Amphibian thrombocyte-derived extracellular vesicles, including microRNAs, induce angiogenesis-related genes in endothelial cells.

Thrombocytes circulate in the blood of nonmammalian vertebrates and are involved in hemostasis; however, many detailed characteristics of thrombocytes remain unclear. Recently, we established an amphibian thrombocyte cell line. Here, we report the finding that thrombocytes produce integrin alpha IIb (CD41)-positive extracellular vesicles (EVs), which include microRNAs (miRs). Flow cytometric analysis showed the expression of CD41 and phosphatidylserine on the surface of EVs. Nanotracking analysis showed that these CD41 EVs were approximately 100 nm in diameter. As CD41 EVs were also observed from African clawed frogs, the production of CD41 EVs might be common to amphibians. Microarray analysis showed that the CD41 EVs contain many kinds of miRs. These CD41 EVs were phagocytosed by endothelial cells and macrophages. qPCR analysis showed that many angiogenesis-related genes were up-regulated in CD41 EV-treated endothelial cells. Over-expression of some miRs in the CD41 EVs increased the proliferation of endothelial cells. These results indicated that thrombocytes produced CD41 EVs, including miRs, that were received by endothelial cells to induce the expression of angiogenesis-related genes. These results indicated that the CD41 EVs produced from thrombocytes act as signaling molecules to repair damaged blood vessels.