Blair L A, Heitmeier M R, Scarim A L, Maggi L B, Corbett J A
Edward A. Doisy Department of Biochemistry and Molecular Biology, St Louis University School of Medicine, Missouri 63104, USA.
Diabetes. 2001 Feb;50(2):283-90. doi: 10.2337/diabetes.50.2.283.
Environmental factors, such as viral infection, have been implicated in the destruction of beta-cells during the development of autoimmune diabetes. Double-stranded RNA (dsRNA), produced during viral replication, is an active component of a viral infection that stimulates antiviral responses in infected cells. Previous studies have shown that treatment of rat islets with dsRNA in combination with gamma-interferon (IFN-gamma) results in a nitric oxide-dependent inhibition of glucose-stimulated insulin secretion. This study examines the role of nuclear factor-kappaB (NF-kappaB) and the dsRNA-dependent protein kinase (PKR) in dsRNA + IFN-gamma-induced nitric oxide synthase (iNOS) expression and nitric oxide production by rat, mouse, and human islets. Treatment of rat and human islets with dsRNA in the form of polyinosinic-polycytidylic acid (poly IC) and IFN-gamma resulted in iNOS expression and nitric oxide production. Inhibitors of NF-kappaB activation-the proteasome inhibitor MG-132 and the antioxidant pyrrolidine-dithiocarbamate (PDTC)-prevented poly IC + IFN-gamma-induced iNOS expression and nitric oxide production. Incubation of rat islets for 3 h or human islets for 2 h with poly IC alone or poly IC + IFN-gamma resulted in NF-kappaB nuclear translocation and degradation of the NF-kappaB inhibitor protein, IkappaB, events that are prevented by MG-132. PKR has been shown to participate in dsRNA-induced NF-kappaB activation in a number of cell types, including mouse embryonic fibroblasts. However, poly IC stimulated NF-kappaB nuclear translocation and IkappaB degradation to similar levels in islets isolated from mice devoid of PKR (PKR-/-) and wild-type mice (PKR+/+). Furthermore, the genetic absence of PKR did not affect dsRNA + IFN-gamma-induced iNOS expression, nitric oxide production, or the inhibitory actions of these agents on glucose-stimulated insulin secretion. These results suggest that 1) NF-KB activation is required for dsRNA + IFN-gamma-induced iNOS expression, 2) PKR is not required for either dsRNA-induced NF-kappaB activation or dsRNA + IFN-y-induced iNOS expression by islets, and 3) PKR is not required for dsRNA + IFN-gamma-induced inhibition of glucose-stimulated insulin secretion by islets.
环境因素，如病毒感染，被认为与自身免疫性糖尿病发展过程中β细胞的破坏有关。病毒复制过程中产生的双链RNA（dsRNA）是病毒感染的一种活性成分，可刺激受感染细胞产生抗病毒反应。先前的研究表明，用dsRNA联合γ干扰素（IFN-γ）处理大鼠胰岛会导致一氧化氮依赖性抑制葡萄糖刺激的胰岛素分泌。本研究探讨了核因子κB（NF-κB）和dsRNA依赖性蛋白激酶（PKR）在dsRNA + IFN-γ诱导大鼠、小鼠和人类胰岛一氧化氮合酶（iNOS）表达及一氧化氮产生中的作用。用多聚肌苷酸-多聚胞苷酸（poly IC）形式的dsRNA和IFN-γ处理大鼠和人类胰岛，可导致iNOS表达及一氧化氮产生。NF-κB激活抑制剂——蛋白酶体抑制剂MG-132和抗氧化剂吡咯烷二硫代氨基甲酸盐（PDTC）——可阻止poly IC + IFN-γ诱导的iNOS表达及一氧化氮产生。单独用poly IC或poly IC + IFN-γ孵育大鼠胰岛3小时或人类胰岛2小时，会导致NF-κB核转位及NF-κB抑制蛋白IkappaB降解，而MG-132可阻止这些事件发生。在包括小鼠胚胎成纤维细胞在内的多种细胞类型中，PKR已被证明参与dsRNA诱导的NF-κB激活。然而，poly IC在缺乏PKR的小鼠（PKR-/-）和野生型小鼠（PKR+/+）分离的胰岛中刺激NF-κB核转位及IkappaB降解的程度相似。此外，PKR基因缺失并不影响dsRNA + IFN-γ诱导的iNOS表达、一氧化氮产生，或这些试剂对葡萄糖刺激的胰岛素分泌的抑制作用。这些结果表明：1）dsRNA + IFN-γ诱导的iNOS表达需要NF-κB激活；2）胰岛中dsRNA诱导的NF-κB激活或dsRNA + IFN-γ诱导的iNOS表达不需要PKR；3）胰岛中dsRNA + IFN-γ诱导的葡萄糖刺激的胰岛素分泌抑制不需要PKR。
