Donor specific bone marrow cells suppress lymphocyte reactivity to donor antigens and differentially modulate TH1 and TH2 cytokine gene expression in the responder cell population.

Previous studies have shown that post-transplantation infusion of donor specific bone marrow following a non-specific potent immunosuppressive agent such as antilymphocyte globulin (ALG) can significantly enhance graft survival compared to ALG alone. This enhancement remains variable and is thought to occur through the induction of specific partial tolerance to the renal allograft, but the underlying cellular mechanisms have not been clearly identified. In order to improve the efficacy of this specific immunosuppressive treatment and to study the events leading to enhanced allograft survival, we sought to establish a simple in vitro model based on a mixed lymphocyte reaction (MLR). We show that cellular proliferation seen in a normal MLR can be suppressed by addition of donor specific bone marrow cells (BMC). Significantly, this suppression is not observed with either third party BMC or donor specific peripheral blood mononuclear cells (PBMC). We have defined the optimum conditions of bone marrow infusion regarding number of BMC, their handling and culture, and simple enrichment procedures. Using a semiquantitative polymerase chain reaction assay, we have studied the cytokine gene expression in MLR modulated by donor specific BMC. In an unmodified allogeneic response, the responder cells show increased expression of interleukin-2 (IL-2) gamma-interferon IFN-gamma and receptor (IL-2R) mRNA, and no IL-10 mRNA. When responder cells are cultured with BMC of the stimulator, there is a 256-fold decrease in IL-2 mRNA, and a 64-fold decrease in IFN-gamma and IL-2R mRNA. There is also a 64-fold increase in IL-10 mRNA. This effect is even more marked when the BMC are depleted of CD3+ cells. The kinetics of addition of donor specific BMC to the normal allogeneic MLR culture and specificity of the action of BMC are also elucidated. Our data suggest that the enhancement of graft survival observed with donor BMC may operate through decreased proliferation of reactive T cell clones (due to decreased IL-2/IL-2R) and suppressed monocyte functions (due to decreased IFN-gamma and increased IL-10 gene expression).