Figure 3.

Melanoma-derived extracellular vesicles (EVs) suppress dendritic cell (DC) maturation in vitro. CD14+ human monocytes were matured to DC fate with CD40L in the absence or presence of SKMEL28 melanoma EVs, empty liposomes, or PBS. On day 7, phenotyping by flow cytometry evaluated DC expression of MHC molecules (HLA Class I, HLA-DR) and costimulatory molecules (CD83 and CD86) by histogram or the percent positive cells (mean ± SEM) (n = 8) (A,B). Representative image of reduced CD86 expression by imaging flow cytometry in DCs cocultured with fluorescently labeled melanoma EVs (n = 3) (C). Multiplex results of four analytes significantly decreased (Flt3L, IL-15, MIP-1α, and MIP-1β) in the presence of melanoma EVs compared to liposome-treated control (n = 5) (D). Cytokine and chemokine concentrations were reported in picogram per milliliter (mean ± SEM). Statistical significance was determined by two-tailed t-test analysis (*p < 0.05, **p < 0.01, p < 0.001). [EV-treated DCs are denoted by dotted line and dark gray shading on histograms for CD83 and CD86 expression in panel (A).]