Abstract 13424: Dendritic Cell Surface Proteoglycan Binding-mediated Internalization of the Apob-100 Peptide P210 Increases Cd8+ T Cell Cytolytic Function

Abstract

Background: We have reported that immunization with the apoB-100 derived peptide p210 reduces atherosclerosis and aortic aneurysm formation/rupture, through modulation of CD8+ T cells. It remains unclear how an exogenous peptide like p210 enters dendritic cells for antigen presentation by MHC-I to CD8+ T cells. The p210 peptide sequence is located within the proteoglycan (PTG) binding site of apoB-100, which is a known alternative mechanism for cell internalization of LDL. Antigens internalized by cell surface PTG binding are cross-presented on the MHC-I molecule. We therefore tested the mechanism of p210 internalization by dendritic cell (DC) surface PTG binding and investigated the CD8+ T cell function in response to p210 antigen presentation. Methods and Results: Bone marrow derived DCs from apoE-/- mice were primed with fluorescently tagged p210 (FITC-p210) for 4 hours in vitro. There was significantly increased FITC internalization in DCs treated with FITC-p210 compared to FITC alone (24.3±2.7% vs. 6.6±0.8%; P<0.001; N=8 each) assessed by flow cytometry. Heparin significantly reduced internalization of FITC-p210 compared to no treatment (14.0±4.0%, N=4 vs. 29.9±6.7% N=9; P<0.01). Treatment of DCs with p-nitrophenyl-β-D-xylopyranoside, an inhibitor of cell surface heparan sulfate chain addition, significantly reduced FITC-p210 internalization (10.5±1.5% vs. 22.8±10.0% P<0.05, N=5 each) confirming the role of PTG binding. Adoptive transfer of p210-primed DCs into recipient apoE-/- mice significantly reduced CD8+CD25+FoxP3+ regulatory cells compared to unprimed DC controls (0.26±0.04% vs. 0.37±0.05%; N=5 each; P<0.01). Cytolytic activity against target DCs was significantly higher in CD8+ T cells from p210-primed DC recipient mice compared to controls (1.6±0.3% vs. 1.0±0.4% specific lysis; N=5 each; P<0.05). There were no significant differences in CD8+ T cell activation markers. Conclusion: Internalization of p210 is mediated in part by DC surface PTG binding, resulting in significantly reduced CD8+CD25+FoxP3+ regulatory T cells and increased cytolytic activity, suggesting unopposed CD8+ T cell effector function. Our results support the mechanism of p210 internalization by DC surface PTG increasing CD8+ T cell function.