Bcl-2家族蛋白在Ad.mda-7介导HepG2细胞凋亡中的变化

The change of Bcl-2 protein family by the Ad.mda-7-induced apoptosis in HepG2 cells

目的观察Bcl-2家族蛋白在Ad.mda-7介导HepG2细胞凋亡中的变化,初步探讨Ad.mda-7介导肝癌细胞的凋亡机制。方法将携带人MDA-7/IL-24基因的腺病毒转染HepG2和L02,通过四甲基偶氮哩蓝染色法(MTT)及Hoechst染色观察MDA-7/IL-24对肝癌细胞HepG2和正常肝胚细胞L02的生长抑制和杀伤作用。Annexin V和PI双染后流式细胞仪检测细胞的凋亡。蛋白免疫印迹法(Western blot)检测Bcl-2家族蛋白的变化。结果Ad.mda-7能明显抑制肝癌细胞HepG2的生长,但对正常肝胚细胞L02没有明显抑制增殖作用。Hoechst染色和流式细胞仪提示Ad.mda-7能明显诱导肝癌细胞HepG2细胞的凋亡,但对正常肝胚细胞L02没有明显毒副作用。Western blot说明抑制凋亡的蛋白Bcl-2和Bcl-xl的表达在HepG2明显下降,在L02中则没有变化;而促凋亡蛋白Bax在HepG2和L02都升高。结论Ad.mda-7转染HepG2后,促凋亡(Bax)与抑凋亡蛋白(Bcl-2和Bcl-xl)的比率发生改变,进而发生凋亡。

Objective To study the relation between the Bcl- 2 protein family and the apoptosis of HepG2 induced by Ad. mda - 7. Methods The MDA - 7/IL - 24 gene was transfected into HepG2 and L02 cells with a replication - incompetent adenovirus vector. MTT assay and Hoechst staining were used to observe the growth inhibitory and killing effects of MDA - 7/IL - 24 on HepG2 and L02. After double staining with Annexin - V and PI,the apoptosis was detected by using flow cytometry. The changes of Bcl - 2 protein family were observed by Western blot. Results MTT revealed that Ad. mda - 7 could suppress the grwoth of HepG2 cells, but could not inhibit the proliferation of L02 cells. Hoechst stai- ning and flow cytometry indicated Ad. mda - 7 could obviously induce the apoptosis of HepG2, but had no cytotoxic effects on L02 cells. Western blot showed the expression of protein Bcl - 2 and Bcl - xl was significantly decreased in HepG2 cells after infected with Ad. mda -7 ,but not in L02 cells ,while the expres- sion of Bax protein was increased in both HepG2 and L02. Conclusion The apoptosis of HepG2 induced by Ad. mda-7 had something to do with the change in the ratio of pro - （Bax） to antiapoptoptic （ Bcl-2 and Bcl - xl） proteins.