Interleukin-1 beta-induced nitric oxide production in isolated rat pancreatic islets requires gene transcription and may lead to inhibition of the Krebs cycle enzyme aconitase

Abstract

The aim of this study was to characterize the dynamics and functional relevance of interleukin-1 beta (IL-1 beta)-induced nitric oxide production in isolated pancreatic islets. Thus, islets were isolated from adult rats, precultured for 3-5 days in medium RPMI-1640 plus 10% fetal calf serum, and then exposed to IL-1 beta for different time periods, after which islet nitrite production and aconitase activity were determined. IL-1 beta (5 ng/ml) did not increase islet nitrite production during the first hour of incubation. Moreover, the nitric oxide synthase inhibitor NG-monomethyl-L-arginine (Meth-arg; 5 mM) failed to prevent the initial (90 min) IL-1 beta-induced increase in islet insulin release. After 4, 7, and 24 h, however, nitrite production was increased by 50%, 93%, and 139%, respectively. Islet aconitase activity and glucose oxidation rates were decreased by 70% after incubation for 24 h with IL-1 beta. Both Meth-arg and N alpha-p-tosyl-L-lysine chloromethyl ketone (0.1 mM), a protease inhibitor, could completely counteract the IL-1 beta-induced increases in nitrite production and inhibition of aconitase activity and glucose oxidation rates. In a separate series of experiments, islets were incubated for 60 min with or without IL-1 beta and the RNA synthesis inhibitor actinomycin-D (5 micrograms/ml) and subsequently incubated for another 9 h without any additions. The presence of actinomycin-D during the 1-h IL-1 beta incubation period prevented the IL-1 beta-induced rise in nitrite production and the IL-1 beta-induced inhibition of aconitase activity and insulin release. It is concluded that IL-1 beta-induced nitric oxide production is a late event which requires gene transcription and does not mediate the initial stimulatory effects of IL-1 beta on beta-cell function. However, the gradually augmented rate of nitric oxide production may inhibit the enzyme aconitase, leading to a suppressed mitochondrial activity and a defective insulin release in response to nutrient secretagogues.