乳腺癌组织中ERα mRNA和PR mRNA检测方法的建立和应用  

The establishment of ERa and PR mRNA assays and their application in breast cancer

作　　者：叶伟民[1] 何金[1] 何铭珺[1] 杨洁[1] 徐毅[1] 陆慧琦[1] 

机构地区：[1]第二军医大学长征医院实验诊断科,上海200003

出　　处：《现代免疫学》2012年第5期412-416,共5页Current Immunology

基　　金：上海市科委基金资助项目(08411962100)

摘　　要：建立测定雌激素受体α（ERa）tuRNA和孕激素受体（PR）mRNA的实时荧光定量RTPCR，探讨两者在乳腺癌组织及良性乳腺肿瘤组织中的表达水平。分别以pMDl8－ERα和pMDl8－PR质粒为定量模板，用循环阂值（Ct）定量起始模板，在荧光TaqMan方法的基础上建立了测定ERamRNA和PRmRNA的实时荧光定量RT—PCR，并分别测定48例乳腺癌组织及28例良性乳腺肿瘤组织中ERamRNA和PRmRNA的表达水平。ERamRNA和PRmRNA测定的线性范围为103～108copy／μgRNA；ERamRNA和PRmRNA高值和低值的批内和批间变异系数（cⅥ在5．07％～11．28％之间。ERmRNA在乳腺癌组织中的表达量为6．76×10。copy／μgRNA（4．17×10。，9．34×10。），良性乳腺肿瘤组织中的含量为1．54×10μcopy／／μgRNA（1．02×10。，2．06×10。），乳腺癌组织的表达量高于良性组织（P％0．05）；PRmRNA在乳腺癌组织中的表达量为1．02×10。copy／μgRNA（6．81×10。，1．36×10。），良性乳腺肿瘤组织中的含量为4．93×10。copy／／μgRNA（3．21×10。，6．65×10。），两者表达无明显差异（P〉0．05）。ERαmRNA和PRmRNA在乳腺癌和良性乳腺肿瘤组织中的表达存在相关性。我们建立的测定ERamRNA和PRmRNA的实时荧光定量RT—PCR灵敏、稳定、重复性好，可供临床检测和研究。ERamRNA和PRmRNA基因表达水平可作为预测治疗效果及判断预后的重要指标。To establish a specific fluorogenic quantitative method for detecting the oestrogen receptoralpha （ERa） and proges terone receptor（PR） gene, and to explore their expression both in benign and cancer tissues, based on fluorescent Taq Man methodology, a real time quantitative RTPCR was set up. In this method, cloning vector pMD18ERa and pMD18PR were constructed as standard plasmids. RNA quantification was based on the threshold cycle（Ct） values when using GeneAmp ？500 Sequence Detection Systems to examine the specific expression of ERα and PR in 38 cases with breast cancer and 32 cases with benign tumor（the controls）. The detection linear range of the assay was 10a N 108copy/μg RNA. The intra and interassay co efficient of variation of the ERa and PR for low value and high value was 5. 07% and 11. 28% respectively. The ERα mRNA copy number was 6.76 X l0s copy/gg RNA （4.17× 105 , 9.34 ×105 ） in breast cancers and 1.54 × 105 copies/gg RNA（ 1.02 × 105, 2.06X10s） in controls. ThePRmRNA copy number was 1.02X10scopy/vg RNA （6.81X10s, 1.36X106） in cancer and 4.93 X 105 copy/pμg RNA （3.21 ）〈 105 , 6.65 ）〈 l0s ） in controls, respectively. ERα mRNA levels were significantly elevated in breast cancer tissues when compared to control tissues （P〈0.05）, and were well associated with the clinicopathological lea tures of breast cancer. PR mRNA did not reveal significant difference between cancers and controls （P〉0.05）. Thus,a real time quantitative RTPCR method for detecting the expression of ERRα and PR gene in the breast tissue has been successfully es tablished. , ERRα and PR detection is a very important clinical test in breast cancer, which might not only be an important prog nostic and predictive marker but also be used to determine optimal treatment strategies.

关 键 词：雌激素受体Α 孕激素受体 乳腺癌 实时荧光定量RT-PCR 

分 类 号：R392.12[医药卫生—免疫学]