白头翁皂苷B4和PD-L1 siRNA共递送cRGD修饰靶向脂质体的制备及体外细胞摄取研究  

Preparation and cellular uptake study of anemoside B4 and PD-L1 siRNA co-delivered cRGD-modified targeting liposomes

作　　者：万安平 张婧 周雄 冯育林[1] 刘骏 何瑶 李翔[1] WAN Anping;ZHANG Jing;ZHOU Xiong;FENG Yulin;LIU Jun;HE Yao;LI Xiang(National Pharmaceutical Engineering Center for Solid Preparation in Chinese Herbal Medicine,Jiangxi University of Chinese Medicine,Nanchang 330006,China;Key Laboratory of Modern Preparation of TCM of Ministry of Education,Jiangxi University of Chinese Medicine,Nanchang 330004,China)

机构地区：[1]江西中医药大学中药固体制剂制造技术国家工程研究中心,南昌330006 [2]江西中医药大学现代中药制剂教育部重点实验室,南昌330004

出　　处：《中国药房》2023年第1期18-22,共5页China Pharmacy

基　　金：国家自然科学基金资助项目(No.82260695);江西省自然科学基金项目(No.20212ACB206004);江西中医药大学科技创新团队(No.CXTD22001);江西省青年井冈学者项目(No.QNJG2018077);大学生创新创业训练计划项目(No.双创院发[2022]2号)。

摘　　要：目的制备白头翁皂苷B4(AB4)和程序性死亡配体1(PD-L1)小干扰RNA(siP)共递送环精氨酰甘氨酸天冬氨酸序列(cRGD)修饰靶向脂质体(AB4/siP-c-L),并考察其体外细胞摄取行为。方法采用乙醇注入法制备cRGD修饰的AB4靶向脂质体(AB4-c-L),将AB4-c-L与20 nmol/L的siP等体积混合,通过静电吸附得到AB4/siP-c-L。分别采用激光散射粒径测定仪、透射电子显微镜、超滤离心法、透析法、琼脂糖凝胶电泳法考察AB4/siP-c-L的粒径、Zeta电位、形态、包封率、药物含量、体外释放行为、血清稳定性。分别采用流式细胞术和共聚焦激光扫描技术评价小鼠Lewis肺癌细胞LLC对AB4/siP-c-L的摄取及其在细胞内的分布情况。结果AB4/siP-c-L的平均粒径为(187.4±3.1)nm,Zeta电位为(33.5±1.4)mV,形态呈类球形,AB4的包封率和含量分别为(95.2±0.4)%、(1.0±0.2)mg/mL;AB4/siP-c-L可较好地包裹siP,并具有较好的血清稳定性、pH敏感性以及缓释特性。LLC细胞对AB4/siP-c-L的摄取率显著高于游离药物,并能实现细胞质内富集。结论AB4/siP-c-L可有效实现AB4与基因药物siP共载,且对LLC细胞具有一定的体外靶向性。OBJECTIVE To prepare anemoside B4(AB4)and programmed cell death ligand 1(PDL1)siRNA(siP)codelivered cRGD-modified targeting liposomes(AB4/siP-c-L),and to study the cellular uptake in vitro.METHODS The cRGDmodified AB4-loaded targeted liposomes(AB4-c-L)were prepared by ethanol injection.AB4-c-L was mixed with 20 nmol/L siP in the same volume and AB4/siP-c-L was obtained through electrostatic adsorption.The particle size,Zeta potential,morphology,encapsulation efficiency and drug content,in vitro release behavior and serum stability of AB4/siP-c-L were investigated by laser scattering particle size tester,transmission electron microscopy,ultrafiltration centrifugation,dialysis and agar-gel electrophoresis block test.Cellular uptake of AB4/siP-c-L by Lewis lung cancer cells LLC and its intracellular localization were evaluated by flow cytometry and confocal laser scan technique.RESULTS The average particle size of AB4/siP-c-L was(187.4±3.1)nm,and the Zeta potential was(33.5±1.4)mV.AB4/siP-c-L was spheroidal in shape.The encapsulation efficiency and content of AB4 were(95.2±0.4)%and(1.0±0.2)mg/mL,respectively.AB4/siP-c-L could better package siP,and exhibited good serum stability,obvious pH sensitivity and sustained release property.The uptake rate of AB4/siP-c-L by LLC cells was significantly higher than that of free drug,and was able to accumulate in cytoplasm.CONCLUSIONS AB4/siP-c-L can effectively realize the co-loading of AB4 and gene drug siP,which has certain in vitro targeting to LLC cells.

关 键 词：白头翁皂苷B4 程序性死亡配体1 小干扰RNA 脂质体 细胞摄取 

分 类 号：R943[医药卫生—药剂学]