Purpose

Exosomes have the potential to create new therapeutics as drug loading carriers. Their important inherent roles are stem from structures (also known natural liposomes) and strong biological stability in many cellular processes. The formation of a new drug delivery system in cervical cancer by loading a chemotherapeutic agent docetaxel into exosomes obtained from HELA cells is intended.

Methods

In this study, exosomes were isolated from HELA cells by ultracentrifugation method. Exosome membrane proteins CD9 and CD63 were analyzed by western blotting. Docetaxel was loaded into the exosomes by electroporation. Structures and sizes of docetaxel-loaded exosomes (Exo-Doc) were analyzed by scanning electron microscopy and dynamic light scattering technique. Cell viability with MTT assay and cell migration with scratch assay were measured. Apoptotic changes were measured by Annexin-V binding, Bax, Bcl-2, Caspase-3 protein, and gene expressions by western blotting and qPCR.

Results

It was observed that cell viability and cell migration were decreased, and Annexin-V binding increased after 1µg/mL Exo-Doc application to HELA cells by 24-h incubation. Bax and caspase-3 protein and gene expression were increased and Bcl-2 protein and gene expressions were decreased in cells. Exo-Doc induced mitochondrial apoptosis in HELA cells.

Conclusion

As a result, docetaxel-loaded HELA-derived exosome was generated as a new drug carrier and delivery system which was used to treat the cell itself with low doses of chemotherapeutic. Exo-Doc may be preferred for treating cervical patients in the clinic due to its low toxicity.

中文翻译：

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从 HELA 衍生的外泌体开发新的药物递送系统以及负载多西紫杉醇的外泌体对线粒体凋亡的影响

目的

外泌体有潜力创造新的疗法作为载药载体。它们重要的内在作用源于许多细胞过程中的结构（也称为天然脂质体）和强大的生物稳定性。通过将化疗药物多西紫杉醇加载到从 HELA 细胞获得的外泌体中，在宫颈癌中形成一种新的药物递送系统。

方法

在这项研究中，外泌体是通过超速离心法从 HELA 细胞中分离出来的。通过蛋白质印迹分析外泌体膜蛋白 CD9 和 CD63。通过电穿孔将多西紫杉醇加载到外泌体中。通过扫描电子显微镜和动态光散射技术分析了负载多西紫杉醇的外泌体 (Exo-Doc) 的结构和大小。用 MTT 分析测量细胞活力，用划痕分析测量细胞迁移。通过膜联蛋白-V 结合、Bax、Bcl-2、Caspase-3 蛋白和蛋白质印迹和 qPCR 的基因表达来测量细胞凋亡变化。

结果

观察到细胞活力和细胞迁移降低，并且在将 1μg/mL Exo-Doc 应用到 HELA 细胞 24 小时后，膜联蛋白-V 结合增加。细胞中Bax和caspase-3蛋白和基因表达增加，Bcl-2蛋白和基因表达降低。Exo-Doc 在 HELA 细胞中诱导线粒体凋亡。

结论

结果，产生了负载多西紫杉醇的 HELA 衍生的外泌体，作为一种新的药物载体和递送系统，用于用低剂量的化疗药物治疗细胞本身。由于其低毒性，Exo-Doc 可能是临床治疗宫颈癌患者的首选。