梓醇调控自噬和凋亡促进H_2O_2诱导的PC12细胞存活

Catalpol Promotes H_2O_2-Injured PC12 Cell Survival by Regulating Autophagy and Apoptosis

探讨梓醇对过氧化氢(H_2O_2)诱导的PC12细胞氧化损伤后调控凋亡和自噬相关分子的机制,采用0.01 mmol,0.1 mmol,1 mmol的梓醇预处理PC12细胞2h后,再用250μmol/L H_2O_2损伤细胞12h.MTT法检测细胞存活率,设置梓醇低、中、高3个剂量组(0.01 mmol,0.1 mmol,1 mmol),自噬抑制剂3MA阳性对照组,阴性对照组,H_2O_2诱导的PC12细胞损伤模型组,采用荧光单标检测LC3Ⅱ的表达,Western blot检测凋亡和自噬相关蛋白Bcl2/Bax,LC3Ⅱ/Ⅰ和Cleaved-caspase 3的表达情况.结果显示,梓醇呈剂量依赖性地提高了H_2O_2诱导的PC12细胞的存活,并对H_2O_2诱导的PC12细胞的LC3Ⅱ/Ⅰ,Bcl2/Bax升高蛋白水平表达,降低了Cleaved-caspase 3的表达;随着梓醇浓度的增加,Cleaved-caspase 3的表达减少,而LC3Ⅱ/Ⅰ和Bcl2/Bax的表达在升高.得出梓醇通过恢复凋亡和自噬相关蛋白Bcl2/Bax,LC3Ⅱ/Ⅰ的平衡能够保护PC12细胞免于H_2O_2诱导的氧化损伤.

This study aimed to explore the molecular mechanism for the regulation of autophagy or apoptosis by catalpol after the H2O2-induced oxidative injure in PC12 cells.PC12 cells were pretreated with catalpol at different concentrations（0.01,0.1 and 1 mmol）,and then the viability of H2O2-injured PC12 cells was measured by MTT assay.There were four groups：catalpol of different concentrations（0.01,0.1 or1 mmol）,autophagy inhibitor 3-MA positive control,negative control and H2O2-injured PC12 cell model.Fluorescent single staining was used to detect the expression of LC3 II,and Western blot to detect the expression of autophagy-related proteins LC3Ⅱ/Ⅰ,Bcl2/BAX and cleaved-caspase 3.Compared with the model group,catalpol dose-dependently promoted the survival of H2O2-injured PC12 cells,increased the expression of LC3Ⅱ/Ⅰ and Bcl2/BAX,and reduced the expression of cleaved-caspase 3.Interestingly,as catalpol concentration increased,the reduction degree of cleaved-caspase 3 reduced,while LC3Ⅱ/Ⅰ and Bcl2/BAX increased.Catalpol can protect H2O2-injured PC12 cells and promote their survival through restoring the balance between Bcl2/Bax and LC3Ⅱ/Ⅰ-the two genes related to autophagy and apoptosis.