HIV-Infection and Cocaine Use Regulate Semen Extracellular Vesicles Proteome and miRNAome in a Manner that Mediates Strategic Monocyte Haptotaxis Governed by mIR-128 Network

Abstract

Background: Extracellular vesicles (EVs) are mediators of horizontal transfer of bioactive molecules between cells and regulators of cell-cell interaction. EV-mediated cell-cell interactions play roles in physiological and pathophysiological conditions. However, the effect of pathogens and cocaine use on EV composition and function are not fully understood.  Methods: We obtained 64 seminal plasma specimens from HIV-1-uninfected (HIV—and HIV-1-infected (HIV+) participants who used cocaine (COC—) or did not use cocaine (COC+). The participants were divided into 4 clinical groups (16 per group) denoted as HIV–COC–, HIV–COC+, HIV+COC–, and HIV+COC+ were used. The seminal plasma was used to isolate EVs (SEVs) by differential centrifugation and PPLC-based size exclusion chromatography (SEC) as previously described. We used the SEV to study how HIV infection and cocaine regulate the repertoire and functions of EV-associated proteins, miRNA, and their interactome. After paired proteomics and small RNA-Sequencing (sRNA-Seq) analyses, we employed an in vitro two-dimensional-substrate single cell haptotaxis assay for quantitative assessment of the effect of different SEV on monocyte haptotaxis. Finally, we functionalized SEV with exogenous miR-128 to directly address the role of SEV-associated miR-128 on monocyte haptotaxis.  Findings: Systems biology and multi-omics analysis showed that HIV infection (HIV+) and cocaine (COC) use (COC+) promote the release of SEV with dysregulated extracellular proteome (exProtein), miRNAome (exmiR), and exmiR networks. Integrating SEV proteome and miRNAome revealed a significant decrease in the enrichment of disease-associated, brain-enriched, and HIV-associated miR-128-3p (miR-128) in HIV+COC+ SEV along with a concomitant increase in miR-128 targets—PEAK1 and RND3/RhoE. Two-dimensional-substrate single cell haptotaxis showed that in the presence of HIV+COC+ SEVs, contact guidance provided by the extracellular matrix (ECM, collagen type 1) network facilitated far-ranging haptotactic cues that guided monocytes over longer distances. Functionalizing SEVs with miR-128 mimic revealed that the strategic changes in monocyte haptotaxis are in large part the result of SEV-associated miR-128.  Interpretation: Compositionally and functionally distinct HIV+COC+ and HIV–COC– SEVs and their exmiR networks may provide cells relevant but divergent haptotactic guidance in the absence of chemotactic cues, under both physiological and pathophysiological conditions. Funding Statement: This work was supported by grants from the National Institute on Drug Abuse (NIDA), grants DA042348, DA050169 and DA053643 to C.M.O. Declaration of Interests: None to declare. Ethics Approval Statement: This study was conducted according to University regulations approved by Stony Brook University Institutional Review Boards (IRB # 201608703) using de-identified human specimens obtained through the Multicenter AIDS Cohort Study (MACS).