Nitric oxide synthase generates superoxide and nitric oxide in arginine-depleted cells leading to peroxynitrite-mediated cellular injury.

Y Xia, VL Dawson, TM Dawson… - Proceedings of the …, 1996 - National Acad Sciences
Y Xia, VL Dawson, TM Dawson, SH Snyder, JL Zweier
Proceedings of the National Academy of Sciences, 1996National Acad Sciences
Besides synthesizing nitric oxide (NO), purified neuronal NO synthase (nNOS) can produce
superoxide (. O2-) at lower L-Arg concentrations. By using electron paramagnetic resonance
spin-trapping techniques, we monitored NO and. O2-formation in nNOS-transfected human
kidney 293 cells. In control transfected cells, the Ca2+ ionophore A23187 triggered NO
generation but no. O2-was seen. With cells in L-Arg-free medium, we observed. O2-
formation that increased as the cytosolic L-Arg levels decreased, while NO generation …
Besides synthesizing nitric oxide (NO), purified neuronal NO synthase (nNOS) can produce superoxide (.O2-) at lower L-Arg concentrations. By using electron paramagnetic resonance spin-trapping techniques, we monitored NO and .O2- formation in nNOS-transfected human kidney 293 cells. In control transfected cells, the Ca2+ ionophore A23187 triggered NO generation but no .O2- was seen. With cells in L-Arg-free medium, we observed .O2- formation that increased as the cytosolic L-Arg levels decreased, while NO generation declined. .O2- formation was virtually abolished by the specific NOS blocker, N-nitro-L-arginine methyl ester (L-NAME). Nitrotyrosine, a specific nitration product of peroxynitrite, accumulated in L-Arg-depleted cells but not in control cells. Activation by A23187 was cytotoxic to L-Arg-depleted, but not to control cells, with marked lactate dehydrogenase release. The cytotoxicity was largely prevented by either superoxide dismutase or L-NAME. Thus, with reduced L-Arg availability NOS elicits cytotoxicity by generating .O2- and NO that interact to form the potent oxidant peroxynitrite. Regulating arginine levels may provide a therapeutic approach to disorders involving .O2-/NO-mediated cellular injury.
National Acad Sciences