亚砷酸对人喉癌Hep-2细胞RECK基因表达及去甲基化的影响

Effects of Arsenic Trioxide on Expression and Demethylation of RECK Gene in Laryngocarcinoma Cells

目的:探讨不同浓度亚砷酸对人喉癌Hep-2细胞中伴Kazal域的富含半胱氨酸的逆转诱导蛋白(RECK)基因表达及去甲基化的影响。方法:分别将0.5、1.0、2.0、4.0μmoL/L的亚砷酸作用于体外培养的Hep-2细胞24～72 h,MTT法计算细胞生长抑制率,流式细胞术检测细胞周期,并用甲基特异性PCR检测RECK基因甲基化情况。RT-PCR和W estern b lot分别检测RECK mRNA和蛋白表达水平。结果:0.5～4.0μmoL/L亚砷酸能抑制Hep-2细胞增殖,使Hep-2细胞阻滞于S期和G2/M期;RECK基因甲基化水平随亚砷酸浓度增高而降低,非甲基化水平则逐渐增高;亚砷酸能增加RECK mRNA和蛋白表达。结论:亚砷酸能促进Hep-2细胞中RECK基因去甲基化并提高其表达水平,从而发挥抑制细胞增殖。

Objective To study the effects of arsenic trioxide on reversion-inducing cysteine-rich protein with Kazal motif(RECK) gene expression and demethylation in laryngocarcinoma cell line Hep-2.Methods The Hep-2 cells were cultured with different concentrations of arsenic trioxide in vitro.The growth inhibitroy rates of Hep-2 cells were detected by MTT assay,the cell cycle was analyzed by flow cytometry,the methylation status of RECK gene was determined with methylation specific PCR.The mRNA and protein expression were detected with RT-PCR and Western blot,respectively.Results Arsenic trioxide could inhibit the proliferation of Hep-2 cells in dose range from 0.5 to 4.0 μmoL/L,arrest the cell cycle at G2/M phase.The methylation of RECK gene was decreased following the increased dosage of arsenic trioxide.The mRNA and protein expression of RECK gene could be heightened with arsenic trioxide.Conclusion Arsenic trioxide could induce RECK gene expression,thus to inhibit the proliferation of laryngocarcinoma cells.