C/EBP-α基因诱导肝细胞和肝星状细胞自噬和凋亡的差异研究

Research on the difference of C/EBP-α gene on hepatocyte and hepatic stellate cells in the autoph-agy and apoptosis

目的：通过体外研究，探讨 CCAAT 增强子结合蛋白-α（C/ EBP-α）诱导的肝细胞和肝星状细胞（HSCs）自噬和凋亡的差异。方法构建腺病毒介导的 C/ EBP-α基因（Ad-C/ EBP-α），分别以50、100、200 MOI 感染大鼠 BRL-3A 细胞系和 HSC-T6细胞系，采用流式细胞仪检测细胞凋亡情况；应用 Western blot 方法检测自噬标志蛋白 LC3Ⅰ/ LC3Ⅱ的表达，探讨 C/ EBP-α诱导2种细胞的自噬及凋亡途径有无差异。结果流式细胞仪显示HSC-T6细胞系从50 MOI 时即出现显著凋亡，且随剂量上升，凋亡率进一步升高，上升趋势差异有统计学意义（P〈0.05）；BRL-3A 细胞系并无明显变化。Western blot 显示 HSC-T6LC3Ⅰ/ LC3Ⅱ从50 MOI 时水平明显下降，且剂量越高，下降幅度越大，下降趋势差异有统计学意义（P〈0.05）；BRL-3A 细胞系并无明显变化。结论 C/ EBP-α基因对肝星状细胞的自噬有剂量依赖的抑制作用，对其凋亡有剂量依赖的促进作用；但对肝细胞的自噬及凋亡无明显影响。

Objective To investigate the difference of the autophagy and apoptosis hepatocyte and hepatic stellate cells inducted by CCAAT enhancer binding protein -α（C / EBP-α）by vitro study. Methods The adenovirus mediated C / EBP-α gene（Ad-C / EBP-α）were constructed,the BRL-3A cell lines and HSC-T6 cell lines were infected with 50,100,200 MOI,and the cell apoptosis situation was de-tected by the flow cytometry. The expression of autophagy marker protein LC3Ⅰ/ LC3Ⅱ were detected by＆amp;nbsp;application of Western blot method. Then,the differences in the autophagy and apoptosis pathways of the 2 cells induced by C / EBP-α were investigated. Results Flow cytometry showed that HSC-T6 cell line showed significant apoptosis in 50 MOI cell line,and with the dose increased,the apoptosis rate was fur-ther increased,the upward trend was significant（P〈 0. 05）. There was no significant change in BRL-3A cell line. Western blot showed that the HSC-T6LC3Ⅰ/ LC3Ⅱ from the 50 MOI level was significantly decreased,and the dose was higher,the decline was greater,the downward trend was significant（P〈0. 05）. There was no significant change in BRL-3A cell line. Conclusions The C / EBP-α gene has a dose-dependent inhibitory effect on the autophagy of hepatic stellate cells,which has a dose-dependent effect on apoptosis,but it has no significant effect on the autophagy and apoptosis of hepatocytes.