Insulin-induced peroxynitrite production in human platelet-rich plasma

Abstract

Recent data support the possible role of nitric oxide (NO^•) in the development of insulin signalling. The aim of this study was to examine the effect of insulin on NO^• production by platelets. The chemiluminescence of platelet-rich plasma prepared from the blood of healthy volunteers was measured in the presence of luminol. Indirect detection of NO^• by luminol is possible in the form of peroxynitrite produced in the reaction of NO^• with a superoxide free radical. Luminol oxidation induced by hydroxyl free radical and lipid peroxidation was prevented by 150 µmol/l of desferrioxamine mesylate. Insulin, in the range of 0.084–840 nmol/l, induced a concentration-dependent increase in chemiluminescence, which was inhibited both by the competitive antagonist of the NO^• synthase enzyme, Nω-nitro-L-arginine methyl ester (at concentrations of 2.0–4.0 mmol/l, P <0.001), and by the elimination of superoxide free radicals using superoxide dismutase (72–144 IU/ml, P <0.001). In conclusion, we assume that the insulin-induced increase in chemiluminescence of platelet-rich plasma was due to increased production of NO^• and superoxide free radicals forming peroxynitrite. The data are consistent with production of peroxynitrite from human platelets under insulin stimulation.