The Role of FXR on the Intestinal Barrier in Acute Injury.

项目摘要/摘要 这项提议的目标是成为一名外科医生兼科学家，成为一名独立的研究员 在一名杰出人员的监督下完成拟议的研究项目和职业发展 胃肠道和FXR信号专家的指导团队。所描述的职业发展将提供 在我设计实验方法和评估研究结果时，具有基础性的洞察力。该奖项还将 支持在肠道生理学领域建立强有力的出版记录所需的保护时间 并为竞争性R级提交制定初步数据。 法尼醇X受体(FXR)是一种重要的胆汁酸受体，影响肠上皮屏障。而当 我们的初步数据显示，激活FXR似乎对慢性肠道损伤模型有利 在急性炎症性损伤中，FXR的激活是有害的。相比之下，FXR基因敲除(KO)的动物 保护免受急性伤害。在这项提案中，我们将定义FXR在肠道屏障功能中的作用 急性损伤。我们假设FXR的激活通过上调Small来损害肠道屏障功能。 异二聚体蛋白(SHP)，同时干扰EGFR信号，导致紧密连接中断。我们会 用以下具体目标检验这一假说：1)确定FXR调节的机制 肠上皮屏障功能；2)检测组织特异性FXR抑制对肠道屏障的影响。 采用体内急性损伤模型。 我们将在体外和体外研究FXR及其下游效应因子SHP在肠屏障功能中的作用。 活着。为了探索这一点，我们将使用来自野生型、FXR KO、SHO KO和EGFR的肠样衍生单分子层 显性阴性‘天鹅绒’小鼠对肠道屏障和紧密连接蛋白的影响 无论有没有受伤。我们将使用qPCR和Western印迹分析来确定FXR和EGFR信号的激活。 好的。我们将使用转基因骨髓培养来确定巨噬细胞的作用。 动物。这些体外发现将在体内使用脂多糖注射急性损伤模型进行测试，使用FXR (全身、肠道和巨噬细胞特异性)和SHP KO小鼠。盲肠结扎和穿刺会 用于测试FXR通路在疾病进展中的药理操作。 该项目将开始确定FXR对肠上皮屏障的调节在设置 急性损伤。从长远来看，这些发现将为预防和预防艾滋病的发展提供必要的洞察力 肠源性败血症、炎症性肠病或其他胃肠道疾病的治疗策略。这项研究 有可能通过更好的治疗改善患有影响肠道完整性的疾病患者的预后 了解FXR介导的肠屏障功能的机制。

Project Summary/Abstract The goal of this proposal is to advance as a surgeon-scientist to become an independent investigator through completion of the proposed research project and career development under the oversight of an outstanding mentoring team of gastrointestinal and FXR signaling experts. The career development described will provide foundational insight as I design experimental approaches and assess study outcomes. This award will also support the protected time needed to establish a strong publication record in the field of intestinal physiology and develop preliminary data for a competitive R level submission. The farnesoid X receptor (FXR) is a key bile acid receptor that influences the intestinal epithelial barrier. While activation of FXR seems to be beneficial in chronic injury models of the intestine, our preliminary data show that in acute, inflammatory injury, FXR activation is deleterious. FXR knock-out (KO) animals, in contrast, are protected from acute injury. In this proposal, we will define the role of FXR in intestinal barrier function during acute injury. We hypothesize that FXR activation compromises intestinal barrier function by upregulating small heterodimer protein (SHP), while interfering with EGFR signaling, leading to tight junction disruption. We will test this hypothesis with the following specific aims: 1) Determine the mechanisms involved in FXR regulating intestinal epithelial barrier function, and 2) Test the effect of tissue-specific FXR inhibition on the intestinal bar- rier using acute injury models in vivo. We will determine the role of FXR and its downstream effector SHP in intestinal barrier function in vitro and in vivo. To explore this, we will use enteroid-derived monolayers from wild type, FXR KO, SHO KO, and EGFR dominant-negative ‘Velvet’ mice to determine the effects on the intestinal barrier and tight junctional proteins with and without injury. We will determine FXR and EGFR signal activation using qPCR and Western blot anal- yses. We will determine the role of macrophages using bone marrow-derived culture from genetically-modified animals. These in vitro findings will be tested in vivo using an LPS-injection model of acute injury, utilizing FXR (whole body, intestinal-specific, and macrophage-specific) and SHP KO mice. Cecal ligation and puncture will be used to test pharmacologic manipulation of the FXR pathway on disease progression. This project will begin to establish the role of FXR regulation of the intestinal epithelial barrier in the setting of acute injury. Long term, these findings will provide needed insight into the development of preventative and therapeutic strategies in gut origin sepsis, inflammatory bowel disease, or other GI pathologies. This research has the potential to improve the outcomes of patients with diseases that affect intestinal integrity through better understanding of the mechanisms of FXR-mediated intestinal barrier function.