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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 effects of 5-Aza-CdR on proliferation and apoptosis of human colorectal cancer cell line Lovo and the expression of tumor suppressor gene RUNX3.Methods: -Aza-CdR0.4, 4 and 40μmol / L were treated with Lovo3d and the cells were cultured for 5 days. MTT assay was used to observe the growth activity of the cells before and after treatment by semi-quantitative The expression of RUNX3 mRNA was detected by RT-PCR before and after treatment. The methylation status of RUNX3 was detected by methylation-specific PCR (MSP), and the apoptosis was detected by flow cytometry The relative expression of RUNX3 mRNA in cells treated with 0.4,4,40μmol / L of 5-Aza-CdR (0.46 ± 0.06,0.71 ± 0.06,0.84 ± 0.07vs0, P <0.01) and apoptosis rate (10.95% ± 2.09%, 17.61% ± 1.51%, 26.60% ± 1.89% vs 2.92% ± 0.93%, P <0.01) , F = 145.7). The growth of Lovo cells decreased and RUNX3 mRNA was re-expressed in colorectal cancer cells, and the gene promoter region was partially methylated.Conclusion: 5-Aza-CdR can reverse the hypermethylation of RUNX3 promoter and inhibit the growth of cells Long, apoptosis inducing portion.