JNK信号通路在丙泊酚减轻人脐静脉内皮细胞氧化应激损伤中的作用

Role of JNK signaling pathway in reduction of oxidative stress injury to human umbilical vein endothelial cells by propofol preconditioning

目的探讨不同浓度丙泊酚预处理对人脐静脉内皮细胞(HUVECs)氧化应激损伤的作用,探讨其与c-Jun氨基端激酶(JNK)信号通路的关系。方法体外培养的HUVECs分为八组:对照组(C1组)细胞行常规培养;DMSO对照组(C2组)细胞的培养基中加入DMSO(终浓度0.05%);丙泊酚组(P组)细胞的培养基中加入丙泊酚(终浓度50μmol/L)处理4h;H2O2处理组(H组)细胞的培养基中加入H2O2(终浓度400μmol/L)处理4h;超低浓度丙泊酚组(HP1组)的细胞以1μmol/L丙泊酚预处理HUVECs 30min后加入H2O2400μmol/L处理4h;余在HP1组基础上增加预处理丙泊酚浓度分别为5μmol/L(HP5组)、25μmol/L(HP25组)和50μmol/L(HP50组)。各组细胞培养后,用CCK-8法测定细胞活力,流式细胞仪检测细胞凋亡情况,Western blot法检测HUVECs磷酸化JNK(p-JNK)、Bcl-2及Bax蛋白表达水平。结果与C1组比较,H组和HP1、HP5、HP25、HP50组细胞活力明显降低,细胞凋亡率明显升高,p-JNK蛋白表达以及Bax蛋白表达明显升高,而Bcl-2/Bax比值明显降低(P<0.05);HP1组和HP5组p-JNK/JNK比值明显升高,Bcl-2蛋白表达明显下降(P<0.05)。与H组比较,HP25组和HP50组的细胞活力明显升高,HP5组、HP25组和HP50组细胞凋亡率明显降低,p-JNK/JNK和Bax表达明显下调,Bcl-2和Bcl-2/Bax比值明显升高(P<0.05)。结论 25μmol/L和50μmol/L浓度的丙泊酚预处理可减轻氧化应激对HUVECs的损伤,其机制与抑制JNK信号通路有关。

Objective To evaluate the role of c-Jun N-terminal kinase （JNK） signaling in the protective effect of different concentrations of propofol preconditioning against oxidative stress injury in human umbilical vein endothelial cells（HUVECs）. Methods Cultured HUVECs in vitro were divided into eight groups （n=5） as follows., control group （group C1, cells cultured with no further treatment）, DMSO group （group C2, cells treated with 0.05% DMSO）, propofol group（group P, cells with propofol 50umol/L for 4 h）, H2O2 group（group H, H2O2 400umol/L for 4 h）, H2O2 ＋ propofol 1 umol/L preconditioning group（group HP1, propofol 1 umol/L preconditiong for 30 min followed by H2O2 400 umol/L for 4 h）, H2 O2 ＋ propofol 5, 25, and 50 umol/L preconditioning group（group HP5, HP25, and HP50, corresponding concentration of propofol preconditiong for 30 min followed by H2O2 400 umol/L for 4 h）. Cell viability was measured by CCK-8, cell apoptosis was assessed by flow cytometry, and levels of JNK phosphorylation（p-JNK）, Bcl-2, and Pax in HU- VECs were determined by Western blot. Results Compared with group C1, cells in group H and groups HP1,HPS, HP25, HP50 had a lower viability, an increased apoptosis rate, a higher p-JNK and Bax, as well as a decreased Bcl-2/Bax ratio （P〈0. 05） ; cells in groups HP1 and HP5 had an in- creased p-JNK/JNK ratio, 13cl-2 was down-regulated （P〈0. 05）. Compared with group H, the cell viability was significantly increased in group HP25 and group HP50 （P〈0.05）, meanwhile, the apoptosis rate decreased, the p-JNK/JNK ratio and Bax down-regulated, and Bcl-2/Bax ratio and Bcl-2 increased significantly in group HPS, HP25 and HP50 （P〈0.05）. Conclusion Propofol 25 umol/L or 50 umol/L preconditioning may protect aganist the oxidative stress injury in HUVECs with underlying possible mechanism may be related to the JNK signaling pathway inhibition.