Arterial antibodies promote atherosclerotic plaque development in mice

Abstract

Abstract Atherosclerosis is characterized by the progressive buildup of cholesterol-rich plaques within the arterial wall. Plaques activate the immune system, of which B cells play a major role. Antibodies bind autoantigens released from necrotic cells and form immune complexes to increase inflammation. We isolated aorta-associated B cells from atherosclerotic ApoE-deficient mice on a high-fat diet and performed single cell sequencing of the B cell receptors to obtain antibody pairs. 32 over-represented antibodies were identified and then expressed in Expi293F cells. Epitope determination of these antibodies was performed against a human proteome array, and 15 potential autoantigens bound the monoclonal antibodies with significant affinity and specificity. The 12 most-promising antigens were scrutinized for their location in plaque tissue by immunohistochemistry using the cognate antibodies. Nine of these relevant antigens were then successfully synthesized using the pET bacterial expression system, and purified proteins were used to interrogate antibodies in human and mouse sera by ELISA. Mice were then immunized with the 5 best antigens to confirm their immunogenicity. The relevant antigens and antibodies are now being tested to see if they will protect or promote atherosclerosis development, and if they can be used as biomarkers in human disease. This research was supported in part by the Intramural Research Program of the NIH, National Institute on Aging.