Fig. 6

EVs derived from cocaine treated HIV-1 infected Macrophages induce inflammation and altered endothelial barrier integrity. A Quantitative analysis of TNF-α, IL-6 and IL-1β levels by ELISA in supernatants of Human Brain Microvascular Endothelial Cells (HBMECs) treated with EVs derived from cocaine treated or untreated, HIV-1 infected or uninfected macrophages. B Fluorescence microscopic image showing adhesion of PKH26-labelled cocaine treated or untreated, HIV-1 infected or uninfected DC derived and C macrophage derived EVs on HBMECs. Respective bar diagrams indicate number of EVs adhered to the HBMEC monolayer. D TEER was measured at indicated time intervals until 72 h after treatment with macrophage derived EVs by using Electric Cell-Substrate Impedance Sensing (ECIS) Ztheta 96 well array station. The impedance measurements were plotted as a percentage, with the reading of untreated as reference. E Fluorescence intensity of FITC-Dextran beads permeabilized through a HBMEC monolayer untreated or treated with 10 µM cocaine or treated with EVs derived from macrophages treated with or without cocaine, infected with or without HIV-1. VEGF was used as positive control. Data represent the mean ± SD of 3 independent experiments, and p-values were calculated relative to untreated controls (*p ≤ 0.05, **p ≤ 0.01, ***p < 0.001)