Peripheral Tolerance Regulates RBC-specific CD4+ T Cells in a Novel Mouse

Abstract

Abstract T cell tolerance mechanisms towards tissue-restricted antigens are well described and include deletion, anergy, or regulatory cells; however tolerance mechanisms against RBC-restricted antigens are poorly understood. Herein, we utilized the HOD mouse to investigate T cell tolerance to RBC-specific antigens. The HOD mouse expresses a fusion protein consisting of hen egg lysosyme (HEL), ovalbumin (OVA), and human blood group, Duffy driven by an RBC-specific promoter. The HOD mouse was crossed to TCR transgenic OTII mouse to generate HOD mice with autoreactive CD4+ T cells (OTII+HOD+). Young OTII+HOD+ mice do not have detectable autoantibodies, suggesting immune tolerance. Analysis of thymii 8 week-old mice revealed similar numbers of OTII CD4+ SP thymocytes between OTII+HOD+ and OTII+HOD−, suggesting that central tolerance plays little role in RBC specific CD4+ T cells. However, in the periphery, several tolerance mechanisms were identified in OTII+HOD+ mice. Autoreactive splenic CD4+ T cells in OTII+HOD+ mice underwent deletion, expressed higher levels of inhibitory markers PD-1, LAG3, CD5, and had higher numbers of CD25+Foxp3+ regulatory T cells, when compared to OTII+HOD−. Furthermore, CD4+ T cells from OTII+HOD+ had significantly reduced proliferation in response to cognate antigen, suggesting anergy or functional inactivation. Analysis of antigen presenting cells in OTII+HOD+ revealed higher frequencies of myeloid DC, known to express high levels of MHC II and propensity to activate CD4+ T cells. Finally, a break in tolerance was observed in 30% of OTII+HOD+ at 6 months, as indicated by anti-HOD antibodies. Future studies will focus on identifying defective tolerance mechanism that contributed to autoantibody genesis.