地塞米松联合IL-2对供鼠体内调节性T细胞选择性扩增作用及对急性移植物抗宿主病反应的抑制作用

目的 通过地塞米松(Dex)联合IL-2处理小鼠,建立体内扩增调节性T细胞(Treg细胞)的方法 .观察用该方法 处理的小鼠脾细胞移植后受鼠急性移植物抗宿主病(aGVHD)的发生与转归.方法 使用Dex和IL-2处理雄性C57BL/6N供鼠3 d后提取脾单个核细胞(MNC),用流式细胞术(FCM)分析CD4~+ CD25~+、CD25~+ FOXP3~+细胞的变化.,以以上方法 处理的雄性C57BL/6N小鼠为供者,对雌性BALB/c受鼠进行非清髓异基因淋巴细胞移植.移植后观测受鼠的生存率、生存时间、组织病理学变化,以及用PCR和FCM分析测定受鼠嵌合体等来评价aGVHD的发生.结果 经过Dex和IL-2联合处理,供鼠脾CD4~+ CD25~+ FOXP3~+ Treg细胞数量(24.2±7.6)%]明显高于对照组(4.0±0.8)%](P=0.01).Dex联合IL-2组供鼠Treg细胞与效应T细胞(Teff)的比值(0.43±0.15)显著高于对照组(0.14±0.01)(P=0.01).经过Dex联合IL-2处理供鼠后进行异基因淋巴细胞移植,受鼠aGVHD明显减轻,中位生存时间＞60 d,与对照组的中位生存时间12 d相比明显延长(P=0.0045).对照组出现典型的aGVHD表现,移植后2周Dex联合IL-2组和对照组的总体死亡率分别为29.4%和71.4%(P＜0.05).结论 经IL-2和糖皮质激素处理供鼠后进行主要组织相容性抗原复合物完全不相合脾淋巴细胞移植能明显减轻aGVHD,显著延长生存时间,可能与供者体内诱导扩增的Treg细胞增加有关.

Combination of dexamethasone with IL-2 selectively induces the expansion of CD4~+ CD25~+ FOXP3~+ regulatory T cells in vivo and suppresses graft versus host disease

Objective To establish a method for increasing T regulatory cells (Treg) in the graft by using in vivo treatment with dexamethasone( Dex) plus IL-2 and observe its suppressing effect on graft-versus-host-disease(GVHD) in mice. Methods After treatment of male C57BL/6N mice (donor) with Dex (5 mg ·kg~(-1)·d~(-1) ) combined with IL-2 (300 000 IU· mouse~(-1)·day~(-1)) for three days, spleen mononuclear cells were isolated for flow cytometry analysis of CD4~+ CD25~+ POXP3~+ Treg cells. The allogeneic lymphocytes were transplanted from male C57BL/6N mice to female BALB/c mice aged 8-12 weeks. GVHD and survival time were investigated after transplantation. Donor-derived hematopoiesis reconstituted in recipient mice was detected by Y-chromosome-specific PCR and H-2K~b by flow cytometry. Results Administration of Dex and IL-2 markedly expanded functional CD4~+ CD25~+ FOXP3~+ Treg cells in murine spleen, the number of which in treated group was (24.2±7.6)% while in control group was (4.0±0.8)% (P =0.01). The ratio of Treg to effector T eells( Teff) increased obviously in the treated group (0.43±0. 15 vs 0. 14±0. 01 , P = 0. 01). In a murine allogeneic lymphocyte transplantation model, the grafts from donor with combined treatment of Dex and IL-2 led to a longer survival time than that from the control group( median survival time >60 d vs 12 d, P = 0. 0045 ) , while the mortality rate was decreased ( 29.4% vs 71.4% , P <0.05). Conclusion Costimulation with Dex and IL-2 can selectively expand the functional CD4~+ CD25~+ FOXP3~+ Treg in vivo, which can suppress acute GVHD.