Efficacy and limitations of cyclophosphamide-induced tolerance against alphaGal antigen

In the present study, we have elucidated the efficacy of two cyclophosphamide (CP)-induced tolerance protocols for the induction of B-cell tolerance against Galalpha1-3Galbeta1-4GlcNAc (alphaGal) antigens. alpha1,3-galactosyltransferase-deficient (GalT-/-; H-2(b/d)) mice received with 1 x 10(8) AKR (alphaGal+/+ H-2k) spleen cells (SC) followed by 200 mg/kg CP, or alternatively followed by 200 mg/kg CP, 30 mg/kg Busulfan (BU) and 1 x 10(8) T-cell-depleted AKR bone marrow cells (BMC). The generation of both anti-alphaGal and anti-donor antibodies were completely suppressed, but normal antibody production against third party antigens was observed after BALB/c skin grafting in both groups of GalT-/- mice. In GalT-/- mice, treated with SC and CP, mixed chimerism was not observed. Cellular rejection was observed in grafted donor AKR hearts with an absence of humoral rejection, whereas humoral rejection was observed in untreated GalT-/- mice. On the other hand, long-term mixed chimerism and permanent acceptance of donor AKR skin graft and heart graft were achieved in GalT-/- mice treated with SC, CP, BU and BMC. These results demonstrate the efficacy of classical drug-induced tolerance in the induction of B-cell tolerance against alphaGal antigens. However, induction of stable mixed chimerism was required for the suppression of cellular rejection.