Impaired membrane translocation of thrombin stimulated PKC epsilon by high glucose.
Impaired membrane translocation of thrombin stimulated PKC epsilon by high glucose.
复制标题
高葡萄糖刺激 PKC epsilon，凝血酶膜易位受损。
DOI：
10.1677/joe.0.157r007
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发表时间：
1998
影响因子：
4
通讯作者：
W. Waldhäusl
中科院分区：
文献类型：
--
作者：
G. Reining;S. Baumgartner‐Parzer;W. Waldhäusl
众所周知，高血糖会导致内皮功能障碍，并促进糖尿病血管病变。因此，本研究旨在评价30 mM和5 mM葡萄糖长期孵育(16+/-1天)对配对培养的人脐静脉内皮细胞(HUVECs)配体诱导的蛋白激酶C(PKC)转位的影响。增加凝血酶浓度(0.01、0.1、1、10和100 nM)分别在5 mM和30 mM葡萄糖中刺激细胞30s，并用免疫印迹法检测PKC亚型α和epsilon。融合培养细胞在凝血酶刺激下，膜结合的PKCα和epsilon呈浓度依赖性升高。与正常葡萄糖相比，高糖对凝血酶对PKCα的转位没有影响，而在30 mM葡萄糖中生长的细胞中，与在5 mM葡萄糖中培养的对照细胞相比，在最大凝血酶浓度(曲线下面积，AUC90.4+/-7%；p<；0.008；n=6)下生长的细胞中PKCepsilon的转位减少。在相同的分离株中，30 mM甘露醇作为渗透压对照并不能减少PKC epsilon的易位。静息细胞在30 mM葡萄糖和5 mM葡萄糖长期孵育后，其总PKCα和膜结合PKCα或PKC epsilon无明显变化。这些数据表明，30 mM葡萄糖以一种异构体特异性的方式诱导PKC易位，而在没有凝血酶刺激的情况下，PKC异构体的亚细胞分布仍然不受30 mM葡萄糖的影响。
Hyperglycaemia is known to cause endothelial dysfunction and to promote diabetic angiopathy. Therefore, this study was designed to evaluate the effect of long term incubation (16 +/- 1 days) in 30 mM vs 5 mM glucose on ligand induced translocation of protein kinase C (PKC) in paired cultures of individual isolates of human umbilical vein endothelial cells (HUVECs). Cells were stimulated with increasing concentrations of thrombin (0.01, 0.1, 1, 10 and 100 nM) for 30 seconds in the presence of 5 mM and 30 mM glucose, respectively, and analyzed by immunoblotting for PKC-isoforms alpha and epsilon. Stimulation by thrombin of confluent cultures displayed a concentration dependent rise in membrane bound PKC alpha and epsilon. Translocation of PKC alpha by thrombin remained unaffected by high versus normal ambient glucose, whereas translocation of PKC epsilon in cells grown in 30mM glucose was reduced at maximal thrombin concentrations (area under the curve, AUC: 90.4 +/- 7% of control cells; p < 0.008; n = 6) versus control cultures kept in 5mM glucose. In the identical isolates translocation of PKC epsilon was not reduced by 30 mM mannitol used as osmotic control. No change was induced by long term incubation of resting cells with 30 mM vs 5 mM glucose as to total and membrane bound PKC alpha or PKC epsilon. The obtained data suggest modulation by 30 mM glucose of ligand induced PKC translocation in an isoform specific manner, whereas subcellular distribution of PKC isoforms in the absence of thrombin stimulation remains unaffected by 30 mM glucose.