Arafat Hwyda A, Katakam Anand K, Chipitsyna Galina, Gong Qiaoke, Vancha Ajith R, Gabbeta Jagadeesh, Dafoe Donald C
Department of Surgery, Thomas Jefferson University, 1015 Walnut Street, Philadelphia, Pennsylvania 19107, USA.
Endocrinology. 2007 Feb;148(2):575-84. doi: 10.1210/en.2006-0970. Epub 2006 Nov 16.
Osteopontin (OPN), a phosphorylated glycoprotein that binds to an integrin-binding motif, has been shown to regulate nitric oxide (NO) production via inhibition of induced NO synthase (iNOS) synthesis. In the transplanted islets, iNOS and toxic amounts of NO are produced as a result of islets infiltration with inflammatory cells and production of proinflammatory cytokines. Here, we demonstrate that addition of OPN before IL-1beta in freshly isolated rat islets improved their glucose stimulated insulin secretion dose-dependently and inhibited IL-1beta-induced NO production in an arginine-glycine-aspartate-dependent manner. Transient transfection of OPN gene in RINm5F beta-cells fully prevented the toxic effect of IL-1beta at concentrations that reduced the viability by 50% over 3 d. OPN prevention of IL-1beta-induced toxicity was accompanied by inhibited transcription of iNOS by 80%, resulting in 50% decreased formation of the toxic NO. In OPN-transfected cells, the IL-1beta-induced nuclear factor-kappaB activity was significantly reduced. Islets exposed to IL-1beta revealed a naturally occurring early up-regulated OPN transcription. OPN promoter activity was increased in the presence of IL-1beta, IL-1beta-induced NO, and an inducer of NO synthesis. These data suggest the presence of a cross talk between the IL-1beta and OPN pathways and a unique trans-regulatory mechanism in which IL-1beta-induced NO synthesis feedback regulates itself through up-regulation of OPN gene transcription. Our data also suggest that influencing OPN expression represents an approach for affecting cytokine-induced signal transduction to prevent or reduce activation of the cascade of downstream devastating effects after islet transplantation.
骨桥蛋白（OPN）是一种与整合素结合基序结合的磷酸化糖蛋白，已被证明可通过抑制诱导型一氧化氮合酶（iNOS）的合成来调节一氧化氮（NO）的产生。在移植的胰岛中，由于胰岛被炎性细胞浸润和促炎细胞因子的产生，会产生iNOS和有毒量的NO。在此，我们证明，在新鲜分离的大鼠胰岛中，在白细胞介素-1β（IL-1β）之前添加OPN可剂量依赖性地改善其葡萄糖刺激的胰岛素分泌，并以精氨酸-甘氨酸-天冬氨酸依赖性方式抑制IL-1β诱导的NO产生。在RINm5Fβ细胞中瞬时转染OPN基因可完全预防IL-1β的毒性作用，该浓度在3天内可使细胞活力降低50%。OPN预防IL-1β诱导的毒性伴随着iNOS转录被抑制80%，导致有毒NO的形成减少50%。在转染OPN的细胞中，IL-1β诱导的核因子-κB活性显著降低。暴露于IL-1β的胰岛显示出自然发生的早期OPN转录上调。在存在IL-1β、IL-1β诱导的NO和NO合成诱导剂的情况下，OPN启动子活性增加。这些数据表明IL-1β和OPN途径之间存在相互作用，以及一种独特的反式调节机制，其中IL-1β诱导的NO合成通过上调OPN基因转录来反馈调节自身。我们的数据还表明，影响OPN表达代表了一种影响细胞因子诱导的信号转导的方法，以预防或减少胰岛移植后下游破坏性效应级联反应的激活。
