Donor-derived CD8 ⁺ CD122 ⁺ Tregs generated in mixed donor chimeric NOD mice delete autoreactive T cells

The establishment of mixed hematopoietic chimerism is a promising way to induce immune tolerance for islet replacement therapy and to treat the underlying autoimmunity in Type 1 diabetes (T1D). Mixed chimerism not only promotes effective thymic negative selection of autoreactive cells but also restores regulatory T cell (Treg) function and peripheral tolerance. In the current study, we determined that a novel class of donor-derived CD8 ⁺ CD44 ⁺ CD122 ⁺ Tregs (d-CD8 ⁺ CD122 ⁺ Tregs) plays a crucial role in controlling autoimmunity in non-obese diabetic (NOD) mice with induced mixed chimerism. Using adoptive T cell transfer experiments, we showed that d-CD8 ⁺ CD122 ⁺ Tregs abrogate autoimmunity by selectively depleting the exogenously injected diabetogenic T cells in Recombination-Activating Gene deficient NOD mice. These d-CD8 ⁺ CD122 ⁺ Tregs from NOD chimeras show upregulation of Helios, Programmed cell death protein 1, perforin, granzyme-B, CD39, Folate receptor 4, and downregulation of proinflammatory markers like Scart1 and Scart2. Using <i>in vitro</i> assays, we show that d-CD8 ⁺ CD122 ⁺ Tregs respond specifically to a Complementarity-Determining Region-3 peptide sequence derived from T cell receptors of islet antigen-specific autoreactive T cells. Thus, mixed chimerism might be a method to revitalize CD8 ⁺ CD122 ⁺ Tregs which are decreased in number and functionality in NOD mice. Similarly, we found that individuals with T1D have a deficiency in CD8 ⁺ CD122 ⁺ Tregs, suggesting a potential loss of regulatory function accompanies disease onset. Revitalizing CD8 ⁺ CD122 ⁺ Tregs may offer a new therapeutic strategy of restoring immune tolerance in autoimmune diabetes. One sentence summary Inducing mixed donor chimerism in NOD mice generates donor-derived CD8 ⁺ CD122 ⁺ Tregs that suppress autoimmunity and restore immune tolerance by selectively eliminating autoreactive T cells.