Hettich Britta F, Ben-Yehuda Greenwald Maya, Werner Sabine, Leroux Jean-Christophe
Institute of Pharmaceutical Sciences Department of Chemistry and Applied Biosciences ETH Zurich Zurich 8093 Switzerland.
Institute of Molecular Health Sciences Department of Biology ETH Zurich Zurich 8093 Switzerland.
Adv Sci (Weinh). 2020 Oct 27;7(23):2002596. doi: 10.1002/advs.202002596. eCollection 2020 Dec.
Human mesenchymal stem cell exosomes have been shown to promote cutaneous wound healing. Their bioactivity is most often attributed to their protein and nucleic acid components, while the function of exosomal lipids remains comparatively unexplored. This work specifically assesses the involvement of lipids and the transmembrane enzyme CD73 in the exosomes' biological activity in stimulating the cutaneous wound healing process. Since exosome preparation processes are not harmonized yet, certain production and purification parameters are first systematically investigated, enabling the optimization of a standardized protocol delivering high exosome integrity, yield, and purity. An in situ enzymatic assay is introduced to specifically assess the vesicle functionality, and quantitative proteomics is employed to establish the cell culture conditions yielding a stable exosome protein profile. Using a combination of in vitro approaches, CD73 and constitutional lipids of HPV-16 E6/E7 transformed human bone marrow stromal cell-derived exosomes are identified as key bioactive components promoting the exosome-driven acceleration of processes required for wound repair. A pilot wound healing study in mice indeed suggests a role of lipids in the exosomes' biological activity. Strikingly, the extent of the bioactivity of these exosomal components is found to be dependent on the target cell type.
人骨髓间充质干细胞外泌体已被证明可促进皮肤伤口愈合。其生物活性通常归因于其蛋白质和核酸成分，而外泌体脂质的功能仍相对未被探索。这项工作专门评估脂质和跨膜酶CD73在外泌体刺激皮肤伤口愈合过程中的生物活性中的作用。由于外泌体制备过程尚未统一，因此首先系统地研究了某些生产和纯化参数，从而优化了一种标准化方案，可提供高外泌体完整性、产量和纯度。引入了一种原位酶测定法来专门评估囊泡功能，并采用定量蛋白质组学来确定产生稳定外泌体蛋白质谱的细胞培养条件。使用多种体外方法，CD73和人乳头瘤病毒16 E6/E7转化的人骨髓基质细胞衍生外泌体的组成脂质被确定为促进外泌体驱动的伤口修复所需过程加速的关键生物活性成分。在小鼠中进行的一项初步伤口愈合研究确实表明脂质在外泌体生物活性中起作用。令人惊讶的是，发现这些外泌体成分的生物活性程度取决于靶细胞类型。
