority of whom had been adopted by the age of 1 between 1927 and They found that, despite having shared an environment with their adoptive parents, the adoptees' body-mass indexes approximated those of their biologic parents rather than their adoptive parents. Stunkard AJ et al. NEJM 1986

21 Summary of loci found by genome-wide association studies to be associated with obesity Total variance of adiposity explained ranges from 0.34% to 1.34% according to the study Fall T et al. Mol Cell Endocrinol Jan 25;382(1):

22 Outline of this presentation Clinical characteristics and complications of Pediatric Obesity Role of the gut microbiome in the pathogenesis of obesity Evidence of the effect of gut microbiome on obesity complications: type 2 diabetes, cardiovascular diseases. Future Perspectives

23 What is the role of Microbiota in human obesity? Human Genome ~ Protein coding genes ~ 6 billion base pairs per diploid cell Microbiome The combined genomes of the gut microbiota the microbiome contain 100-fold more genes than the human genome Microbiota Microbial species Microbiome Microbial gene repertoire

24 Factors influencing the composition of the human gut microbiota Graf D et al. Microb Ecol Health Dis. 2015; 26: /mehd.v

25 Next Generation Sequencing 16S gene sequencing METAGENOME Phyla and genera Species and Pathways

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27 Germ free mice

28 Obesity is associated with a significant decrease in the level of diversity Lean Obese Turnbaugh PJ Nature 457: ; 2009

29 The gut microbiota as an environmental factor that regulates fat storage Conventional Transplanted with Microbiota of donors Germ Free Mice allowed to acquire a microbiota from birth to adulthood CONV-R donors Germ free Fredrik Bäckhed et al. PNAS 2004;101:44:

30 The gut microbiota as an environmental factor that regulates fat storage CONV-R = Mice allowed to acquire a microbiota from birth to adulthood CONV-D =CONV-R donors GF = germ free Fredrik Bäckhed et al. PNAS 2004;101:44:

31 A 14-d conventionalization of WT GF B6 mice increases circulating leptin levels and decreases sensitivity to insulin. CONV-D =CONV-R donors GF = germ free Fredrik Bäckhed et al. PNAS 2004;101:44:

32 Conventionalization induces hepatic lipogenesis and nuclear import of the basic helix-loop-helix transcription factor ChREBP. Liver sections CONV-D =CONV-R donors GF = germ free Liver gene expression Fredrik Bäckhed et al. PNAS 2004;101:44:

33 GF mice are protected against diet-induced obesity Fredrik Bäckhed et al. PNAS 2007;104:3:

34 Common Phyla associated with obesity Germ Free mice Fat Firmicutes Bacteroidetes Conventionalized Mice Fat Firmicutes Bacteroidetes

35 Correlation between body-weight loss and gut microbial ecology Ley RE et al. Nature Dec 21;444(7122):

36 Ridaura VK et al. Science 2013

37 Replication of human donor microbiota phenotype in gnotobiotic mice. Ridaura VK et al. Science 2013

38 Effect of Low Saturated Fat (SF) -High Fruit and Vegetables (FV) intake and High SF-Low FV intake on Metabolic Phenotype SF= saturated fat; FV= Fruit and Vegetables Ridaura VK et al. Science 2013

39 Does the gut microbiome play any role in the development of pediatric obesity and in the accumulation of fat in canonical (visceral and subcutaneous) and ectopic (liver) depots?

40 Roles of Birth Mode and Infant Gut Microbiota in Intergenerational Transmission of Overweight and Obesity From Mother to Offspring Tun HM JAMA Pediatr Apr 1;172(4):

41 Association of Exposure to Formula in the Hospital and Subsequent Infant Feeding Practices With Gut Microbiota and Risk of Overweight in the First Year of Life Forbes JD, et al. JAMA Pediatr. 2018

42 Risk of Overweight in Adolescence by Duration of Breastfeeding in Infancy Gillman MW et al. JAMA May 16;285(19):2461-7

43 Association of Exposure to Formula in the Hospital and Subsequent Infant Feeding Practices With Gut Microbiota and Risk of Overweight in the First Year of Life Forbes JD, et al. JAMA Pediatr. 2018

44 Association of Exposure to Formula in the Hospital and Subsequent Infant Feeding Practices With Gut Microbiota and Risk of Overweight in the First Year of Life Breastfeeding may be protective against overweight, and gut microbiota may contribute to this effect. Formula feeding appears to stimulate changes in microbiota that are associated with overweight, whereas other complementary foods do not. Subtle microbiota differences emerge after brief exposure to formula in the hospital. Forbes JD, et al. JAMA Pediatr. 2018

45 84 children and adolescents studied; the region V4 of the 16S rrna gene was sequenced with Illumina MiSeq. This heatmap shows the abundance (shades of grey) of common Operational Taxonomic Units (OTU) with samples as rows and OTU as columns. Goffredo M et al JCEM 2017

46 Relative abundance of Phyla among BMI categories 15 non-obese, 7 overweight, 27 obese and 35 severely obese Goffredo M et al JCEM 2017

47 Association between the major Phyla composing the human gut microbiota and BMI F/B Rank 100 r 2 = 0.11; P = Bacteroidetes Rank 100 r 2 =0.14; P = Actinobacteria Rank 100 r 2 =0.07; P = B M I (kg/m 2) B M I (kg/m 2) B M I (kg/m 2) Goffredo M et al JCEM 2017

48 Differences in gut flora capability to ferment carbohydrates between lean and obese 13 C-Fructose was prepared at 250 µm. One hundred microliters of each 13 C-substrate was added to the diluted fecal sample. Goffredo M et al JCEM 2017

