| Aim Allogeneic bone marrow transplantation (Allo-BMT) is the most effective therapeutic modalitity for the cure of hematological disorders, such as aplastic anemia and leukemia. However, the leading cause of transplant failure and the most common complication in Allo-BMT is graft versus host disease (GVHD) . The immunopathgenesis of acute GVHD (aGVHD) is mainly mediated by Th1 cells stemmed from the bone marrow donors. Tim-3 is a transmembrane protein marker which is preferentially expressed on Th1 cells. The ligation of Tim-3 with its ligand galectin-9 downregulates Th1 responses and promotes the periphereal tolerance. This study sought to explore the expression level of Tim-3 and galectin-9 in various organs of aGVHD models in mice. Our results may provide information on the roles of Galectin-9-Tim-3 pathway in the process of aGVHD. Activation of Galectin-9-Tim-3 pathway may beneficial to the prevention of aGVHD and induction of tolerance.Method Lethally (8.0 Gy) irradiated female BALB/c (H-2b) recipients were transplanted with bone marrow cells and splenic cells from male C57BL/6(H-2d)donors. The recipients were observed for the features of GVHD, the dynamics of engraftment. The evidence of complete chimerism was shown by the amplification of mice Y chromosome specific DNA by PCR from the recipient bone marrow. HE stain histology was also performed for livers, intestines and skins in recipients at different time points. The percentage of CD4~+/CD8~+-Tim-3~+ T cells in the spleens was detected by flow cytometry. Western Blot was performed to analyze the expression level of Tim-3 and Galectin-9 in livers and intestines in the recipients suffered from aGVHD.Results (1)Typical features were seen in the mice aGVHD models, such as piloerection, hunched posture, diarrhea and depilation. Pathological evaluation of intestine, liver, skin and kidney 21 days post transplantation suggested that all the affected organs were graded as GVHD IV. (2) PCR assays confirmed that complete chimerism was successfully established in all mice. The hematopoietic disorder was restored 14 days post transplantation, while relapsed 21 days post transplantation. (3) The percentage of CD4~+Tim-3~+ and CD8~+Tim-3~+ cells were elevated 14 days and 21 days post transplantation, then their percentage was downregulated to basal levels. (4) The Tim-3 protein level was higher in liver and intestine tissues of aGVHD mice than that of normal mice, while the galectin-9 protein level was lower in these tissue of aGVHD mice than that of nomal mice. Conclusion Stable and reliable aGVHD model was established. In this model, the expression of Tim-3 on T cells was correlated with the pathogenesis of aGVHD. In the tissues of aGVHD mice, the elevated expression of Tim-3 was accompanied by lowered expression of galectin-9. This phenomenon suggests that the galectin-9-Tim-3 pathway was not sufficient for suppression of Th1 responses which culminated in tissue injury. |
