Alloantigen-Pulsed Host Dendritic Cells Induce T-Cell Regulation and Prolong Allograft Survival in a Rat Model of Hindlimb Allotransplantation

Background: Composite tissue allotransplantation is restricted due to the risks presented by long-term therapeutic immunosuppression. This study is conducted to investigate whether treatment with recipient immature dendritic cells (DCs) pulsed with donor alloantigens can prolong allograft survival and induce T-cell regulation in a rodent model. Materials and methods: Orthotopic hindlimb transplants from Brown-Norway (RT1ⁿ) to Lewis (RT1¹) rats were performed (day 0). DCs were propagated from the recipient bone marrow and pulsed with the donor alloantigen lysate. Group 1 (control group) did not receive any treatment. Groups 2 and 3 received cyclosporine A (CsA) at a concentration of 10 and 16 mg · kg^-1 · day^-1, respectively, on days 0-20 following composite tissue allotransplantation. Group 4 received antilymphocyte serum (i.p. administered 4 d before and 1 d after transplantation) therapy. Group 5 received combined treatment with CsA (10 mg · kg^-1 · day^-1, days 0-20) and donor alloantigen-pulsed recipient DCs (i.v. administered on days 7, 14, and 21). Group 6 received combined treatment with CsA (10 mg · kg^-1 · day^-1 on days 0-20), antilymphocyte serum (administered i.p. 4 d before and 1 d after transplantation), and DCs (administered i.v. on days 7, 14, and 21). Graft rejection was defined as epidermolysis/desquamation of the donor skin. The mixed lymphocyte reaction was performed to determine the donor T-cell reactivity. Tissue samples were biopsied to analyze the histological changes, and flow cytometry was performed to quantify the donor T-cells. Results: Allograft survival was significantly prolonged (>200 d) in Group 6 when compared with the other groups (P < 0.001). The mixed lymphocyte reaction performed for Group 6 revealed hyporesponsiveness of the T-cells to donor alloantigens. Flow cytometric analysis in Group 6 revealed a significant increase in the percentage of CD4⁺/CD25⁺ and CD4⁺/foxP3⁺ T-cells expression, and significant increase in the percentage of donor cells (RT1ⁿ) in the recipient peripheral blood. Immunohistochemical staining of allo-skin revealed a significant increase in the proportion of CD25⁺ cells in the subcutaneous and dermis layers in Group 6, as compared to other groups. Conclusion: Treatment with donor alloantigen-pulsed recipient immature DCs in combination with transient immunosuppression prolongs allograft survival and induced tolerance by inducing T-cell hyporesponsiveness to donor alloantigens and increasing the CD4⁺/CD25⁺ T-cell population.