Mechanisms of alteration of gastrointestinal physiology by gut microbes

项目摘要/摘要 肠易激综合征(IBS)是一种全球流行的疾病(约11%)，其特征是肠易激综合征 大便形式/次数与腹部不适或疼痛有关。IBS分为以下几类 便秘、腹泻或混合(IBS-C、IBD-D、IBS-M)，基于主要的大便形式/频率。这个 IBS的病理生理学是复杂的，针对IBS潜在的病理生理学的治疗方案 是有限的。最近的研究支持肠道微生物代谢物在维持正常状态中的作用 胃肠(GI)功能，但不同微生物代谢物的变化和相互作用 这些代谢物影响IBS病理生理学中涉及的分子通路仍然是一个关键知识 差距。因此，目前经验性设计的微生物疗法(益生菌)具有 很大程度上被证明对IBS无效。为了解决这一差距，在前一个赠款周期中，我们专注于 细菌代谢物色胺，并发现色胺增加分泌和粘液释放在5- HT4R依赖方式，加速转运，并在啮齿动物模型中保护炎症。这个 我们在人类研究中发现IBS-D中色胺水平升高，这支持了我们的观察结果。在……里面 同样的纵向多组学人类研究，多组学中在IBS-C中最一致的发现 平台组的大便次黄嘌呤和丁酸盐显著减少。这样做的总体目标是 建议通过鉴定分子来确定这些代谢物的生理相关性。 受这些与IBS-C相关的代谢物影响的通路。我们的中心假设基于 根据先前的研究和我们的初步数据，次黄嘌呤是一种效应代谢物， 通过增加肠嗜铬细胞5-羟色胺的释放来加速胃肠道转运，而丁酸盐是一种调节剂 增强效应代谢物生物活性的代谢物。这将在两个目标上进行测试：在AIM 1，我们将确定次黄嘌呤促进EC细胞5-羟色胺释放和 加速胃肠道转运，在目标2，我们将确定丁酸调节EC细胞的机制 对效应剂代谢物的反应及其对胃肠功能的影响。我们将使用钙离子成像技术 来自新型转基因小鼠的有机物/原代EC细胞培养，异源受体表达与位点- 定向突变，表观基因组学和转录组数据，结合体外结肠 制剂、诺生菌和EC细胞耗尽的小鼠模型、同基因细菌突变体和新的 解决上述目标的封装方法。我们的发现将揭示具体的途径，通过 这些微生物代谢物影响胃肠道转运，并允许开发基于新机制的微生物 IBS-C的治疗方法

PROJECT SUMMARY/ABSTRACT Irritable bowel syndrome (IBS) is a globally prevalent disorder (~11%) characterized by an alteration in stool form/frequency in association with abdominal discomfort or pain. IBS is categorized into constipation, diarrhea or mixed (IBS-C, IBD-D, IBS-M) based on the predominant stool form/frequency. The pathophysiology of IBS is complex and therapeutic options targeting the underlying pathophysiology in IBS are limited. Recent studies support a role for gut microbial metabolites in maintaining normal gastrointestinal (GI) function, but how changes in different microbial metabolites and interactions among these metabolites affect molecular pathways involved in IBS pathophysiology remains a critical knowledge gap. Hence, it is not surprising that the current empirically designed microbial therapies (probiotics) have largely proven ineffective in IBS. To address this gap, in the previous grant cycle we focused on the bacterial metabolite tryptamine and found tryptamine increases secretion and mucus release in a 5- HT4R dependent manner, accelerates transit, and protects against inflammation in rodent models. The observations were supported by our finding of elevated levels of tryptamine in IBS-D in our human study. In the same longitudinal multi-omics human study, the most consistent finding in IBS-C across multiple -omics platforms were significant decreases in stool hypoxanthine and butyrate. The overall objective of this proposal is to determine the physiologic relevance of these metabolites by identifying the molecular pathways affected by each of these metabolites that are relevant to IBS-C. Our central hypothesis based on prior research and our preliminary data is that hypoxanthine is an effector metabolite that accelerates GI transit by increasing enterochromaffin (EC) cell serotonin release while butyrate is a regulatory metabolite that augments the biologic activity of effector metabolites. This will be tested in two Aims: In Aim 1, we will determine the mechanism by which hypoxanthine increases EC cell serotonin release and accelerates GI transit and in Aim 2, we will determine the mechanism by which butyrate regulates EC cell responses to effector metabolites and the resultant effects on GI function. We will use Ca2+ imaging in organoids/primary EC cell culture from novel transgenic mice, heterologous receptor expression with site- directed mutagenesis, and epigenomic and transcriptomics data, combined with ex vivo colon preparations, gnotobiotic- and EC cell-depleted mouse models, isogenic bacterial mutants, and novel encapsulation methods to address the above aims. Our findings will uncover specific pathways by which these microbial metabolites affect GI transit and allow development of novel mechanism-based microbial therapies for IBS-C.