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目的:探讨腺病毒介导的Runx3基因(Ad-Runx3)与化疗药物顺铂(DDP)联合应用对肝癌细胞HLE增殖的抑制效果。方法:检测肝癌细胞HLE中Runx3基因启动子区域甲基化状态及表达情况。通过Ad-Runx3感染肝癌细胞HLE,利用RT-PCR方法检测Runx3基因在肝癌细胞中的转录,用Ad-Runx3联合化疗药物顺铂处理培养的肝癌细胞HLE,利用Western-blot法检测RUNX3在肝癌细胞中高效表达,MTT法检测细胞增殖抑制率,流式细胞术检测细胞周期和凋亡率。结果:在肝癌细胞HLE中Runx3基因启动子区域存在甲基化异常,RT-PCR结果证实HLE中Runx3基因不表达;Ad-Runx3感染HLE细胞后,RT-PCR结果显示有目的基因的转录,并在感染48 h后,RUNX3高效表达。MTT结果显示,100 MOI Ad-Runx3与6.25 mg/L DDP联合应用后5 d,HLE细胞增殖抑制率达(92.46±1.13)%,显著高于单用Ad-Runx3组的(59.28±1.37)%和DDP组的(46.37±2.51)%(均P<0.05)。流式细胞术结果显示,Ad-Runx3与DDP联合应用明显导致细胞S期减少,G2/M期阻滞;Ad-Runx3联合DDP组细胞凋亡率为(18.62±2.48)%,显著高于单用Ad-Runx3组的(8.66±0.78)%和DDP组的(7.48±0.32)%(均P<0.05).结论:Ad-Runx3与DDP联合应用能显著提高对肝癌细胞HLE增殖的抑制作用。
Objective: To investigate the inhibitory effect of adenovirus-mediated Runx3 gene (Ad-Runx3) combined with cisplatin (DDP) on hepatocellular carcinoma cell HLE proliferation. Methods: The methylation status and expression of Runx3 promoter region in HLE cells were detected. The expression of RUNX3 in HCC cells was detected by RT-PCR and Ad-Runx3 combined with cisplatin. The expression of RUNX3 was detected by Western-blot in hepatoma cells The cell proliferation rate was detected by MTT assay and the cell cycle and apoptosis rate by flow cytometry. Results: The promoter region of Runx3 gene was abnormal in HLE cells. Runx3 gene was not expressed in HLE cells by RT-PCR. After transfection of HLE cells with Ad-Runx3, RT-PCR results showed that the gene was transcribed After 48 h of infection, RUNX3 was highly expressed. The results of MTT showed that the inhibition rate of HLE cell proliferation was (92.46 ± 1.13)% at 5 days after combination of 100 MOI Ad-Runx3 and 6.25 mg / L DDP, significantly higher than that of Ad-Runx3 alone (59.28 ± 1.37)% And (46.37 ± 2.51)% in DDP group (all P <0.05). The results of flow cytometry showed that the combination of Ad-Runx3 and DDP significantly reduced the S phase and G2 / M phase. The apoptosis rate of Ad-Runx3 combined with DDP group was (18.62 ± 2.48)%, which was significantly higher than that of single (8.66 ± 0.78)% in Ad-Runx3 group and (7.48 ± 0.32)% in DDP group (all P <0.05) .Conclusion: Combination of Ad-Runx3 and DDP can significantly improve the inhibition of HLE proliferation in hepatocellular carcinoma cells.