Stem Cell Dynamics in Colonic Epithelial Repair

项目摘要 成体器官再生的研究经历了一场重大的革命， 成体干细胞的分子标记物和在体外培养器官样结构的技术的发展， 身体在小鼠小肠中，发现了多个明显标记的上皮干细胞群， 塑造了我们对自我更新种群如何相互作用以及在生理上的差异利用的看法。 例如稳态和损伤。在这里，我们重点关注小鼠结肠中的干细胞群（大 肠）。结肠上皮在正常组织维持期间由Lgr 5阳性干细胞维持， 但用于损伤的干细胞群及其相互作用的规则尚未阐明。的 小肠中损伤激活的干细胞群的标记物不标记结肠中的干细胞。我们 目的是表征小鼠结肠上皮修复状态期间的干细胞动力学和信号传导， 特别是那些伴随或跟随在患有炎症性疾病的人类患者中所见的炎性损伤的疾病。 肠道疾病。这在小鼠结肠中是可能的，因为损伤模型相对丰富，我们最近的研究表明， 开发成像平台，使干细胞能够在特定组织中可视化 在大范围内修复结构。这项应用获得的初步数据表明，干细胞 修复中的群体动态与稳态和损伤诱导的群体动态有根本不同 干细胞具有独特的分子特性。这些干细胞来源于肠道和鳞状细胞 组织和它们的修复动力学导致克隆多样性的急剧丧失。该项目的具体目标是： (1)分析远端结肠隐窝再生的细胞来源和动力学，（2）确定远端结肠隐窝再生的细胞来源， 损伤诱导的小鼠远端结肠鳞状上皮化生的起源和生长机制，以及（3） 表征表皮生长因子受体（EGFR）信号传导在干细胞对损伤的反应中的作用。 在我们的方法中使用的主要方法包括谱系追踪、来自小鼠的结肠类器官培养和 人组织、高通量测序和EGFR功能障碍的先进模型的利用。这项工作 将有助于确定特定的细胞群体和过程，以针对下一代治疗，重点是 炎症性肠病中未充分利用的粘膜愈合途径。

PROJECT SUMMARY The study of the regeneration of adult organs has undergone a major revolution with the discovery of specific molecular markers of adult stem cells and development of techniques to grow organ-like structures outside the body. In the mouse small intestine, the finding of multiple, distinctly marked populations of epithelial stem cells has shaped our view of how self-renewing populations interact and are differentially utilized in physiological contexts such as homeostasis and injury. Here we focus on stem cell populations in the mouse colon (large intestine). The colonic epithelium is maintained by Lgr5-positive stem cells during normal tissue maintenance, but the stem cell populations utilized in injury and their rules of interaction have not been elucidated. The markers of injury-activated stem cell populations in the small intestine do not label stem cells in the colon. Our goal is to characterize stem cell dynamics and signaling during mouse colonic epithelial repair states, especially those that accompany or follow inflammatory injuries seen in human patients with inflammatory bowel disease. This is possible in mouse colon because of the relative wealth of injury models and our recent development of an imaging platform that allows stem cells to be visualized in the context of specialized tissue repair structures over a large region. Preliminary data obtained for this application suggest that stem cell population dynamics in repair are fundamentally different from those in homeostasis and that injury-induced stem cells have a distinct molecular identity. These stem cells originate from both intestinal and squamous tissues, and their repair dynamics result in a dramatic loss in clonal diversity. Specific aims for this project are: (1) To analyze the cellular origins and dynamics of crypt regeneration in distal colon, (2) to determine the origins and growth mechanisms of injury-induced squamous metaplasia in distal mouse colon, and (3) to characterize the role of epidermal growth factor receptor (EGFR) signaling in stem cell responses to injury. Primary methodologies used in our approach include lineage tracing, colonic organoid culture from mouse and human tissue, high throughput sequencing, and utilization of advanced models of EGFR dysfunction. This work will help identify specific cellular populations and processes to target for next generation therapies focusing on the underutilized mucosal healing pathway in inflammatory bowel disease.