5-杂氮-2′-脱氧胞苷诱导鼻咽癌细胞株Syk基因启动子去甲基化的研究  

Study on 5-aza-2'-deoxycytidine Induces Syk Gene Promoter Demethylation in Nasopharyngeal Carcinoma Cell Lines

作　　者：金巧智[1] 鄢冲[1] 陶宝鸿[2] 李志海[2] 蔡志毅[2] 

机构地区：[1]温州医学院第一临床医学院,温州325000 [2]浙江省台州市立医院耳鼻咽喉-头颈外科,台州318000

出　　处：《中国细胞生物学学报》2012年第9期865-871,共7页Chinese Journal of Cell Biology

基　　金：台州市科技计划(No.08KY23);台州学院青年基金(No.2011-QN31)资助项目~~

摘　　要：利用5-杂氮-2′-脱氧胞苷(5-aza-2′-deoxycytidine,5-aza-CdR)处理体外培养的鼻咽癌细胞株CNE-1、CNE-2及永生化非癌性人鼻咽上皮细胞株NP-69,采用BS-PCR、Q-RT-PCR及Westernblot方法分别检测经5μmol/L的5-aza-CdR处理前后,各细胞株中Syk基因启动子甲基化状况及SykmRNA和蛋白质表达情况。探讨去甲基化药物5-杂氮-2′-脱氧胞苷(5-aza-CdR)对鼻咽癌细胞株中脾酪氨酸激酶(spleen tyrosine kinase,Syk)启动子甲基化水平及其表达的影响。结果显示,Syk基因启动子甲基化水平与鼻咽癌细胞分化程度呈负相关,两种鼻咽癌细胞株的Syk mRNA和蛋白质表达水平显著低于NP-69细胞(P<0.01);经5-aza-CdR处理后两种鼻咽癌细胞株的Syk基因启动子甲基化水平降低,Syk mRNA及蛋白质表达升高(P<0.05);高分化鼻咽癌细胞株对药物敏感性高于低分化鼻咽癌细胞株(P<0.01)。由此可见,两种鼻咽癌细胞株中存在不同程度的Syk基因启动子甲基化状态,5-aza-CdR能有效逆转鼻咽癌细胞株Syk基因启动子的甲基化状态,升高Syk mRNA及蛋白质表达,同时鼻咽癌细胞分化程度越高恢复Syk基因表达的比率越高。The nasopharyngeal carcinoma cell lines,CNE-1,CNE-2 and the non-cancerous immortalized nasopharyngeal epithelial cell lines NP-69 were treated with 5 μmol/L 5-aza-2＇-deoxycytidine（5-aza-CdR） in vitro.BS-PCR,Q-RT-PCR,Western blot were used to detect Syk promoter methylation and the level of Syk mRNA and protein.This study purposed to investigate the effect of 5-aza-CdR on the degree of promoter methylation and the expression of spleen tyrosine kinase（Syk） in nasopharyngeal carcinoma cell lines.It suggests that the mRNA and protein level of Syk in the two nasopharyngeal carcinoma cell lines were significantly lower than in NP-69 cell line（P0.01）,and shows negative relation with the methylation level of Syk promoter.The promoter methylation level of Syk gene was significantly decreased and the expression of Syk mRNA and protein were up-regulated（P0.05） after treatment with 5-aza-CdR in the cell lines of CNE-1 and CNE-2,which was not observed in NP-69 cell line（P0.05）.Compared with poorly differentiated nasopharyngeal carcinoma cell line,well-differentiated nasopharyngeal carcinoma cell line was more sensitive to 5-aza-CdR（P0.01）.Taken together,Syk promoter methylation directly leads to the decreased expression of Syk in nasopharyngeal carcinoma cell lines.5-aza-CdR induces the demethylation of the Syk promoter,restores the expression of Syk both in mRNA and protein levels.Meanwhile,the cytotoxicity of 5-aza-CdR did not affect the transcriptional activity of Syk gene in nasopharyngeal epithelial cells.Moreover,the different sensitivity of nasopharyngeal carcinoma cell lines to 5-aza-CdR was related with the methylation of Syk promoter.

关 键 词：鼻咽癌 去甲基化 5-杂氮-2′-脱氧胞苷 SYK基因 

分 类 号：R739.63[医药卫生—肿瘤]