摘要
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。
Objectives： We are currently investigating the role of Bak in apoptotic pathway and are evaluating its potential as a target for therapeutic intervention in hepatocellular carcinoma (HCC). Methods： Liver biopsy samples from 50 cases HCC including peritumorous tissues were immunohistostained by using ABC method. TUNEL assay was also perform to evaluate apoptotic rate in HCC. We used an inducible system, MT Ⅱ regulatory system, which allowed controlled expression of Bak protein upon addition of ZnSO4(100 μ mol/L) as external inducer. Rapidly growing HCC 9204 cell line was chosen as a target and stable transfected with inducible expression vector containing Bak gene. Results： Bak antigen was found mainly in the cytoplasm of the cancer or noncancer cells in the tissues of HCC. The specific signals were occasionally observed in the nuclei of cancer cells. The prevalence of Bak antigen expression was closely correlated with TUNEL LI in HCC. HCC 9204 cells overexpressing Bak showed extensive cell death with nucleus fragmentation detected by TUNEL assay and loss of asymmetry by Annexin V assay. FACS analyses showed Bak induced apoptosis in 19.29 ％ cells 24 h after induction. Conclusions： Apoptosis in the HCC tissues correlates with Bak expression. Bak may lead to apoptosis in HCC 9204 cells. Therefore, Bak may be a good candidate for cancer gene therapy.
出处
《癌症》
SCIE
CAS
CSCD
北大核心
2000年第9期891-895,共5页
Chinese Journal of Cancer
