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Title:Microbial and genetic influences on the development of obesity in children
Author(s):Wang, Anthony An
Director of Research:Teran-Garcia, Margarita
Doctoral Committee Chair(s):Donovan, Sharon
Doctoral Committee Member(s):Gaskins, H; Miller, Michael
Department / Program:Nutritional Sciences
Discipline:Nutritional Sciences
Degree Granting Institution:University of Illinois at Urbana-Champaign
Degree:Ph.D.
Genre:Dissertation
Subject(s):childhood obesity
genetics
gut microbiome
Abstract:Childhood obesity is a nutrition-related disease with multiple underlying etiologies. The gut microbiota is thought to be a contributor in the development of obesity by fermentation of non-digestible polysaccharides to short chain fatty acids (SCFA), which increases host capacity for energy harvest and storage. Several genes encoding SCFA receptors and transporters, as well as other host responders to gut microbiota have been described. However, the collective impact of common genetic variations (single nucleotide polymorphisms [SNP]) in these genes on obesity phenotypes has yet to be examined in humans. Our hypothesis is that genetic variation in SCFA recognition pathways are related to excess weight gain and that the SCFA producing capacity of the fecal microbiota are associated with overweight and obesity in children. These studies provide a unique approach in studying the relationship between gut microbiota and child obesity through identification of gut microbe-host adiposity relationships measured by dual energy X-ray absorptiometry (DXA), assessment of the SCFA-producing capacity of fecal inoculates, and evaluation of associations between SNPs within genes with known interaction with SCFA and obesity-related phenotypes. Study 1 Microbial dysbiosis is implicated in obesity in adults, but less in known about its association with childhood obesity. Herein, fecal microbiota, BMI and fat distribution were investigated in 4-7 year-old children to test the hypothesis that abundance of butyrate-producing bacteria is associated with overweight and obesity in children. RT-qPCR was conducted on fecal DNA for Clostridium cluster XIVa, Clostridium cluster IV, Lactobacillus spp., Bifidobacterium spp., Bacteroides-Prevotella group, Faecalibacterium prausnitzii, Roseburia intestinalis, and Eubacterium rectale. BMI was calculated from height and weight measures and percent body fat was measured by dual-energy x-ray absorptiometry (DXA). Associations between bacterial abundance and obesity phenotypes were tested using t-tests, correlations, and regressions. Overweight (n=6) children harbored lower Clostridium cluster IV abundance compared to lean (n=12) (P = 0.03). Relationships between percent body fat and bacterial abundance were impacted by sex. In girls, Bifidobacterium spp. were negatively associated with trunk fat, while F. prausnitzii was negatively correlated with BMI. In boys, Clostridium cluster IV was positively correlated with both trunk and visceral fat (P < 0.05). These findings demonstrate associations between fecal bacterial composition and overweight in children. Gender-specific effects are apparent as early as 4-7 years-of-age, and DXA measurements provide insight into the relationship between bacterial quantities and regional fat depots beyond BMI alone. Study 2 The majority of the current work in the gut microbiome and childhood obesity has focused on associations between the gut microbial composition and BMI, and little work to date has addressed the metabolic capacity of fecal microbiome from preschool age children. Building upon data from study 1, our goal was to further characterize the gut microbiome-host relationship by conducting an in vitro experiment to measure the SCFA-production of fecal inoculates from preschool age children. Fresh stool samples were collected and processed within 45 minutes of defecation (n = 6, female = 2) and were used to inoculate media containing inulin substrate. Fermentation was conducted and SCFA quantities including acetate, propionate, and butyrate were measured at 0 and 12 hours by gas chromatography. Dietary fiber intake was assessed with 24-hour recall and whole body fat was measured with DXA. Dichotomous groups were established based on high and low total fiber intake and whole body fat separated by the median. Differences in SCFA quantities were tested using Kruskal-Wallis for nonparametric data and T-test. Children in the high body fat group produced more acetate and total SCFAs than children in the low body fat group (1.591 v. 3.775 mmol/g feces, P = 0.0495; 2.926 v. 5.541 mmol/g feces, P = 0.0495 respectively). Children consuming high total fiber had higher butyrate production compared to those consuming low total fiber (0.417 v. 1.615 mmol/g feces, P = 0.0495). Propionate as a percent of total SCFAs was lower in all fermentation experiments. Within the low total fiber group, there was a significant increase in acetate as a percent of the total from 0 to 12 hours (30.9% ± 10.4 v. 68.0% ± 8.2, P = 0.0439). Current and past dietary exposures may play a role in SCFA production of the gut microbiome. Further analysis of the microbiome by sequencing will increase our understanding of the metabolic capabilities of the microbiome, which are shaped by dietary influences. Study 3 This study investigated the cumulative association of single nucleotide polymorphisms (SNP) of genes involved in SCFA recognition and metabolism with obesity. Study participants were non-Hispanic White children (2-5 yrs.) from the STRONG Kids Illinois and Michigan cohorts (n=270). Height and weight were measured to calculate obesity-related phenotypes. Genomic DNA was extracted from saliva and genotyped using the Fluidigm® platform. Statistical analyses were performed in SAS 9.4. Ten SNP variables (PPARG, ANGPTL3/4, LPL, PYY, NPY2R, SLC5A8, SLC16A3, SLC16A1, and IL6) were dichotomized according to dominant or recessive inheritance models with the effect size of each SNP variable on BMI Z-score established using β-coefficients estimated from general linear models. A weighted genetic risk score (GRS) was generated by summing the β-coefficients. The GRS was significantly associated with BMI Z-score with the model explaining 12.4% of the variance using linear regression (R2 = 0.124, P <0.0001). Similarly, the GRS was associated with weight-for-age Z-score (R2 = 0.045, P = 0.002) but not with height-for-age Z-score. This initial analysis suggests the cumulative association of the genetic variants studied with early-onset obesity. Our data supports the concept that gut microbiota influences obesity development through key host genes interacting with SCFA, warranting further investigation into the mechanisms driving these associations. Overall, these data demonstrate distinct genetic and metabolic relationships between the gut microbiome and adiposity of preschool age children. Study 1 and 2 present key methodological considerations for future gut microbiome studies including the need to collect data on antibiotic use and dietary intake, to stratify analysis by sex when considering regional body fat, and to collect and preserve fresh stool samples for in vitro experiments. Study 3 represents a novel approach to generating a cumulative genetic risk score from the selection of candidate genes and SNPs to the construction of the risk score. These findings provide avenues for new lines of research in genetic epidemiology and the gut microbiome. As both fields rapidly advance, the translation of this work to clinical and public health settings hinges on our ability to harness an understanding of the biological mechanisms underlying obesity to develop practical strategies to treat and prevent childhood obesity.
Issue Date:2015-12-04
Type:Thesis
URI:http://hdl.handle.net/2142/89228
Rights Information:Copyright 2015 Anthony An Wang
Date Available in IDEALS:2016-03-02
Date Deposited:2015-12


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