去泛素化酶USP14调控H_2O_2诱导的H9c2细胞氧化应激损伤

USP14 regulates H_2O_2 induced oxidative stress in H9c2 cells

目的:观察抑制泛素特异性蛋白酶14(ubiquitin-specific protease 14,USP14)的活性对H_2O_2诱导的H9c2细胞氧化应激损伤的影响。方法:体外培养H9c2心肌细胞,用H_2O_2(25μmol/L)处理2 h,建立心肌细胞氧化应激损伤模型。将细胞分为对照(control,CON)组、模型组(H_2O_2组)、USP14抑制剂IU1处理组(IU1组)和IU1处理后建模组(IU1+H_2O_2组)。MTS法检测H9c2心肌细胞活力;流式细胞术检测细胞内活性氧簇(ROS)的产生和细胞存活率;Western blot法检测丝裂原活化蛋白激酶家族相关蛋白的表达变化。结果:与CON组相比,H_2O_2组细胞活力、细胞存活率显著降低,细胞内ROS含量、Bax/Bcl-2、P53、p-ERK1/2、p-JNK和p-P38的蛋白水平显著增加(P<0.05);与H_2O_2组比较,IU1+H_2O_2组细胞活力、细胞存活率显著增加,细胞内ROS含量、Bax/Bcl-2、P53、p-ERK1/2、p-JNK和p-P38蛋白水平显著降低(P<0.05)。结论:抑制USP14活性可以减轻氧化应激条件下H9c2心肌细胞的损伤,其机制可能与抑制丝裂原活化蛋白激酶信号活化和下调凋亡相关蛋白有关。

AIM： To evaluate the effect of inhibiting ubiquitin-specific protease 14 （USP14） activity on oxida-tive stress induced by H2O2 of H9c2 cells. METHODS： The H9c2 cells were incubated with H2O2 at 25 jjimol/L for 2 h to establish the oxidative stress injury model. The cells were divided into control group, H202 group, IU1 group （25 jjimol/ L or 50 jjimol/L） and IU1 ＋ H202 group. The H9c2 cells activity was measured by MTS assay. The level of intracellular reactive oxygen species （ROS） and cell survival rate were analyzed by flow cytometry assay. The changes of the mitogen- activated protein kinase （ MAPK） family related proteins were detected by Western blot. RESULTS ： Compared with con-trol group, the cell activity and the viability rate in H202 group were decreased （P 〈0. 05） , while the intracellular ROS, the protein levels of Bax/Bcl-2, P53, p-ERKl/2, p-JNK and p-P38 were increased （ P 〈 0 . 0 5 ） . Compared with H2O2 group, the cell activity and the viability rate of the H9c2 cells in IU1 ＋ H202 group were increased （ P 〈 0. 05 ） , while the intracellular ROS, the protein levels of Bas/Bcl-2, P53, p-ERKl/2, p-JNK and p-P38 were decreased （ P 〈 0. 05 ） . CONCLUSION： Inhibition of USP14 activity reduces the oxidative stress injury of the H9c2 cells. The mechanism may be related to inhibition of the MAPK signaling and down-regulation of apoptosis related proteins.