5-氮杂-2’-脱氧胞苷对人胰腺癌细胞系MiaPaca2生长和RUNX3基因甲基化的影响  

Effect of 5-aza-2＇-deoxycyfidine on growth and methylation of RUNX3 gene in human pancreaticcancer cell line MiaPaca2

作　　者：韩旭[1,2] 谭志军[1,2] 郭仁德[1,2] 李照晋[1,2] 王玉亮[3] 

机构地区：[1]天津市第一中心医院普通外科,300192 [2]淮北市矿工总医院普通外科,235000 [3]卫生部危重病急救医学重点实验室

出　　处：《中华肿瘤杂志》2013年第1期17-21,共5页Chinese Journal of Oncology

基　　金：天津市卫生局重点项目（2010KR04）

摘　　要：目的探讨去甲基化制剂5-氮杂-2’-脱氧胞苷（5-Aza-CdR）对胰腺癌细胞系MiaPaca2生长的影响，以及对RUNX3抑癌基因表达和甲基化的影响。方法光镜观察5-Aza-CdR处理前后MiaPaca2细胞形态学变化，四甲基偶氮唑蓝法检测5-Aza-CdR对MiaPaca2细胞增殖活性的影响，半定量逆转录聚合酶链反应检测RUNX3mRNA表达的变化，甲基化特异性聚合酶链反应检测RUNX3基因甲基化的变化。结果经2．5、5、10和20μmol／L5-Aza-CdR处理24h后，MiaPaca2细胞的抑制率分别为（9．17±2．15）％、（10．754-2．04）％、（12．574-1．64）％和（18．70±1．51）％；48h后分别为（14．94±1．68）％、（18．60±1．57）％、（22．84±1．58）％和（33．24±1．53）％；72h后分别为（21．46±1．60）％、（28．62±1．72）％、（35．14±1．64）％和（45．06±1．47）％；96h后分另0为（26．35±1．71）％、（34．48±1．69）％、（40．05±1．60）％和（49．99±1．61）％。各实验组与对照组抑制率比较，差异均有统计学意义（均P〈0．05）。在同一时点，各浓度组间MiaPaca2细胞的抑制率差异均有统计学意义（均P〈0．05）。在48、72、96h时，各浓度组抑制率两两比较，差异均有统计学意义（均P〈0．05）。在药物处理24h以后，5-Aza．CdR对MiaPaca2细胞的生长抑制作用随着-Aza-CdR浓度的增加而增强。5-Aza-CdR能够逆转RUNX3基因的甲基化状态，恢复RUNX3mRNA的表达。结论RUNX3基因甲基化状态与胰腺癌的发生发展密切相关，异常甲基化可能是引起RUNX3基因表达缺失的重要机制。5-Aza-CdR可以逆转MiaPaca2细胞RUNX3基因的甲基化状态，重新激活其表达，从而抑制肿瘤细胞的生长，为胰腺癌的诊断和治疗提供了一条新途径。Objective To investigate the effect of demethylating agent 5-aza-2＇-deoxycytidine （5-Aza-CdR） on the growth of human pancreatic cancer cell line MiaPaca2 and the expression and methylation of tumor suppressor gene RUNX3. Methods Human pancreatic cancer cell line MiaPaca2 cells were treated with different concentrations of 5-Aza-CdR. Morphological changes of MiaPaca2 cells were observed by light microscopy. The activity of cell proliferation was analyzed by MTT assay. The changes of RUNX3 mRNA expression were detected by semi-quantitative reverse transcription-polymerase chain reaction （RT-PCR）. Changes of RUNX3 gene methylation was detected by methylation-specific polymerase chain reaction. Results MiaPaca2 cells were treated with 2.5, 5, 10 and 20 ttmol/L 5-Aza-CdR, respectively. The inhibition rates of MiaPaca2 cells treated for 24 h were （9. 17 ±2. 15）%, （10.75 ±2.04）%, （12.57± 1.64）% and （18.70 ±1.51）%, respectively. The inhibition rates were （14.94 ± 1.68）%, （18.60 ± 1.57）%, （22.84 ± 1.58）% and （33.24 ± 1.53）%, respectively, after 48 h treatment; （21.46 ±1.60）%, （28.62 ± 1.72）%, （35. 14± 1.64）% and （45.06 ±1.47）%, respectively, after 72 h treatment; and （26.35±1.71）%, （34.48 ±1.69）%, （40.05 ± 1.60）% and （49.99 ±1.61）%.respectively, after 96 h treatment. The differences between inhibition rates of each experimental and control groups （0.00 ± 0.00） % were statistically significant （ P 〈 0.05 ）. At the same time, the inhibition rates of different concentration groups showed significant differences （ P 〈 0.05 ）. At 48 h, 72 h and 96 h, the inhibition rates of each pair concentration groups showed significant differences （ P 〈 0.05 ）. 5-Aza-CdR inhibited the growth of MiaPaca2 cells, and the higher the concentration, the stronger the inhibition after 24 h. 5-Aza-CdR also reversed the methylation status of RUNX3 gene, and restored the expression of RUNX3 mRNA with a dose-effect relationship. Conclusions

关 键 词：5-氮杂-2’-脱氧胞苷 胰腺肿瘤 RUNX3基因 甲基化 

分 类 号：R735.9[医药卫生—肿瘤]