从纤支镜刷落细胞提取微量RNA并检测Foxp3基因表达  

Isolation of total RNA from bronchofibroscopic brush-off cells and detection of Foxp3 gene expression

作　　者：王琦[1] 张行[1] 周畔[1] 张敷彪[1] 曹江[1] 

机构地区：[1]浙江大学邵逸夫医院临床医学研究所,浙江杭州310016

出　　处：《全科医学临床与教育》2008年第3期204-206,210,F0002,共5页Clinical Education of General Practice

摘　　要：目的建立纤维支气管镜刷落细胞提取微量核糖核酸(RNA)的方法,检测分析调节性T细胞(Treg)Foxp3基因在肺癌及肺良性疾病患者中的表达水平。方法以磁珠吸附法提取154例肺癌及肺良性疾病患者纤支镜刷脱细胞的微量RNA,以实时荧光定量RT-PCR的方法检测Treg特异性转录因子Foxp3基因在肺癌及肺良性疾病患者中的表达水平。结果平均总RNA提取量为5.08μgRNA/例,最低为3.20μg,最高为15.25μg。肺癌患者组中Foxp3 mRNA的表达水平(0.0000～13682.0805,四分位数间距0.0011)显著高于肺良性疾病组(0.0000～0.0061,四分位数间距0.0000)(Z=-3.508,P<0.01),肺癌患者组中Foxp3的阳性率(29.34%,27/92)亦明显高于肺良性疾病组(3.23%,2/62)(χ2=16.535,P<0.01)。结论利用纤支镜毛刷刷取病灶部位涂片后毛刷中的剩余细胞,采用磁珠吸附的方法可以提取微量RNA,并用于基因表达水平的检测。Objective To setup protocol for RNA isolation from bronchofibroscopic brush-off cells and analyze the expression of Foxp3 in samples of lung carcinoma and benign disease patients. Methods Total RNA was isolated from bronchofibroscopic brush-off cell samples of 154 patients with lung carcinoma and benign lung diseases by magnetic-bead total RNA isolation method. Real-time fluorescent quantitative RT-PCR was used to detect the expression of Treg-specific transcription factor Foxp3, Results The mean yield of total RNA was 5.08μg/sample, with the lowest as of 3.20μg/sample and the highest as of 15.25μg/sample. The expression level of Foxp3 in lung cancer patient samples （0.0000-13682.0805, interquartile range 0.0011） was significantly higher than that in benign disease samples （0.0000-0.0061, interquartile range 0.0000） （Z=-3.508, P〈0.01）, and the positive rate of Foxp3 in lung cancer patient samples （29.34%, 27/92） was also higher that that in benign disease samples （3.23%, 2/62）（X^2=16.535, P〈0.01）. Conclusions Total RNA can be isolated from bronchofibroscopic brush-off cells by magnetic-bead method and used for further detection of gene expression.

关 键 词：纤维支气管镜 刷落细胞 微量RNA提取 FOXP3 

分 类 号：R450[医药卫生—治疗学]