Xin Xin, Ingrid Dijkgraaf, Tilman M. Hackeng, Rory R. Koenen
{"title":"冻干血小板衍生的细胞外囊泡类似物的生成和表征","authors":"Xin Xin, Ingrid Dijkgraaf, Tilman M. Hackeng, Rory R. Koenen","doi":"10.1016/j.vesic.2025.100095","DOIUrl":null,"url":null,"abstract":"<div><div>Platelet-derived extracellular vesicles (P-EV) are thought to facilitate the transfer of information from platelets to target cells, playing a role in both physiologic and pathophysiologic processes, particularly in regulating immune responses and healing processes. In addition, P-EV show promise as drug carriers and biomarkers for disease. However, the procedures for isolation, purification and fluorescent labeling of P-EV remain unstandardized. Moreover, the requirement to use freshly obtained platelets for generating EV presents a logistical challenge for their study. In this study, we isolated, characterized, and compared P-EV analogues by sonication of freshly obtained and lyophilized platelets, investigated fluorescent labeling methods, and monitored cellular uptake. We found that P-EV analogues derived from fresh or lyophilized platelets showed similar characteristics regarding size, surface proteins and content. Among the fluorescent labeling methods, CFSE and DiO-C6 were most effective in labeling P-EV analogues from both fresh and lyophilized platelets. All labeling methods led to an increase in P-EV analogue's size, with CFSE and DiO-C6 resulting in the smallest increase. The addition of P-EV analogues to cultured immortal endothelial cells revealed that P-EV analogues were effectively internalized and directed to the lysosomal compartment. The results indicate that P-EV analogues from lyophilized platelets have similar functional properties as those from freshly isolated platelets and these are retained after labeling with CFSE. Thus, lyophilized platelets can serve as a source of P-EV analogues for functional studies.</div></div>","PeriodicalId":73007,"journal":{"name":"Extracellular vesicle","volume":"6 ","pages":"Article 100095"},"PeriodicalIF":0.0000,"publicationDate":"2025-09-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Generation and characterization of platelet-derived extracellular vesicle analogues from lyophilized platelets\",\"authors\":\"Xin Xin, Ingrid Dijkgraaf, Tilman M. Hackeng, Rory R. Koenen\",\"doi\":\"10.1016/j.vesic.2025.100095\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><div>Platelet-derived extracellular vesicles (P-EV) are thought to facilitate the transfer of information from platelets to target cells, playing a role in both physiologic and pathophysiologic processes, particularly in regulating immune responses and healing processes. In addition, P-EV show promise as drug carriers and biomarkers for disease. However, the procedures for isolation, purification and fluorescent labeling of P-EV remain unstandardized. Moreover, the requirement to use freshly obtained platelets for generating EV presents a logistical challenge for their study. In this study, we isolated, characterized, and compared P-EV analogues by sonication of freshly obtained and lyophilized platelets, investigated fluorescent labeling methods, and monitored cellular uptake. We found that P-EV analogues derived from fresh or lyophilized platelets showed similar characteristics regarding size, surface proteins and content. Among the fluorescent labeling methods, CFSE and DiO-C6 were most effective in labeling P-EV analogues from both fresh and lyophilized platelets. All labeling methods led to an increase in P-EV analogue's size, with CFSE and DiO-C6 resulting in the smallest increase. The addition of P-EV analogues to cultured immortal endothelial cells revealed that P-EV analogues were effectively internalized and directed to the lysosomal compartment. The results indicate that P-EV analogues from lyophilized platelets have similar functional properties as those from freshly isolated platelets and these are retained after labeling with CFSE. Thus, lyophilized platelets can serve as a source of P-EV analogues for functional studies.</div></div>\",\"PeriodicalId\":73007,\"journal\":{\"name\":\"Extracellular vesicle\",\"volume\":\"6 \",\"pages\":\"Article 100095\"},\"PeriodicalIF\":0.0000,\"publicationDate\":\"2025-09-12\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Extracellular vesicle\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/S2773041725000332\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"\",\"JCRName\":\"\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Extracellular vesicle","FirstCategoryId":"1085","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S2773041725000332","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
Generation and characterization of platelet-derived extracellular vesicle analogues from lyophilized platelets
Platelet-derived extracellular vesicles (P-EV) are thought to facilitate the transfer of information from platelets to target cells, playing a role in both physiologic and pathophysiologic processes, particularly in regulating immune responses and healing processes. In addition, P-EV show promise as drug carriers and biomarkers for disease. However, the procedures for isolation, purification and fluorescent labeling of P-EV remain unstandardized. Moreover, the requirement to use freshly obtained platelets for generating EV presents a logistical challenge for their study. In this study, we isolated, characterized, and compared P-EV analogues by sonication of freshly obtained and lyophilized platelets, investigated fluorescent labeling methods, and monitored cellular uptake. We found that P-EV analogues derived from fresh or lyophilized platelets showed similar characteristics regarding size, surface proteins and content. Among the fluorescent labeling methods, CFSE and DiO-C6 were most effective in labeling P-EV analogues from both fresh and lyophilized platelets. All labeling methods led to an increase in P-EV analogue's size, with CFSE and DiO-C6 resulting in the smallest increase. The addition of P-EV analogues to cultured immortal endothelial cells revealed that P-EV analogues were effectively internalized and directed to the lysosomal compartment. The results indicate that P-EV analogues from lyophilized platelets have similar functional properties as those from freshly isolated platelets and these are retained after labeling with CFSE. Thus, lyophilized platelets can serve as a source of P-EV analogues for functional studies.