Yu Ya-Hui, Lyu Tian-Tian, Xue Bing-Quan, Yu Hai-Yan, Huang Hai-Ying
School of Pharmacy, Henan University of Chinese Medicine Zhengzhou 450046, China.
Zhongguo Zhong Yao Za Zhi. 2021 Jun;46(11):2824-2829. doi: 10.19540/j.cnki.cjcmm.20210303.301.
A drug delivery system of forsythoside A-loaded exosomes(FTA-Exos) with high biocompatibility and low immunogenicity was established to investigate its impact on the migration of human lung epithelial adenocarcinoma A549 cells. The exosomes from A549 cells were extracted and purified by ultra-high speed centrifugation and ultrafiltration. FTA-Exos were prepared by ultrasonic incubation, and characterized by particle size analysis, transmission electron microscopy, and Western blot assay. The uptake of FTA-Exos by A549 cells was observed under the laser confocal microscope, and the impact of FTA-Exos on the migration of A549 cells was investigated by cell scratch assay. The results showed that the average particle size of the prepared FTA-Exos was(138.90±2.37) nm, which increased slightly after drug loading. The PDI was 0.291±0.013, and the average potential was(-10.1±0.66) mV. The FTA-Exos were spheroidal in appearance as observed by transmission electron microscope, with an obvious saucer-like double-layer membrane. Western blot assay indicated that the specific proteins CD63 and Alix were both expressed in exosomes. The laser confocal microscopy suggested that FTA-Exos were taken up by A549 cells and stably maintained in the cell for 4-8 h, and the fluorescence was significantly enhanced at 4 h. The scratch assay showed that the inhibitory effect of FTA-Exos on the migration of A549 cells was significantly stronger than that of forsythoside A(P < 0.05). In conclusion, the drug delivery system of FTA-Exos established in this study had good stability, reliable preparation process, and potent inhibitory effect on the migration of A549 cells in vitro, which can provide an important reference for subsequent in-depth research and application.
建立了一种具有高生物相容性和低免疫原性的载连翘酯苷A外泌体(FTA-Exos)药物递送系统,以研究其对人肺上皮腺癌A549细胞迁移的影响。通过超速离心和超滤从A549细胞中提取并纯化外泌体。通过超声孵育制备FTA-Exos,并通过粒度分析、透射电子显微镜和蛋白质免疫印迹分析进行表征。在激光共聚焦显微镜下观察A549细胞对FTA-Exos的摄取,并通过细胞划痕试验研究FTA-Exos对A549细胞迁移的影响。结果显示,制备的FTA-Exos平均粒径为(138.90±2.37)nm,载药后略有增加。PDI为0.291±0.013,平均电位为(-10.1±0.66)mV。透射电子显微镜观察显示,FTA-Exos外观呈球形,具有明显的碟状双层膜。蛋白质免疫印迹分析表明,外泌体中特异性蛋白CD63和Alix均有表达。激光共聚焦显微镜显示,FTA-Exos被A549细胞摄取并在细胞内稳定维持4-8小时,4小时时荧光显著增强。划痕试验表明,FTA-Exos对A549细胞迁移的抑制作用明显强于连翘酯苷A(P<0.05)。综上所述,本研究建立的FTA-Exos药物递送系统具有良好的稳定性、可靠的制备工艺,对体外A549细胞迁移具有较强的抑制作用,可为后续深入研究和应用提供重要参考。