Mancusi G, Hutter C, Baumgartner-Parzer S, Schmidt K, Schütz W, Sexl V
Institute of Pharmacology, University of Vienna, Wien, Austria.
Biochem J. 1996 Apr 1;315 ( Pt 1)(Pt 1):281-7. doi: 10.1042/bj3150281.
Alterations in G-protein-controlled signalling pathways (primarily pathways controlled by Gs and Gi) have been reported to occur in animal models of diabetes mellitus. We have therefore studied the effect of a long-term exposure of human umbilical vein endothelial cells to elevated concentrations of glucose on expression and function of G-protein subunits and endothelial NO synthase. Long-term incubation in high glucose (30 mM for 15 days) did not affect the levels of Gialpha-2, Gqalpha, the splice variants (long and short form) of Gsalpha, and the G-protein beta-subunits or adenylate cyclase activity; basal, as well as isoprenaline-, forskolin- and guanosine 5'-[gamma-thio]triphosphate-stimulated enzyme activities were comparable in high- and low-glucose-treated cells, thus ruling out any functional changes in the stimulatory pathway. Pretreatment of endothelial cells with pertussis toxin blocked a substantial fraction (50%) of the mitogenic response to serum factor(s) which depend(s) of functional Gi2. The sensitivity of cells cultured in high glucose was comparable with that of the paired controls maintained in normal glucose (EC50 = 3.1 +/- 0.5 and 3.3 +/- 0.4 ng/ml respectively). Similarly, we failed to detect any differences in endothelial NO synthase expression, or intracellular distribution and basal activity of the enzyme in endothelial cells cultured in high glucose. Stimulation of NO synthase in intact cells revealed a comparable response to the calcium ionophore (A23187). In contrast, stimulation with histamine (which acts via H1-receptors predominantly coupled to Gq) resulted in a significantly increased response in the cells maintained in high glucose. These data are suggestive of an altered H1-histamine receptor-Gq-phospholipase C pathway in endothelial cells cultured in high glucose concentrations, but rule out any glucose-induced functional changes in Gs- and Gi-controlled signalling pathways.
据报道，在糖尿病动物模型中会出现G蛋白控制的信号通路（主要是由Gs和Gi控制的通路）的改变。因此，我们研究了人脐静脉内皮细胞长期暴露于高浓度葡萄糖对G蛋白亚基和内皮型一氧化氮合酶的表达及功能的影响。在高糖（30 mM，处理15天）条件下长期孵育，并未影响Gialpha - 2、Gqalpha、Gsalpha的剪接变体（长形式和短形式）、G蛋白β亚基的水平或腺苷酸环化酶活性；高糖处理组和低糖处理组细胞的基础活性以及异丙肾上腺素、福斯可林和鸟苷5'-[γ-硫代]三磷酸刺激的酶活性相当，从而排除了刺激通路中的任何功能变化。用百日咳毒素预处理内皮细胞可阻断对依赖功能性Gi2的血清因子的大部分（50%）促有丝分裂反应。在高糖中培养的细胞的敏感性与在正常葡萄糖中培养的配对对照相当（EC50分别为3.1±0.5和3.3±0.4 ng/ml）。同样，我们未能检测到在高糖中培养的内皮细胞中内皮型一氧化氮合酶表达、该酶的细胞内分布及基础活性存在任何差异。完整细胞中一氧化氮合酶的刺激显示对钙离子载体（A23187）的反应相当。相反，用组胺刺激（组胺主要通过与Gq偶联的H1受体起作用）导致在高糖中培养的细胞中的反应显著增强。这些数据表明在高糖浓度下培养的内皮细胞中H1 - 组胺受体 - Gq - 磷脂酶C通路发生了改变，但排除了葡萄糖诱导的Gs和Gi控制的信号通路中的任何功能变化。
