REGULATION OF SURFACTANT PROTEIN B GENE EXPRESSION

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Pulmonary surfactant protein B (SP-B) is a critical surfactant component for normal lung function, involved in formation of lamellar bodies in type II alveolar cells and both surfactant absorption and film stability. Developmental deficiency of SP-B contributes to surfactant deficiency in both human infants and knock-out mice is a newborn lethal condition. In addition, acquired deficiency of SP-B occurs in various animal models of acute lung injury and may contribute to human lung diseases involving inflammation. Thyroid transcription factor-1 (TTF-1) is a key transcription factor for activity of the SP-B promoter. In vitro studies during the past funding found that both phorbol ester and TGFbeta decrease SP-B gene transcription, that responsiveness to both agents maps to a region of the proximal SP-B promoter containing TTF-1 and HNF3 binding sites, and that there was a loss of both transcription factors from nuclei of treated cells. Recent preliminary data indicate that hormone induced differentiation of the surfactant system in human fetal lung epithelial cells is associated with increased TTF-1 gene expression. The overall goal of this renewal project is to further elucidate factors and mechanisms involved in both positive and negative regulation of SP-B gene expression is mediated, at least in part, through modulation of TTF- 1 expression or function. Most experiments will be carried out in a unique primary lung cell culture system in which differentiation of the surfactant system is induced and maintained by glucocorticoid plus cAMP treatment. Aim 1 will characterize hormonal induction of TTF-1 and its relationship to SP-B gene expression in cultured fetal lung epithelial cells examining dose and time course, transcription rate and TTF-1 isoforms. Aim 2 will examine the role of TTF-1 in SP-B gene expression using Aim 3 will address down regulation of SP-B by TGFbeta, investigating the role of Smad signaling, TTF-1 and phosphorylation. Aim 4 will examine the roles of TGFbeta and TTF-1 in SP-B expression in vivo using the mouse bleomycin lung injury model and two approaches to abrogation to TGFbeta signaling. The proposed studies are a logical extension of recent research and will provide new information related to this critical surfactant protein and transcription factor. This project involves interactions and collaborations with most of the other PPG projects and utilizes each core.