Project Grant F31AG099733

IMPACT OF PLATELET-MONOCYTE INTERACTIONS ON HIV-ASSOCIATED VASCULAR DISEASE - ABSTRACT EVEN ON COMBINED ANTIRETROVIRAL THERAPY (CART), PEOPLE LIVING WITH HIV (PLWH) EXPERIENCE A LOW LEVEL OF VIRAL REPLICATION THAT ENGENDERS A CHRONIC INFLAMMATORY STATE AND INCREASES THEIR RISK TO DEVELOP MULTIPLE TYPES OF VASCULAR DISEASE. PREVIOUS WORK BY OUR LAB HAS DEMONSTRATED PLATELET-MONOCYTE INTERACTIONS (PMCS) OCCUR WITH GREATER FREQUENCY IN PLWH DUE TO ABERRANT PLATELET ACTIVATION, AND THESE COMPLEXES ARE ASSOCIATED WITH HIGHER EXPRESSION OF MONOCYTE ACTIVATION MARKERS, AS WELL AS PRO-INFLAMMATORY CD16+ INTERMEDIATE AND NON-CLASSICAL MONOCYTE (NCM) PHENOTYPES. HOWEVER, THE FATE OF PMCS IN THE VASCULAR DISEASE MICROENVIRONMENT, AND THE PLATELET-INDUCED TRANSCRIPTIONAL NETWORKS THAT LEAD TO DYSFUNCTION, ARE NOT WELL CHARACTERIZED. SCRNASEQ ANALYSIS OF PMCS FROM PEOPLE LIVING WITH HIV AND ATHEROSCLEROSIS (AS) SIMULTANEOUSLY REVEALS AN UPREGULATION OF INFLAMMATORY AND DISEASE-ASSOCIATED PATHWAYS IN COMPARISON TO PEOPLE WITH AS ALONE. I HYPOTHESIZE THAT THESE DYSREGULATED PMC PHENOTYPES IN PLWH CONTRIBUTE TO BOTH LARGE VESSEL (ATHEROSCLEROSIS) AND SMALL VESSEL (CEREBRAL SMALL VESSEL DISEASE. CSVD) DISEASE. TO INVESTIGATE THIS, I WILL DESIGN AND OPTIMIZE 3D HYDROGEL MODELS OF THE INTI MA AND NEUROVASCULAR UNIT (NVU) TO RECAPITULATE ATHEROSCLEROSIS AND BLOOD-BRAIN BARRIER (BBB) DYSFUNCTION, RESPECTIVELY. IN AIM 1, I WILL CONSTRUCT THE INTIMA MODEL WITH MONOCYTE, ENDOTHELIAL CELL (EC), AND SMOOTH MUSCLE CELLS (SMC) COMPONENTS. MONOCYTE-EC ADHESION, TRANSMIGRATION, AND FOAM CELL FORMATION WILL BE ANALYZED BY LIVE-CELL CONFOCAL IMAGING. MONOCYTE PHENOTYPES POST-EXTRAVASATION, AS ALTERED BY PLATELET INTERACTIONS, WILL BE ANALYZED BY FULL-SPECTRUM FLOW CYTOMETRY. FURTHERMORE, DIFFERENTIALLY EXPRESSED MONOCYTE GENES AND PATHWAYS IDENTIFIED BY SCRNASEQ WILL BE PERTURBED TO DETERMINE THEIR RELEVANCE IN AN ATHEROSCLEROTIC SETTING. IN AIM 2, I WILL USE WELL-ESTABLISHED FLOW-BASED NVU MODEL TO FIRST EXAMINE HOW THE KINETICS OF PMCBBB ADHERENCE AND MIGRATION DIFFER IN COMPARISON TO NON-PMCS. THEN, I WILL UTILIZE THE INTIMA MODEL FRAMEWORK ESTABLISHED IN AIM 1 TO DESIGN A NVU HYDROGEL SYSTEM (COMPRISED OF BRAIN ECS, PERICYTES, AND INFILTRATING MONOCYTES) TO DETERMINE IF AND HOW PMCS DIFFERENTIATE INTO INFLAMMATORY PERIVASCULAR MACROPHAGES (PVMS) OVER TIME AND CONTRIBUTE TO BBB INFLAMMATION, AS ASSESSED BY A LOSS OF EC TIGHT JUNCTION INTEGRITY AND AN INFLAMMATORY CYTOKINE MILIEU. PERICYTE CELL DEATH WILL BE ASSESSED AS A SURROGATE OF ADVERSE PVM IMMUNE ACTIVITY. AS IN AIM 1, GENES AND GENE PATHWAYS THAT ARE DIFFERENTIALLY REGULATED BASED ON HIV OR CSVD STATUS IN OUR SCRNASEQ ANALYSIS WILL BE MANIPULATED TO ISOLATE THEIR EFFECTS ON NEUROINFLAMMATORY PROCESSES. COLLECTIVELY, THESE EXPERIMENTS WILL IMPROVE OUR UNDERSTANDING OF HIV-ASSOCIATED IMMUNE HYPERACTIVATION, SPECIFICALLY THE EFFECTS OF PLATELET-MONOCYTE COMPLEXES ON VASCULAR PATHOLOGIES AND THE DYSREGULATED GENE NETWORKS THAT UNDERLY THESE CLINICAL OUTCOMES. MOREOVER, MY RESULTS WILL ALSO HAVE IMPLICATIONS FOR OTHER CONDITIONS ASSOCIATED WITH HEIGHTENED PLATELET ACTIVATION, SUCH AS SARS-COV-2 INFECTION.