? This translational research proposal seeks to apply recent novel insights into the mechanisms of cell signaling at the level of the plasma membrane (the caveola/raft signaling hypothesis and the interleukin-6-raft-STAT3 signaling model) to an understanding of the pathogenesis of PH. Caveolin-1-containing detergent-resistant plasma membrane rafts are now recognized as specialized signaling organelles, including cytokine signaling. There is now growing evidence for a role of cytokines in the pathogenesis of lung diseases. As examples, elevated serum levels of IL-6 have been observed in primary pulmonary hypertension (PH) and in PH associated with autoimmune diseases and AIDS. In a rat model, a single injection of the plant alkaloid monocrotaline (MCI) results within 48 hrs in endothelial cell damage, membrane leakage, upregulation of IL-6 mRNA and bioactivity but a marked downregulation of caveolin-1 in the lung, followed by development of PH 10-14 days later. ? ? The focus of the proposed studies is two-pronged: (a) to evaluate the hypothesis that pulmonary endothelial-cell raft/caveolar disruption by MCT is an initiating event in the pathogenesis of PH (Specific Aim I), and (b) to investigate the function of membrane rafts and of the newly discovered cytosolic caveolin-containing Palade complexes in IL-6-induced STAT3 signaling in lung-specific cells (Specific Aims II and III).
Aim I will include investigations of the time-course, histologic location, and cellular and molecular mechanisms for the downregulation of caveolin proteins and gene expression, and of the integrity of caveolar/raft function in pulmonary vascular and parenehymal tissues of MCT-treated rats.
Aim II includes molecular studies of the mechanisms of association of STAT3 with caveolin-1 and of STAT3 activation in plasma membrane rafts in pulmonary endothelial cells, alveolar type II-like epithelial cells and lung fibroblasts.
Aim I ll includes studies of the protein components of STAT3-containing cytosolic Palade complexes and their function in ferrying signaling molecules from the plasma membrane rafts to the cell interior. ? ? Mechanistic insights derived from this project are likely to suggest novel therapeutic approaches in the management of pulmonary hypertension. Moreover, the proposed studies are of particularly broad significance in that insights into the molecular mechanisms involved in raft-STAT signaling are likely to be applicable to cytokine-mediated activation of STAT transcription factors in perhaps all cell types, as well as to other signaling pathways localized in raft microdomains (eNOS and angiotensin II signaling). ? ?
