摘要
目的:探讨5-氮-2'-脱氧胞苷(5-Aza-CdR)对人结肠癌Lovo细胞增殖凋亡及抑癌基因RUNX3表达的影响.方法:用特异性甲基转移酶抑制剂5-Aza-CdR0.4,4,40μmol/L处理人结肠癌细胞株Lovo3d,继续常规培养5d后,采用四唑盐法(MTT)比色法观察细胞经药物处理前后的生长活性,以半定量RT-PCR检测细胞处理前后抑癌基因RUNX3 mRNA的表达,以甲基化特异性PCR(methylation-specific PCR,MSP)检测细胞处理前后RUNX3的甲基化状态,应用流式细胞仪进行细胞凋亡率的检测.结果:与对照组相比,0.4,4,40μmol/L的5-Aza-CdR处理细胞后,细胞RUNX3 mRNA的相对表达量(0.46±0.06,0.71±0.06,0.84±0.07vs0,P<0.01)和细胞凋亡率均增高(10.95%±2.09%,17.61%±1.51%,26.60%±1.89%vs2.92%±0.93%,P<0.01),呈剂量依赖性(F=168.4,F=145.7),结肠癌Lovo细胞生长速率下降,RUNX3 mRNA重新表达,其基因启动子区域部分甲基化.结论:5-Aza-CdR可逆转RUNX3启动子高甲基化状态,抑制细胞生长,诱导部分细胞凋亡。
AIM： To investigate the expression and methylation status of tumor suppressor gene RUNX3 in human colon cancer cell line Lovo and explore the effects of 5-aza-2＇-deoxycytidine （5-Aza-CdR） on the proliferation and apoptosis of Lovo cells and the expression of RUNX3 gene, METHODS： Human colon cancer cell line Lovo was treated with 5-Aza-CdR, a specific methyltransferase inhibitor, at the concentrations of 0.4, 4 and 40 μmol/L for 3 d, and then cultured in RPMI 1640 medium for 5 d. The activation of Lovo cells was respectively observed by Tetrazolium salt colorimetric （MTT） assay before and after 5-Aza-CdR treatment. The change in expression of RUNX3 mRNA was observed by semi-quantitative reverse transcription-polymerase chain reaction （RT-PCR）. The apoptosis was analyzed by flow cytometry. The methylation status of gene promoter was determined by methylation-specific PCR （MSP）. RESULTS： Lovo cells treated with 5-Aza-CdR （0.4, 4, 40 μmol/L） displayed a slowed growth rate in different degrees in contrast with those in the control group and their growth rates decreased accordingly with the increase of 5-Aza-CdR concentration. There were significant increases in RUNX3 mRNA expression （0.46 ± 0.06, 0.71 ± 0.06, 0.84 ± 0.07 vs 0, P 〈 0,01） and apoptotic rates of Lovo cells （10.95% ± 2.09%, 17.61% ± 1.51%, 26.60% ± 1.89% vs 2.92% ± 0.93%, P 〈 0.01） after 5-Aza-CdR treatment in comparison with those in the control group, The level of RUNX3 mRNA expression and the apoptotic rates of Lovo cells were increased in correlation with 5-Aza-CdR concentration （F= 168.4, F =145.7, P 〈 0.01）. Methylation of RUNX3 promoter region was confirmed in Lovo cells of control group and detected partly in 5-Aza- CdR-treated group. CONCLUSION： 5-Aza-CdR is able to reverse the methylation status of RUNX3 promoter region. The re-expression of RUNX3 gene can inhibit Lovo cell growth and partly induce Lovo cell apoptosis.
出处
《世界华人消化杂志》
CAS
北大核心
2008年第7期711-715,共5页
World Chinese Journal of Digestology
