Fig. 7
EV subpopulations increase the production of key pro-inflammatory markers in hCMEC/D3 cells. (A) mRNA expression of inflammatory candidate genes (IL6, IL1β, ICAM-1 and VCAM-1) in hCMEC/D3 treated with (normalized to 109) of the size-based tEV subpopulations (2 K,10 and 100 K), TNFα/IFNγ treated cells and untreated cells. Data were analyzed using two stable reference genes and the fold changes were plotted as the mean ± SEM and normalized to non-stimulated control cells (n = 3 biological replicates per group). (B–C) Representative flow cytometry data (left) and quantification (right) of ICAM-1 (B) and VCAM (C) level hCMEC/D3 in response to size-based EV subpopulations (2 K,10 and 100 K) and in comparison with TNFα/IFNγ (10 ng/mL) and untreated hCMEC/D3 as positive and negative controls, respectively. (D–E) Protein levels from IL-10 (D) and CXCL10 (E) produced by hCMEC/D3 in response to size-based EV subpopulations (2 K,10 and 100 K) and in comparison, to TNFα/IFNγ (10 ng/mL) and untreated hCMEC/D3 as positive and negative controls, respectively. Data are presented as mean ± SEM (*p < 0.05)