Targeting macrophage retention in the extracellular matrix during atherosclerosis

The National Heart, Lung, and Blood Institute awarded East Tennessee State University $536,447 under the Cardiovascular Diseases Research program (CFDA 93.837) on August 15, 2026, to investigate mechanisms of macrophage retention in the extracellular matrix during atherosclerosis development.

The project examines how oxidation of polyunsaturated fatty acids—including docosahexaenoic acid, arachidonic acid, and linolenic acid—generates adhesive substrates in vascular tissue that anchor macrophages through a newly identified ligand-receptor interaction between carboxyalkylpyrrole-modified extracellular matrix proteins and integrin αdβ2. The research pursues three aims: evaluating the role of different carboxyalkylpyrroles in macrophage adhesion and migration within three-dimensional matrices; identifying binding sites within integrin αdβ2 for carboxyalkylpyrrole-modified proteins; and validating the identified binding site using in vitro adhesion and migration assays and in vivo atherosclerosis models.

Performance occurs in Johnson City, Tennessee. The award runs through April 30, 2029. This project grant funds basic and translational research in cardiovascular disease mechanisms under NHLBI's competitive research portfolio, which supports prevention and treatment of heart, lung, and blood diseases through investigation in basic, translational, clinical, and population sciences.

Abstract

Macrophage retention at the site of inflammation is a critical event for the development of different inflammatory diseases including atherosclerosis. Fatty acid oxidation is an important mechanism that is facilitated by inflammation and contributes to its further development. We recently showed that the end-product of docosahexaenoic acid (DHA) oxidation, carboxyethylpyrrole, forms an adduct with ECM proteins and serves as a ligand for the macrophage adhesive receptor, integrin ΑDΒ2. Similar products are formed during the oxidation of arachidonic and linolenic acids, which accumulate in inflamed tissues. The goal of our investigation is to determine the mechanism of macrophage retention at the site of chronic inflammation, focusing on the potential pathological role of polyunsaturated fatty acids oxidation. We hypothesize that inflammation-mediated oxidation of different polyunsaturated fatty acids generates a unique adhesive substrate in the vascular wall for a strong anchorage of macrophages by the formation of new ligand-receptor pair (carboxyalkylpyrroles (CAPS) modified extracellular matrix proteins - integrin ΑDΒ2), which is critical for macrophage retention during atherosclerosis. Guided by our published results and strong preliminary data, this hypothesis will be tested by pursuing three aims: 1. Evaluate and compare the role of different carboxyalkylpyrroles in macrophage adhesion and migration in the extracellular matrix; 2. Identify the binding site(S) within integrin ΑDΒ2 for carboxyalkylpyrrole-modified proteins; 3. Validate the CAPS-binding site within ΑDΒ2 utilizing in vitro adhesion/migration assays and in vivo models of atherosclerosis. Under the first aim, the contribution of different CAPS to macrophage adhesion and migration in 3D matrixes will be evaluated. Under the second aim, the binding interface between cap and ΑDΒ2 will be defined using mutagenesis approaches and protein-protein binding assays. Under the third aim, the mutation of potential critical amino acids in ΑDΒ2 structure will be introduced in HOXB8 hematopoietic progenitor cells using crispr/CAS9. Macrophages will be generated from HOXB8 progenitor cells and tested in 3D macrophage migration assays and in vivo model of atherosclerosis. The contribution of our research will be significant for understanding the process of chronic inflammation during atherogenesis and other low-grade chronic inflammatory diseases. We propose a universal mechanism of pro-inflammatory substrate generation, which links the oxidation of polyunsaturated fatty acids with integrin ΑDΒ2-mediated macrophage retention within the CAPS-modified extracellular matrix of inflamed tissue. Our innovation resides in proposing a new concept of macrophage retention at the site of inflammation. We propose a critical role of pufa end-products in the retention of pro-inflammatory macrophages in atherosclerotic lesions. The obtained information will provide a new strategy for the treatment of chronic inflammatory diseases. The blocking of CAPS in tissues can prevent macrophage accumulation and the progression of atherosclerosis and other chronic inflammation. The development of anti-inflammatory treatments is an objective of our further investigations.