乳铁蛋白修饰的长循环脂质体对肝癌细胞的体外靶向性评价  

In vitro targeting effect of lactoferrin modified PEGylated liposomes for hepatoma cells

作　　者：韦敏燕[1,2] 邹奇[2] 吴传斌[2] 徐月红[2] 

机构地区：[1]广州医科大学药学院,广东广州511436 [2]中山大学药学院,广东广州510006

出　　处：《药学学报》2015年第10期1272-1279,共8页Acta Pharmaceutica Sinica

基　　金：国家自然科学基金资助项目(81402881);广东省自然科学基金资助项目(S2013040014348);广州医科大学博士启动资助项目(2012C74)

摘　　要：本研究构建了包载香豆素-6荧光探针的乳铁蛋白修饰肝癌靶向脂质体(Lf-PLS),并对其体外肝癌靶向性进行评价。采用薄膜分散法与后插入法制备长循环脂质体(PLS)。乳铁蛋白(Lf)的氨基与脂质体膜表面DSPE-PEG2000-COOH共价结合,组装得到Lf-PLS,并进行表征。采用激光共聚焦显微镜与流式细胞术,考察无唾液酸糖蛋白受体(ASGPR)阳性细胞Hep G2对Lf-PLS的摄取。所制得包载香豆素-6的Lf-PLS为圆形或椭圆形囊泡,粒径约为130 nm,zeta电位约为-30 m V,包封率>80%。细胞摄取实验结果表明,Hep G2细胞对Lf-PLS的摄取显著高于PLS,且随孵育浓度、时间及温度的增加而增大。该摄取可被大量游离的Lf所抑制。结果提示,经Lf修饰后Lf-PLS对Hep G2细胞具有较好的靶向性。其靶向作用可能是Lf与Hep G2细胞膜蛋白ASGPR相结合,Lf-PLS通过受体介导的主动转运进入肝癌Hep G2细胞。上述体外实验证明,Lf-PLS是一个具有潜在肝癌靶向性的药物传递系统,为进一步探索该脂质体在体内的肝癌靶向性、抗肿瘤活性和代谢规律提供了实验基础。A lactoferrin-containing PEGylated liposome system （Lf-PLS） was developed and tested in vitro as a hepatoma-targeting drug delivery system. PEGylated liposomes （PLS） were successfully prepared using the thin film hydration method with peglipid post insertion. Lf was covalently conjugated onto the carboxyl terminal of DSPE-PEG200o-COOH on liposomes. Coumarin-6 was used to trace Lf-PLS with fluorescence. The cellular uptake of this system was carried out in asialoglycoprotein receptor （ASGPR） positive HepG2 cells via confocal microscopy and flow cytometry. The Lf-PLS liposome was observed as spherical or oval vesicles with the particle size around 130 nm, zeta potential about -30 mV and encapsulation efficiency more than 80%. The confocal microscopy images and flow cytometry data demonstrated that Lf-PLS resulted in significantly higher cell association by ASGPR positive HepG2 cells compared to PLS. The association between Lf-PLS and cells were dependent on the concentration, time and temperature, which was inhibited by pre-incubation with excessive free Lf. The results suggest that Lf-PLS has a good targeting effect on HepG2 cells in vitro. The targeting mechanism may be related to the specific binding of Lf and ASGPR on HepG2 cells, which guidesLf-PLS to the cell surface to induce an active endocytosis process. All these results demonstrated that Lf-PLS might be a potential drug delivery system in targeting hepatocellular carcinoma, which deserves more research on its targeting ability, antitumor efficiency, and metabolism in vivo for treatment of hepatomacellular carcinoma.

关 键 词：乳铁蛋白 无唾液酸糖蛋白受体 肝癌 长循环脂质体 主动靶向 

分 类 号：R943[医药卫生—药剂学]