Induction of nitric oxide synthase by protein synthesis inhibition in aortic smooth muscle cells

British Journal of Pharmacology
N MarczinJ D Catravas

Abstract

1. The role of de novo protein synthesis in inducible NO synthase (iNOS) activation was investigated in vitro by evaluating the effects of protein synthesis inhibitors cycloheximide (CH) and anisomycin (ANI) on iNOS activity, protein and mRNA levels in rat aortic smooth muscle cells (RASMC). 2. As determined by cyclic GMP accumulation, substrate (L-arginine)- and inhibitor (N(G)-monomethyl-L-arginine, NMMA)-sensitive iNOS activity was significantly elevated in CH- or ANI-treated RASMC after 24 h. 3. Lipopolysaccharide (LPS) produced a time-dependent increase in cyclic GMP levels with maximal stimulation at 6 h and a decline to near baseline at 24 h. CH attenuated LPS-induced cyclic GMP accumulation at 3 and 6 h. However, cyclic GMP levels were superinduced at later times by CH. The concentration-dependence of cyclic GMP stimulation by cycloheximide was biphasic both in the absence and presence of LPS, with maximal stimulation at 10 microM and inhibition at higher concentrations. 4. Increased iNOS activity by CH was associated with elevated levels of immunoreactive iNOS protein as judged by Western blotting in LPS- and CH-treated cells. 5. CH-induced iNOS activity and superinduction of iNOS by CH in cells treated with LPS were b...Continue Reading

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