RESPIRATORY EPITHELIAL CELL DIFFERENTIATION (TTF-1, HNF-3B)

(申请人摘要)该项目寻求继续开展活动，重点是 决定前肠承诺的机制的阐明：1)前肠的承诺 内胚层形成胚胎肺；2)细胞增殖分化 发育中的肺中呼吸细胞的不同亚群和3) 肺损伤后呼吸道上皮细胞的增殖和重塑 受伤。这项申请是基于在过去四年中取得的进展 资金表现出三个截然不同的关键、相互作用的作用 转录因子家族，即TTF-1(同源结构域)、HNF-3β和 呼吸道上皮细胞中的HFH-4和GATA-6 血统承诺与肺形态发生。本提案将 确定其中两个基因的精确时间/空间需求 (HNF-3β和TTF-1)在肺的形成中的作用，并将确定其 对呼吸道上皮细胞分化、基因表达及功能的影响 分枝形态发生。研究计划是在成功的基础上制定的 肺特异性条件性重组酶技术的研究进展 介导的基因删除和添加，这项技术将使呼吸道 上皮细胞谱系分析和定位基因打靶。基因 编码TTF-1和HNF-3β的重组酶将被介导 在发育过程中的精确时间删除，诱导重组酶与 多西环素控制的肺特异性条件系统。时间和空间 重组介导的TTF-1和HNF-3β缺失对肺组织的影响 形态发生将被辨别出来。呼吸道上皮细胞分化 HNF-3β和TTF-1靶基因的表达将在 体内和体外。Cre重组酶与逆转录病毒介导的细胞标记 祖细胞将被用来定义呼吸系统的谱系图 活体正常发育的肺上皮细胞。TTF-1和HNF-3β的作用 肺形态发生和呼吸道上皮细胞缺失 分化将在产前和出生后辨别出来。的效果 降低TTF-1和HNF-3β对细胞增殖和分化的影响 氧损伤的修复将被识别出来。TTF-1和TTF-1的潜在作用 人神经营养因子-3β在肺畸形和支气管肺发病机制中的作用 不典型增生(BPD)和其他形式的慢性肺部疾病将在 人体研究。

(Applicant's Abstract) This project seeks to continue activities focused to the elucidation of mechanisms which determine, 1) commitment of foregut endoderm to form the embryonic lung, 2) proliferation and differentiation of distinct subsets for respiratory cells in the developing lung and 3) proliferation and remodeling of the respiratory epithelium following lung injury. This application is based on progress made in the last four years of funding which demonstrated critical, interacting roles of three distinct families of transcription factors, namely TTF-1 (homeodomain), HNF-3beta and HFH-4 (winged helix) and GATA-6 (zinc finger) in respiratory epithelial cell lineage commitment and lung morphogenesis. The present proposal will determine the precise temporal/spatial requirements for two of these genes (HNF-3beta and TTF-1) in formation of the lung and will determine their influence on respiratory epithelial cell differentiation, gene expression and branching morphogenesis. The research plan is based on the successful development of technology for conditional, lung-specific cre-recombinase mediated gene deletion and addition, technology that will enable respiratory epithelial cell lineage analysis and site-specific gene targeting. Genes encoding TTF-1 and HNF-3beta will be subjected to cre-recombinase mediated deletion at precise times during development, inducing cre-recombinase with a doxycycline-controlled lung-specific conditional system. Temporal and spatial effects of recombination mediated deletion of TTF-1 and HNF-3beta on lung morphogenesis will be discerned. Respiratory epithelial cell differentiation and the expression of HNF-3beta and TTF-1 target genes will be assessed in vivo and in vitro. Cre-recombinase and retroviral mediated cell marking of progenitor cells will be used to define a lineage map of the respiratory epithelium in the normal developing lung in vivo. Effects of TTF-1 and HNF-3beta deletion on lung morphogenesis and respiratory epithelial cell differentiation will be discerned prenatally and postnatally. Effects of decreased TTF-1 and HNF-3beta on cell proliferation and differentiation during repair from oxygen injury will be discerned. Potential roles of TTF-1 and HNF-3beta in the pathogenesis of lung malformation and bronchopulmonary dysplasia (BPD) and other forms of chronic lung disease will be sought in human studies.