Cunningham J M, Mabley J G, Green I C
Biochemistry Laboratory, School of Biological Sciences, University of Sussex, Brighton, BN1 9QG, UK.
Cytokine. 1997 Aug;9(8):570-6. doi: 10.1006/cyto.1996.0203.
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.
胰岛β细胞中一氧化氮合酶的诱导及一氧化氮的生成可能介导细胞因子诱导的功能障碍，进而导致胰岛素依赖型糖尿病。一氧化氮的生成可受精氨酸底物可用性的调节，而精氨酸底物可用性又可能受到诸如精氨酸酶催化生成鸟氨酸和尿素等竞争途径中底物利用情况的影响。在本研究中，我们调查了大鼠胰岛素瘤来源的细胞系RINm5F中精氨酸酶的活性，以及白细胞介素1β、一氧化氮合酶反应中间体N G -羟基-L-精氨酸和一氧化氮生成化合物3-吗啉代亚硝基胍及S-亚硝基谷胱甘肽对其的影响。用或不用白细胞介素1β（0.1nM，18小时）处理的RINm5F细胞的胞质溶胶与[U-14C]精氨酸一起孵育（45分钟，37℃）。通过高效液相色谱或离子交换色谱分离放射性标记产物（一氧化氮合酶产生的[14C]瓜氨酸、精氨酸酶产生的[14C]鸟氨酸和[14C]尿素）。白细胞介素1β增加了瓜氨酸的生成（从0.01±0.002增加到0.58±0.03 pmol/μg细胞蛋白），表明一氧化氮合酶被诱导，同时显著降低了鸟氨酸（从4.60±0.20降低到3.40±0.2）和尿素（从0.93±立0.05降低到0.69±0.04 pmol/μg）的生成（P<0.001），表明精氨酸酶活性降低。精氨酸酶被N G -羟基-L-精氨酸（IC50 = 50μM）、S-亚硝基谷胱甘肽（500μM：为对照的69±7%）和3-吗啉代亚硝基胍（1 mM：为对照的57±7%）显著抑制（P<0.05）。我们得出结论，在细胞因子介导的β细胞攻击过程中，一氧化氮合酶催化生成的N G -羟基-L-精氨酸和一氧化氮可能抑制精氨酸酶，从而增加用于一氧化氮生成的精氨酸的可用性。
