It is now generally accepted that angiotension converting enzyme (ACE) participates in regulation of vasoactivity and may be important in such varied cellular functions as modulation of inflammation, metabolism of neuropeptides, airway reactivity, electrolyte absorption, granuloma formation and spermatogenesis or reproduction. We have begun to identify intracellular mechanisms for regulation of synthesis of ACE by bovine endothelial cells in culture; intracellular Ca2+ and cAMP appear to be important components of this regulation that is also under hormonal (T3, T4, dexamethasone, aldosterone) control. We now wish to further explore regulation of ACE synthesis in bovine and human endothelial cells and will also extend these studies to macrophages, monocytes and fibroblasts. The mechanism by which hypoxia elevates cellular ACE will be evaluated. A factor we have found to be produced by smooth muscle cells that stimulates ACE synthesis by endothelial cells will be studied. In addition to Ca2+ and cAMP, possible influence of ACE synthesis of inositol polyphosphates will be explored and possible intermediate effects on protein phosphorylations through kinase actions will be examined. We will continue to assess ACE by enzymatic activity, but will also utilize our newly developed ELISA for measuring ACE antigen in cells and radiolabeling with [35S] methionine for turnover studies that should better define kinetics of our observations. Synthetic cDNA probes will be developed by polymerase chain reaction for measurement of ACE mRNA in human and bovine cells and tissue. This methodology will be applied to assessments of the role of transcriptional processes for various factors that influence cellular ACE. We believe the proposed studies should provide the first composite understanding of regulation of ACE at the cellular and molecular level and may allow new insights for the regulation of this important enzyme in more intact organ systems.
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