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目的探讨对氨基水杨酸钠(PAS-Na)对铅诱导PC12细胞凋亡的影响。方法 PC12细胞培养至其对数生长期后,正常对照组和正常+PAS-Na 500μmol·L~(-1)组细胞于正常培养液中培养24 h后弃培养液,前者加入正常培养液、后者加入PAS-Na继续培养24 h。醋酸铅模型组及醋酸铅+PAS-Na 20,100和500μmol·L~(-1)组先与醋酸铅(终浓度10μmol·L~(-1))共培养24 h后弃培养液,再加入相应浓度PAS-Na继续培养24 h。用MTT法测定细胞存活率,试剂盒方法测定细胞内还原型谷胱甘肽(GSH)含量,Hoechst33342荧光染色检测凋亡率,AnnexinⅤ/PI双染流式细胞术检测PC12细胞早期凋亡率,Western蛋白质印迹法检测Bcl-2、Bax和P53蛋白水平。结果与正常对照组比较,醋酸铅模型组细胞存活率降低(P<0.05),细胞凋亡形态变化典型,细胞早期凋亡率增高(P<0.05),细胞内GSH含量降低(P<0.01),P53蛋白水平升高(P<0.01),Bax/Bcl-2比值升高(P<0.01);正常+PAS-Na 500μmol·L~(-1)组上述指标未见明显变化。与醋酸铅模型组比较,醋酸铅+PAS-Na组细胞存活率增高(P<0.05),细胞凋亡率和早期凋亡率均降低(P<0.05),细胞内GSH含量增高(P<0.01),P53蛋白水平和Bax/Bcl-2比值降低(P<0.05)。结论 PAS-Na对铅致PC12细胞凋亡有一定的拮抗作用,其机制可能与PAS-Na抗脂质过氧化损伤和降低Bax/Bcl-2比值有关。
Objective To investigate the effect of sodium salicylate (PAS-Na) on the apoptosis of PC12 cells induced by lead. Methods PC12 cells were cultured until their logarithmic growth phase. The cells in normal control group and normal + PAS-Na 500μmol·L -1 group were cultured in normal medium for 24 h, then the culture medium was discarded. The former was added into normal culture medium, The latter added PAS-Na continue to train for 24 h. Lead acetate group and lead acetate + PAS-Na 20, 100 and 500μmol·L -1 group were incubated with lead acetate (final concentration 10μmol·L -1) for 24 h, then the culture solution was discarded, and the corresponding concentration of PAS- Na continue to train for 24 h. The cell viability was measured by MTT assay, the content of glutathione (GSH) in cells was determined by kit method, the apoptosis rate was detected by Hoechst33342 fluorescent staining, the apoptosis rate of PC12 cells was detected by AnnexinⅤ / PI double staining flow cytometry, Western blotting was used to detect the protein levels of Bcl-2, Bax and P53. Results Compared with the normal control group, the cell viability of lead acetate group was decreased (P <0.05), the morphological changes of apoptotic cells were typical, the early apoptotic rate was increased (P <0.05) and the content of intracellular GSH was decreased (P <0.01) P53 protein level increased (P <0.01), Bax / Bcl-2 ratio increased (P <0.01); normal + PAS-Na 500μmol·L -1 group had no significant change. Compared with lead acetate group, the cell viability in lead acetate + PAS-Na group increased (P <0.05), the apoptosis rate and early apoptosis rate decreased (P <0.05) and the content of intracellular GSH increased (P <0.01) P53 protein and Bax / Bcl-2 ratio decreased (P <0.05). Conclusion PAS-Na can antagonize the apoptosis of PC12 cells induced by lead, and its mechanism may be related to the anti-lipid peroxidation injury and the decrease of Bax / Bcl-2 ratio of PAS-Na.