In the pulmonary artery (PA), the earliest and most dramatic growth in response to chronic hypoxia occurs in the adventitial compartment where the fibroblast resides. The adventitial thickening in response to hypoxia is especially impressive during the neonatal period and clearly contributes to the marked increase in pulmonary vascular resistance observed. Recent studies suggest the existence of multiple phenotypically and functionally distinct fibroblast subpopulations in adventitia. We have found that one subpopulation of fibroblasts proliferates rapidly in response to hypoxia, while another group lacks this response. Interestingly we have also demonstrated that the number of subpopulations which have the unique capability to replicate in response to hypoxia are higher in adventitia of hypoxic hypertensive calves compared to the control calves. Our overall goal is to define the signaling mechanisms that confer the ability to proliferate under hypoxic conditions to only specific PA adventitial fibroblast subpopulations. We have demonstrated that hypoxia-induced growth of PA adventitial fibroblasts is dependent on protein kinase C (PKC) activation. We have also shown that hypoxia can activate PKC isozymes in fibroblasts in a manner that mimics other growth-promoting stimuli. Our preliminary studies have shown 1) the expression of seven out of twelve described PKC isozymes in PA adventitial fibroblasts and 2) unique nuclear localization patterns of PKC isozymes in fibroblasts consistent with other observations demonstrating that isozyme location has specific functional consequences. We propose to evaluate the hypothesis that hypoxia-induced differential expression, localization and activation pattern of PKC isozymes will allow discrimination between """"""""hypoxia-responsive"""""""" and """"""""hypoxia-nonresponsive"""""""" fibroblast subpopulations in adventitia and that hypoxia-induced proliferation is mediated through the activation of only selective PKC isozymes. Our preliminary data have also demonstrated that hypoxia activates mitogen activated protein kinases, ERK1 and ERK2, and that activation is necessary for hypoxic proliferation of fibroblasts. Therefore, we will also test the hypothesis that hypoxia-induced activation of ERK1 and ERK2 is mediated through specific PKC isozymes and this selective interaction of PKC/ERK leads to proliferation in cells. Successful completion of the proposed experiments is likely to provide new insights into the role of distinct fibroblast subpopulations in pulmonary vascular disease. A better understanding of the signaling mechanisms contributing to cell proliferation in select fibroblast subpopulations during hypoxia-induced pulmonary hypertension could lead to the development of improved therapeutic strategies to treat chronic hypoxic pulmonary diseases.
