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Principal Investigator: Gabor Csanyi
Organization: AUGUSTA UNIVERSITY
Fiscal Year: 2022
Award: $385,000
Funding agency: National Heart Lung and Blood Institute
Project Summary
Atherosclerosis, the main underlying cause of death worldwide, is characterized by chronic inflammation and
accumulation of lipids in the arterial wall. Excessive lipid accumulation by macrophages (MΦs) and vascular
smooth muscle cells (SMCs) plays a key role in the initiation and progression of atherosclerosis. It is generally
accepted that atherosclerosis arise from LDL modification in the arterial wall and its subsequent internalization
by MΦs and SMCs through a variety of scavenger receptors. The role of scavenger receptor-independent lipid
uptake in atherosclerosis, the physiological factors stimulating this process, and the signaling mechanisms
involved remain poorly characterized. We reported that matrix protein thrombospondin-1 (TSP1) stimulates
direct, scavenger receptor-independent uptake of unmodified, native LDL (nLDL) in MΦs. TSP1 via its cognate
receptor CD47 activates actin-binding protein cofilin, leading to macropinocytosis of nLDL, and excessive
cholesterol accumulation. The signaling mechanisms downstream of CD47 that stimulate macropinocytosis are
unknown. Although phospholipase C (PLC) and slingshot phosphatase 1 (SSH1) have been shown to activate
cofilin, their roles in TSP1-induced macropinocytosis have not yet been investigated. Moreover, the ability of
TSP1-CD47 signaling to stimulate MΦ macropinocytosis in atherosclerotic vessels in vivo and the significance
of lipid macropinocytosis in the pathogenesis of atherosclerosis remain to be determined. Our novel preliminary
data show that TSP1 and CD47 knockout mice are protected from atherosclerosis and the macropinocytosis
inhibitor EIPA decreases atherosclerotic lesion formation in hypercholesterolemic mice. We hypothesize that
TSP1 via CD47 promotes lipid macropinocytosis in the arterial wall, contributing to lipid accumulation and the
pathogenesis of atherosclerosis. The hypothesis will be tested via the following aims: (1) examining for the first
time whether CD47 receptor signaling in MΦs stimulates macropinocytosis via PLC- and SSH1-mediated cofilin
activation and contributes to atherosclerosis development; (2) exploring whether MΦs internalize lipoproteins via
macropinocytosis in atherosclerotic arteries in vivo and that deletion of NHE1 (major target of EIPA) specifically
in MΦs attenuates atherosclerosis, and (3) investigating whether TSP1 binding to CD47 stimulates
macropinocytosis in MΦ-like SMCs via Nox1-mediated cofilin activation. The proposal will employ global and
cell-specific knockout mice, and other genetic tools to test the hypothesis. Specific targeting of CD47 via multiple
approaches (antibody blockade, siRNA/morpholino silencing, and CD47 activating peptide sequences) will
provide confirmation of results obtained in genetic mutants. Multiple complementary techniques will be used to
study macropinocytosis in vitro (pharmacological, genetic, fluorescence/high-resolution microscopy) and in vivo
(cofilin mutants, AngioSPARK 680, MΦ-specific NHE1 knockouts). This innovative proposal has the potential to
reveal important new mechanisms of lipid internalization and provide a paradigm shift in our knowledge about
how atherosclerosis develops.
Terms: <Actin-Binding Protein><Affinity><Antibodies><Antigen Presentation><Aorta><Apo-E><ApoE><Apolipoprotein E><Area><Arterial Fatty Streak><Arterial Fatty Streaks><Arteries><Atheroma><Atheromatous><Atheromatous degeneration><Atheromatous plaque><Atheroscleroses><Atherosclerosis><Atherosclerotic Cardiovascular Disease><Attenuated><Binding><Blocking Antibodies><CD36><CD36 gene><CD47><CD47 Antigen><CD47 Glycoprotein><CD47 gene><Cardiovascular><Cardiovascular Body System><Cardiovascular Organ System><Cardiovascular system><Cause of Death><Cell Body><Cell Communication and Signaling><Cell Fractionation><Cell Signaling><Cell membrane><Cells><Cholesterol><Chronic><Confocal Microscopy><Cytoplasmic Membrane><Data><Dephosphorylation><Development><Dissociation><Exhibits><Extracellular Matrix Proteins><Flow Cytofluorometries><Flow Cytofluorometry><Flow Cytometry><Flow Microfluorimetry><Flow Microfluorometry><Fluorescence><Foam Cells><GP3B><GP4><GPIV><Genetic><HPLC><Healthcare Delivery><Heart Vascular><High Performance Liquid Chromatography><High Pressure Liquid Chromatography><High Speed Liquid Chromatography><Human><Immunoblotting><In Vitro><Inflammation><Inflammatory><Integrin-Associated Protein><Intracellular Communication and Signaling><Isoforms><KO mice><Knock-out><Knock-out Mice><Knockout><Knockout Mice><Knowledge><LDL><LDL Lipoproteins><LDL Receptors><LDL oxidation><Lecithinase C><Leiomyocyte><Link><Lipid-Laden Macrophage><Lipids><Lipoprotein LDL Receptors><Lipoproteins><Low Density Lipoprotein Receptor><Low Density Lipoprotein oxidation><Low-Density Lipoproteins><MER6><Mediating><Membrane><Mice><Mice Mammals><Microscopy><Modern Man><Modification><Molecular Interaction><Murine><Mus><Mφ><NHE1><Null Mouse><Other Genetics><PDGF-BB><Pathogenesis><Pathway interactions><Pharmacology><Phenotype><Phosphatases><Phosphohydrolases><Phospholipase C><Phosphomonoesterases><Phosphoric Monoester Hydrolases><Physiologic><Physiological><Plasma Membrane><Play><Process><Protein Dephosphorylation><Protein Isoforms><Proteins><Receptor Protein><Receptor Signaling><Reporting><Resolution><Role><SCARB3><Short interfering RNA><Signal Transduction><Signal Transduction Systems><Signaling><Small Interfering RNA><Smooth Muscle Cells><Smooth Muscle Myocytes><Smooth Muscle Tissue Cell><Subendothelial Layer><Surface Antigen Identified by Monoclonal Antibody 1D8><TSP-1><TSP1><Techniques><Testing><Thrombospondin 1><Time><United States><Vascular Smooth Muscle><Western Blotting><Western Immunoblotting><Woman><acetyl-LDL receptor><acetylated LDL receptor><atheromatosis><atherosclerosis plaque><atherosclerotic disease><atherosclerotic lesions><atherosclerotic plaque><atherosclerotic vascular disease><beta-Lipoproteins><biological signal transduction><circulatory system><cofilin><cost><developmental><flow cytophotometry><health care delivery><health delivery systems><health services delivery><in vivo><inhibitor><innovate><innovation><innovative><lipophosphodiesterase I><macrophage><membrane structure><men><men's><mouse genetics><mutant><nano particle><nano-sized particle><nanoparticle><nanosized particle><novel><overexpress><overexpression><pathway><peptide aminoacid sequence><peptide sequence><phosphatidylcholine cholinephosphohydrolase><plasmalemma><platelet-derived growth factor BB><protein aminoacid sequence><protein blotting><receptor><scavenger receptor><siRNA><social role><subcellular fractionation><therapeutic target><tool><uptake><vascular inflammation><vulnerable plaque>