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目的：本文旨在研究bcl-2基因家族促凋亡蛋白Bak在肝癌细胞凋亡中的作用，同时评价它作为肿瘤治疗的潜在靶基因的可能性。方法：免疫组织化学法研究Bak在HCC组织中的分布，并结合TUNEL染色，分析Bak在肝癌细胞凋亡中潜在的作用。采用MTⅡ调节系统，通过加锌离子(ZnSO4，100μmol/L）诱导Bak基因表达。HCC-9204肝癌细胞系作为靶细胞，获得表达Bak基因的稳定转染子。结果：在HCC组织中，Bak抗原的表达主要位于肝（癌）细胞的胞浆，呈细颗粒状均匀分布，在癌细胞中偶有核内表达。TUNELLI的分析结果显示，低TUNELLI组Bak表达的细胞阳性率显著低于高TUNELLI组的细胞阳性率(P<0.01)。可诱导性Bak基因过表达的HCC9204细胞显示广泛的细胞死亡。TUNEL染色证实细胞核的碎片化，Annexin-V实验证实细胞膜不对称丧失，从而提示这种细胞死亡是凋亡。流式细胞仪显示在诱导后24h，有19.29％的细胞发生凋亡。结论：肝癌细胞的凋亡与bcl2家族促凋亡成员Bak抗原的表达有关；Bak基因显著诱导HCC－9204细胞凋亡。因此Bak基因可能成为肿瘤基因治疗的靶基因。
OBJECTIVE: To investigate the role of bcl-2 gene family pro-apoptotic protein Bak in apoptosis of hepatoma cells and to evaluate its potential as a potential target gene for tumor therapy. METHODS: Immunohistochemistry was used to study the distribution of Bak in HCC tissues and TUNEL staining was used to analyze the potential role of Bak in apoptosis of HCC cells. Using the MTII regulatory system, Bak gene expression was induced by adding zinc ions (ZnSO4, 100 μmol/L). The HCC-9204 hepatoma cell line serves as a target cell to obtain stable transfectants expressing the Bak gene. RESULTS: In HCC tissues, the expression of Bak antigen was mainly localized in the cytoplasm of liver (cancer) cells and distributed evenly in fine particles. Occasionally, the expression of Bak antigen was expressed in cancer cells. TUNELLI analysis showed that the positive rate of Bak expression in the low TUNELLI group was significantly lower than that in the high TUNELLI group (P<0.01). Inducible Bak gene overexpressed HCC9204 cells exhibit extensive cell death. TUNEL staining confirmed the fragmentation of the nucleus, Annexin-V experiments confirmed the loss of cell membrane asymmetry, suggesting that this cell death is apoptosis. Flow cytometry showed that 19.29% of cells developed apoptosis 24 h after induction. CONCLUSION: The apoptosis of hepatocellular carcinoma cells is related to the expression of Bak2 family-promoting Bak antigen. Bak gene significantly induces apoptosis of HCC-9204 cells. Therefore, the Bak gene may become a target gene for tumor gene therapy.