二氢叶酸还原酶在两种不同方式诱导的甲氨蝶呤耐药绒癌细胞系中的动态表达  

Dynamic profiling of dihydrofolate reductase transcript level in two different series of MTX-inducing JeG-3 sub-lines.

作　　者：韩冰[1,2] 向阳[2] 王峥[3,4] 王云[3,4] 张浩[2] 黄尚志[3,4] 

机构地区：[1]苏州大学附属第一医院妇产科,苏州215006 [2]中国医学科学院北京协和医学院北京协和医院妇产科 [3]中国医学科学院基础医学研究所医学遗传学系 [4]北京协和医学院基础学院WHO遗传病社区控制合作中心

出　　处：《现代妇产科进展》2010年第8期561-564,共4页Progress in Obstetrics and Gynecology

基　　金：国家自然基金资助项目(No:30973178)

摘　　要：目的:用间断和连续两种诱导方式建立对甲氨蝶呤(MTX)耐药的绒毛膜癌细胞系,并动态检测二氢叶酸还原酶(DHFR)在两种耐药细胞系建立过程中的表达。方法:采用间断和连续诱导法诱导人绒癌细胞系JeG-3,分别建立绒癌MTX耐药细胞系MTXA1(间断诱导浓度为1.1×10-3mol/L)和MTXC2(连续诱导浓度为1.1×10-5mol/L)。细胞计数法(CCK-8)观察耐药细胞的耐药指数;化学发光法和荧光定量PCR技术分别检测不同诱导方法和不同浓度MTX诱导的JeG-3细胞中β人绒毛膜促性腺激素(β-HCG)的分泌量和DHFR mRNA的表达水平。结果:(1)历时1年成功建立了绒癌耐药细胞系MTXA1和MTXC2,其耐药指数分别为21.36和28.84;(2)β-HCG分泌和DHFR mRNA表达水平在间断和连续两种不同诱导方式诱导的绒癌细胞株中呈相同的动态变化趋势:经低浓度MTX诱导后,JeG-3细胞中β-HCG分泌和DHFR mRNA的表达水平上调;但当MTX诱导浓度增加到一定剂量时,两者不再继续上调而呈下降趋势。结论:成功建立了间断和连续两种方式诱导的耐药绒癌细胞系MTXA1和MTXC2。绒癌细胞β-HCG分泌和DHFR mRNA的表达与MTX作用浓度以及耐药性无明显的正相关关系。目前的研究结果表明,DHFR mRNA的表达水平不适宜作为绒癌细胞是否对MTX耐药的指标。Objective：To assess dynamic profiling of dihydrofolate reductase transcript level in MTX-resistant JeG-3 sub-lines by using intermittent and consecutive feeding methods respectively.Methods：Intermittent and consecutive feeding methods were used to establish MTX-resistance JeG-3 sub-lines：MTXA1 and MTXC2.β subunit human chorionic gonadotropin（β-HCG）in the culture medium was detected by chemo-luminescence method.The resistant index（RI） was measured by cell counting Kit-8（CCK-8） assay.Quantitative real-time PCR（qRT-PCR） was used to detect the mRNA expression level of dihydrofolate reductase（DHFR） in the sub-lines during the different concentration of MTX-exposure.Results：The RI of MTXA1 and MTXC2 were 21.36 and 28.84 respectively.Interestingly,by using two different kinds of MTX-exposure,the same dynamic profiling of β-HCG secretion and DHFR transcript level were found：At certain range of concentration,the upward mRNA level of DHFR transcript level and the β-HCG secretion were correlated with that of MTX-exposing.But,once the concentration of MTX exceeded this range,they would not go up sequentially but decrease sharply.Conclusions：We established the MTX-resistant sub-lines of JeG-3 by using two different kinds of inducing methods;just like the β-HCG secretion,the DHFR mRNA expression level is not in accordance with the concentration of MTX-exposure.Our data suggested that DHFR mRNA expression level may not be used as a biomarker to predict the chemo-resistant in MTX-based chemotherapy.

关 键 词：绒毛膜癌 JeG-3细胞系 抗药性 肿瘤 甲氨蝶呤 二氢叶酸还原酶 人绒毛膜促性腺激素 荧光定量PCR 

分 类 号：R737.33[医药卫生—肿瘤]