Abstract 20122: HIV Protein Negative Factor (Nef) - Induced Release of Endothelial Monocyte Activating Polypeptide II (EMAPII) Mediates Endothelial Dysfunction in HIV Patients

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HIV patients have increased risk of vascular pathologies including cardiovascular diseases and pulmonary hypertension. A likely candidate is the endothelial dysfunction causing HIV-Nef protein, which persists in peripheral blood mononuclear cells (PBMCs) and plasma derived exosomes even in HIV patients virologically-suppressed on anti-retroviral therapy (ART). We sought to determine the role of EMAPII induced endothelial cell death in HIV-related endothelial dysfunction. PBMCs from HIV patients on ART, HIV-Nef+ T cell lines and HIV-1 NL4-3 infected PBMCs were co-cultured with primary human endothelial cells including human coronary artery endothelial cells. Apoptosis was measured through TUNEL and cleaved caspase-3 staining, reactive oxygen species (ROS) with 5uM dihydroethidium (DHE) staining and mitochondrial membrane potential determined as a ratio of aggregate/monomer of JC-1. Bronchoalveolar lavage cells (BAL) and PBMCs from HIV patients on ART showed persistence of HIV-Nef protein and positive correlation between HIV-Nef positive cells and EMAPII surface positive cells (n=17, Spearman correlation coefficient = 0.74, p = 0.0006 for BAL cells). HIV-Nef expression and HIV infection induced a 4-8 fold increase in EMAPII surface expression in T cells. Co-culture with HIV-Nef+ T cells induced a 3 fold increase of cleaved caspase 3 in endothelial cells which was ameliorated with an EMAPII neutralizing antibody, M7/1. Endothelial mitochondrial membrane potential was decreased by HIV-Nef+ T cells (0.16 vs 0.49, p<0.05) but preserved by adding M7/1 mAB. (0.39, p<.05). HIV NL4-3 infected PBMCs induced a 4-fold increase in Cleaved Caspase-3 and 2.5 fold increase in ROS which was alleviated by M7/1 mAB (p<0.05). Finally, when PBMC from HIV+ patients were co-cultured with arterial endothelial cells, a trend towards increased apoptosis (TUNEL) was seen. BAL cells and PBMCs of HIV patients on ART show persistence of HIV-Nef protein which strongly correlates with EMAPII expressing cells. In conclusion, our data indicates that endothelial dysfunction in HIV patients is likely to be induced by HIV-Nef in an EMAPII mediated fashion. This suggests the possibility of using EMAPII neutralizing mAB as a potential therapeutic in HIV patients.

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