MnTMPyP通过抑制线粒体凋亡通路对百草枯诱导肺上皮细胞损伤的保护作用

MnTMPyP protects paraquat- induced lung epithelial -like cell injury through inhibition ofoxidative stress and mitochondrial apoptosis pathway

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RPMI1640培养液；②Mn组：加入Mn10μmol／L；③PQ组：加入PQ750μmol／L；④PQ＋Mn组：Mn10μmol／L预处理90min，再加入PQ750μmol／L。各组细胞处理24h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，其机制是和拈抗PQ引起的氧化应激诱导的线粒体凋亡途径有关。

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 MTY 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, themitochondrial 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.