Introduction: The effector function of gamma-delta (γδ) T cells are well characterized in several infections, autoimmunity, and tumor model. The role of γδ T cells and their regulatory function in transplantation is well characterized.

Method: C57BL/6 mice (H-2^b) were given donor-specific transfusion (DST; 10 X 10⁶ cells/mouse on day -7) and anti-CD40L mAb (clone MR1; 250 μg/injection/mouse on -7, -4, 0, and +4 day relative to transplantation) and transplanted BALB/c mice (H-2^d) tail-skin allografts. Graft survival was monitored. To deplete the total γδ T cells or Vγ2⁺ T cells, recipient mice were given intravenous injection (200 μg/mouse/injection) of anti-mouse γδ TCR mAb (clone UC7-13D5) or anti-mouse TCR Vγ2 mAb (clone UC3-10A6) on -3, +1, +5, +10 day relative to transplantation.

Results: Our results showed that tolerogen (DST plus anti-CD40L mAb) treatment significantly prolonged the survival of skin allografts compared to control mice (Mean survival time, MST 23.5 days vs. 13 days, p=0.008), and depletion of γδ T cells with anti-γδ TCR mAb significantly reduced the tolerogen-induced survival of allografts (MST 16 days; p=0.068). Tolerogen treatment did not significantly change the frequency or the absolute number of total γδ T cells or secretion of inflammatory cytokines (IL-17 and IFN-γ) or regulatory cytokine IL-10 in total γδ T cells in the thymus, spleen, and lymph nodes. Further analysis showed that tolerogen treatment significantly increased the frequency of Vγ2⁺ γδ T cells in the thymus, spleen, and lymph nodes. Tolerogen treatment promoted CD39⁺Vγ2⁺ γδ T cells and suppressed IFN-γ-production in Vγ2⁺ γδ T cells in the spleen, lymph nodes, and allografts. Vγ2⁺ γδ T cells isolated from tolerized mice suppress in vitro differentiation of Th1 cells. Depletion of only Vγ2⁺ subsets of γδ T cells with anti-Vγ2-specific mAb prevented tolerogen-induced survival of allografts (MST 15.5 days). Flow cytometry analysis showed that depleting Vγ2⁺ T cells reduced the frequency of Foxp3⁺CTLA4⁺ regulatory T cells in the secondary lymphoid organs and allografts. This rejection was specific to only allografts but not to the syngenic grafts in the same recipients. Furthermore, the adoptive transfer of only purified Vγ2⁺ T cells from the tolerogen-treated mice to naïve TCR δ^-/- (H-2^b) recipient mice prolonged the survival of BALB/c skin allografts.

Conclusion: Vγ2⁺ subset of γδ T cells promotes the survival of allografts and can be used as an adoptive cellular therapy to prolong the survival of allografts in a transplantation setting.