受者来源的PIR-B转染的树突状细胞对小鼠异基因骨髓移植GVHD的保护作用

发布时间：2018-03-20 02:11

本文选题：异基因骨髓移植　切入点：小鼠　出处：《华中科技大学》2010年博士论文　论文类型：学位论文

【摘要】： 目的：建立一个稳定可靠的异基因骨髓移植(allogeneic bone marrow transplantation, allo-BMT)小鼠急性移植物抗宿主病(acut graft-versus-host disease, aGVHD)动物模型,为后期实研究验allo-BMT后aGVHD和移植耐受提供理想的实验平台。方法：将供者C57BL/6J (H-2b)雄性小鼠骨髓细胞和脾细胞按不同比例(1:1和1：2)混合,输入接受致死性全身照射(8.5Gy)的受鼠雌性BALB/C (H-2d)小鼠以引起不同程度的aGVHD,检测嵌合体情况,根据受鼠的一般情况、临床表现、生存期和组织病理学等判断aGVHD程度。结果：混合不同比例的供鼠骨髓细胞和脾细胞allo-BMT的小鼠均发生了aGVHD,但出现aGVHD的时间和程度均有差异。其中骨髓与脾细胞1：2组小鼠可在相对集中的时间内观察到典型的aGVHDI临床和病理改变,所有小鼠均在14-24d死亡。骨髓与脾细胞1:1组aGVHD程度较轻,移植后4周小鼠均全部存活。结论：骨髓细胞和脾细胞比例为1:2组比1:1组出现更典型的aGVHD,其发病和死亡时间较集中。选定TBI8.5Gy及细胞移植的混合比例(1:2)构建小鼠aGVHD模型,为后续实验提供理想模型。目的：探讨不同方法诱导的受者来源树突状细胞(dendritic cells, DCs)输注对异基因骨髓移植(allogeneic bone marrow transplantation, allo-BMT)小鼠急性移植物抗宿主病(acut graft-versus-host disease,aGVHD)和造血重建的影响。方法：受鼠为雌性BALB/C (H-2d)小鼠,供鼠为雄性C57BL/6J (H-2b)小鼠。无菌分离受鼠骨髓细胞(bone marrow cell, BMC),与GM-CSF共培养得到未成熟树突状细胞(immature dendritic cells, imDCs);受鼠BMC与GM-CSF、IL-10共培养得到IL-10-DCs;用配对性免疫球蛋白样受体B (Paired immunoglobin-like receptor B, PIR-B)慢病毒载体转染imDCs得到PIR-B-DCs。受鼠随机分为四组,在移植骨髓后分别予尾静脉注射imDCs、IL-10-DCs、PIR-B-DCs和RPMI1640培养液。以移植后aGVHD临床表现,肝脏、小肠、皮肤病理组织改变,生存期为观察指标并进行组间比较。结果：PIR-B-DCs移植组、IL-10-DCs移植组、imDCs移植组和单纯移植组受鼠平均生存时间为(46.0±13.6)天、(36.4±13.0)天、(21.6±2.8)天和(17.4±3.6)天,p＜0.01；4组小鼠在移植后15天GVHD临床评分为(5.28±0.27)、(5.26±0.31)、(2.46±0.18)和(0.86±0.21),p0.05。单纯移植组病理显示肝脏汇管区大量淋巴细胞浸润,小胆管塌陷破坏,皮肤基底层连续性中断,有大量淋巴细胞,肠粘膜下层淋巴细胞浸润,肠绒毛萎缩变性,PIR-B-DCs移植组病理组织检查仅有轻微GVHD表现。结论：受者来源PIR-B-DCs联合骨髓移植能够明显延长受鼠生存时间,减轻移植后aGVHD反应,促进造血重建。
[Abstract]:Objective: to establish a stable and reliable animal model of acute graft-versus-host disease graft-versus-host disease (aGVHD) in allogeneic bone marrow transfer (allo-BMTT) mice, and to provide an ideal experimental platform for the later study of aGVHD and transplantation tolerance after allo-BMT. Methods: bone marrow cells and spleen cells of donor C57BL / 6J / H-2b) mice were mixed at different proportions of 1: 1 and 1: 2) and injected into murine female BALB/C / H-2d mice receiving lethal whole-body irradiation for different degrees of aGV HDD, and chimerism was detected. The degree of aGVHD was judged according to the general condition, clinical manifestation, survival time and histopathology of recipient mice. Results: the aGV HDD was found in mice mixed with different proportions of donor bone marrow cells and splenocytes allo-BMT, but the time and degree of aGVHD appeared were different, among which the bone marrow and spleen cells 1: 2 group could be observed in a relatively concentrated time. Typical clinical and pathological changes of aGVHDI, All the mice died from 14 to 24 days. The aGVHD degree of bone marrow and spleen cells 1: 1 group was mild, and all the mice survived 4 weeks after transplantation. Conclusion: the ratio of bone marrow cells to spleen cells in group 1: 2 is more typical than that in group 1: 1, and the onset and death time is more concentrated. The aGVHD model of mice was constructed by selecting the mixed ratio of TBI8.5Gy and cell transplantation (1: 2) to provide an ideal model for further experiments. Aim: to investigate the effects of dendritic cells (DCs) induced by different methods on allogeneic bone marrow transfer (allo-BMTM) and hematopoietic reconstitution in mice with acute graft-versus-host disease, acut graft-versus-host disease, aGVHD. Methods: the recipient mice were female BALB/C (H-2d) mice. The donor mice were male C57BL / 6J ~ (H-2b) mice. Bone marrow cells were isolated from bone marrow cells and co-cultured with GM-CSF to obtain immature dendritic cells dendritic cells, imDCsN; recipient BMC co-cultured with GM-CSFFIL-10 to obtain IL-10-DCs; matched immunoglobulin-like receptor B Paired immunoglobin-like receptor. The imDCs vector was transfected with B, PIR-B-DCs.The recipient mice were randomly divided into four groups. After bone marrow transplantation, we injected imDCssil IL-10-DCssil PIR-B-DCs and RPMI1640 culture medium into caudal vein, respectively. The clinical manifestations of aGVHD, pathological changes of liver, small intestine and skin, survival time were observed and compared among the groups. Results the average survival time of the two groups were 46.0 卤13.6) and 36.4 卤13.0) days and 21.6 卤2.8 days and 17.4 卤3.6 days, respectively. The GVHD clinical scores were 5.28 卤0.27 + 5.26 卤0.31 卤0.18) and 0.86 卤0.21 p0.05respectively. The clinical scores of GVHD were 5.28 卤0.27, 2.46 卤0.18) and 0.86 卤0.21p0.05in the 15 days after transplantation, respectively. The clinical scores of GVHD were 5.28 卤0.27 (2.46 卤0.18) and 0.86 卤0.21 (p0.05) respectively in the 10 ~ (-10) -DCs transplantation group and in the control group. The clinical scores of GVHD were 5.28 卤0.27 (2.46 卤0.18) and 0.86 卤0.21 (p0.05) respectively. A large number of lymphocytic infiltration in the canal area, In the small bile duct collapse, the continuity of the basal layer of the skin was interrupted, there were a large number of lymphocytes and submucous lymphocytes infiltrated in the intestinal submucosa. The pathological findings of PIR-B-DCs transplantation group showed only slight GVHD findings. Conclusion: recipient PIR-B-DCs combined with bone marrow transplantation can significantly prolong the survival time of recipient mice, alleviate aGVHD response after transplantation, and promote hematopoietic reconstitution.
【学位授予单位】：华中科技大学
【学位级别】：博士
【学位授予年份】：2010
【分类号】：R392

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