Objective:To investigate the effect of donor lymphocyte infusion (DLI) by intra-bone marrow (IBM) or intravenous (IV) on the incidence of graft-versus-host disease(GVHD) after allogeneic peripheral hematopoietic stem cell transplantation (allo-PBSCT).Methods:Female C57BL/6 mice as recipients received total body irradiation (TBI) on day 0,followed by injection of peripheral hematopoietic stem cells from mobilized male BALB/c with granulocyte-colony stimulating factor (rhG-CSF),and DLI was performed via IV or IBM.The extent of GVHD was compared in recipients received allogeneic IBM-DLI with those received IV-DLI.The percentage of donor-derived cells and CD4~+CD25~+ regulatory T cells (Tregs) was detected by flow cytometry.14 days after DLI,the levels of IL-4 and interferon (IFN)-γ were tested by ELISA.Results:It was found that the frequency and severity of GVHD were reduced in IBM-DLI compared with that of IV-DLI (P＜0.01).7 days after transplantation,the percentage of H-2~d-positive cells was over 95% in all surviving transplanted mice;and 14 days after transplantation,the percentage of Tregs detected as CD4~+CD25~+ was significantly higher in recipients treated with IBM-DLI than those treated with IV-DLI(P＜0.01).Compared with that of the recipients in IBM-DLI group,the level of IL-4 was significantly decreased,while the level of IFN-γ were elevated in group IV-DLI (P＜0.01).Conclusion:IBM-DLI could induce the proliferation of Tregs and the Th polarizing to Th2,resulting in decreasing the incidence and alleviating severity of GVHD after allo-PBSCT.