DACT2基因启动子甲基化与宫颈癌细胞化疗敏感性的相关性研究

The Relationship between DACT2 Gene Promoter Methylation and Chemotherapy Sensitivity of Cervical Cancer Cells

目的:探讨与研究DACT2基因启动子甲基化与宫颈癌细胞化疗敏感性的相关性。方法:人宫颈癌顺铂耐药细胞系SIHA/DDP根据实验目的分为三组-对照组、DACT 1组与DACT 2组,组分别加入含0.0μmol/L、1.0μmol/L、10.0μmol/L DACT2基因启动子甲基化抑制剂-5-aza-dC进行治疗,采用MTT法检测细胞增殖指数,流失细胞法检测细胞凋亡指数,PCR法检测甲基化水平,流式细胞仪检测胞周期,Western Blot检测Wnt蛋白与TGF-β1蛋白表达情况。结果:治疗后24 h、36 h的DACT 1组与DACT 2组DACT2基因启动子甲基化相对水平低于对照组(P<0.05),DACT 2组低于DACT 1组(P<0.05)。治疗后24 h、36 h的DACT 1组与DACT 2组细胞增殖指数低于对照组(P<0.05),细胞凋亡指数高于对照组(P<0.05),DACT 2组与DACT1组对比差异都有统计学意义(P<0.05)。治疗后24 h、36 h的DACT 1组与DACT 2组G2/M期细胞比例高于对照组(P<0.05),G0/G1期细胞比例低于对照组(P<0.05),DACT 1组与DACT 2组对比差异有统计学意义(P<0.05)。治疗后24 h、36 h的DACT 1组与DACT 2组的Wnt蛋白与TGF-β1蛋白相对表达水平低于对照组(P<0.05),DACT 2组低于DACT 1组(P<0.05)。结论:抑制DACT2基因启动子甲基化能抑制宫颈癌细胞的Wnt/TGF-β1信号通路的激活,能调节宫颈癌细胞周期平衡,能抑制顺铂耐药性宫颈癌细胞增殖,促进细胞凋亡,并增强化疗敏感性,且在本研究设置范围内,作用剂量越高,效果越显著。

Objective:To explore and study the correlation of Homo sapiens dapper,antagonist of beta-catenin(DACT)2 gene promoter methylation and chemotherapy sensitivity of cervical cancer cells.Methods:The cisplatin-resistant human cervical cancer cell line SIHA/DDP was divided into three groups according to the experimental purpose-control group,DACT 1 group and DACT 2 group.Submethylation inhibitor-5-aza-dC were used for treatment.MTT method were used to detect cell proliferation index,loss cell method to detect cell apoptosis index,PCR method to detect methylation level,flow cytometry to detect cell cycle,Western Blot The expression of Wnt protein and TGF-β1 protein were detected.Results:At 24h and 36h after treatment,the relative levels of DACT2 gene promoter methylation in DACT 1 and DACT 2 groups were lower than those in the control group(P<0.05),and the DACT 2 group were lower than that in the DACT 1 group(P<0.05).The cell proliferation index in DACT 1 group and DACT 2 group were lower than that in the control group(P<0.05),and the apoptosis index were higher than that in the control group(P<0.05).There were also statistical significance compared between the DACT 2 group and the DACT 1 group(P<0.05).At 24h and 36h after treatment,the proportion of cells in G2/M phase in DACT 1 group and DACT 2 group were higher than that in control group(P<0.05),and the proportion of cells in G0/G1 phase were lower than that in control group(P<0.05).There were also statistical significance compared between the DACT 2group and the DACT 1 group(P<0.05).The relative expression levels of Wnt protein and TGF-β1 protein in DACT 1 group and DACT 2group at 24h and 36h after treatment were lower than those in control group(P<0.05),and DACT 2 group were lower than DACT 1group(P<0.05).Conclusion:Inhibition of DACT2 gene promoter methylation can inhibit the activation of Wnt/TGF-β1 signaling pathway in cervical cancer cells,regulate the balance of cervical cancer cell cycle,inhibit the proliferation of cisplatin-resistant cervical cancer cells,and promote cell apoptosis,thereby increasing chemosensitivity.And within the setting range of this study,the higher the dose,the more significant the effect.