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. 2016 Mar 16;25(10):774–787. doi: 10.1089/scd.2016.0009

FIG. 4.

FIG. 4.

DC-derived EV are internalized by DC in an autocrine/paracrine manner mediated by EV surface CTLA-4. (A) Staining pattern of CFSE-labeled DC indicating some colocalization with CTLA-4+ structures. (B) Cultured supernatants from CFSE-loaded DC were subsequently depleted of all cells and incubated with unlabeled DC for various time points. Recipient, unlabeled DC could be visualized binding and (C) internalizing CFSE+ microvesicles. (D) Cultured CFSE-loaded DC supernatants were incubated with protein G-plus beads coated with various concentrations of αCTLA-4, and treated supernatants were subsequently incubated with unlabeled DC for 6 h at 37°C before flow cytometric analysis of CFSE+ microvesicle uptake. (E) Recipient DC were also analyzed for their ability to still bind αCD86 and αCD80 antibodies after 6 and 12 h incubations with CFSE+ CTLA-4+ microvesicles. A significant log-fold decrease in B7 expression was apparent among DC that internalized CFSE+ microvesicles. This decrease was specific to B7 and not observed among other markers such as CD11c. Error bars at 6 and 12 h = ±SD of four independent experiments. *P < 0.05.