Bubolz Aaron H, Wu Qingping, Larsen Brandon T, Gutterman David D, Liu Yanping
Department of Medicine, The Medical College of Wisconsin, Milwaukee, Wisconsin, USA.
Am J Physiol Heart Circ Physiol. 2007 Oct;293(4):H2231-7. doi: 10.1152/ajpheart.00717.2007. Epub 2007 Aug 3.
Small coronary arteries (SCA) from diabetic rats exhibit enhanced peroxynitrite (ONOO(-)) formation and concurrent impairment of voltage-dependent potassium (K(v)) channel function. However, it is unclear whether ONOO(-) plays a causative role in this impairment. We hypothesized that functional loss of K(v) channels in coronary smooth muscle cells (SMC) in diabetes is due to ONOO(-) with subsequent tyrosine nitration of K(v) channel proteins. Diabetic rats and nondiabetic controls were treated with or without ebselen (Eb) for 4 wk. SCA were prepared for immunohistochemistry (IHC), immunoprecipitation (IP) followed by Western blot (WB), videomicroscopy, and patch-clamp analysis. IHC revealed excess ONOO(-) in SCA from diabetic rats. IP and WB revealed elevated nitration of the K(v)1.2 alpha-subunit and reduced K(v)1.2 protein expression in diabetic rats. Each of these changes was improved in Eb-treated rats. Protein nitration and K(v)1.5 expression were unchanged in SCA from diabetic rats. Forskolin, a direct cAMP activator that induces K(v)1 channel activity, dilated SCA from nondiabetic rats in a correolide (Cor; a selective K(v)1 channel blocker)-sensitive fashion. Cor did not alter the reduced dilation to forskolin in diabetic rats; however, Eb partially restored the Cor-sensitive component of dilation. Basal K(v) current density and response to forskolin were improved in smooth muscle cells from Eb-treated DM rats. We conclude that enhanced nitrosative stress in diabetes mellitus contributes to K(v)1 channel dysfunction in the coronary microcirculation. Eb may be beneficial for the therapeutic treatment of vascular complications in diabetes mellitus.
糖尿病大鼠的小冠状动脉(SCA)表现出过氧亚硝酸盐(ONOO⁻)生成增加以及电压依赖性钾(Kv)通道功能同时受损。然而,尚不清楚ONOO⁻是否在这种损伤中起因果作用。我们假设糖尿病时冠状动脉平滑肌细胞(SMC)中Kv通道的功能丧失是由于ONOO⁻导致Kv通道蛋白随后发生酪氨酸硝化。糖尿病大鼠和非糖尿病对照大鼠接受或不接受依布硒啉(Eb)治疗4周。制备SCA用于免疫组织化学(IHC)、免疫沉淀(IP)后进行蛋白质印迹(WB)、视频显微镜检查和膜片钳分析。IHC显示糖尿病大鼠SCA中存在过量的ONOO⁻。IP和WB显示糖尿病大鼠中Kv1.2α亚基的硝化增加且Kv1.2蛋白表达减少。在接受Eb治疗的大鼠中,这些变化均得到改善。糖尿病大鼠SCA中蛋白质硝化和Kv1.5表达未发生变化。福斯可林是一种直接的cAMP激活剂,可诱导Kv1通道活性,它能以对可雷洛肽(Cor;一种选择性Kv1通道阻滞剂)敏感的方式扩张非糖尿病大鼠的SCA。Cor并未改变糖尿病大鼠对福斯可林扩张反应的降低;然而,Eb部分恢复了对Cor敏感的扩张成分。接受Eb治疗的糖尿病大鼠的平滑肌细胞中基础Kv电流密度以及对福斯可林的反应均得到改善。我们得出结论,糖尿病中氧化应激增强导致冠状动脉微循环中Kv1通道功能障碍。Eb可能对糖尿病血管并发症的治疗有益。