【摘要】 目的 探讨MnTMPyP(Mn)通过抑制氧化应激和线粒体凋亡通路对百草枯(PQ)诱导肺上皮细胞损伤的保护作用。方法 体外培养人肺泡Ⅱ型上皮样细胞A549细胞,以超氧化物歧化酶(SOD)的模拟物Mn为干预因素。实验分为四组:①对照组:加入RPMI 1640培养液;②Mn组:加入Mn 10μmol/L;③PQ组:加入PQ 750μmol/L;④PQ+Mn组:Mn 10μmol/L预处理90min,再加入PQ 750μmol/L。各组细胞处理24 h后检测相关指标。MTT法检测细胞活性;流式细胞术检测细胞凋亡率、线粒体膜电位、活性氧(ROS);分光光度法检测Caspase-3;Western blotting检测线粒体凋亡蛋白Bcl-2、Bax蛋白的表达。结果 与对照组比较,Mn组检测指标无统计学意义;PQ组细胞活性降低,细胞凋亡率增加有统计学意义(P <0. 01),线粒体膜电位降低有统计学意义(P<0.01),ROS的产生显著增加,Caspase-3活性增高,Western blotting显示线粒体凋亡途径中抗凋亡蛋白Bcl-2表达量降低,促凋亡蛋白Bax表达增强。与PQ组比较,PQ+Mn组细胞活性增加,细胞凋亡率降低,线粒体膜电位增强,ROS的产生减少,Caspase-3活性降低,Western blotting显示Bcl-2蛋白表达量增强,Bax蛋白表达降低。结论 Mn有效减轻PQ诱导肺上皮细胞的损伤,其机制是和拮抗PQ引起的氧化应激诱导的线粒体凋亡途径有关。更多还原

【Abstract】 Objective To investigate the protective effect and mechanisms of MnTMPyP( Mn)on paraquat( PQ)-induced lung alveolar epithelial-like cell injury. Methods Lung alveolar epithelial-like cells( A549) were cultured in vitro. Superoxide dismutase mimic Mn was an intervention factor. The experiments were divided into four groups: control group: RPMI 1640 medium was added; Mn group: Mn 10 μmol/L; PQ group: PQ 750 μmol/L; PQ + Mn group: Mn 10 μmol/L was pretreated for 90 min, and then add PQ 750 μmol/L. After each group of cells was treated for 24 h,relevant indicators were detected; the cell survival rate was assessed by MTT assay. Cellular apoptosis,mitochondrial transmembrane potential, ROS were assayed by flow cytometry. The activities of Caspase-3 were assayed by spectrophotometry. Western blotting was used to analyze the expression of proteins, such as Bcl-2, Bax. Results Compared with the control group, there was no statistically significant change in Mn group; in PQ group, the cell viability was decreased, the apoptosis rate was increased, the mitochondrial membrane potential was reduced, the production of ROS was increased, Caspase-3 activity was increased. Western blotting showed that the expression of anti-apoptotic protein Bcl-2 decreased and the expression of pro-apoptotic protein Bax increased in mitochondrial apoptotic pathway;Compared with PQ group, in the Mn +PQ group, the cell viability increased, apoptosis rate decreased,mitochondrial membrane potential increased, production of ROS decreased, Caspase-3 activity decreased. Western blotting showed that the expression of Bcl-2 protein was increased, the expression of Bax protein was decreased. Conclusion MnTMPyP can effectively reduce PQ-induced lung epithelial-like cell injury, and its mechanism is related to antagonism of PQ-induced mitochondrial apoptosis pathway.更多还原