异基因造血干细胞移植后慢性移植物抗宿主病小鼠模型的建立和评价  

Establishment and evaluation of mouse model of chronic graft-versus-host disease after allogeneic hematopoietic stem cell transplantation

作　　者：伊文芳[1] 郭坤元[2] 贺信[2] 林广[1] 张成[1] 王静[2] YI Wen-fang GUO Kun-yuan HE Xin LIN Guang ZHANG Cheng WANG Jing(Department of Pediatrics,Zhuhai People＇s Hospital,Zhuhai 519000,Guangdong,China Department of Hematology,Zhujiang Hospital,Southern Medical University,Guangzhou 510280,Guangdong,China)

机构地区：[1]珠海市人民医院儿科,珠海519000 [2]南方医科大学珠江医院血液内科,广州510280

出　　处：《第二军医大学学报》2016年第12期1501-1505,共5页Academic Journal of Second Military Medical University

基　　金：珠海市医学科研基金(2015J030)~~

摘　　要：目的采用化疗药物预处理方案建立异基因造血干细胞移植的慢性移植物抗宿主病小鼠模型,并进行评价。方法以BALB/c^(H-2kd)小鼠作为供鼠,C57BL/6^(H-2kb)小鼠为受鼠。对受鼠使用不同的化疗药物进行预处理[方案1:白消安20mg/(kg·d)×4d+环磷酰胺150mg/(kg·d)×2d;方案2:白消安20 mg/(kg·d)×4d+环磷酰胺100 mg/(kg·d)×2d],然后经尾静脉注射不同剂量的供鼠脾细胞(6×10~7或4×10~7个)和(或)相同剂量的骨髓细胞(2×10~7个),建立慢性移植物抗宿主病小鼠模型;采用嵌合体分析、临床评分、组织病理学等进行评价。结果采用白消安20mg/(kg·d)×4d+环磷酰胺150mg/(kg·d)×2d的方案进行预处理、2×10~7个骨髓单个核细胞+6×10~7个脾单个核细胞进行移植可形成较高水平的供受者混合嵌合体。移植物抗宿主病发生时间多集中在供鼠脾细胞输注后30~90d,化疗药物剂量大的预处理方案及移植细胞数高的移植组小鼠的临床评分和慢性移植物抗宿主病的发生率高于化疗药物剂量小的预处理方案及移植细胞数少的移植组(P<0.05)。模型小鼠肠、肝脏、皮肤、脾脏等器官出现细胞和结构异常、炎症细胞浸润等病理改变。结论采用白消安和环磷酰胺预处理、给予2×10~7个骨髓单个核细胞+6×10~7或4×10~7个脾单个核细胞可形成较稳定的慢性移植物抗宿主病小鼠模型,为进一步指导临床治疗慢性移植物抗宿主病奠定了实验基础。Objective To establish a mouse model of chronic graft-versus-host disease（GVHD）after allogeneic hematopoietic stem cell transplantation using chemotherapy preconditioning regimens.Methods The donor mouse was BALB/c H-2kd and the recipients mouse was C57BL/6H-2kb.The pretreatment of recipient mouse was performed using different doses of chemotherapy drugs,busulfan（BU）20mg/（kg·d）×4d＋cyclophosphamide（CTX）150mg/（kg·d）×2d,or BU 20mg/（kg·d）×4d ＋ CTX 100mg/（kg·d）×2d,and then the recipient mouse was injected with different doses of spleen cells（6×10^7 or 4×10^7 cells）and/or same dose of bone marrow cells（2×10^7 cells）to establish the model of chronic GVHD.Hematopoietic chimerism analysis,clinical score and histopathology were used to evaluate the model.Results The high level of recipient-donor mixture chimera was established by pretreated by BU 20mg/（kg·d）×4d ＋CTX 150mg/（kg·d）×2dand2×10^7 bone marrow mononuclear cells＋6×10^7 spleen mononuclear cells.The onset of GVHD was concentrated at the 30-90 d after injecting spleen cells of the donor mouse.The clinical scores and incidence rate of chronic GVHD were significantly increased in the large numbers of cells and high doses of chemotherapy transplantation group compared with those in smaller numbers of cells and lower doses of chemotherapy groups（P〈0.05）.Pathological changes such as cellular and structural abnormalities and inflammatory cell infiltration were noted in the intestines,liver,skin and spleen of the model mice.Conclusion Bone marrow mononuclear cells（2×10^7 cells）＋ spleen mononuclear cells（6×10^7 or 4×10^7 cells）can be used to establish a stable chronic GVHD mouse model via preconditioning with BU and CTX,which paves a way for clinical treatment of chronic GVHD.

关 键 词：动物模型 移植物抗宿主病 同种移植 造血干细胞移植 

分 类 号：R457.7[医药卫生—治疗学]