MiR-27a modulates MDR1/P-glycoprotein expression by targeting HIPK2 in human ovarian cancer cells
MiR-27a modulates MDR1/P-glycoprotein expression by targeting HIPK2 in human ovarian cancer cells
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DOI：
10.1016/j.ygyno.2010.06.004
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发表时间：
2010-10-01
影响因子：
4.7
通讯作者：
Wang, Zehua
中科院分区：
文献类型：
--
作者：
Li, Zhimin;Hu, Sha;Wang, Zehua
Objective. microRNA（miRNAs）是一类非编码的单链小RNA，负调控基因表达，参与细胞的基本过程和人类肿瘤的发生、发展和进展。在这项研究中，我们研究了miR-27 a在卵巢癌细胞耐药发展中的作用。采用茎环实时荧光定量PCR检测卵巢癌细胞株A2780和A2780/Taxol中miR-27 a的表达。通过Lipofectamine 2000用miR-27 a的模拟物或抑制剂或阴性对照RNA（NC）转染A2780和A2780/Taxol细胞。实时定量PCR检测MDR 1 mRNA表达，Western blot检测P-糖蛋白（P-gp）和同源结构域相互作用蛋白激酶2（HIPK 2）蛋白表达。MTT法检测药物敏感性，流式细胞仪检测细胞凋亡及细胞内Rhodamine 123（Rh-123）荧光强度。卵巢癌紫杉醇耐药细胞株A2780/Taxol中miR-27 a和P-gp的表达水平较其亲本细胞株A2780明显上调。转染miR-27 a抑制剂后，A2780/Taxol细胞MDR 1 mRNA和P-gp蛋白表达降低，HIPK 2蛋白表达增加，紫杉醇敏感性增强，紫杉醇诱导的细胞凋亡增加，细胞内Rh-123荧光强度增强。miR-27 a模拟物处理A2780细胞后，MDR 1 mRNA表达增加，而对紫杉醇的敏感性降低。miR-27 a的失调可能参与卵巢癌细胞耐药的发展，至少部分通过靶向HIPK 2调节MDR 1/P-gp的表达。(C)2010年爱思唯尔公司All rights reserved.
Objective. microRNAs (miRNAs) are non-coding, single-stranded small RNAs that regulate gene expression negatively, which is involved in fundamental cellular processes and the initiation, development and progression of human cancer. In this study, we investigated the role of miR-27a in the development of drug resistance in ovarian cancer cells.Methods. Expression of miR-27a in ovarian cancer cell lines A2780 and A2780/Taxol were detected by stem-loop real-time PCR. A2780 and A2780/Taxol cells were transfected with the mimics or inhibitors of miR-27a or negative control RNA (NC) by Lipofectamine 2000. The expression levels of MDR1 mRNA, P-glycoprotein (P-gp) and Homeodomain-interacting protein kinase-2 (HIPK2) proteins were assessed by real-time PCR and western blot respectively. Drug sensitivity was analyzed by MTT assay while apoptosis and the fluorescence intensity of intracellular Rhodamine 123 (Rh-123) were measured by FACS.Results. The expression levels of miR-27a and P-gp were up-regulated in paclitaxel-resistant ovarian cancer cell line A2780/Taxol as compared with its parental line A2780. Transfection of A2780/Taxol cells with the inhibitors of miR-27a decreased the expression of MDR1 mRNA and P-gp protein, increased HIPK2 protein expression, enhanced the sensitivity of A2780/taxol cells to paclitaxel, increased paclitaxel-induced apoptosis and the fluorescence intensity of intracellular Rh-123. Expression of MDR1 mRNA was increased while the sensitivity to paclitaxel was decreased in A2780 cells management with the mimics of miR-27a.Conclusions. The deregulation of miR-27a may be involved in the development of drug resistance, regulating the expression of MDR1/P-gp, at least in part, by targeting HIPK2 in ovarian cancer cells. (C) 2010 Elsevier Inc. All rights reserved.