白头翁皂苷B4和PD-L1siRNA共递送cRGD修饰靶向脂质体的制备及体外细胞摄取研究
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篇名: 白头翁皂苷B4和PD-L1siRNA共递送cRGD修饰靶向脂质体的制备及体外细胞摄取研究
TITLE: Preparation and cellular uptake study of anemoside B4 and PD-L1 siRNA co-delivered cRGD-modified targeting liposomes
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20nmol/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。
ABSTRACT: OBJECTIVE To prepare anemoside B4 (AB4) and programmed cell death ligand 1 (PDL1) siRNA (siP) co- delivered cRGD-modified targeting liposomes (AB4/siP-c-L), and to study the cellular uptake in vitro. METHODS The cRGD- modified 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.
期刊: 2023年第34卷第01期
作者: 万安平;张婧;周雄;冯育林;刘骏;何瑶;李翔
AUTHORS: WAN Anping,ZHANG Jing,ZHOU Xiong,FENG Yulin,LIU Jun,HE Yao,LI Xiang
关键字: 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
KEYWORDS: anemoside B4; PD-L1; siRNA; liposome; cellular uptake
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