阿霉素对肝癌细胞核因子κB活化及细胞周期基因D_1表达的影响

Effect of adriamycin on activation of nuclear factor-κB and gene expression of cyclin D_1 in BEL-7402 cell

目的探讨阿霉素诱导肝癌细胞凋亡过程中的表达及其对细胞周期的影响。方法体外培养人肝癌细胞株BEL-7402,应用流式细胞术检测阿霉素对BEL-7402细胞周期和凋亡的影响;细胞免疫组织化学检测不同浓度阿霉素处理BEL-7402细胞后核因子κB(NF-κB)p65亚基入核情况;反转录聚合酶链反应(RT-PCR)方法检测细胞周期基因D1(cyclin D1)表达的差异。结果阿霉素诱导BEL-7402细胞凋亡具有时效和量效关系,1、10、100 mg/L作用24小时后,其诱导细胞凋亡率分别为(13.80±1.20)%、(23.73±3.01)%和(30.02±3.85)%,与对照组(5.20±0.98)%相比差异有统计学意义(P<0.05);随着阿霉素用药浓度的增加,S期细胞比例和细胞增殖指数增加,NF-κBp65被激活并入核,其下游cyclin D1基因的表达逐渐增强。结论阿霉素诱导肝癌BEL-7402细胞凋亡的同时可激活NF-κB;其下游基因cyclin D1过度表达可能是促进细胞增殖、对抗细胞凋亡的重要因素之一。

Objective To detect the effect of adriamycin（ADR） on expressions of cyclin D1 ,nuclear factor kappa B（NF-κB） and their effects on apoptosis in BEL-7402 cell. Methods The BEL-7402 cell of human liver cancer used in the experiment was cultured in vitro,flow cytometry （FCM） was used to examine the effect of ADR on the apoptosis and cell cycle of BEL-7402 cell. The mRNA expression of cyclin D1 gene was determined by reverse transcriptase polymerase chain reaction（RT-PCR）in BEL-7402 cell treated with different concentrations of ADR for 24 hours,NF-κB activity of BEL-7402 was measured by immunohistochemistry. Results ADR could induce apoptosis in a concentrationdependent manner, ADR of 1,10,100 rag/L induced apoptotic rate of （ 13.80 ± 1.20） %, （23.73 ± 3.01） % and （30.02 ± 3.85） %, respectively; the percentage of S phase cells and proliferation index（H） increased with the concentration of adriamycin, cyclin D1 mRNA expression and NF-JB activity were also enhanced. Conclusion ADR can induce apoptosis in BEL-7402 cell of liver cancer; treatment with ADR for 24 horus causes activation of NF-kappa B; overexpression of cyclin D1 mRNA may be an important factor that promotes cell proliferation and takes part in the process of anti-apoptosis.