Enteric CRF Receptor Signaling in Stress-Induced Intestinal Barrier Dysfunction

描述(由申请人提供)： 应激引起的肠道疾病，如肠易激综合征，是已知的经济负担最大的胃肠道疾病。我在K08申请中提出的这项研究的长期目标是研究心理应激导致肠屏障功能崩溃的基本机制，这是导致肠道疾病激活的关键潜在事件。在以往的动物研究中，我们证明了肠道促肾上腺皮质激素释放因子(CRF)及其受体在应激引起的以肠道通透性增加为特征的肠屏障功能破坏中起主要作用。我最近的发现揭示了肠道肥大细胞(MC)作为关键效应细胞在这种粘膜应激反应中的重要作用。我们假设心理应激刺激肠道释放CRF，通过CRF受体与肠系膜细胞结合，触发MC类胰蛋白酶的释放和肠屏障功能的破坏。我们将在三个具体目标上验证这一假说：1)确定CRF是否通过CRF受体激活MC，从而导致肠道通透性增加；2)确定MC类胰蛋白酶是否是触发肠道通透性增加的关键MC介质；以及3)确定CRF-MC信号通路是否在体内介导心理应激诱导的屏障功能障碍。在特定的目标1和2中，我们将利用体外MC-肠上皮共培养系统来研究CRF和表达在MC上的CRF受体之间的相互作用，以及这些相互作用如何启动导致肠上皮屏障功能紊乱的信号事件。我们将在这个模型中使用各种分子、遗传和药理学方法来检验我的假设。在具体目标3中，我们将利用早期心理应激的小鼠模型来诱导结肠粘膜屏障功能的永久性障碍，从而模拟人类的应激相关疾病。我们将利用MC缺陷小鼠结合基于分子的方法来确定CRF-MC信号在心理应激诱导的屏障功能障碍中的确切作用。 公共卫生相关性：这项研究将提高我们对CRF-MC信号的基本机制的理解，并将对未来设计有针对性的治疗策略以治疗这些代价高昂的疾病具有重要意义。

DESCRIPTION (provided by applicant): Stress-induced intestinal disorders such as Irritable Bowel Syndrome are the most economically burdensome gastrointestinal diseases known. The long-term objective of the research proposed in my K08 application is to study the basic mechanisms of psychological stress-induced breakdown of intestinal barrier function which is a key underlying event responsible for activation of intestinal disease. In previous animal studies, we demonstrated that enteric corticotrophin releasing factor (CRF) and its receptors play a major role in stress-induced breakdown of intestinal barrier function characterized by increased intestinal permeability. My recent findings revealed an important role of intestinal mast cells (MCs) as key effector cells in this mucosal stress response. We hypothesize that psychological stress stimulates enteric release of CRF that binds to intestinal MCs, via CRF receptors, triggering MC tryptase release and breakdown in intestinal barrier function. We will test this hypothesis in three Specific Aims: 1) Determine if CRF activates MCs via CRF receptors resulting in increased intestinal permeability, 2) Determine if MC tryptase is the key MC mediator triggering increases in intestinal permeability, and 3) Determine if CRF-MC signaling pathways mediate psychological stress-induced barrier dysfunction in vivo. In Specific Aims 1 and 2, we will utilize in vitro MC-intestinal epithelial coculture systems to study the interactions between CRF and CRF receptors expressed on MCs and how these interactions initiate signaling events that lead to disturbed intestinal epithelial barrier function. We will employ a variety of molecular, genetic, and pharmacological approaches in this model to test my hypothesis. In Specific Aim 3, we will utilize a mouse model of early life psychological stress to induce permanent disturbances in colonic mucosa barrier function thus mimicking stress-related disease in humans. We will utilize MC-deficient mice combined with molecular-based approaches to determine the definitive role of CRF-MC signaling in psychological stress induced barrier dysfunction. PUBLIC HEALTH RELEVANCE: This research will improve our understanding of the basic mechanisms of CRF-MC signaling and should have important implications in the future design of targeted therapeutic strategies to treat these costly disorders.