This SCOR application seeks to determine the genetic mechanisms controlling: (l) the commitment and differentiation of foregut endoderm to form the respiratory epithelial cells of the embryonic lung buds; (2) the mechanisms by which the nuclear transcription proteins, TTF-1 and HNF family members, regulate the growth and differentiation of the respiratory epithelium; and (3) to determine the roles of TTF-1 and HNF family members in the re-epithelialization and differentiation of the lung following oxidant injury in the neonatal period. Previous work from this laboratory identified TTF-1 and HNF family members as critical factors involved in the regulation of surfactant proteins A, B, C and CCSP gene expression. The present application will determine the mechanisms by which TTF-1 and HNF family members interact to regulate respiratory epithelial cell differentiation during embryogenesis of the fetal mouse lung and later in the regulation of surfactant protein gene transcription. The mechanisms, transcriptional and autocrine/paracrine, that mediate transcription of TTF-1 gene will be determined identifying the nuclear proteins and cis-active elements that control TTF-1 synthesis including the HNF and HOX family members of nuclear transcription proteins bind to and activate the TTF-1 gene in respiratory cells. Stoichiometric relationships between TTF-1, HNFs and cellular markers will be delineated using specific markers for Clara, non-ciliated bronchiolar Type ll and Type I epithelial cells. The role of the nuclear transcription factors in cell differentiation will be assessed in vitro and in transgenic mice in vivo. The factors controlling TTF-1 gene transcription and respiratory epithelial cell differentiation will be assessed in developing, injured and recovering respiratory epithelium. Transgenic mice will be made in which the TTF-1 and HNF family members, will be expressed in subsets of respiratory epithelial cells to determine the role of these proteins in the commitment and maintenance of respiratory epithelial cell phenotype. Promoter elements which can be regulated externally (by tetracycline) will be utilized to discern the role of HNFs and TTF-1 in the differentiation of respiratory epithelial cells. Clinical correlations between TTF-1, HNF family members and respiratory epithelial cell differentiation will be assessed in the human lung. We will identify genetic defects in TTF-1 and HNFs that might be related to lung disease and determine the role of these transcription factors in the pathogenesis of lung injury and recovery in patients with RDS, ARDS and BPD. These studies seek to extend our knowledge of control of respiratory epithelial cell differentiation and function during development and in the injured lung and should have important impact on our understanding the pathogenesis of common lung diseases and congenital anomalies of lung development including cystadenomatoid malformation, congenital alveolar hypoplasia and the aberrant remodeling of the lung parenchyma associated with bronchopulmonary dysplasia and other lung diseases caused by lung injury in premature infants.
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