Humoral immune response to carbamyl-epitopes in atherosclerosis

Carbamylation of proteins in vivo occurs by cyanate, non-enzymatically when urea is dissociated, and by myeloperoxidase-catalyzed oxidation from thiocyanate. Carbamylation of low-density lipoprotein is suggested to enhance atherogenesis in patients with with chronic kidney disease and uremia. This thesis study assessed the questions of whether healthy humans or uremic patients under enhanced carbamylation have antibodies recognizing carbamyl-epitopes in plasma, and what is their role in vivo. Also, humoral immune response to carbamyl-LDL immunization and its impact on atherogenesis in LDLR-/mice was investigated. In this thesis study, plasma antibodies to carbamylated proteins were detected in humans, and IgG antibodies to carbamylated proteins were associated with uremia and smoking, conditions with enhanced carbamylation. The human IgG and IgM antibodies binding to carbamyl-epitopes were associated with oxidation-specific epitopes in plasma. Monoclonal Fab antibodies with characteristics of a natural antibody and ability to bind both carbamyland malondialdehydederived epitopes were cloned from healthy humans. An investigated Fab antibody was able to bind epitopes found in atherosclerotic lesions and inhibit the uptake of modified LDL by macrophages. Human plasma antibodies and the monoclonal Fab bound to epitopes found on apoptotic cells. Human B-cells secreted antibodies with similar cross-reactive binding properties between carbamyland malondialdehyde adducts and apoptotic cells in vitro. Immunization with mouse carbamyl-LDL without adjuvant resulted in specific IgG immune response in LDLR-/mice, but also a cross-reaction with malondialdehyde-adducts was observed. Carbamyl-LDL immunized mice had enhanced plasma antibody binding to apoptotic cells. Carbamyl-LDL immunization did not affect atherogenesis in mice. This thesis demonstrates that IgG antibodies to carbamyl-epitope might serve as a novel indicator of carbamylation in vivo in uremic patients or smokers. The cross-reactivity between antibodies binding to carbamylated and oxidation-specific epitopes, and apoptotic cells may have a role in explaining the link between enhanced atherogenesis and kidney disease.