通过小干扰RNA沉默mdr-1基因逆转卵巢癌耐药细胞株SKOV3/TAXOL多药耐药的研究  

Study on silencing of endogenetic mdr-1 gene expression by RNA interference in ovarian cancer resistant strain SKOV3/TAXOL

作　　者：冯继良[1] 姚焕铃 王红梅[2] 杜秀平[2] 

机构地区：[1]徐州市妇幼保健院妇科,210009 [2]徐州医学院肿瘤研究所

出　　处：《肿瘤研究与临床》2010年第10期686-688,共3页Cancer Research and Clinic

基　　金：徐州市社会发展基金（X200523113）

摘　　要：目的 构建靶向于卵巢癌耐药细胞株中高表达的多药耐药-1（mdr-1）基因的短发夹状RNA重组质粒,探讨RNAi沉默技术逆转卵巢上皮性癌（卵巢癌）细胞多药耐药的可行性.方法运用基因克隆技术构建靶向于mdr-1基因的重组质粒PGPU6/GFP/Neo-mdr-1并转染至卵巢癌耐药细胞株SKOV3/TAXOL,反转录聚合酶链反应检测到mdr-1基因表达.CCK-8检测对SKOV3/TAXOL细胞增殖的抑制作用.结果构建的重组质粒PGPU6/GFP/Neo-mdr-1能明显抑制mdr-1基因的表达.蛋白质印迹结果表明,转染重组质粒PGPU6/GFP/Neo-mdr-1的SKOV3/TAXOL靶细胞的抑制率显著提高.RT-PCR检测mdr-1基因mRNA转录水平下降;pPGPU6/GFP/Neo-mdr-1明显抑制SKOV3/TAXOL细胞的增殖.结论重组质粒PGPU6/GFP/Neo-mdr-1可有效地抑制SKOV3/TAXOL细胞内源mdr-1基因的表达和mRNA的转录,并抑制细胞的增殖和促进凋亡的发生,应用RNAi技术,能够逆转卵巢癌细胞对化疗药物的多药耐药.为化疗耐受的肿瘤细胞提供了新的基因治疗手段.Objective To construct the small hairpinRNA recombinant plasmids targeting mdr-1 gene which expresses highly in ovarian cancer resistance strain SKOV3/TAXOL to silence endogenetic mdr-1 gene expression and investigate the role of mdr-1 gene in the development of resistant ovarian cancer. Methods The pPGPU6/GFP/Neo-mdr-1 were constructed by gene clone technology. The influence on proliferation and apoptosis were investigated by CCK-8 in SKOV3/TAXOL after transfected. pPGPU6/GFP/Neo-mdr-1. Results The expression against mdr-1 proteins were inhibited by pPGPU6/GFP/Neo-mdr-1. The cell proliferation were inhibited after transfected pPGPU6/GFP/Neo-mdr-1 by CCK-8. The apoptosis were observed in DAB experiments and the apoptosis rate incised. Conclusion mdr-1 plays an important role in proliferation of resistant ovarian cancer and the short hairpinRNA of mdr-1 can efficiently suppress mdr-1expression and enhance the apoptosis in SKOV3/TAXOL, which provides a novel method for chemotherapy resistant tumors.

关 键 词：RNA干扰 卵巢肿瘤 多药耐药蛋白1 P糖蛋白 基因 细胞增殖 细胞凋亡 

分 类 号：R737[医药卫生—肿瘤]