Objective: To establish the aGVHD mouse model，and investigate the regulatory effect and its mechanism of low-dose GSI combined with BMSC on aGVHD mice.
Methods: C57BL/6 (H-2b) and BALB/c (H-2d) were selected as donor and recipient of allogeneic transplantation to establish the aGVHD mouse model. BALB/c mice were randomly divided into 6 groups, which were the bone marrow cell infusion after irradiation (BM) group; the bone marrow cells + spleen cells after irradiation (BM+SC) group; the bone marrow cells + spleen cells + DMSO (BM+SC+DMSO) (transplant control) group; bone marrow cells + splenocytes +GSI after irradiation (BM+SC+GSI) group; bone marrow cells + spleen cells + bone marrow mesenchymal stromal infusion after irradiation cell (BM+SC+BMSC) group; bone marrow cells + spleen cells + bone marrow mesenchymal stromal cells +GSI infused after irradiation (BM+SC+BMSC+GSI) group. The mice in the two groups containing GSI were intraperitoneally injected with GSI at 5 μmol/kg on day 1, 2, and 3 after transplantation with DMSO as a control. The general conditions, survival time and hematopoietic recovery of mice were observed, cytokines were detected by ELISA, and histopathological changes were detected by immunohistochemistry. The effects of low-dose GSI combined with BMSC on hematopoietic reconstruction and aGVHD development after allo-BMT were investigated.
Results: The survival rate of the mice in BM+SC+BMSC+GSI combination group was 80% during the observation period, which was significantly higher than that in the other groups; the incidence of aGVHD was reduced in the BMSC GSI or their combination groups after 21 days of transplantation. GSI could partly promote the recovery of leukocytes, and show no significant delayed effect on the recovery platelets. Moreover, the level of Th1 cytokines (IFN-γ) in BM+SC+BMSC+GSI combined group was lower than that in BM+SC+GSI group (P<0.01), the level of Th2 cytokines (IL-4) in the combination group was higher than that in BM+SC+GSI group (P<0.01), also the level of IL-17 was significantly lower than that in the corresponding control group (P<0.001).
Conclusion: Low dose GSI combined with BMSC can promote hematopoietic reconstruction and regulate cytokines secretion including IFN-γ, IL-4 and IL-17. GSI combined with BMSC achieve the goal of synergistically inhibiting the occurrence and progression of aGVHD.
题目: γ分泌酶抑制剂联合BMSC对小鼠急性移植物抗宿主病的影响.
目的: 建立aGVHD小鼠模型，探讨低剂量γ分泌酶抑制剂（GSI）联合BMSC对小鼠aGVHD的调控作用及其机制.
方法: 选择C57BL/6（H-2b）→BALB/c（H-2d）作为异基因移植供、受体，建立aGVHD模型。BALB/c受鼠随机分为6组，分别为照射后输注骨髓细胞组（BM）, 照射后输注骨髓细胞与脾细胞组（BM+SC）, 照射后输注骨髓细胞与脾细胞+DMSO组（BM+SC+DMSO）（移植对照组）, 照射后输注骨髓细胞与脾细胞+GSI组（BM+SC+GSI）, 照射后输注骨髓细胞与脾细胞及骨髓基质细胞组（BM+SC+BMSC）, 照射后输注骨髓细胞与脾细胞及骨髓基质细胞+GSI组（BM+SC+BMSC+GSI）。含GSI的两组小鼠在移植后1、2、3 d以腹腔注射GSI 5 μmol/kg，以DMSO为对照。观察小鼠一般状况、生存时间及造血恢复情况，ELISA法测定细胞因子，免疫组化检测病理组织学改变等，研究其对allo-BMT后造血重建和aGVHD发生发展的影响.
结果: BMSC与GSI联合组的小鼠在观察期内存活率80%，明显高于其他各组小 鼠, BMSC、GSI或者两者联合治疗组在移植后21 d以上时，aGVHD的发生率均降低。GSI部分促进白细胞恢复，且对血小板的恢复无明显延迟作用。同时，BMSC与GSI联合组小鼠的外周血清中Th1型细胞因子（IFN-γ）含量明显低于GSI组小鼠（P<0.01），联合组Th2型细胞因子（IL-4）含量明显高于GSI组小鼠（P<0.01），IL-17含量显著低于相应对照组（P<0.001）.
结论: 低剂量GSI联合BMSC促进造血重建并调节细胞因子IFN-γ、IL-4及IL-17的分泌。GSI联合BMSC达到协同抑制aGVHD发生及进展的目的.