OBJECTIVETo investigate the effect of iNOS gene on cell apoptosis and insulin secretion of pancreas islet in rats by RNA inference (RNAi).

METHODSIslets obtained from thirty Wistar rats were randomly divided into five groups, and siRNA oligo was purchased from Genepharma in Shanghai. The cultured islets were transfected with iNOS siRNA, and then were divided into five groups. Islet cultured only was taken as blank control group, and cultured with TNF-alpha + IL-1 beta as cytokine group. Islet transfected with negative or iNOS siRNA were taken as negative transfection control group or RNAi group, while that transfected with iNOS siRNA and cultured with TNF-alpha + IL-1 beta as RNAi + cytokine group. Expression of iNOS mRNA was evaluated by RT-PCR and iNOS protein was evaluated by Western blot to detect the effect of RNAi. The expression of apoptosis correlated gene, Bax, Fas were analyzed, and the apoptotic cells were identified by TUNEL method meanwhile. Insulin secretion index assay the function of the islets.

RESULTS500 - 600 IEQ islets could be extracted from every rat. RNAi attenuated the expression of iNOS and restrained the synthesis of iNOS protein.With treatment of cytokines IL-1 beta and TNF-alpha, the level of iNOS increased remarkably, the expression of Bax and Fas ascended distinctly, and insulin secretion index decreased strikingly. While, the expression of apoptosis gene and amount of apoptotic cells descended in group of RNAi + cytokine, and insulin secretion index were satisfying.

CONCLUSIONThe apoptosis from cytokines to islets mediated by iNOS could be suppressed by RNAi, which leaded to favorable function and survival of islets.