Interleukin 1beta-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 1beta, the nitric oxide synthase reaction intermediate NG-hydroxy-l-arginine and the nitric oxide-generating compounds 3-morpholinosydnonimine and S-nitrosoglutathione. Cytosols from RINm5F cells treated with or without interleukin 1beta (0.1nM, 18h) were incubated (45min, 37 degrees C) with [U-14C]arginine. Radiolabelled products ([14C]citrulline from nitric oxide synthase, [14C]ornithine and [14C]urea from arginase) were separated by high-performance liquid chromatography or ion-exchange chromatography. Interleukin 1beta increased citrulline production (from 0.01+/-0.002 to 0.58+/-0.03 pmol/microg 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/microg) and urea (0.93+/-0.05 to 0.69+/-0.04 pmol/microg) (P<0.001), indicating decreased activity of arginase. Arginase was significantly inhibited by NG-hydroxy-l-arginine (IC50=50 microM), S-nitrosoglutathione (500 microM: 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 NG-hydroxy-l-arginine and nitric oxide may inhibit arginase thereby increasing the availability of arginine for nitric oxide production.
Cunningham
J M
JM
Biochemistry Laboratory, School of Biological Sciences, University of Sussex, Brighton, BN1 9QG, UK.
Mabley
J G
JG
Green
I C
IC
D000818:Animals; D001119:Arginase / Q000037:antagonists & inhibitors*; D001120:Arginine / Q000302:isolation & purification; D002956:Citrulline / Q000302:isolation & purification; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005978:Glutathione / Q000031:analogs & derivatives / Q000494:pharmacology; D007340:Insulinoma; D007375:Interleukin-1 / Q000494:pharmacology*; D007515:Islets of Langerhans / Q000187:drug effects / Q000201:enzymology; D008981:Molsidomine / Q000031:analogs & derivatives / Q000494:pharmacology; D019001:Nitric Oxide Synthase / Q000378:metabolism; D009603:Nitroso Compounds / Q000494:pharmacology; D009952:Ornithine / Q000302:isolation & purification; D051381:Rats; D026422:S-Nitrosoglutathione; D014407:Tumor Cells, Cultured
D004791:Enzyme Inhibitors; D007375:Interleukin-1; D009603:Nitroso Compounds; D002956:Citrulline; D026422:S-Nitrosoglutathione; C002385:linsidomine; D001120:Arginine; D008981:Molsidomine; D009952:Ornithine; D019001:Nitric Oxide Synthase; D001119:Arginase; D005978:Glutathione