REGULATION OF INTERLEUKIN-1-BETA-STIMULATED INDUCIBLE NITRIC-OXIDE SYNTHASE EXPRESSION IN CULTURED VASCULAR SMOOTH-MUSCLE CELLS BY HEMOSTATIC PROTEINS

Experiments were performed to examine the mechanism by which specific hemostatic proteins regulate the release of nitric oxide (NO) from int erleukin-1 beta (IL-1 beta) stimulated cultured rat aortic smooth musc le cells. Treatment of smooth muscle cells with IL-beta stimulated ind ucible nitric oxide synthase (iNOS) mRNA expression, which preceded th e release of NO (as measured by the accumulation of nitrite in the cul ture media). The cytokine-stimuiated production of nitrite was blocked by the protein synthesis inhibitor cycloheximide, the transcriptional inhibitor actinomycin D, and the competitive inhibitor of NOS nitro-L -arginine. However, only actinomycin D inhibited IL-1 beta-stimulated iNOS mRNA expression. Treatment of smooth muscle cells with IL-1 beta in the presence of platelet derived growth factor or thrombin resulted in the inhibition of cytokine-stimulated expression of iNOS mRNA and NO release. The inhibitory effect of thrombin was reversed by hirudin and was mimicked by a 14 amino acid thrombin receptor activating pepti de. In contrast, the concomitant exposure of smooth muscle cells to IL -1 beta and plasmin resulted in the potentiation of both IL-1 beta-sti mulated iNOS expression and NO generation. Finally, treatment of smoot h muscle cells with IL-1 beta in the presence of the hemostatic protei ns did not affect the half-life of iNOS mRNA. These results demonstrat e that specific protein components of the hemostatic system regulate I L-1 beta-stimulated iNOS mRNA expression in vascular smooth muscle cel ls. The capacity of hemostatic proteins to modulate the induction of v ascular iNOS activity may play an important role in governing the rele ase of NO and regulating thrombogenesis in vivo.