Interleukin-1 beta-mediated inhibition of arginase in RINm5F cells

Induction of nitric oxide synthase and generation of nitric oxide in pancreatic islet beta-cells may mediate cytokine-induced dysfunction leading to insulin-dependent diabetes mellitus. Nitric oxide generation can be regulated by availability of arginine substrate which, in turn, may be affected by substrate utilization in competing pathways such as the arginase-catalysed formation of ornithine and urea. In this study we have investigated the activity of arginase in the rat insulinoma-derived cell line RINm5F and the effect on this of interleukin 1 beta, the nitric oxide synthase reaction intermediate N-G-hydroxy-L-arginine and the nitric oxide-generating compounds 3-morpholinosgdnonimine and S-nitrosoglutathione. Cytosols from RINm5F cells treated with or without interleukin 1 beta (0.1 nM, 18 h) were incubated (45 min, 37 degrees C) with [U-C-14]arginine. Radiolabelled products ([C-14]citrulline from nitric oxide synthase, [C-14]ornithine and [C-14]urea from arginase) were separated by high-performance liquid chromatography or ion-exchange chromatography. Interleukin 1 beta increased citrulline production (from 0.01 +/- 0.002 to 0.58 +/- 0.03 pmol/mu g cell protein), indicating induction of nitric oxide synthase, and significantly decreased production of both ornithine (from 4.60 +/- 0.20 to 3.40 +/- 0.20 pmol/mu g) and urea (0.93 +/- 0.05 to 0.69 +/- 0.04 pmol/mu g) (P < 0.001), indicating decreased activity of Arginase was significantly inhibited by N-G-hydroxy-L-arginine (IC50 = 50 mu M), S-nitrosoglutathione (500 mu M: 69 +/- 7% of control) and 3-morpholinosydnonimine (1 mM: 57 +/- 7% of control) (P < 0.05). We conclude that during cytokine-directed beta-cell assault nitric oxide synthase-catalysed production of N-G-hydroxy-L-arginine and nitric oxide may inhibit arginase thereby increasing the availability of arginine for nitric oxide production. (C) 1997 Academic Press Limited.