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目的研究不同剂量的水飞蓟宾(silibinin)对锰致人神经母细胞瘤SH-SY5Y细胞毒性的对抗作用及可能机制。方法体外培养SH-SY5Y细胞,加入不同浓度的水飞蓟宾(5~20μmol/L),继续培养24 h,吸出后加入MnCl2(终浓度为0.6 mmol/L),MTT法检测细胞活力,Western blot检测自噬标志蛋白LC3和Beclin-1的表达和活化情况,DCFH-DA染色检测活性氧(ROS)含量,同时检测水飞蓟宾的体内外抗氧化活性。结果 MTT结果显示(5~20μmol/L)的水飞蓟宾对MnCl2诱导的细胞损伤有显著的拮抗作用。Western blot结果显示水飞蓟宾能剂量依赖性地抑制MnCl2处理诱导的自噬性损伤的发生。与对照组相比,0.6 mmol/L的MnCl2处理组ROS含量明显升高,而水飞蓟宾给药则能显著抑制ROS的产生,同时体内外的抗氧化能力检测显示水飞蓟宾抗具有较强的总抗氧化能力和抗超氧阴离子自由基活力,能显著恢复细胞内GSH和SOD的活力。结论水飞蓟宾能剂量依赖性地对抗MnCl2诱导的SH-SY5Y细胞损伤,这一作用可能是通过清除细胞内ROS继而抑制自噬性死亡的发生来实现的。
Objective To investigate the antagonistic effect of different doses of silibinin on the cytotoxicity of SH-SY5Y induced by manganese and its possible mechanism. Methods SH-SY5Y cells were cultured in vitro. Silybin at different concentrations (5 ~ 20μmol / L) was added to culture SH-SY5Y cells for 24 h. After adding MnCl2 (final concentration 0.6 mmol / L), the cell viability was measured by MTT assay blot was used to detect the expression and activation of autophagy markers LC3 and Beclin-1. DCFH-DA staining was used to detect the content of reactive oxygen species (ROS) and the anti-oxidative activity of silybin in vitro and in vivo. Results The results of MTT showed that silibinin (5 ~ 20μmol / L) had a significant antagonistic effect on MnCl2-induced cell injury. Western blot results showed that silybin inhibited the occurrence of autophagic injury induced by MnCl2 in a dose-dependent manner. Compared with the control group, the ROS content in 0.6 mmol / L MnCl2 treatment group was significantly increased, while silibinin administration could significantly inhibit the production of ROS, at the same time in vitro and in vivo antioxidant capacity test showed silibinin anti Strong total antioxidant capacity and superoxide anion free radical activity, can significantly restore intracellular GSH and SOD activity. Conclusion Silybin can antagonize MnCl2-induced injury of SH-SY5Y cells in a dose-dependent manner. This effect may be achieved through the elimination of intracellular ROS and then the inhibition of autophagic death.