Mechanisms of action of lipoxygenase and cytochrome P-450-mono-oxygenase inhibitors in blocking endothelium-dependent vasodilatation.

1. Acetylcholine, ionophore A23187 and melittin induced endothelium-dependent relaxations of preconstricted strips of rabbit aorta. These relaxations are likely to be mediated by endothelium-derived relaxing factor (EDRF). 2. Relaxations in response to acetylcholine (1 microM) were inhibited by the following lipoxygenase inhibitors, with the approximate IC50 values indicated in parentheses: gossypol (1.5 microM), nordihydroguairetic acid (NDGA, 5 microM), AA 861 (20 microM), phenidone (30 microM), quercetin (40 microM), BW 755C (300 microM), and piriprost (500 microM); with cirsiliol 50% inhibition was not achieved. Acetylcholine-induced relaxations were also blocked by the cytochrome P-450-mono-oxygenase inhibitors proadifen (SKF 525A, 4 microM), metyrapone (300 microM), and cimetidine (300 microM); 7,8 benzoflavone had no effect up to 100 microM. 3. The more potent inhibitors were also tested against relaxations induced by A23187 (0.1 microM) and melittin (1 microM) and produced partial inhibition of these relaxations. 4. The mechanism of action of the more potent inhibitors was investigated in a bioassay system. EDRF was produced in columns filled with cultured human endothelial cells. The factor was bioassayed with endothelium denuded segments of rabbit femoral artery. When added to effluent of the column, NDGA, AA861, proadifen and metyrapone inhibited the EDRF-induced vasodilatation, whereas gossypol had no effect. Gossypol, however, blocked EDRF production when infused through the column. 5. The more potent inhibitors were also tested to determine their effect on purified soluble guanylate cyclase. While gossypol, NDGA and proadifen had no appreciable effects, basal and nitroprusside (50 microM)-stimulated guanylate cyclase activity was inhibited by AA861 and metyrapone. 6. These data suggest that many of the above compounds inhibit EDRF by mechanisms other than lipoxygenase- or cytochrome P-450-mono-oxygenase inhibition.