CYTOSTATIC ROLE OF THE ARGININE-NITRIC OXIDE PATHWAY

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Nitric oxide (NO) is believed to interfere with cell proliferation, although its mechanism of cytostatic action is largely unknown. This view is based on the observations that NO donor agents or induction of NO synthase in cultured cells is associated with the concomitant attenuation of cell proliferation. We have found that N/G-hydroxy-L- arginine (NOHA), an intermediate in the oxidation of arginine to NO + citrulline, is synthesized and released from cells in culture. NOHA is a potent competitive inhibitor or arginase (Ki = 10 muM) and turns off production of ornithine + urea in cells. Ornithine is the precursor to putrescine and other polyamines required for cell proliferation. We have also found that NO inhibits ornithine decarboxylase activity. The principal objective of the proposed research is to determine whether NO synthase, via the biologic actions of NOHA and NO, plays a physiological or pathophysiological role in regulating the growth of vascular smooth muscle cells and tumor cells, and to elucidate the mechanisms by which cell growth is inhibited. The central hypothesis that drives this proposal is that two products of the NO synthase reaction, NOHA and NO, function biologically to slow cell proliferation by inhibiting two sequential enzymatic steps (NOHA inhibits arginase; NO inhibits ornithine decarboxylase) in the pathway leading to the production of polyamines from arginine. These additional cytostatic mechanisms resulting from increased NO synthase activity would complement other known cytostatic mechanisms of NO such as guanylyl cyclase activation and cyclic GMP-mediated impairment of DNA synthesis. The two specific aims that will be taken to rigorously test the proposed hypothesis are: (a) to ascertain the mechanisms by which NOHA, NO and NO synthase activity interfere with cell proliferation under physiological or pathophysiological conditions and (b) to elucidate the mechanism by which NO inhibits ornithine decarboxylase. The proposed research should advance our understanding of the biological role of NO synthase in regulating cell proliferation.