49 Association between obesity, body fat partitioning and SCFA Acetate (mm/l) Propionate (mm/l) Butyrate (mm/l) Visceral fat (cm 2 ) r2=0.108, P=0.002 r2=0.246, P<0.001 r2=0.207, P<0.001 Subcutaneous fat (cm 2 ) r2=0.157, P=0.001 r2=0.326, P<0.001 r2=0.290, P<0.001 Hepatic fat content (%) r2=0.001, P=0.611 r2=0.040, P=0.075 r2=0.068, P=0.019 Goffredo M et al JCEM 2017

50 Assessment of Hepatic De Novo Lipogenesis In vivo DNL can be measured by stable isotopes like 2 H2O Malonyl CoA Isolate large VLDL after 20 UC Measure D2 incorporated in Palmitate Palmitate (C16H32O2)

51 Relationship between SCFA and hepatic de novo lipogenesis A B C Fasting DNL% r=0.67; p= Acetate (um/l) Fasting DNL% r=0.38; p= Proprionate (um/l) Delta hepatic DNL r=0.55; p= Butyrate (um /L) Goffredo M et al JCEM 2017

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53 High fat fed rats exhibit increased whole-body acetate turnover Perry R et al Nature 2016

54 The contents of the colonic lumen are the primary source of acetate in high fat fed rats Whole Body Acetate turnover (micmol/kg/*min) Perry R et al Nature 2016

55 Acetate turnover drives Glucose Stimulated insulin Secretion Perry R et al Nature 2016

56 Acetate drives increased GSIS via parasympathetic activation Perry R et al Nature 2016

57 Perry R et al Nature 2016

58 Outline of this presentation Clinical characteristics and complications of Pediatric Obesity Role of the gut microbiome in the pathogenesis of obesity Evidence of the effect of gut microbiome on obesity complications: cardiovascular diseases. Future Perspectives

59 Tang WH et al. NEJM Apr 25;368(17):

60 The importance of methylamines is increasingly being recognized with respect to liver, cardiometabolic and, more recently, neurological disorder. Choline is processed into phosphatidylcholine (lecithin) by the host, which assists in excretion of VLDL particles from the liver. This process prevents hepatic accumulation of triglycerides and hypertriglyceridemia In the intestine choline can also be converted to trimethylamine (TMA) by bacteria. TMA is converted in trimethylamine N-oxide (TMAO) in the liver by the hepatic flavin-containing monooxygenases.

61 Plasma TMAO levels depend on the gut flora Tang WH et al. NEJM Apr 25;368(17):

62 Kaplan Meier Estimates of Major Adverse Cardiovascular Events, According to the Quartile of TMAO Level. Tang WH et al. NEJM Apr 25;368(17):

63 Tang WH et al. NEJM Apr 25;368(17):

64 Bidirectional communication between gut and liver Stanhope KL Obes Rev May 14. doi: /obr.12699

65 Outline of this presentation Clinical characteristics and complications of Pediatric Obesity Role of the gut microbiome in the pathogenesis of obesity Evidence of the effect of gut microbiome on obesity complications: type 2 diabetes, cardiovascular diseases and fatty liver disease. Future Perspectives

66 Chen Z et al J Clin Invest 2014; 124:

67 Although a number of therapeutic molecules could be potentially biosynthesized by bacteria for the treatment of obesity, this study focused on N-acyl-phosphatidylethanolamines (NAPEs), the immediate precursors of N-acylethanolamides (NAEs), a family of the potent anorexigenic lipids To determine whether NAPEs synthesized by intestinal E. coli could markedly alter food intake, a daily bolus of CFU pnape-ec or pec bacteria by gavage was administered to lean male C57BL/6J mice for 7 days Chen Z et al J Clin Invest 2014; 124:

68 Incorporation of therapeutically modified bacteria into gut microbiota inhibits obesity Chen Z et al J Clin Invest 2014; 124:

69 Incorporation of therapeutically modified bacteria into gut microbiota inhibits obesity Chen Z et al J Clin Invest 2014; 124:

70 Treatment with pnape-ecn induces expression of genes encoding for fatty acid oxidation, but not fatty acid synthesis, and reduces expression of inflammatory genes in the liver. Chen Z et al J Clin Invest 2014; 124:

71 Incorporation of therapeutically modified bacteria into gut microbiota inhibits obesity Chen Z et al J Clin Invest 2014; 124:

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73 from lean male donors with a body mass index 23 kg/m2; n=9 reinfusion of own collected feces; n=9 Vrieze A et al. Gastroenterology 2012

74 Vrieze A et al. Gastroenterology 2012

75 Acknowledgments YALE PEDIATRIC OBESITY Sonia Caprio Bridget Pierpont Mary Savoye Michelle Van Name Mariana Mata Jessica Nouws Jennifer Dawiczyk Alfonso Galderisi YALE PO FORMER MEMBERS Ram Weiss, Anna Cali, Ebe D Adamo, Grace Kim, Martina Goffredo, Cosimo Giannini, Chao Zheng, Anna Di Sessa, Giuseppina Umano, Domenico Trico MIZZOU Elizabeth J. Parks YALE PUBLIC HEALTH Hongyu Zhao YALE GI/H Udeme Ekong Pam Valentino UCONN Joerg Graf Kendra Mass Jenn Huang Patients and their families HRU Staff Yale New Haven Hospital YALE GENETICS Allen E. Bale Daniel J. Dykas YALE INTERNAL MEDICINE Gary Cline INDIANA UNIVERSITY Naga Chalasani UC SAN DIEGO Ariel E. Feldstein LUND UNIVERSITY Leif Groop METABOLIC SOLUTIONS INC. David Wagner Allen Foundation Inc. R01DK A1