Studies On Graft Versus Host Disease: Establishment Of Animal Models And Comparison Of Clinical Prevention Regimens

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is by now the most efficient treatment for hematological malignancies and some solid tumors. Allogeneic donor T lymphocytes are also used for treatment of relapse or lymphoma associated with Epstein-Barr virus infection post transplantation. In recent years, great progress has been made in related disciplines, from which the long-term survival has been achieved, however, graft versus host disease (GVHD) induced by allogeneic immune cells remains to be one of the major causes of mortality and morbidity in the recipients of allo-HSCT. To date, it is generally acknowledged that the major effector of GVHD is T lymphocytes. In clinical settings, acute GVHD occurred in 30-70% patients after all-HSCT, although effective prophylatic measures were already taken. In our former studies, we had established a murine GVHD model in which C57BL/6 mice ware used as a donors and Balb/c mice as recipients. This GVHD animal model was a good platform for the study of GVHD, however, the role of human allogeneic stem cell grafts and T lymphocytes in the mechanism of GVHD and its prevention and treatment should be based on a model closer to clinical settings. In the present study, we established a NOD/SCID-Hu-GVHD xenograft chimera model based on NOD/SCID mice. The distribution of human T lymphocyte in NOD/SCID mice and its relation with GVHD, as well as the application of novel suicide YCD/5-FC in T lymphocytes, were investigated. In the other hand, once the severe acute GVHD occurred, it is troublesome for its treatment. Efficient prophylaxis for GVHD is the most important measure to be taken in clinical practice. To further optimize the regimen of GVHD prevention, we design a novel regimen including low dose cyclosporin A (CsA) plus short course methotrexate (MTX) and mycophenolate mofetil (MMF), and compared its efficiency with traditional prevention regimen, CsA+MTX. Part one The establishment of NOD/SCID-hu chimera GVHD model To establish a NOD/SCID-hu chimera GVHD model, 2×107 (A group, n=4) and 5×107 (B group, n=4) mononuclear cells(MNC)from G-CSF mobilized peripheral blood (PB) of healthy donor were intravenously injected into a lateral tail vein of NOD/SCID mice, respectively. Control mice (n=4) received 0.9% sodium chloride without cells. The manifestation and survival time of mice were observed. Peripheral 11PDF 文件使用 "pdfFactory" 试用版本创建 www.fineprint.com.cn第二军医大学博士研究生学位论文 英文摘要 blood was collected from the retro-orbital plexus every 7 days, MNCs isolated, resuspended in phoshate-buffered saline (PBS), and the percentage of human CD45+, CD3+, and CD4+ cells were measured by FACS. Sixty days after transplantation, or when dying, mice were sacrificed by CO2 anesthesia, liver, spleen, bone marrow (BM) and PB were collected, MNCs isolated, and the percentages of CD3+, CD4+, CD45+ cells measured by FACS. The occurrence and severity of GVHD was evaluated by pathology and manifestations of the mice. Results The decrease of weight, hunched back , and pale were observed in both A and B groups, and no diarrhea showed up. The average survival time were 42.7±12.0 days and 40.4±7.7 days, respectively, while mice in control group was all lived up to 60 days. The difference in survival time was statistically significant between the study groups and control group, but not between Aand B groups. The percentages of CD45+/CD3+/CD4+cells in liver, spleen, BM and PB of the mice were detected by FACS. In control group, no CD45+/CD3+/CD4+ were detected. In A group, the percentages of CD45+,CD3+ and CD4+ cells were 80.69±16.38%, 73.32±11.06% and 22.49±2.37% in liver, 75.88±14.58%, 70.53±15.69% and 20.46±1.49% in spleen, 9.64±1.83%, 8.76?...