Prevention of the expression of inducible nitric oxide synthase by a novel positive inotropic agent, YS 49, in rat vascular smooth muscle and RAW 264.7 macrophages.

1 The effects of a novel positive inotropic isoquinoline compound, YS 49, on NO production and iNOS protein expression were investigated in cultured rat aortic vascular smooth muscle cells (RAVSMC) and RAW 264.7 cells exposed to lipopolysaccharide (LPS) plus interferon-gamma (IFN-gamma). In addition, the effects of YS 49 on vascular hyporeactivity in vitro and ex vivo, and on survival rate (mice) and serum NOx (rat) levels, were also investigated in LPS-treated animals. 2 Pre- or co-treatment of YS 49 with LPS plus IFN-gamma, concentration-dependently reduced NO production in RAVSMC and RAW 264.7 cells (IC50 values, 22 and 30 microM, respectively). Although the inhibitory effect on NO production was reduced when YS 49 was applied 2 and 4 h after cytokine in RAW 264.7 cells, it was still statistically significant (P<0.05). 3 YS 49 reduced iNOS mRNA expression in LPS-treated rat aorta in vitro, an effect which was associated with restoration of contractility to the vasoconstrictor, phenylephrine (PE), and reduction in L-arginine-induced relaxation. 4 Serum NOx levels were significantly (P<0.01) reduced by YS 49 (5 mg kg-1, i.p.) in LPS-treated rats (10 mg kg-1, i.p.). Administration of YS 49 (10 and 20 mg kg-1) 30 min prior to LPS (10 mg kg-1) also significantly (P<0.01) increased the subsequent survival rates in mice. 5 Finally, expression of iNOS protein induced by LPS plus IFN-gamma in RAVSMC and RAW 264.7 cells was suppressed by YS 49, in a concentration-dependent manner. 6 These data strongly suggest that YS 49 suppresses iNOS gene expression induced by LPS and/or cytokines in RAVSMC and RAW 264.7 cells at the transcriptional level. YS 49 could therefore be beneficial in septic shock and other diseases associated with iNOS over-expression.