Modulation of the Intestinal Microbiome in Obesity by a High Protein Diet

摘要 在临床前啮齿动物模型和临床试验中，高蛋白饮食已被证明是一种有效的减肥方法。 与对照组相比， 等热量标准蛋白质饮食。然而，这种反应的机制尚未完全阐明。我 最近在啮齿动物模型中证明，高蛋白饮食会诱导肠道微生物组的变化， 包括一种被报道具有抗肥胖作用的微生物--嗜粘蛋白阿克曼氏菌。基于 根据这些初步研究，我假设高蛋白饮食会导致肠道微生物组的改变， 介导其对肥胖症的临床疗效。为了验证这一点，我将研究216名超重和肥胖的退伍军人（BMI 27-40），将其1：1随机分配至等热量高蛋白（30%）或正常蛋白（15%）1500卡路里饮食 利用西洛杉矶VA医疗中心的现有临床基础设施， 最近完成的高蛋白饮食临床试验在目标1中，研究了高蛋白饮食对 肠道微生物组的组成和功能将通过16 S rRNA测序、鸟枪法和基因测序来评估。 宏基因组学和代谢组学。我预测，在高蛋白饮食之后， 肠道微生物组的组成有利于阿克曼氏菌和其他微生物的定植， 与正常蛋白质饮食相比，促进脂肪减少。我预计微生物组成的这种变化 与宏基因组的改变有关-包括相关细菌基因丰度的减少 碳水化合物的利用和代谢组学。研究期间共采集11份粪便样本 期间，以确定微生物变化的动力学。在目标2中，粪便微生物、宏基因组和 将鉴定对高蛋白饮食反应的代谢组学预测因子。将通过以下方式测量响应： 体重减轻、体脂减少、肝脂肪变性减少、血脂改变、血红蛋白A1 c减少， 降低高敏C反应蛋白，增加饱腹感。将使用 基线微生物组数据以及在治疗期间的每个时间点收集的样品。我还将评估 特定的微生物、基因和代谢物是否与影响血液循环的激素水平有关， 饱腹感（瘦素、胃饥饿素、胰高血糖素、胰高血糖素样肽-1、肽YY和胰多肽）。在目标3中， 无菌小鼠将用在基线或16周饮食后获得的新鲜冷冻粪便定殖 干预，以建立高蛋白饮食中微生物组的改变与 易患肥胖症。这些人源化的gnotobiotic小鼠将被置于西方饮食中以诱导肥胖 并比较体重增加、体脂积累、胰岛素敏感性和饱腹感/饥饿激素水平。在 平行地，具有预先存在的饮食诱导的肥胖症的小鼠将用饮食干预后的人 微生物群并置于标准饮食中。体重减轻、体脂减少、胰岛素敏感性和激素 将比较两组之间的水平。拟议研究的结果将提供深入了解 特定的微生物，驱动高蛋白饮食的临床反应，并可能确定候选抗肥胖 这些微生物可以进一步开发成新的微生物疗法。

ABSTRACT A high protein diet has been shown in preclinical rodent models and clinical trials to be an effective obesity treatment that is associated with greater loss of body weight and fat mass and increased satiety compared to isocaloric standard protein diets. However, the mechanisms of this response have not been fully elucidated. I recently demonstrated in a rodent model that a high protein diet induces shifts in the intestinal microbiome including a bloom of Akkermansia muciniphila, a microbe reported to have an anti-obesity effect. Based on these preliminary studies, I hypothesize that a high protein diet induces alterations in the intestinal microbiome that mediate its clinical efficacy for obesity. To test this, I will study 216 overweight and obese Veterans (BMI 27-40) who will be randomized 1:1 to isocaloric high protein (30%) or normal protein (15%) 1500 calorie diets for 16 weeks utilizing existing clinical infrastructure at the West Los Angeles VA Medical Center established for a recently completed clinical trial of a high protein diet. In Aim 1, the effect of a high protein diet on the composition and function of the intestinal microbiome will be assessed by 16S rRNA sequencing, shotgun metagenomics, and metabolomics. I predict that following a high protein diet, there will be a major shift in the composition of the intestinal microbiome to favor colonization with Akkermansia and other microbes that promote fat loss compared to the normal protein diet. I anticipate that this change in microbial composition will be associated with alterations in the metagenome – including reduced abundance of bacterial genes involved in carbohydrate utilization – and the metabolome. A total of 11 fecal samples will be collected during the study period to determine the kinetics of microbial changes. In Aim 2, fecal microbial, metagenomic, and metabolomic predictors of response to a high protein diet will be identified. Response will be measured by weight loss, reduced body fat, decreased hepatic steatosis, altered lipid profiles, reduced hemoglobin A1c, decreased high sensitivity C-reactive protein, and increased satiety. Predictors will be generated using the baseline microbiome data as well as samples collected at each time point during treatment. I will also evaluate whether specific microbes, genes, and metabolites are associated with circulating levels of hormones affecting satiety (leptin, ghrelin glucagon, glucagon-like peptide-1, peptide YY, and pancreatic polypeptide). In Aim 3, germ-free mice will be colonized with fresh frozen feces obtained at baseline or after 16 weeks of dietary intervention to establish a causal relationship between alteration of the microbiome on a high protein diet and susceptibility to obesity. These humanized gnotobiotic mice will be placed on a Western diet to induce obesity and compared for weight gain, body fat accumulation, insulin sensitivity, and satiety/hunger hormone levels. In parallel, mice with pre-existing diet-induced obesity will be colonized with post-dietary intervention human microbiota and placed on a standard diet. Weight loss, body fat reduction, insulin sensitivity, and hormone levels will be compared between the two groups. The results of the proposed study will provide insight into the specific microbes that drive the clinical response to a high protein diet and may identify candidate anti-obesity microbes that could be further developed into novel microbial therapeutics.