The long term goal of the research is to gain insight into how vascular tone and structure are regulated at the cellular and molecular level. Important insights into the pathogenesis of diseases of disrupted vascular homeostasis, such as atherosclerosis and pulmonary hypertension, would be gained by an elucidation of the cellular and molecular vascular homeostatic mechanisms. Complex circuits involving vascular cell interactions and feedback regulatory loops are known to control vascular tone and structure. The central hypothesis of this proposal is that the production of endogenous carbon monoxide (CO) in the vasculature represents an adaptive response to hypoxia. Endogenous CO is largely the product of the enzymatic activity of heme oxygenase (HO). The inducible form of this enzyme, HO-1, is expressed by hypoxic vascular smooth muscle cells (SMCs) which release CO in the vasculature. Smooth muscle cell-derived CO may affect vascular cell function in autocrine and paracrine ways. The applicant proposes to investigate the role of CO in vascular homeostasis using cell culture and transgenic animal techniques. The specific aims of the proposed project are: (I) to study the molecular mechanisms by which CO regulates gene expression (ii) to determine the role of SMC-derived CO in endothelial-smooth muscle cell interactions in the setting of hypoxia and (iii) to validate the role of CO in the regulation of pulmonary vascular tone and structure in vivo. The experimental design involves use of in vitro single-cell type and co- culture systems in conjunction with regulated overexpression of HO-1 by transfected SMCs. Aspects of endothelial cell function to be studied include gene expression, proliferation and barrier function. Similarly, SMC gene expression, growth and proliferative response to mitogenic stimuli will be examined. A transgenic mouse model approach will be used to examine the role of CO in pulmonary vascular homeostasis under hypoxia in vivo. The human Surfactant Protein-C (SP-C) promoter will be used to target expression of the HO-1 transgene to the lung and the animals will be assessed as to the development of pulmonary hypertension in response to hypoxia. The above studies should enhance the understanding of vascular homeostasis and the role of CO.