吡格列酮对内皮细胞功能影响及其机制的体外研究

in vitro Study of the Effects of Pioglitazone on Endothelial Cell Functions of Human Umbilical Vein and the Mechanisms Involved

目的了解吡格列酮(PIO)对内皮细胞(EC)功能指标的影响及其可能的作用机理。方法体外原代培养人脐静脉内皮细胞(HUVECs),分别加入5.5mmol/L葡萄糖(对照组)、30mmol/L葡萄糖(高糖组)以及30mmol/L葡萄糖+PIO(10-9mol/L、10-7mol/L、10-5mol/L)(PIO+高糖组),检测各组内皮细胞一氧化氮(NO)和可溶性细胞间粘附分子-1(sICAM-1)及细胞凋亡率,用激光共聚焦显微镜及Westernblotting杂交分别观察PKCα、PKCδ的位置变化及蛋白表达。结果高糖可诱导EC凋亡增加,NO浓度降低,sICAM-1水平增加;PIO可抑制高糖诱导的上述变化;PIO可抑制高糖诱导的HUVECs的PKCα由胞浆向胞核转移,并抑制PKCδ在高糖作用下由胞核向胞浆及胞膜转移;PIO可抑制高糖诱导的HUVECsPKCδ表达增强的作用,而高糖作用对HUVECs的PKCα表达影响不明显。结论PIO可纠正高糖诱导的EC功能异常,其保护作用可能部分是通过抑制PKCα及PKCδ的激活来实现。

Objective To determine whether pioglitazone (PIO) has effects on human umbilical vein endothelial cells (HUVECs) in vitro and to clarify the possible mechanisms therein involved. Methods The HUVECs were selected to be a model, which was cultured with 5.5 mmol/L glucose (control group), 30 mmol/L glucose (high glucose group), and 30 mmol/L glucose+PIO at 10^(-9),10^(-7),10^(-5) mol/L respectively (PIO+high glucose groups). The effects of the drugs on the endothelial dysfunction induced by high glucose were studied. The role of protein kinase C (PKC) α and δ in the endothelial dysfunction induced by high glucose and the effects of PIO were assessed. The translocation of PKCδ or PKCα in a single HUVEC was observed by Laser-Scanning Confocal Microscope, and the expression analysis was conducted quantificationally by Western blotting. Results High glucose could induce HUVECs apoptosis; the concentration of NO reduced and the level of sICAM-1 increased in high glucose group. PIO could inhibit the increasing apoptosis peaks induced by high glucose and could reverse the concentration of NO and sICAM-1 to normal level. PIO could inhibit the translocation of PKCα from plasm to nucleus in HUVECs induced by high glucose; it also could inhibit the translocation of PKCδ from nucleus to plasm and membrane in HUVECs. PIO could inhibit the increasing expression of PKCδ in HUVECs induced by high glucose. The expression level of PKCα in the high glucose group was not significantly different from that in the control. Conclusion PIO could correct the endothelial cell dysfunction induced by high glucose, and this drug action of PIO may be effected via the inhibition of PKCα and PKCδ.