Role of Aryl Hydrocarbon Receptor in Microbiota-Colon Stem Cell Interactions

 描述（由申请人提供）：该项目的总体目标是更好地了解成体结肠干细胞群如何在干细胞相关表型、能量代谢以及体内暴露于 AhR 配体后结肠癌的发生和进展方面对芳基烃受体（AhR）活性毒物作出反应。结肠的“成体”体干细胞特别令人感兴趣，因为它们维持自我更新并且是癌症引发突变的靶细胞。人们普遍认为结肠干细胞分化/可塑性、干细胞定位/增殖和Wnt/β-连环蛋白信号传导的扰动代表了结肠肿瘤发生的最早步骤。由于已知 AhR 及其配体会影响造血干细胞和胃肠道生物学，因此我们建议研究通过 AhR 的信号传导对结肠干细胞动力学和功能的作用。我们将特别关注 AhR 活性毒物、原型膳食 AhR 配体和微生物群从膳食色氨酸产生的代谢物（分别为 AhR 的外源性和内源性配体）。由于大多数 AhR 配体以组织和浓度特异性方式是 AhR 的激动剂和/或拮抗剂，因此我们假设 AhR 配体的联合激动剂和拮抗作用是结肠干细胞动力学和反应的重要决定因素。我们的初步研究支持了这一点，该研究首次证明 AhR 配体显着影响结肠干细胞稳态和基因表达。所提出的实验是新颖且相关的，因为暴露于外源性 AhR 配体及其与 AhR 活性肠道内源性 AhR 配体的相互作用对成体肠道干细胞生物学的影响尚未确定。将解决以下具体目标：（1）使用结肠癌细胞系和离体类器官培养系统确定 AhR 活性毒物和膳食 AhR 配体对肠道干细胞反应的激动剂和/或拮抗剂活性； (2) 研究微生物群衍生的AhR配体及其与环境毒物和膳食配体的相互作用对体内和离体肠道干细胞反应的影响； (3) 量化体内暴露于 AhR 配体后结肠癌发生的起始和进展阶段的结肠隐窝中干细胞的数量和时空位置、DNA 损伤和靶向缺失。利用体内和离体模型将使我们能够在存在或不存在 AhR 的情况下剖析 AhR 配体的作用。

 DESCRIPTION (provided by applicant): The overall goal of this project is to better understand how the adult colonic stem cell population responds to Aryl hydrocarbon receptor (AhR)-active toxicants in terms of stem cell-related phenotypes, energy metabolism, and the initiation and progression of colon cancer following exposure to AhR ligands in vivo. "Adult" somatic stem cells of the colon are of particular interest because they sustain self-renewal and are target cell for cancer initiating mutations. Perturbations in colon stem cell differentiation/plasticity, stem ell location/proliferation, and Wnt/beta-catenin signaling are generally believed to represent the earliest step towards colon tumorigenesis. Since the AhR and its ligands are known to affect hematopoietic stem cells and gastrointestinal biology, we propose to investigate the role of signaling through the AhR on colon stem cell dynamics and function. We will specifically focus on AhR-active toxicants, prototypical dietary AhR ligands and metabolites generated from dietary tryptophan by the microbiota (exogenous and endogenous ligands for the AhR, respectively). Since a majority of the AhR ligands are agonists and/or antagonists for the AhR in a tissue- and concentration- specific manner, we hypothesize that the combined agonist and antagonistic role of AhR ligands are important determinants in colon stem cell dynamics and responses. This is supported by our preliminary studies, which demonstrate for the first time that AhR ligands significantly influence colonic stem cell homeostasis and gene expression. The proposed experiments are novel and relevant because the impact of exposures to exogenous AhR ligands and their interactions with AhR-active intestinal endogenous AhR ligands on adult intestinal stem cell biology has not been determined. The following specific aims will be addressed: (1) Determine the agonist and/or antagonist activity of AhR-active toxicants and dietary AhR ligands on intestinal stem cell responses using colon cancer cell lines and an ex vivo organoid culture system; (2) Investigate the effect of microbiota-derived AhR ligands and their interactions with environmental toxicants and dietary ligands on intestinal stem cell responses in vivo and ex vivo; and (3) Quantify the number and spatio-temporal location of stem cells, DNA damage and targeted deletion in the colonic crypt at the initiation and progression stages of colon carcinogenesis following exposure to AhR ligands in vivo. Utilization of both in vivo and ex vivo models will allow us to dissect the effects of AhR ligands in the presence or absence of the AhR.