Regulation of adult stem cell homeostatic response to inflammatory injury

描述（由申请人提供）：微生物群或感染等环境因素会引发免疫介导的组织损伤，从而导致遗传易感受试者出现慢性粘膜疾病，例如炎症性肠病（IBD）。近年来，干细胞疗法在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 激活被认为分别促进成体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，STAT5 信号传导可预防结肠炎小鼠模型中的肠道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，并抑制了成人 IESC 衍生类器官中的 NOTCH 激活和细胞连接成熟。因此，我们假设 STAT5 信号传导通过调节干细胞基因特征来控制成人 IESC 活性，从而促进细胞间连接的形成； STAT5 信号传导的激活可增强 IEC 类器官植入，对抗感染引起的肠道屏障功能障碍。我们将通过以下两个目标来检验我们的假设。我们的目标 1 将确定 STAT5 信号传导在成人 IESC 分化中的作用。该目标的研究首先将在小鼠中诱导删除 IEC STAT5 或过度激活 IEC STAT5，以表征成年小鼠肠道中的 IESC 稳态、干细胞基因谱和 Wnt/Notch 信号传导。其次，利用人类多能干细胞衍生的肠道类器官，我们将对 STAT5 表达进行基因操作，以确定 STAT5 在 IESC 分化过程中 Wnt/Notch 信号传导调节中的作用。我们的目标 2 将定义 STAT5 信号传导对 IEC 单层分化相关开发的要求。首先，我们将使用 STAT5 缺陷型和 STAT5 诱导型 IESC 衍生类器官来表征紧密连接的发育。其次，我们将培养这些小鼠类器官以植入受体对照小鼠，以评估 IESC 完整性和紧密连接形成。总的来说，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特征，以促进细胞间连接重建，从而治愈粘膜炎症。我们的研究将证明 STAT5 在通过干细胞基因特征调节成体干细胞稳态中的重要作用，并将探索一种新型工程组织疗法，可直接治愈粘膜炎症期间上皮屏障的破坏。我们建议该研究领域进行范式转变，从将 STAT5 信号传导理解为干细胞生物学的重要调节因子，转向开发潜在的治疗工具。我们相信，这种探索性的 R21 机制将允许基础科学家和临床研究人员的合作团队启动一个全新的项目，并探索一种恢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.