Caveolin-1在脂多糖致大鼠肺微血管内皮细胞损伤中作用的初探

Study on role of caveolin-1 in lipopolysaccharide-induced rat pulmonary microvascular endothelial cell injury

目的观察脂多糖（lipopolysaccharide，LPS）对大鼠肺微血管内皮细胞（rat pulmonary microvascular endothelial cell，RPMVEC）小窝蛋白-1（caveolin-1，Cav-1）表达的影响。方法体外培养RPMVEC，分别采用WI、PCR、原位杂交及免疫荧光检测Cav-1mRNA及蛋白的表达，原位杂交检测LPS和蛋白激酶C（PKC）抑制剂双吲哚基顺丁烯二酰亚胺（bis—indolylmaleimide，BIM）对Cav-ltuRNA表达的影响。结果RT—PCR和原位杂交同步检测出RPMVEC表达Cav-1基因；免疫荧光检测出RPMVEc表达Cav-1蛋白；LPS以浓度依赖方式诱导Cav-1mRNA表达增加：0．1、1、10mg／L LPS分别孵育RPMVEC6h后，Cav-1mRNA表达量随其浓度增加逐渐升高，与正常组比较差异均有统计学意义（P〈0．05）；10mg／L LPS刺激RPMVEC不同时间（1h、2h、3h、6h）后，RPMVEC中Cav-1mRNA表达水平呈动态变化：1h开始上升，2h达峰值，之后逐渐下降，但至6h仍高于正常水平（P〈O．05）；10μmol／LBIM预孵育RPMVEClh后，可显著下调10mg／L LPS对Cav-1mRNA表达的诱导效应（P〈0．05）。结论RPMVEC表达Cav-1，LPS上调RPMVECCav-1基因转录，PKC信号通路参与LPS对RPMVECCav-1基因转录的调控。

Objective To observe the effect of lipopolysaccharide （LPS） on the production of Caveolin-1 （Car-1） mRNA in rat pulmonary microvaseular endothelial cells （RPMVEC）. Methods RPMVEC were isolated and cultured in vitro, then RT-PCR, in situ hybridization and immunofluorescenee were used to observe Cav-1 expression in RPMVEC. Cellular in situ hybridization was performed to detect Caw1 mRNA expression under various stimulations. Results We observed that LPS challenge of RPMVEC induced concentration-dependent manner （0.1,1,10 rag/L） increases in expression of Car-1 mRNA. After challenge of RPMVEC with 10 mg/L LPS, the expression of Car-1 mRNA rose at 1 h, peaked at 2 h, then decreased gradually, but still higher than the control level at 6 h （ P （0.05）. Pretreated RPMVEC with protein kinase C （PKC） inhibitor bis-indolylmaleimide （BIM） significantly reduced the expression of Car-1 mRNA which induced by LPS （ P （0.05）. Conclusions These results indicate that there exit Cav-1 in RPMVEC. LPS can up-regulate the level of RPMVEC Cav-1 gene transcription and the PKC signaling pathway involved in modulation of this intracellular signal process.