A Simple and Quick Method for Loading Proteins in Extracellular Vesicles.
A Simple and Quick Method for Loading Proteins in Extracellular Vesicles.
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一种简单而快速的方法，用于在细胞外囊泡中加载蛋白质。
DOI：
10.3390/ph14040356
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发表时间：
2021-04-13
期刊：
影响因子：
--
通讯作者：
Wolfram J
中科院分区：
文献类型：
--
作者：
Busatto S;Iannotta D;Walker SA;Di Marzio L;Wolfram J
细胞外囊泡（EV）介导体内生物分子货物的细胞间转运，使其成为生物活性化合物的有前途的递送载体。生产细胞的基因工程已经使得能够在EV中包封治疗性蛋白质。然而，基因工程方法可能是昂贵的、耗时的，并且与某些EV来源（例如人血浆和牛乳）不相容。本研究的目的是开发一种快速，通用，简单的方法，用于在分离后的EV中加载蛋白质。包括CRISPR相关蛋白9（Cas9）在内的蛋白质与阳离子脂质结合，阳离子脂质通过被动孵育与MDA-MB-231细胞衍生的EV进一步复合。使用尺寸排阻色谱法除去未与EV复合的组分。将EV介导蛋白质的细胞内递送的能力与常规方法如电穿孔和商业蛋白质转染试剂进行比较。结果表明，EV在蛋白负载后保留了天然特征，并获得与常规方法相似的细胞内蛋白递送水平，但显示出较低的毒性。这种方法为快速探索用于蛋白质递送的EV开辟了机会。
Extracellular vesicles (EVs) mediate intercellular transport of biomolecular cargo in the body, making them promising delivery vehicles for bioactive compounds. Genetic engineering of producer cells has enabled encapsulation of therapeutic proteins in EVs. However, genetic engineering approaches can be expensive, time-consuming, and incompatible with certain EV sources, such as human plasma and bovine milk. The goal of this study was to develop a quick, versatile, and simple method for loading proteins in EVs post-isolation. Proteins, including CRISPR associated protein 9 (Cas9), were bound to cationic lipids that were further complexed with MDA-MB-231 cell-derived EVs through passive incubation. Size-exclusion chromatography was used to remove components that were not complexed with EVs. The ability of EVs to mediate intracellular delivery of proteins was compared to conventional methods, such as electroporation and commercial protein transfection reagents. The results indicate that EVs retain native features following protein-loading and obtain similar levels of intracellular protein delivery as conventional methods, but display less toxicity. This method opens up opportunities for rapid exploration of EVs for protein delivery.
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DOI：
10.1016/j.jconrel.2015.07.030
发表时间：
2015-12-10
期刊：
Journal of controlled release : official journal of the Controlled Release Society
影响因子：
--
作者：
Batrakova EV;Kim MS
通讯作者：
Kim MS
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影响因子：
16
作者：
Aspe JR;Diaz Osterman CJ;Jutzy JM;Deshields S;Whang S;Wall NR
通讯作者：
Wall NR
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DOI：
10.1016/j.jconrel.2015.03.033
发表时间：
2015-06-10
期刊：
Journal of controlled release : official journal of the Controlled Release Society
影响因子：
--
作者：
Haney MJ;Klyachko NL;Zhao Y;Gupta R;Plotnikova EG;He Z;Patel T;Piroyan A;Sokolsky M;Kabanov AV;Batrakova EV
通讯作者：
Batrakova EV
影响因子：
5.8
作者：
Fukuhara T;Tani H;Shiokawa M;Goto Y;Abe T;Taketomi A;Shirabe K;Maehara Y;Matsuura Y
通讯作者：
Matsuura Y
影响因子：
10.2
作者：
Busatto S;Yang Y;Walker SA;Davidovich I;Lin WH;Lewis-Tuffin L;Anastasiadis PZ;Sarkaria J;Talmon Y;Wurtz G;Wolfram J
通讯作者：
Wolfram J