异基因骨髓移植小鼠Treg、NK细胞及相关细胞因子与aGVHD的关系
Relationship of aGVHD with Treg, NK cells as well as their related cytokines in mice with allogeneic bone marrow transplantation

摘要：目的 探讨异基因骨髓移植小鼠体内调节性T细胞(Treg)、NK细胞以及与其功能相关的细胞因子白介素-10(IL-10)、转化生长因子(TGF-β)、穿孔素与急性移植物抗宿主病(aGVHD)的关系。方法 建立H-2完全不合的C57BL/6→BALB/c纯种小鼠间异基因骨髓移植后发生aGVHD模型。流式细胞仪检测移植后存活的aGVHD小鼠脾脏CD4+CD25+Treg细胞和NK-1.1+NK细胞的比例；ELISA方法检测环孢素A干预aGVHD小鼠血清IL-10、TGF-β和穿孔素的水平，以正常的C57BL/6小鼠为对照。结果 与正常小鼠相比，移植后发生aGVHD小鼠体内Treg细胞比例下降(3.6% vs. 1.55%)，NK细胞比例上升(3.3% vs. 11.5%)；环孢素A治疗aGVHD组小鼠，与未干预组相比，血清IL-10水平明显上升[(125.79±0.27)pg/mL vs. (103.09±3.27)pg/mL，P<0.01]，TGF-β水平上升但无统计学差异[(252.05±7.84)pg/mL vs. (241.61±15.41)pg/mL，P>0.05]，穿孔素水平明显上升[(186.97±4.68)pg/mL vs. (144.35±14.42)pg/mL，P<0.01]。结论 ①小鼠移植后发生aGVHD与Treg细胞比例下降有关，Treg功能相关的细胞因子IL-10、TGF-β参与环孢素A介导的免疫抑制治疗；②NK细胞参与异基因骨髓移植后aGVHD的发生过程，穿孔素的水平上升与抑制aGVHD有关。
ABSTRACT: Objective To investigate the relationship of acute graft versus host disease (aGVHD) and the regulatory T (Treg) cells, NK cells as well as their function-related cytokines such as IL-10, TGF-β, and perforin in mice with allogeneic bone marrow transplantation. Methods H-2 completely mismatched C57BL/6→BALB/c aGVHD mice model was constructed. Flow cytometry analysis was used to detect the proportion of CD4+CD25+ Treg cells and NK-1.1+ NK cells in the spleen of aGVHD mice after transplantation. ELISA method was used to detect the serum levels of IL-10, TGF-β and perforin in the aGVHD mice intervened with CSA prophylactic or not. The normal C57BL/6 mice were used as controls. Results Compared with those of normal mice, the proportion of Treg cells in aGVHD mice after transplantation was decreased (mean 3.6% vs. 1.55%) and the proportion of NK cells increased (mean 3.3% vs. 11.5%). In the aGVHD mice treated with cyclosporine A, serum IL-10 expression level was significantly increased (treated group (125.79±0.27)pg/mL, untreated group [(103.09±3.27)pg/mL, P<0.01)], TGF-β expression level was increased [(252.05±7.84)pg/mL vs. (241.61±15.41)pg/mL, P>0.05], perforin expression level was significantly increased [(186.97±4.68)pg/mL vs. (144.35±14.42)pg/mL, P<0.01]. Conclusion ① The occurrence of aGVHD is correlated with the decreased number of Treg cells after transplantation in mice. Treg cell function-related cytokines IL-10 and TGF-β are involved in the treatment of aGVHD by cyclosporine A-mediated immunosuppression. ②NK cells are involved in the occurrence of aGVHD after allogeneic bone marrow transplantation, and the increased level of perforin is related to the inhibition of aGVHD