Regulatory T cells inhibit autoantigen-specific CD4⁺ T cell responses in lupus-prone NZB/W F1 mice

Introduction: Progressive loss of regulatory T cell (Treg)-mediated control over autoreactive effector T cells contributes to the development of systemic lupus erythematosus (SLE). Accordingly, we hypothesized that Treg may also have the capacity to suppress the activation of autoreactive CD4⁺ T cells that are considered to drive autoimmunity. Methods: To investigate whether Treg are involved in the control of autoreactive CD4⁺ T cells, we depleted CD25⁺ Treg cells either in vivo or in vitro, or combined both approaches before antigen-specific stimulation with the SLE-associated autoantigen SmD1(83-119) in the NZB/W F1 mouse model either after immunization against SmD1(83-119) or during spontaneous disease development. Frequencies of autoantigen-specific CD4⁺ T cells were determined by flow cytometry using the activation marker CD154. Results: Both in vitro and in vivo depletion of CD25⁺ Treg, respectively, increased the frequencies of detectable autoantigen-specific CD4⁺ T cells by approximately 50%. Notably, the combined in vivo and in vitro depletion of CD25⁺ Treg led almost to a doubling in their frequencies. Frequencies of autoantigen-specific CD4⁺ T cells were found to be lower in immunized haploidentical non-autoimmune strains and increased frequencies were detectable in unmanipulated NZB/W F1 mice with active disease. In vitro re-addition of CD25⁺ Treg after Treg depletion restored suppression of autoantigen-specific CD4⁺ T cell activation. Discussion: These results suggest that the activation and expansion of autoantigen-specific CD4⁺ T cells are partly controlled by Treg in murine lupus. Depletion of Treg therefore can be a useful approach to increase the detectability of autoantigen-specific CD4⁺ T cells allowing their detailed characterization including lineage determination and epitope mapping and their sufficient ex vivo isolation for cell culture.