Title

CD47 Blockade Enhances Macrophage Efferocytosis Via a Process Requiring Low-Density Lipoprotein Receptor-Related Protein 1

Author

Richard Anthony Maldonado, Concordia University - PortlandFollow

Date of Award

Spring 4-1-2019

Document Type

Thesis

College

College of Arts & Sciences

Department

Math & Science

Degree Name

Biology, BA

First Advisor

Mihail Iordanov, Ph.D.

Abstract

Vulnerable atherosclerotic plaques are characterized by large necrotic cores caused by impaired clearance of apoptotic cells (efferocytosis) in the artery wall. Targeted antibody blockade of CD47 (CD47), an anti-efferocytosis protein, enhances efferocytosis and reduces plaque area in mouse models of atherosclerosis. LDL receptor-related protein-1 (LRP1) is a pro-efferocytosis receptor whose deletion from macrophages accelerates atherogenesis. We recently found that CD47 requires macrophage LRP1 to limit atherosclerosis. Thus, we hypothesize that the enhanced efferocytosis caused by CD47 also requires macrophage LRP1. We conducted in vitro studies using wildtype (WT) and LRP1-/- macrophages as efferocytes and either Jurkat lymphocyte cells or WT and LRP1-/- murine macrophages as apoptotic cell substrates (ACs). To stimulate efferocytosis, violet fluorescent-labeled apoptotic cells were co-incubated with green fluorescent-labeled efferocytes in the presence of CD47 (10μg/mL) or IgG control. Phagocytic index (PI) is determined using confocal microscopy and flow cytometry as the percent of efferocytes with internalized ACs. In experiments using Jurkat cells as ACs, CD47 increased PI 2.4-fold and 2.0-fold in WT and LRP1-/- efferocytes, respectively, compared to IgG control. No differences in PI were observed between WT and LRP1-/- efferocytes treated with CD47 or IgG control. In experiments using WT and LRP1-/- macrophages as ACs, we observed no differences in PI between WT and LRP1-/- efferocytes. Interestingly, the use of LRP1-/- macrophages as ACs reduced the PI of WT efferocytes by 44.7% relative to WT macrophage ACs and independently of CD47. These data suggest that the loss of LRP1 on ACs, not the efferocyte, impairs efferocytosis independently of CD47 by rendering the dying cells a poor substrate for clearance.

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