Expression of the citrulline-nitric oxide cycle in rodent and human pancreatic beta-cells: induction of argininosuccinate synthetase by cytokines.

Nitric oxide (NO) may be a mediator of beta-cell damage in insulin-dependent diabetes mellitus. beta-Cells express the inducible form of NO synthase (iNOS) and produce large amounts of NO upon exposure to cytokines. iNOS requires the amino acid arginine for NO formation. It has been shown in other cell types that interferon-gamma (IFN gamma) and bacterial lipopolysaccharide induce the enzyme argininosuccinate synthetase (AS), enhancing the capacity of these cells to regenerate arginine from citrulline and maintain NO production in the presence of low arginine concentrations. To characterize the messenger RNA (mRNA) expression of AS in insulin-producing cells, RINm5F cells (RIN cells) were exposed to interleukin-1 beta (IL-1 beta) or to tumor necrosis factor-alpha plus IFN gamma. After 4-6 h, there was a significant and parallel induction of AS and iNOS mRNA. IL-1 beta-induced AS and iNOS mRNA expression was prevented by an inhibitor of the activation factor NF-kappa B pyrrolidine diaminoguanidine, an inhibitor of gene transcription (actinomycin D), and a blocker of protein synthesis (cycloheximide), suggesting coregulation of AS and iNOS by cytokines. RIN cells exposed to IL-1 beta in the presence of citrulline but the absence of arginine had increased AS enzyme activity and produced NO, demonstrating that cytokine-induced AS mRNA expression is accompanied by increased AS activity. Both adult rat islets exposed to IL-1 beta and human pancreatic islets cultured in the presence of IL-1 beta, tumor necrosis factor-alpha, and IFN gamma were able to use citrulline to regenerate arginine and produce NO. Taken as a whole, the present data suggest that regulation of AS activity may play a role in modulation of NO production in both rodent and human insulin-producing cells.