Mature exosomes induce strong T-cell activation in vivo and fast skin graft rejection. (A) Exosomes (15 μg) were injected subcutaneously into female C57Bl/6 mice containing adoptively transferred CFSE-labeled, CD45.1⁺ Marilyn lymph node cells. Draining lymph node cells were analyzed by FACS 4 days after injection. CFSE staining of Marilyn cells, identified as CD4⁺CD45.1⁺, from mice treated with 15 μg of HY-free mature exosomes (Control Exo), HY-bearing immature exosomes (Imm HY Exo), or HY-bearing mature exosomes (Mat HY Exo). The percentage of cells that have undergone more than 5 divisions cycles is shown. One representative of 5 independent experiments is shown. (B) Percent of Marylin T cells that underwent more than 5 division cycles, upon injection of various doses of control (▵), immature (⋄), or mature (▪) exosomes. Results obtained from 8 mice per condition, in 3 to 5 independent experiments, were pooled. (C) Male skin grafts were transplanted onto female recipients that had been previously injected with PBS (▴), 5 μg immature HY-bearing exosomes (⋄), or 5 μg mature HY-bearing exosomes (▪). Graft rejection was examined every other day. Results obtained from 9 to 11 mice per group, in 4 independent experiments, are shown. Differences observed between mature exosome–treated mice and control group, and mature exosome–treated mice and immature exosome–treated mice are significant in Log rank test (P<.001 and P=.01 respectively).