Fig. 5

Novel-miR-7 is a key hepatic sEV cargo that promotes microvascular endothelial hyperpermeability by targeting LAMP1. Hepatic sEV miRNAs were profiled by the small RNA sequencing analysis. MVECs were incubated with 120 μg/mL hepatic sEVs for 24 h (D) or transfected with 50 nmol/L novel-miR-7 mimics or negative control (NC) mimic (E–K). A The heat-map of the ten miRNAs with the most significant (fold change > 10, FDR < 0.05) abundance differences in hepatic sEVs derived from the MCD group versus the MCS group, n = 3 per group. B, C Quantitative PCR analysis of novel-miR-7 expression levels in hepatic sEVs and mouse plasma-derived sEVs, n = 6 per group. D Quantitative PCR analysis of novel-miR-7 expression levels in hepatic sEVs-treated MVECs, n = 4 per group. E Transfection efficiency of 5-FAM labeled novel-miR-7 mimics. Scale bar, 20 μm. F Representative images of acridine orange (AO) and magic red (MR) staining, n = 4 per group. Scale bar, 20 μm. G Representative Western blot bands and the summarized data determined by densitometric analysis, n = 4 per group. H Representative flow cytometry images of caspase-1 FLICA staining and the summarized data of the positive cells, n = 4 per group. The fold changes were obtained by calculating the ratio of the positive cells of the treated groups to the NC group. I Relative permeability of the endothelial monolayer to FITC-dextran, n = 4 per group. J LAMP1 reporter gene activity was performed using a dual-luciferase reporter assay system and normalized relative to Renilla luciferase activity, n = 4 per group. K Representative western blot bands and the summarized data determined by densitometric analysis, n = 4 per group. Data are expressed as the mean ± SEM. Statistics: Student t-test (B–D, G–I, K) and One-way ANOVA (J), *P < 0.05, **P < 0.01 vs. hepatic sEVs or plasma-sEVs derived from MCS group, or MVEC treated with MCS-sEVs, or NC mimic; ^&P < 0.05, ^&&P < 0.01 vs. hepatic sEVs or plasma-sEVs derived from ND group, or MVEC treated with ND-sEVs