Salehi A, Carlberg M, Henningson R, Lundquist I
Department of Pharmacology, University of Lund, Sweden.
Am J Physiol. 1996 Jun;270(6 Pt 1):C1634-41. doi: 10.1152/ajpcell.1996.270.6.C1634.
Recent immunohistochemical findings suggested that a constitutive nitric oxide synthase (cNOS) resides in endocrine pancreas. Here we provide direct biochemical evidence for the presence of cNOS activity in isolated islets. The regulating influence of this nitric oxide synthase (NOS) activity for islet hormone release was also investigated. We observed that cNOS activity could be quantitated in islet homogenates by monitoring the formation of L-citrulline from L-arginine using an Amprep CBA cation-exhange minicolumn before derivatization with o-phthaldialdehyde and subsequent high-performance liquid chromatography analysis. The islet NOS was dependent on both Ca2+ and calmodulin and suppressed by the NOS inhibitor NG-nitro-L-arginine methyl ester (L-NAME). This effect was enantiomerically specific. Islet insulin release induced by a mixture of L-arginine and glucose was enhanced by L-NAME, whereas L-arginine-induced glucagon release was inhibited. The effect of L-NAME on insulin release was dose dependently potentiated by increasing glucose concentrations, suggesting that glucose is an important regulator of islet NO production. Complementary in vivo studies showed similar results, i.e., the insulin secretory response to a mixture of glucose and L-arginine was extremely enhanced by pretreatment with L-NAME, whereas L-arginine-stimulated glucagon response was suppressed. Finally, in isolated islets, the intracellular nitric oxide (NO) donor hydroxylamine suppressed insulin release and increased glucagon release. In summary, the islets of Langerhans contain a constitutive, Ca2+/calmodulin-dependent isoform of NOS. Islet NO suppressed insulin but enhanced glucagon secretion. The data also suggest a negative feedback by NO on glucose-induced insulin release. The islet NO system is a novel and important regulatory factor in insulin and glucagon secretion.
最近的免疫组织化学研究结果表明，组成型一氧化氮合酶（cNOS）存在于内分泌胰腺中。在此，我们为分离的胰岛中存在cNOS活性提供了直接的生化证据。我们还研究了这种一氧化氮合酶（NOS）活性对胰岛激素释放的调节作用。我们观察到，通过在邻苯二甲醛衍生化和随后的高效液相色谱分析之前，使用Amprep CBA阳离子交换微型柱监测从L-精氨酸形成L-瓜氨酸，可对胰岛匀浆中的cNOS活性进行定量。胰岛NOS依赖于Ca2+和钙调蛋白，并被NOS抑制剂NG-硝基-L-精氨酸甲酯（L-NAME）抑制。这种作用具有对映体特异性。L-NAME可增强L-精氨酸和葡萄糖混合物诱导的胰岛胰岛素释放，而L-精氨酸诱导的胰高血糖素释放则受到抑制。L-NAME对胰岛素释放的作用随葡萄糖浓度增加呈剂量依赖性增强，这表明葡萄糖是胰岛NO产生的重要调节因子。补充的体内研究显示了类似的结果，即L-NAME预处理可极大地增强对葡萄糖和L-精氨酸混合物的胰岛素分泌反应，而L-精氨酸刺激的胰高血糖素反应则受到抑制。最后，在分离的胰岛中，细胞内一氧化氮（NO）供体羟胺抑制胰岛素释放并增加胰高血糖素释放。总之，胰岛中含有一种组成型的、依赖Ca2+/钙调蛋白的NOS同工型。胰岛NO抑制胰岛素分泌但增强胰高血糖素分泌。这些数据还表明NO对葡萄糖诱导的胰岛素释放存在负反馈。胰岛NO系统是胰岛素和胰高血糖素分泌中的一种新的重要调节因子。
