We propose to study on a molecular basis the control of production of angiotensin-converting enzyme (ACE) by cultured glomerular endothelial cells as a way of understanding the control of glomerular pressure. This is based on a growing body of evidence suggesting that the modulation of glomerular pressure is important in the pathogenesis of chronic renal failure in diabetes. Evidence suggests that endothelium is the most likely source of glomerular ACE (ACE is not produced in vitro by human glomerular mesangial or epithelial cells) and that this tissue is not a stationary producer of this enzyme but may vary its production in response to external stimuli such as hormone levels. We hypothesize that local variations of ACE level influence glomerular pressure. We further hypothesize that endothelial cell up- or down-regulation of ACE levels reflect regulation of the rate of transcription of the gene encoding ACE. Parts I and II of this proposal discuss experiments designed to isolate a cDNA encoding ACE and its use in identifying the ACE promoter-enhancer region. Part III of this proposal discusses experiments to establish that cell confluence, ACE inhibitors, dexamethasone and insulin up-or down-regulate ACE expression by cultured glomerular endothelial cells and that this is mediated through up- or down-regulation of ACE mRNA transcription. Part IV of this proposal combines these experimental approaches to functionally dissect the ACE promoter-enhancer region by establishing what DNA sequences are responsible for the transcriptional regulation of the individual stimuli known or suspected to regulate ACE levels. This work will provide a greater understanding of how on a molecular basis ACE production is regulated by glomerular endothelium and may lead to new insights or treatment modalities for the kidney disease of diabetes mellitus.
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