Regulation of adult stem cell homeostatic response to inflammatory injury

描述(申请人提供)：环境因素，如微生物区系或感染，触发免疫介导的组织损伤，导致遗传易感对象的慢性粘膜疾病，如炎症性肠病(IBD)。近年来，干细胞治疗在IBD个体化治疗方面取得了许多成功案例，但肠上皮干细胞(IESC)治疗仍处于早期阶段。JAK-STAT激活可能分别促进成人肠干细胞分裂和分化，从而驱动肠上皮细胞(IEC)的更新。我们最近报道了STAT5信号通路可预防结肠炎小鼠模型的肠屏障功能障碍，并促进粘膜愈合。此外，我们的初步数据表明，STAT5的缺失降低了干细胞标记物Lgr5、Ascl2和Olfm4的表达，并抑制了IESC来源的成年器官中的缺口激活和细胞连接成熟。因此，我们假设STAT5信号通过调节干细胞基因信号来控制成年IESC的活动，以促进细胞间连接的形成；激活STAT5信号可以增强IEC有机物的植入，对抗感染诱导的肠屏障功能障碍。我们将用以下两个目标来检验我们的假设。我们的目标1将确定STAT5信号在成人IESC分化中的作用。这一目标的研究将首先在小鼠中诱导缺失IEC STAT5或高激活IEC STAT5，以表征IESC在成年小鼠肠道中的稳态、干细胞基因谱以及Wnt/Notch信号转导。其次，利用人类多能干细胞来源的肠道器官，我们将从基因上操纵STAT5的表达，以确定STAT5‘S在IESC分化过程中对Wnt/Notch信号的调控作用。我们的目标2将定义IEC单层分化相关发育所需的STAT5信号。首先，我们将使用STAT5缺陷和可诱导IESC衍生的有机化合物来表征紧密连接的发育。其次，我们将培养这些小鼠器官类化合物来植入受体对照组小鼠，以评估IESC的完整性和紧密连接的形成。总体而言，我们将确定上皮STAT5通过与Notch信号相互作用来调节干细胞基因签名，以促进细胞间连接重组，从而治愈粘膜炎症。我们的研究将展示STAT5通过干细胞基因签名在调节成人干细胞动态平衡中的重要作用，并将探索一种新的工程化组织疗法，直接修复黏膜炎症过程中的上皮屏障破坏。我们建议这一研究领域的范式转变，从理解STAT5信号作为干细胞生物学的基本调节因子，转向开发潜在的治疗工具。我们相信，这种探索性的R21机制将允许一个由基础科学家和临床研究人员组成的合作团队启动一个全新的项目，并探索一条修复粘膜炎症中上皮屏障功能障碍的新的潜在治疗途径。

DESCRIPTION (provided by applicant): Environmental factors such as microbiota or infection trigger immune-mediated tissue injury, which leads to chronic mucosal diseases in the genetically susceptible subjects, such as Inflammatory Bowel Diseases (IBD). Stem cell therapy has recently achieved many successful cases in the personalized treatment of IBD, but intestinal epithelial stem cell (IESC) treatment is still at an early stage. JAK-STAT activation wa suggested to promote adult intestinal stem cell division and differentiation, respectively, and thereby could drive the renewal of intestinal epithelial cells (IEC). We recently reported that STAT5 signaling prevents intestinal barrier dysfunction and promotes mucosal healing in colitic mouse models. Furthermore, our preliminary data indicated that deletion of STAT5 decreased the expression of stem cell markers Lgr5, Ascl2, and Olfm4, and inhibited NOTCH activation and cellular junction maturation in adult IESC-derived organoid. We therefore hypothesize that STAT5 signaling controls adult IESC activity via regulation of a stem cell gene signature to promote intercellular junction formation; activation of STAT5 signaling enhances IEC organoid engraftment against infection-induced intestinal barrier dysfunction. We will test our hypotheses with the following two Aims. Our Aim 1 will determine the role of STAT5 signaling for adult IESC differentiation. The studies in this Aim will, first, inducibly delete IEC STAT5 or hyperactivate IEC STAT5 in mice to characterize IESC homeostasis, stem cell gene profiles, and Wnt/Notch signaling in adult mouse intestines. Second, using human pluripotent stem cells-derived intestinal organoids, we will genetically manipulate STAT5 expression to define STAT5's role in the regulation of Wnt/Notch signaling during IESC differentiation. Our Aim 2 will define the requirement of STAT5 signaling for differentiation-associated development of IEC monolayers. First, we will characterize tight junction development using both STAT5 deficient and STAT5 inducible IESC-derived organoids. Second, we will culture these mouse organoids to implant the recipient control mice to assess IESC integrity and tight junction formation. Overall, we will determine that epithelial STAT5 regulates a stem cell gene signature via interplay with NOTCH signaling to promote intercellular junction reformation for healing mucosal inflammation. Our studies will demonstrate an essential role for STAT5 in the regulation of adult stem cell homeostasis via a stem cell gene signature, and will explore a novel engineered tissue therapy that directly heals the epithelial barrier disruption during mucosal inflammation. We propose a paradigm shift for this research field from understanding STAT5 signaling as an essential regulator of stem cell biology toward developing the potential therapeutic tool. We believe this exploratory R21 mechanism will permit a collaborative team of basic scientists and clinical investigators to start a brand-new project, and explore a novel potential therapeutic avenue for restoring epithelial barrier dysfunction in mucosal inflammation.