Inflammation-mediated oxidation of polyunsaturated fatty acids generates a new adhesive substrate for integrin αMβ2- and αDβ2-dependent macrophage migration and retention
Location
D.P. Culp Center Ballroom
Start Date
4-5-2024 9:00 AM
End Date
4-5-2024 11:30 AM
Poster Number
30
Name of Project's Faculty Sponsor
Valentin Yakubenko
Faculty Sponsor's Department
Biomedical Sciences
Competition Type
Competitive
Type
Poster Presentation
Presentation Category
Health
Abstract or Artist's Statement
Oxidation of polyunsaturated fatty acids (PUFA) contributes to different aspects of the inflammatory response due to the variety of generated products. Here, we describe a unique mechanism for different PUFAs which is characterized by the generation of inflammation-specific adhesive substrate for macrophage migration in the extracellular matrix (ECM). Previously we demonstrated that docosahexaenoic acid (DHA) oxidation, particularly during inflammation, generates carboxyethylpyrrole (CEP) that forms an adduct with different proteins via an e-amino group of lysine. Most importantly, we found that CEP-modified proteins become the ligands for macrophage adhesive receptors, integrins αDβ2 and αMβ2. The binding to integrins occurs via a negatively charged carboxyl group in the side chain of pyrrole. The published data demonstrate that peroxidation of other n-3 and n-6 PUFAs, including arachidonic and linoleic acids, forms similar end-products that were detected in plasma and atherosclerotic lesions of patients. Based on structural homology, we hypothesized that carboxypyrroles derived from other PUFA during oxidation can also serve as ligands for macrophage adhesion. We generated a product of arachidonic acid oxidation, carboxypropylpyrrole (CPP)-modified albumin (BSA), and tested its adhesive properties. Mouse macrophages demonstrated strong adhesion to CPP-BSA that adheres in a similar mechanism to the previously described DHA product (CEP). Using αMβ2 - and αDβ2 -transfected HEK 293 cells and anti-integrin antibodies we confirmed the specificity of CPP-BSA binding via integrin αMβ2 - and αDβ2. A similar mechanism of molecular recognition for DHA and arachidonic acid end-products was shown by inhibition of cell adhesion to immobilized CPP-BSA by soluble CEP-BSA. Biolayer interferometry verified the high binding affinity since recombinant αM and αD I-domains demonstrated concentration-dependent interaction with immobilized CPP-BSA. Based on our data, a dominant expression of αMβ2 on monocyte-derived (M0) macrophages and the upregulation of αDβ2 on pro-inflammatory (M1) macrophages serve for different steps of macrophage migration. Particularly, the engagement of carboxypyrroles-modified proteins to αMβ2 supports the recruitment of M0 macrophages, while strong adhesion to αDβ2 on M1 macrophages promoted macrophage retention at the site of inflammation. In summary, our results demonstrate that oxidation of different n-3 and n-6 PUFAs changes a positively charged lysine to a negatively charged carboxyl group in CPP/CEP structure that converts ECM proteins to αDβ2 -and αMβ2 -recognition ligands. These modifications form the new adhesive substrate at the site of inflammation and regulate macrophage migration and retention via two macrophage receptors, integrins αDβ2 and αMβ2.
Inflammation-mediated oxidation of polyunsaturated fatty acids generates a new adhesive substrate for integrin αMβ2- and αDβ2-dependent macrophage migration and retention
D.P. Culp Center Ballroom
Oxidation of polyunsaturated fatty acids (PUFA) contributes to different aspects of the inflammatory response due to the variety of generated products. Here, we describe a unique mechanism for different PUFAs which is characterized by the generation of inflammation-specific adhesive substrate for macrophage migration in the extracellular matrix (ECM). Previously we demonstrated that docosahexaenoic acid (DHA) oxidation, particularly during inflammation, generates carboxyethylpyrrole (CEP) that forms an adduct with different proteins via an e-amino group of lysine. Most importantly, we found that CEP-modified proteins become the ligands for macrophage adhesive receptors, integrins αDβ2 and αMβ2. The binding to integrins occurs via a negatively charged carboxyl group in the side chain of pyrrole. The published data demonstrate that peroxidation of other n-3 and n-6 PUFAs, including arachidonic and linoleic acids, forms similar end-products that were detected in plasma and atherosclerotic lesions of patients. Based on structural homology, we hypothesized that carboxypyrroles derived from other PUFA during oxidation can also serve as ligands for macrophage adhesion. We generated a product of arachidonic acid oxidation, carboxypropylpyrrole (CPP)-modified albumin (BSA), and tested its adhesive properties. Mouse macrophages demonstrated strong adhesion to CPP-BSA that adheres in a similar mechanism to the previously described DHA product (CEP). Using αMβ2 - and αDβ2 -transfected HEK 293 cells and anti-integrin antibodies we confirmed the specificity of CPP-BSA binding via integrin αMβ2 - and αDβ2. A similar mechanism of molecular recognition for DHA and arachidonic acid end-products was shown by inhibition of cell adhesion to immobilized CPP-BSA by soluble CEP-BSA. Biolayer interferometry verified the high binding affinity since recombinant αM and αD I-domains demonstrated concentration-dependent interaction with immobilized CPP-BSA. Based on our data, a dominant expression of αMβ2 on monocyte-derived (M0) macrophages and the upregulation of αDβ2 on pro-inflammatory (M1) macrophages serve for different steps of macrophage migration. Particularly, the engagement of carboxypyrroles-modified proteins to αMβ2 supports the recruitment of M0 macrophages, while strong adhesion to αDβ2 on M1 macrophages promoted macrophage retention at the site of inflammation. In summary, our results demonstrate that oxidation of different n-3 and n-6 PUFAs changes a positively charged lysine to a negatively charged carboxyl group in CPP/CEP structure that converts ECM proteins to αDβ2 -and αMβ2 -recognition ligands. These modifications form the new adhesive substrate at the site of inflammation and regulate macrophage migration and retention via two macrophage receptors, integrins αDβ2 and αMβ2.