INTERLEUKIN-1-BETA-MEDIATED INHIBITION OF ARGINASE IN RINM5F CELLS

Induction of nitric oxide synthase and generation of nitric oxide in p ancreatic islet beta-cells may mediate cytokine-induced dysfunction le ading 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 a s the arginase-catalysed formation of ornithine and urea. In this stud y 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-arginin e and the nitric oxide-generating compounds 3-morpholinosgdnonimine an d S-nitrosoglutathione. Cytosols from RINm5F cells treated with or wit hout interleukin 1 beta (0.1 nM, 18 h) were incubated (45 min, 37 degr ees C) with [U-C-14]arginine. Radiolabelled products ([C-14]citrulline from nitric oxide synthase, [C-14]ornithine and [C-14]urea from argin ase) were separated by high-performance liquid chromatography or ion-e xchange chromatography. Interleukin 1 beta increased citrulline produc tion (from 0.01 +/- 0.002 to 0.58 +/- 0.03 pmol/mu g cell protein), in dicating induction of nitric oxide synthase, and significantly decreas ed production of both ornithine (from 4.60 +/- 0.20 to 3.40 +/- 0.20 p mol/mu g) and urea (0.93 +/- 0.05 to 0.69 +/- 0.04 pmol/mu g) (P < 0.0 01), indicating decreased activity of Arginase was significantly inhib ited 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-direc ted beta-cell assault nitric oxide synthase-catalysed production of N- G-hydroxy-L-arginine and nitric oxide may inhibit arginase thereby inc reasing the availability of arginine for nitric oxide production. (C) 1997 Academic Press Limited.