Regulation of differentiation in esophageal epithelia

描述(申请人提供)：这是我的研究项目资助(R01)“食道上皮分化的调控”(DK069984)的续期申请的重新提交，该申请于2011年3月31日结束。这项建议的首要目标是通过对关键转录调控因子Kr|ppel-like factor4(KLF4)的机制研究，了解食道上皮分层、炎症和癌变的调节。这项建议的意义在于(A)食道疾病的流行，这是美国和世界上最常见的疾病之一，以及(B)KLF4在许多组织和细胞类型(包括食管角蛋白细胞)中控制基本细胞过程中的重要和新出现的作用。PI是一位经验丰富的研究人员，他是Kr|ppel样因子(KLF)、疾病动物模型和食道鳞状细胞生物学方面的专家，并得到一支出色的研究团队的支持，并得到专家合作者的补充。在这个提案中，我们将利用我们已经建立的新的小鼠模型，以及利用二维和三维器官型培养的补充体外系统来验证KLF4是成人食道鳞状上皮内的关键调节因子的假设，控制角质形成细胞迁移和分化、炎症反应和鳞状细胞癌的发展等过程。为了探讨KLF4对食道上皮细胞层化、炎症和癌变的调控作用，我们将进行以下相关的具体研究：(1)我们将通过对KLF4和KLF4靶点Wnt5A的上位性机制的研究，以及(B)通过功能基因组分析寻找其他相关的KLF4靶基因，来探讨KLF4在食道上皮细胞迁移和层化中的作用。(2)我们将通过(A)检测KLF4和p53在体外和体内的食道上皮细胞中的功能相互作用来探讨KLF4缺失在食管癌发生中的作用我们将通过(A)评估Rho stas在KLF4激活NF：B中的作用，(B)检测NF：B和TNF1抑制对ED-L2/KLF4小鼠角质形成细胞增殖的影响，以及(C)将ED-L2/KLF4小鼠与食道特异的Ik2基因缺失小鼠杂交，以确定KLF4促进炎症和非细胞自主增殖反应的机制。这些互补的方法将确认和扩展我们当前资金周期的发现，如初步数据和我们最近在胃肠病学上发表的两篇论文所概述的那样。此外，拟议的研究将得到一流的和合作的知识环境以及为PI和他的团队提供的特殊资源和设施的支持。我们预计这些研究将为维持正常的食道上皮内稳态的因素以及在食道疾病中被破坏的途径提供洞察力，无论是良性的还是恶性的。

DESCRIPTION (provided by applicant): This is a resubmission of the renewal application for my Research Project Grant (R01) entitled "Regulation of Differentiation in Esophageal Epithelia" (DK069984), which concluded on March 31, 2011. The overarching goal of this proposal is to understand the regulation of esophageal epithelial stratification, inflammation, and carcinogenesis through mechanistic studies of the key transcriptional regulator Kr|ppel-like factor 4 (KLF4). The significance of this proposal lies in (a) the prevalence of esophageal disorders, which are among the most common ailments in the United States and the world and (b) the important and emerging roles of KLF4 in the control of essential cellular processes in numerous tissues and cell types, including esophageal keratinocyctes. The PI is an experienced investigator who is an expert in the Kr|ppel-like factors (KLFs), animal models of disease, and esophageal squamous cell biology, and is supported by a superb research team, complemented by expert collaborators. In this proposal, we will take advantage of novel mouse models, which we have generated, and complementary in vitro systems utilizing two-dimensional and three-dimensional organotypic culture to test the hypothesis that KLF4 is a critical regulator within adult esophageal squamous epithelia, controlling processes such as keratinocyte migration and differentiation, the inflammatory response, and squamous cell cancer development. To explore the regulation of esophageal epithelial stratification, inflammation, and carcinogenesis by KLF4, we will undertake the following interrelated Specific Aims: (1) We will explore the function of KLF4 in esophageal epithelial migration and stratification (a) through mechanistic, epistatic studies of KLF4 and the KLF4 target WNT5A, a non-canonical Wnt, and (b) by identification of additional, relevant KLF4 target genes through functional genomics analyses~ (2) We will investigate the role of KLF4 loss in esophageal carcinogenesis by (a) examining the functional interaction of KLF4 and p53 in esophageal epithelial cells in vitro and in vivo, and (b) identifying the mechanisms of KLF4 loss in esophageal dysplasia and squamous cell cancer~ and (3) We will determine the mechanisms by which KLF4 promotes inflammation and the non-cell autonomous proliferative response by (a) evaluating the role of Rho Stases in NF:B activation by KLF4, (b) examining the effects of NF:B and TNF1 inhibition on proliferation of keratinocytes from ED-L2/KLF4 mice~ and (c) crossing ED-L2/KLF4 mice with esophageal-specific Ikk2 null mice. These complementary approaches will confirm and extend the findings from our current funding cycle, as outlined in Preliminary Data and our two recent publications in Gastroenterology. Moreover, the proposed research will be supported by the superb and collegial intellectual environment and the exceptional resources and facilities available to the PI and his team. We anticipate that these studies will provide insight into the factors that maintain normal esophageal epithelial homeostasis and the pathways that are disrupted in esophageal diseases, both benign and malignant.