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

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.