Protein composition of extracellular vesicles from skin secretions of the amphibian Bombina maxima

DEAR EDITOR, Extracellular vesicles(EVs)are important for the transport of biologically active materials and for intercellular communication.As an exposed mucosa,amphibian skin participates in many essential physiological processes.To date,however,little is known about EVs in amphibian skin.Here,we successfully isolated EVs from the skin secretions of Bombina maxima,and characterized the EVs using nanoparticle tracking,western blotting,and electron microscopy.Proteomic analysis using liquid chromatography-tandem mass spectrometry identified 131 proteins in the skin secretion-derived EVs,which were enriched in biological processes and molecular functions such as immune stress,metabolism,oxidoreduction,and transport.Non-EV skin secretions were also analyzed,with 313 proteins(not in EVs)identified in these samples.Comparative analysis revealed that 62 proteins were unique to EVs,244 proteins were unique to non-EVs,and 69 proteins were shared between EVs and non-EVs.Flow cytometry showed that the isolated EVs could be taken up by B.maxima skin-derived fibroblasts.These findings provide the first evidence that EVs containing bioactive molecules are secreted from amphibian skin,suggesting that EVs may play important physiological roles in amphibian skin.