Nitric oxide differentially regulates induction of type II nitric oxide synthase in rat vascular smooth muscle cells versus macrophages

We studied effects of nitric oxide (NO) released by different MO donors on induction of inducible NO synthase (iNOS) in rat aortic smooth muscle cells (RASMC) and rat macrophage cell line NR8383. iNOS protein expression induc ed by a CM (interleukin-1 beta 250 U/mL, interferon-gamma 150 U/mL, and tum or necrosis factor-alpha 150 U/mL) was not affected by the NO donor SNAP (0 .2 to 1 mmol/l,) in RASMC at 24 hours of incubation but was dose-dependentl y decreased by SNAP in macrophages (maximal 60% inhibition). A fully functi onal -3.2-kb rat iNOS promoter was transfected into RASMC and macrophages. The CM-induced promoter activity in transfected macrophages was inhibited b y SNAP (maximal 67% inhibition), but this inhibitory effect by SNAP was not observed in transfected RASMC. Electrophoretic mobility-shift assays demon strated that nuclear factor-kappaB (NF-kappaB) binding patterns were differ ent in 2 cell types and that the ratio of p50:p65 subunits was significantl y lower in macrophages than in RASMC. Furthermore, NF-kappaB activity was n ot affected by SNAP in RASMC but was reduced by SNAP in macrophages. Anothe r putative NO donor, NOR3 (1 mmol/L), completely inhibited iNOS induction b y CM in RASMC, but this was accompanied by severe cytotoxicity, which resul ted in cell death. Similar concentrations of SNAP did not exhibit cytotoxic ity in RASMC, whereas macrophages demonstrated 88% viability compared with cells without SNAP. NO synthase inhibitor N-g-monomethyl-L-arginine signifi cantly inhibited CM-induced nitrite production in both cell types and stimu lated iNOS protein expression in macrophages but did not affect iNOS expres sion in RASMC. These data strongly suggest that NO may affect transcription al regulation of iNOS differently in RASMC versus macrophages, possibly by means of regulation of NF-kappaB activation.