| Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is one of the treatments that can cure hematologic malignancies, but many patients do not have the chance of receiving transplantation by the lack of an appropriate HLA-matched donor in time. Virtually, most young patients have a HLA 2-antigen or 3-antigen incompatible relative who could serve as a donor immediately. However, the complications of haploidentical stem cell transplantation such as graft-versus-host disease (GVHD) increase. So how can we reduce transplantation-related complications and improve the outcomes of stem cell transplantation is an issue that clinicians have to solve. Recently researchers both domestic and abroad demonstrated that nonmyeloablative hematopoietic stem cell transplantations (NST) with decreased conditioning intensity is much safer, with relatively little complications and good quality of life, and it can also enlarge the indication of transplantations. Total lymphoid irradiation (TLI) is one of the nonmyeloablative conditioniong regimens, which can selectively change cytokines and cell subsets. Therefore, TLI maybe an appropriate transplant conditioning regimen. The subject is to explore the feasibility of TLI as one of the transplant conditioning regimens through experimental study of mice haploidentical hematopoietic stem cell transplantation.Objective:To establish a model of mice H-2 haploidentical nonmyeloablative HSCT. To explore the feasibility of TLI as one of the transplant conditioning regimens.Methods:1.Parental mice from C57BL/6(H-2b) and BALB/C(H-2d) are raised together to produce F1(H-2b/d) hybrids.2. (C57BL/6×BALB/C)F1(H-2b/d) mice are divided into two groups, one group received total lymphoid irradiation (TLI, 4MeVβ-ray, 17 times all together, each time with the dose of 240cGy and dose rate of 84cGy ), the other group received total body irradiation (TBI, 6MV X-ray, with the dose of 12Gy and dose rate of 1Gy/min ). 24 hours after irradiation, the IL-4 and IFN-γsecretion in supernate of cultured mice spleen cells were determined by enzyme-linked immunosorbent assay (ELISA). The proportional changes of Th1,Th2,NK,NKT cells in mice spleen were detected by Flow Cytometer.3. Our experiment used a P→F transplant model, donors were C57BL/6(H-2b) mice, recipients were C57BL/6×BALB/C hybridized mice, and with TLI as conditionding regimen, We establish a model of mice H-2 haploidentical nonmyeloablative HSCT. H-2 haploidentical HSCT with TBI as myeloablative conditionding regimen was established as controlled group. And then, body weight,clinical and pathologic GVHD manifestations,chimerisms and life span of recipients were observed.Results:1.The changes of mice spleen cytokines and cell subsets 24 hours after irradiation: (1) Detection of cytokines by ELISA: The concentration of IL-4 in untreated group after stimulation was (205.26±5.22) pg/ml, while in TBI group, the concentration of IL-4 was (16.82±7.40) pg/ml, and in TLI group, the concentration of IL-4 was (198.93±34.20) pg/ml; The concentration of IFN-γin untreated group after stimulation was (448.06±80.22) pg/ml, while in TBI group, the concentration of IFN-γwas (96.24±10.39) pg/ml, and in TLI group, the concentration of IFN-γwas (125.45±23.83) pg/ml; The ratio of IL-4 and IFN-γwas 0.47±0.10 in untreated group, while in TBI group, the ratio was 0.17±0.06, and in TLI group, the ratio was 1.31±0.16, There was significant difference between TBI group and TLI group about the ratio of IL-4 and IFN-γ. (2) Detection of spleen cell subsets by flow cytometer: The ratio of Th1 and Th2 was 3.87±1.70 in untreated group, while 24 hours after irradiation, the ratio was 1.02±0.36 in TBI group, and 1.32±0.45 in TLI group; There was no significant difference between TBI group and TLI group. The proportion of NK cells in nucleated cells of spleen in untreated mice was 5.36±1.70, while the proportion was 10.24±2.39 in TBI group, and 16.85±4.62 in TLI group; There was significant difference between TBI group and TLI group. The proportion of NKT cells in nucleated cells of spleen in untreated mice was 1.52±0.54, while the proportion was 4.64±1.17 in TBI group, and 3.98±1.38 in TLI group; There was no significant difference between TBI group and TLI group.2 . The results of mice H-2 haploidentical hematopoietic stem cell transplantation: (1) The changes of body weight: In TBI group, median body weight was 31g pre-transplant and 17g four weeks post-transplant; while in TLI group, there was no significant difference before and after transplantation. (2) The development of acute GVHD (aGVHD): In TBI group, the incidence of aGVHD was 100% four weeks after transplantation, while in TLI group, there was no obvious development of aGVHD. (3) Chimerisms: Full chimerisms were observed in TBI group 2 weeks post-transplant, while in TLI group, mixed chimerisms were observed 3~5 weeks post-transplant. (4) Life span: In TBI group, mice begin to die 32 days after transplantation, and by the time of 6 weeks post-transplant, mortality was 80%, while there was no death in TLI group 6 weeks post-transplant.Conclusions:Total lymphoid irradiation(TLI)in our experiment is a relatively safe and efficient nonmyeloablative conditioning regimen, which can provide rapid and sustained engraftment with reduced incidence and lethality of aGVHD. |
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