多发性骨髓瘤U266细胞株DNA甲基转移酶和甲基结合蛋白的表达  

The expression of DNA methyltransferases and methyl-DNA binding proteins in multiple myeloma U266 cell line

作　　者：许闪闪[1] 李智[1] 何丽梅[1] 翁文浩[1] 于慧[1] 

机构地区：[1]同济大学附属第十人民医院检验科

出　　处：《检验医学》2010年第8期632-636,共5页Laboratory Medicine

摘　　要：目的检测多发性骨髓瘤（MM）细胞DNA甲基转移酶（DNMTs）及甲基结合结构域（MBD）蛋白[包括甲基CpG结合结构域蛋白（MeCP2）和MBD2]的表达状况,探索其在基因甲基化过程的作用。并观察5-氮杂-2＇-脱氧胞苷（5-Aza-CdR）和/或曲古霉素A（TSA）对DNMTs及MeCP2和MBD2的作用。方法利用荧光定量聚合酶链反应（PCR）分析MMU266细胞株DNMTs（DNMT1、DNMT3a、DNMT3b）及MeCP2和MBD2蛋白的mRNA水平,与10名正常人单个核细胞（PBMC）mRNA比较。4μmol/L5-Aza-CdR和/或0.1μmol/LTSA与U266细胞共培养,分析MBD2和MeCP2蛋白mRNA的变化情况。WesternBlot检测这2种药物对U266细胞MBD2蛋白的影响。结果各甲基化调控蛋白mRNA与PBMCmRNA相比,差异有统计学意义（P均〈0.05）。其中DNMTs和MBD2蛋白在U266细胞中表达升高,而MeCP2蛋白表达下降。经5-Aza-CdR和/或TSA作用后,MBD2表达下降,MeCP2升高。5-Aza-CdR在蛋白水平对MBD2的作用亦不显著;TSA可明显降低MBD2的表达,且具有时间依赖性。结论 DNMTs和MBD2、MeCP2蛋白在MMU266细胞中表达异常,可能与U266细胞多个抑癌基因启动子高甲基化相关。5-Aza-CdR和TSA可以逆转U266细胞MBD2和MeCP2的表达。Objective To investigate the expression of DNA methyltransferases （ DNMTs） and methyl-CpG-binding domain （ MBD） proteins including MeCP2 and MBD2 in multiple myeloma （ MM） cell and the influence on methylation,and observe the effect of 5-aza-2-deoxycitydine （ 5-Aza-CdR） and/or trichostatin A （ TSA） on DNMTs,MeCP2 and MBD2. Methods The mRNA levels of DNMTs （ DNMT1,DNMT3a and DNMT3b） ,MeCP2 and MBD2 proteins in MM U266 cell line and the mRNA level of peripheral blood mononuclear cell（ PBMC） from 10 healthy controls were analyzed and compared by fluorescence quantitative polymerase chain reaction （ PCR）. U266 with 4 μmol/L 5-Aza-CdR and/or 0. 1μmol/L TSA was cultured,and the mRNA of MBD2 and MeCP2 proteins was analyzed. In addition,the influence on MBD2 protein in U266 cell was evaluated by Western blot. Results There were statistical significance between the mRNA levels of methylation regulating proteins and PBMC （ P〈0. 05）. The expressions of DNMTs and MBD2 increased in U266,while the expression of MeCP2 decreased. After treated with 5-Aza-CdR and/or TSA,the expression of MBD2 decreased,and the expression of MeCP2 increased. The effect of 5-Aza-CdR to MBD2 was not obvious. TSA with time dependence reduced the expression of MBD2. Conclusions DNMTs and MBD2,MeCP2 proteins are aberrant in MM U266 cell. It may be related with promoter hypermethylation in multiple tumor suppressor genes. 5-Aza-CdR and TSA can reverse the expression of MBD2 and MeCP2 in U266 cell.

关 键 词：多发性骨髓瘤 表观遗传学 DNA甲基转移酶 甲基结合结构域蛋白 

分 类 号：Q503[生物学—生物化学]