SP94修饰的肝靶向阳离子基因载体的制备及其体外评价

发布时间：2018-05-02 17:32

本文选题：适配体 + 聚阳离子载体　；参考：《中国医院药学杂志》2017年10期

【摘要】：目的:研究适配体SP94修饰的阳离子基因载体H_3R_5,合成新型肝靶向纳米复合物SP94-H_3R_5/miR195,增加对肝癌细胞的靶向性,提高基因的转染效率。方法:经半胱氨酸修饰的SP94与H_3R_5末端的半胱氨酸发生氧化反应,组装得到SP94-H_3R_5,利用~1 H-NMR鉴定SP94-H_3R_5载体的结构,通过电位粒度仪测定纳米复合物的电位和粒径,利用琼脂糖凝胶电泳考察载体对miR195的压缩能力。以体外培养的人肝癌SK-Hep-1为研究对象,CCK8法检测SP94-H_3R_5和H_3R_5对细胞增殖的抑制作用,采用激光共聚焦显微镜考察肝癌细胞对纳米复合物的摄取,以pEGFP为报告基因考察基因转染效率,Western blot实验检测SK-Hep-1细胞VEGF的蛋白表达。结果:SP94-H_3R_5具有生物相容性,可以压缩miR195形成稳定的纳米复合物,SP94-H_3R_5/miR195与H_3R_5/miR195相比可以更多地被SK-Hep-1摄取(P0.01),SP94-H_3R_5转染效率高于非靶向载体H_3R_5,对VEGF的阻滞作用也更高(P0.01)。结论:SP94-H_3R_5兼具纳米材料的被动靶向作用和适配体的主动靶向作用,有潜力成为肝癌治疗中的新型载体。
[Abstract]:Aim: to study the cationic gene vector H3RSTE5 modified by aptamer SP94 and synthesize a novel liver targeting nanocomplex SP94-H3R5 / miR195 to increase the targeting of hepatoma cells and improve the efficiency of gene transfection. Methods: the cysteine modified SP94 reacted with cysteine at the end of H_3R_5, and SP94-H3R5 was assembled. The structure of SP94-H_3R_5 carrier was identified by H-NMR. The potential and particle size of the nanocomposites were measured by potentiometric particle size analyzer. Agarose gel electrophoresis was used to investigate the compression ability of the carrier to miR195. The inhibitory effects of SP94-H_3R_5 and H_3R_5 on the proliferation of human hepatocellular carcinoma (SK-Hep-1) in vitro were detected by CCK8 method. The uptake of nano-complexes in hepatoma cells was investigated by confocal laser microscopy. The efficiency of gene transfection was evaluated by using pEGFP as reporter gene. Western blot assay was used to detect the protein expression of VEGF in SK-Hep-1 cells. Results compared with H_3R_5/miR195, the transfection efficiency of S / SP94-H3RS5 was higher than that of H_3R_5/miR195, and the transfection efficiency was higher than that of non-target carrier HNS3R5. The effect on VEGF was also higher than that on VEGF. Conclusion Snx SP94-H3RSP 5 has both passive targeting effect of nano-materials and active targeting of aptamers, and it has the potential to become a new type of carrier in the treatment of liver cancer.
【作者单位】：上海交通大学附属第一人民医院临床药学科;第二军医大学长海医院药学部;
【基金】：国家自然科学基金资助项目(编号:81302212) 上海市自然科学基金资助项目(编号:16ZR1428000) 上海交通大学医工交叉资助项目(编号:YG2014MS32,YG2015QN14)
【分类号】：R450;R735.7

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