BNIP3表达对结肠癌细胞化疗敏感性的影响  

Effect of BNIP3 expression on chemotherapeutic sensitivity of colon carcinoma cells

作　　者：裴莉[1] 解方为[1] 陈克力[1] 江恒[1] 陈建芳[1] 梁后杰[1] 

机构地区：[1]第三军医大学西南医院肿瘤科,重庆400038

出　　处：《解放军医学杂志》2009年第9期1050-1053,共4页Medical Journal of Chinese People's Liberation Army

基　　金：重庆市自然科学基金重点资助项目(2007BA5008)

摘　　要：目的研究启动子甲基化对人结肠癌SW480细胞BNIP3基因的转录调控作用,以及通过去甲基化作用恢复BNIP3表达对细胞化疗敏感性的影响。方法用DNA甲基转移酶抑制剂5-氮-2′-脱氧胞苷(5-aza-dC)处理SW480细胞72h,应用甲基化特异性PCR(MSP)检测BNIP3基因启动子区CpG岛甲基化状态。用RT-PCR和Western blotting检测BNIP3的mRNA和蛋白表达,流式细胞术分析细胞凋亡,MTT法观察化疗药物氟尿嘧啶(5-FU)和奥沙利铂(L-OHP)对细胞的增殖抑制作用。结果BNIP3基因在SW480细胞中表达沉默,基因启动子区存在异常甲基化。1.0μmol/L的5-aza-dC可以使BNIP3基因启动子明显去甲基化、BNIP3表达和细胞凋亡增加,并可以与5-FU和L-OHP发挥协同作用,使药物对细胞的增殖抑制作用显著增强。5-aza-dC的上述作用在低氧培养条件下更加显著。结论启动子甲基化是导致结肠癌SW480细胞BNIP3基因表达沉默的重要原因。5-aza-dC可通过去甲基化作用恢复BNIP3基因的表达,增加细胞对5-FU和L-OHP的敏感性,且其作用在低氧条件下更加显著。Objective To investigate the transcriptional regulation effect of promoter methylation on BCL2-adenovims E1B 19kD- interacting protein 3 （BNIP3） gene, and the effect of restoration of BNIP3 expression by demethylation on the chemotherapeutic sensitivity of colon carcinoma SW480 cell line. Methods Human colon carcinoma SW480 cell line was treated with 5 aza-2-deoxycytidine （5-aza-dC）, a specific DNA methyltransferase inhibitor, for 72h. The methylation status of the CpG island of BNIP3 gene promoter was detected by methylation specific PeR （MSP）. The expression of BNIP3 mRNA and protein were assessed by RT-PeR and Western blotting respectively. Induction of apoptosis was analyzed by flow cytometry. Growth inhibition of SW480 cells by fluorouraeil （5-FU） or oxaliplatin （L-OHP） treatment was evaluated by MTT. Results Aberrant promoter methylation of BNIP3 was detected in BNIP3 silenced colon carcinoma SW480 cell line. After treatment with 1.0μmol/L 5-aza-dC, the promoter of BNIP3 was demethylated and BNIP3 expressed both in mRNA and protein level. Restoration of BNIP3 expression notably enhanced the apeptosis and drug sensitivity of SW480 cells to 5-FU and L-OHP. The effects of 5 aza-dC were strengthened especially in hypoxic culture condition. Conclusions The aberrant methylation of promoter is an important mechanism leading to silence of BNIP3 expression in colon carcinoma SW480 cell line. Restoration of BNIP3 expression by demethylation with 5 aza-dC may enhance the drug sensitivity to 5-FU and L-OHP of SW480 cells, especially in hypoxic culture condition.

关 键 词：结肠肿瘤 BNIP3蛋白 人 DNA甲基化 抗肿瘤联合化疗方案 细胞低氧 

分 类 号：R735.35[医药卫生—肿瘤]