Background: Fully MHC-mismatched mouse liver allografts are accepted without immunosuppressive therapy and induce donor-specific tolerance. Liver dendritic cells (DC) exhibit an immature phenotype that may contribute to immune regulation and the induction of tolerance after liver transplantation. DAP12 is a transmembrane protein that integrates activating or inhibiting signals through associated co-receptors. We have shown previously that DAP12 regulates liver DC maturation and that DAP12-deficient liver DC have enhanced T cell allostimulatory properties. We hypothesized that DAP12-deficient donor liver DC might induce stronger allogeneic immune responses and break liver allograft tolerance.

METHODS: 10-12 week-old male B6 WT or DAP12 KO (B6 background) livers were transplanted orthotopically into normal 10-12 week-old male C3H mice. Graft rejection was assessed by host survival and confirmed histologically. Donor-derived DC (H2-Kb⁺CD11c⁺) in the spleen and CD8⁺IFN-Γ⁺ T cells in the graft were quantified by flow cytometry. Cytokine expression in the graft were determined by RT-PCR. Anti-donor T cell proliferative responses were evaluated by CFSE-MLR, while IFN-Γ secretion in MLR supernatants was measured by cytometric bead array.

RESULTS: Unlike WT grafts, DAP12 KO livers failed to induce tolerance and were rejected (MST: WT: 75d vs DAP12 KO 12.5d; p<0.01). A greater incidence of donor-derived DC was detected in the spleens of DAP12 KO liver recipients compared with those given WT livers on day 1 post transplant (WT: 1.76±0.25% vs DAP12 KO: 5.67±1.18% of bulk CD11c⁺ cells; p<0.01). Moreover, IFN-Γ gene expression in the liver was significantly higher in DAP12 KO liver allografts (WT: 365±20-fold increase vs DAP12 KO: 425±23-fold increase; p<0.05) and the intensity of IFN-Γ expression by CD8 T⁺ cells was significantly higher in DAP12 KO liver allografts (WT: 288±38 MFI vs DAP12 KO: 932±22 MFI; p<0.05). Splenocytes from DAP12 KO liver recipients also showed greater proliferation against T cell-depleted donor splenocytes. Moreover, IFN-Γ secretion by responder T cells was higher for the DAP12 KO liver-grafted mice (WT: 145±29 pg/ml vs DAP12: 259±15 pg/ml; p<0.05).

CONCLUSIONS: DAP12 expression in donor livers appears to regulate the migration of donor DCs to host lymphoid tissue, the consequent alloimmune and transplant tolerance.

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