Superoxide generation by renal proximal tubule nitric oxide synthase.
Lipid A, the biologically active component of lipopolysaccharide, initiates a specific cytotoxic signaling cascade in the renal proximal tubule that involves a rapid release of intracellular calcium, the activation of nitric oxide synthase (NOS) and NO production. Superoxide (O2-) generation is also a component of this cascade and both NO and O2- are required for the development of oxidant stress and cytotoxicity. Here we examined whether NOS activity was responsible for O2- generation. In renal proximal tubules isolated from the rat, the NOS inhibitor N(G)-monomethyl-L-arginine (L-NMMA) but not D-NMMA blocked lipid A (50 microg/ml)-stimulated O2- generation as measured by the reduction of cytochrome c during a 30-min incubation period. When L-arginine (2 mM) was added to the tubule suspensions, O2- generation was significantly inhibited, while NO2- (a marker of NO generation) was significantly increased. The addition of L-arginine also reduced lipid A-stimulated malondialdehyde formation at 30 min (a marker of lipid peroxidation) and lactate dehydrogenase release at 90 min (a marker of cell death). Thus, lipid A-stimulated the generation of both NO and O2- via NOS activation. Furthermore, increasing L-arginine availability shifted NOS activity toward NO generation and reduced oxidant injury. These results offer an explanation of why scavengers of NO or oxygen radicals ameliorate endotoxin-induced acute renal failure in vivo.
Traylor
L A
LA
Department of Pharmacology and Toxicology, University of Arkansas for Medical Sciences, Little Rock 72205, USA.
Mayeux
P R
PR
D000704:Analysis of Variance; D000818:Animals; D001120:Arginine / Q000494:pharmacology; D066298:In Vitro Techniques; D007687:Kidney Tubules, Proximal / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D007770:L-Lactate Dehydrogenase / Q000032:analysis; D008050:Lipid A / Q000494:pharmacology*; D015227:Lipid Peroxidation / Q000187:drug effects; D008297:Male; D008315:Malondialdehyde / Q000378:metabolism; D009566:Nitrates / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000378:metabolism*; D009573:Nitrites / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D013237:Stereoisomerism; D013481:Superoxides / Q000378:metabolism*; D019323:omega-N-Methylarginine / Q000494:pharmacology*
D008050:Lipid A; D009566:Nitrates; D009573:Nitrites; D013481:Superoxides; D019323:omega-N-Methylarginine; D008315:Malondialdehyde; D001120:Arginine; D007770:L-Lactate Dehydrogenase; D019001:Nitric Oxide Synthase