Ultrasound microbubble-mediated delivery of the siRNAs targeting MDR1 reduces drug resistance of yolk sac carcinoma L2 cells.
Ultrasound microbubble-mediated delivery of the siRNAs targeting MDR1 reduces drug resistance of yolk sac carcinoma L2 cells.
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DOI：
10.1186/1756-9966-30-104
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发表时间：
2011-10-28
期刊：
影响因子：
--
通讯作者：
Wei G
中科院分区：
文献类型：
--
作者：
He Y;Bi Y;Hua Y;Liu D;Wen S;Wang Q;Li M;Zhu J;Lin T;He D;Li X;Wang Z;Wei G
MDR 1基因编码的P-糖蛋白是一种ATP依赖的药物外排转运蛋白，与卵黄囊癌的耐药有关。超声微泡介导的基因递送技术是一种新型有效的基因递送方法。我们推测，微泡联合超声介导的靶向MDR 1基因的siRNA（siMDR 1）可以在体外下调MDR 1的表达，提高卵黄囊癌对化疗药物的敏感性。构建了逆转录病毒敲除载体pSEB-siMDR 1 s，并进行了序列验证。将所得pSEB-siMDR 1质粒DNA用脂质微泡包裹，加入培养细胞中时，超声触发DNA释放。通过流式细胞术计数GFP阳性细胞以确定转染效率。采用实时荧光定量PCR和Western blot检测MDR 1 mRNA和蛋白的表达。P-糖蛋白的功能和药物敏感性进行了分析，柔红霉素积累和MTT法。超声微泡介导的pSEB-siMDR 1 DNA转染大鼠卵黄囊癌L2（L2-RYC）细胞的效率显著提高。超声微泡介导的siMDR 1 s递送在mRNA和蛋白水平上有效地抑制MDR 1表达，并降低P-糖蛋白功能。沉默MDR 1导致细胞活力和阿曲斯汀和更生霉素的IC 50降低。我们的研究结果表明，超声微泡介导的MDR 1 siRNA在L2-RYC细胞中是安全有效的。MDR 1基因沉默导致L2-RYC细胞P-糖蛋白活性降低，耐药性降低，有望成为基因与化疗联合治疗卵黄囊癌的新途径。
MDR1 gene encoding P-glycoprotein is an ATP-dependent drug efflux transporter and related to drug resistance of yolk sac carcinoma. Ultrasound microbubble-mediated delivery has been used as a novel and effective gene delivery method. We hypothesize that small interfering RNA (siRNA) targeting MDR1 gene (siMDR1) delivery with microbubble and ultrasound can down-regulate MDR1 expression and improve responsiveness to chemotherapeutic drugs for yolk sac carcinoma in vitro. Retroviral knockdown vector pSEB-siMDR1s containing specific siRNA sites targeting rat MDR1 coding region were constructed and sequence verified. The resultant pSEB-siMDR1 plasmids DNA were encapsulated with lipid microbubble and the DNA release were triggered by ultrasound when added to culture cells. GFP positive cells were counted by flow cytometry to determine transfection efficiency. Quantitative real-time PCR and western blot were performed to determine the mRNA and protein expression of MDR1. P-glycoprotein function and drug sensitivity were analyzed by Daunorubicin accumulation and MTT assays. Transfection efficiency of pSEB-siMDR1 DNA was significantly increased by ultrasound microbubble-mediated delivery in rat yolk sac carcinoma L2 (L2-RYC) cells. Ultrasound microbubble-mediated siMDR1s delivery effectively inhibited MDR1 expression at both mRNA and protein levels and decreased P-glycoprotein function. Silencing MDR1 led to decreased cell viability and IC50 of Vincristine and Dactinomycin. Our results demonstrated that ultrasound microbubble-mediated delivery of MDR1 siRNA was safe and effective in L2-RYC cells. MDR1 silencing led to decreased P-glycoprotein activity and drug resistance of L2-RYC cells, which may be explored as a novel approach of combined gene and chemotherapy for yolk sac carcinoma.
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DOI：
10.1186/1756-9966-29-152
发表时间：
2010-11-23
期刊：
Journal of experimental & clinical cancer research : CR
影响因子：
--
作者：
Chen ZY;Liang K;Qiu RX
通讯作者：
Qiu RX
影响因子：
4.4
作者：
Peignan, Lilia;Garrido, Wallys;Quezada, Claudia
通讯作者：
Quezada, Claudia
超声微泡增强器转染野生型p53质粒转染视网膜母细胞瘤细胞和组织的实验研究
影响因子：
1.6
作者：
Luo, J.;Zhou, X.;Wang, Z.
通讯作者：
Wang, Z.
影响因子：
3.7
作者：
Caffrey DR;Zhao J;Song Z;Schaffer ME;Haney SA;Subramanian RR;Seymour AB;Hughes JD
通讯作者：
Hughes JD
影响因子：
1.1
作者：
Merchant, Azher;Stewart, Rosalyn W.
通讯作者：
Stewart, Rosalyn W.