GGI/MDR1 siRNA逆转A549/DDP细胞多药耐药的研究  

GGI as a gene carrier delivering MDR1 siRNA to A549/DDP cells for reversal of multidrug resistance

作　　者：王子瑞[1] 白凤[1] 张小英[1] 吴佳敏[1] 郭羚[1] 利智 冯敏[1] 

机构地区：[1]中山大学药学院,广东广州510006

出　　处：《药学学报》2017年第2期309-317,共9页Acta Pharmaceutica Sinica

基　　金：国家自然科学基金资助项目(81373332);广东省自然科学基金资助项目(2014A030313155)

摘　　要：通过RNA干扰(RNAi)技术靶向耐药性人肺腺癌细胞内的MDR1基因实现多药耐药性的逆转作用。构建聚乙二醇-聚谷氨酸-聚乙烯亚胺(PEG-b-PLG-g-PEIs,GGI)阳离子聚合物载体,通过~1H NMR表征确认结构,动态光散射法测定GGI/si RNA的粒径及电位,并以A549敏感株和A549/DDP耐药株细胞系为模型,采用MTT法考察GGI的细胞毒性,以流式细胞术评价新型聚合物载体GGI递送si RNA的效率和强度,RT-PCR法考察GGI转染A549和A549/DDP(cisplatin)后MDR1 m RNA水平的表达;Western blot法测定A549/DDP细胞中P-gp的表达。同时,以MTT法和Annexin V-FITC/PI双染法考察si RNA干扰后抗肿瘤药物对药物敏感性的变化。结果显示,GGI/si RNA复合物粒径为150~200 nm,电位稳定在16~28 m V。GGI细胞毒性远低于PEI 25K,且具有更高运载si RNA至细胞内的效率,并能极大降低A549/DDP细胞内MDR1 m RNA和P-gp的表达,能更大程度地增强耐药细胞株对顺铂的敏感性,说明GGI有望在基因传递系统中成为一种新型的聚合物载体并广泛应用。This study was designed to reverse multidrug resistance of lung cancer cells by downregulating MDR1 genes through RNA interference（RNAi） technology. A novel biodegradable cationic polymer（PEG-b- PLG-g-PEIs,GGI） was synthesized and characterized by ~1H NMR. The particle size and zeta potential were measured by dynamic light scattering（DLS）. The cell viability profile of GGI was tested by MTT method with both A549 and A549/DDP cell lines. Flow cytometry（FCM） technology was used to investigate the efficiency and intensity of delivering si RNA to cells by GGI polymer. RT-PCR and Western blot were used to detect the m RNA and P-gp expression after GGI/MDR1 si RNA transfection assay. The sensitivity of cisplatin administration after transfecting GGI/MDR1 si RNA polyplexs was performed with MTT and Annexin V-FITC/PI methods. The results suggest that the particle size and zeta potential of GGI/si RNA were 150-200 nm and 16-28 m V. GGI exhibited a lower cell cytotoxity than PEI 25 K and higher efficiency of delivering si RNA,which dramatically decreased the expression of MDR1 m RNA and P-gp of A549/DDP cells and increased much sensitivity to cisplatin in A549/DDP cells. GGI holds a great potential in gene delivery as a novel cationic polymer for further investigation.

关 键 词：基因传递系统 RNA干扰 多药耐药性 MDR1 SIRNA A549/DDP细胞 

分 类 号：R943[医药卫生—药剂学]