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目的:探讨喉癌中APAF-1基因的表达及其对5-氮杂-2′-脱氧胞苷(5-az-a-2′-deoxycitydine,5-Aza-CdR)诱导的Hep2细胞系凋亡的影响。方法:不同浓度5-Aza-CdR(5×10-8、10-7、2×10-7和3×10-7mol/L)处理体外培养的Hep2细胞后,用流式细胞仪检测处理72h的细胞周期分布和细胞凋亡率变化;通过RT-PCR及免疫组织化学技术检测APAF-1基因在喉癌中的表达情况,并建立APAF-1瞬时过度表达系统,观察在APAF-1过度表达情况下,5-Aza-CdR对Hep2细胞凋亡的影响。结果:10-7mol/L5-Aza-CdR处理Hep2细胞72h后,凋亡率由对照组的1·07%增加到13·96%,且发生G1期细胞周期阻滞;喉鳞癌组织中APAF-1mRNA和蛋白表达都明显下调,分别为16/42、14/31,且未发现APAF-1表达与肿瘤的分化程度具有相关性,P=0·093;在10-7mol/L浓度5-Aza-CdR诱导下,APAF-1过度表达组Hep2细胞系凋亡率由空白对照组的12·46%及空载体转染组的14·74%增高到28·64%,且发生S期细胞周期阻滞。结论:过度表达APAF-1基因可以明显增加Hep2细胞系对去甲基化药物5-Aza-CdR诱导的凋亡敏感性。
Objective: To investigate the expression of APAF-1 gene in laryngeal carcinoma and its effect on the Hep2 cell line induced by 5-aza- a-2’-deoxycitydine (5-Aza-CdR) The impact of death. Methods: Hep2 cells cultured in vitro were treated with different concentrations of 5-Aza-CdR (5 × 10-8, 10-7, 2 × 10-7 and 3 × 10-7mol / L) for 72 h by flow cytometry APAF-1 gene in laryngeal squamous cell carcinoma was detected by RT-PCR and immunohistochemistry, and transient APAF-1 overexpression system was established to observe the effect of APAF-1 overexpression Effects of 5-Aza-CdR on Hep2 Cell Apoptosis. Results: After treated with 10-7mol / L 5-Aza-CdR for 72h, the apoptotic rate increased from 1. 07% to 13 · 96% in control group, and cell cycle arrest occurred in G1 phase. APAF 1 mRNA and protein expression were significantly down-regulated, respectively, 16 / 42,14 / 31, and found no correlation between APAF-1 expression and tumor differentiation, P = 0 · 093; in the concentration of 10-7mol / L 5- The apoptosis rate of Hep2 cells induced by Aza-CdR in APAF-1 overexpression group was increased from 12.46% in the blank control group and 14.74% in the empty vector transfected group to 28.64%, and the S phase cells Periodic block. Conclusion: Overexpression of APAF-1 gene can significantly increase the sensitivity of Hep2 cell line to 5-Aza-CdR-induced apoptosis induced by demethylation drug.