THYROID HORMONE AND THE GUT

描述：（改编自研究者摘要） 肠道的生长和分化对于维持 正常的肠道功能 肠粘膜生长和分化的紊乱 导致腹泻、吸收不良和屏障功能改变。 甲状腺 激素（T3）是肠上皮细胞最有效的调节因子之一， 生长和分化，但其在胃肠道中的作用机制是 大部分未知。 这项研究计划旨在阐明 T3对胃肠道粘膜产生深远影响的分子机制 结构和功能。 体内和体外研究都表明， T3对小肠上皮有两个主要影响（1） 诱导隐窝细胞增生和（2）改变的模式， 肠细胞基因表达，例如，肠碱性磷酸酶升高 (IAP)表情 T3对靶组织的作用通常是 通过与核受体蛋白结合介导， 直接与特定T3应答基因内的DNA顺式元件（TRE）结合。 多种形式的T3受体（TR）由α或β编码。 β cerbA基因，包括非激素结合变体，c-erbA α-2， 其被认为抑制T3作用。 由于细胞的T3反应性是 根据各种TR的相对丰度，我们将使用 原位杂交和隐窝绒毛分离技术，以确定 小肠内TR表达的细胞类型特异性模式 粘膜 他们已经建立了TR转染的HT-29细胞（HT-29 TR），作为一个优秀的细胞。 体外模型研究T3对肠上皮细胞的作用。 HT-29 TR细胞将用T3处理以检查生长的性质 响应，例如，3 H胸苷掺入、原癌基因诱导和 用c-fox血清反应鉴定核复合物的凝胶迁移试验 元素 直接比较将与良好的特点地穴 细胞生长因子EGF 然后将丁酸钠处理的HT-29 TR细胞用作绒毛的模型 肠上皮细胞和T3对IAP基因表达的影响。 瞬态 用IAP报告基因构建体的转染实验将使它们能够 以确定重要的DNA顺式调控元件。 最后，DNA酶1 将进行足迹法和凝胶迁移试验， T3调节基因的相关DNA-蛋白质相互作用 表情 拟议的研究将加强对T3的作用的理解， 在肠粘膜结构和功能方面发挥作用， 对发育和肿瘤形成的生物学有更广泛的影响。 它 希望这些研究将确定潜在的治疗靶点， 可以在未来用于维持肠道完整性， 病

DESCRIPTION: (Adapted from investigator's abstract) The processes of intestinal growth and differentiation are essential to the maintenance of normal gut function. Derangements in gut mucosal growth and differentiation result in diarrhea, malabsorption, and altered barrier function. Thyroid hormone (T3) is one of the most potent regulators of intestinal epithelial growth and differentiation, but its mechanism of action in the GI tract is largely unknown. This research proposal is designed to elucidate the molecular mechanisms by which T3 exerts its profound effects upon GI mucosal structure and function. Both in vivo and in vitro studies have demonstrated that T3 has two major effects upon the small intestinal epithelium (1) induction of crypt cell hyperplasia and (2) alteration in the pattern of enterocyte gene expression, e.g., increased intestinal alkaline phosphatase (IAP) expression. The effects of T3 on target tissues are generally mediated through binding to a nuclear receptor protein which interacts directly with DNA cis-elements (TRE) within specific T3-responsive genes. Multiple forms of the T3 receptor (TR) are encoded by either the alpha or beta cerbA genes, including a non-hormone binding variant, c-erbA alpha-2, which is thought to inhibit T3 action. Since cellular T3-responsiveness is dependent upon the relative abundance of the various TRs, we will use in situ hybridization and crypt-villus separation techniques to determine the cell type-specific patterns of TR expression within the small intestinal mucosa. They have established TR-transfected HT-29 cells (HT-29TR) as an excellent in vitro model to study the T3 effects on intestinal epithelial cells. HT-29TR cells will be treated with T3 to examine the nature of the growth response, e.g., 3H thymidine incorporation, proto-oncogene induction, and gel shift assays to identify a nuclear complex with the c-fox serum response element. Direct comparisons will be made with the well characterized crypt cell growth factor, EGF. Sodium butyrate-treated HT-29TR cells will then be used as a model of villus enterocytes and the T3 effects upon IAP gene expression examined. Transient transfection experiments with IAP reporter gene constructs will enable them to identify the important DNA cis-regulatory elements. Finally, DNAse 1 footprinting and gel shift assays will be performed to more precisely define the relevant DNA-protein interactions underlying T3-regulated gene expression. The proposed studies will enhance the understanding of the role that T3 plays in regards to intestinal mucosal structure and function, and should have broader implications for the biology of development and neoplasia. It is hoped that the studies will identify potential therapeutic targets which could be used in the future to maintain gut integrity during critical illness.