摘要
目的：诱导及鉴定慢性移植物抗宿主病（cGVHD）狼疮小鼠模型,分析其肾病变特点及狼疮相关免疫学特征。方法：选取6 -8周龄（C57BL/10×DBA/2）F1雌性小鼠,随机分为两组,模型组12只,对照组7只,分别于第0、3、8天经尾静脉注射DBA/2雌鼠胸腺和脾的淋巴细胞（模型组）或磷酸盐缓冲液（对照组）。利用考马斯亮蓝法监测小鼠尿蛋白变化;检测小鼠血清的抗核抗体（ANA）、抗双链DNA抗体、抗核小体抗体及抗可提取核抗原（ENA）抗体;利用流式细胞技术分析CD4＋CD25＋调节T细胞的变化;应用实时定量PCR法测定两组小鼠外周血单个核细胞（PBMC）中Foxp3的表达量;于12周末分别用PAMS染色及直接免疫荧光法检测小鼠的肾病理变化。结果：模型组小鼠的尿蛋白自第6周明显增高,与对照组比较差异有统计学意义（P〈0.05）。模型组小鼠血清中ANA抗体阳性率于第8、12周时明显高于对照组;随小鼠周龄增加,模型组小鼠抗dsDNA抗体及抗核小体抗体的滴度增高,与对照组相比差异有统计学意义（P〈0.05）。12周时模型组外周血CD4＋CD25＋调节T细胞的比例明显低于对照组（P=0.002）,但两组间外周血PBMC Foxp3的表达量差异无统计学意义（P〉0.05）。模型组小鼠肾病理的PASM染色可见典型狼疮肾炎病理改变,直接免疫荧光检测可见IgG广泛沉积;而对照组小鼠肾无以上表现。结论：cGVHD模型鼠不仅出现了类似人类狼疮肾炎的尿蛋白变化,血清中还出现多种自身抗体,CD4＋CD25＋调节T细胞明显异常,其肾病理呈典型的狼疮肾炎改变。因此,cGVHD模型鼠可作为一种比较好的系统性红斑狼疮相关基础研究及新疗法的动物模型。
Objective：To establish the murine systemic lupus erythematosus （SLE） model of chronic graft versus host diseases（cGVHD）. To analyze the pathological changes and serological and immunological features in the animals. Methods： Female （C57BL/10 × DBA/2）F1 hybrids aged 6 -8 weeks were randomly divided into model group and healthy controls （HC）. Lymphocytes from female DBA/2 were injected intravenously to the model group on days 0, 3 and 8,while PBS were injected to the HC under the same condition as a control group. Bradford was applied to monitor the development of albuminuria quantitively. Sera were tested by enzyme linked immunosorbent assay （ELISA） and indirect immunofluorescence （IIF） for the presence of autoantibodies. between the two groups by flow cytometry （FCM） To compare the differences of CD4 ^＋ CD25 ^＋ Treg cells and the differences in the expression of Foxp3 by real time polymerase chain reaction（RT-PCR）. The kidneys of model mice were removed in the 12^th week and were made frozen sections for direct immunofluorescence（DIF） and paraffin imbedding for PASM staining. Results： The titers of proteinuria in model group in the 6^th week, 8^th week, 10^th week, and 12^th week were significantly higher than those of the HC groups（P = 0. 004, 0. 005 ,respectively）. The titers of anti-dsDNA and anti-nucleosome antibodies were significantly increased in the model group compared with the HC （P 〈 0. 05）. And the positive rates curves of ANA, anti-dsDNA Abs and anti-nucleosome Abs in model group were significantly different from those of control group （ P 〈 0.05）. And The proportions of CD4^＋ CD25^＋ regulatory T cells from peripheral blood of model group were significantly lower than those of control group （P = 0. 002）, while the expression of Foxp3, one of the most important biomarkers of Treg cells, was not significant. There were mesangial matrix expansion and mesangial cell proliferation in the nephritic pathology in model group and the depositons of IgG along the glomerular capillary walls and in the mesangium were observed in model group. There were no pathological changes and depositons in HC group. Conclusion： It has been proved that there are not only protienuria and autoantibodies, but also decrease of the regulatory T lymphocytes in murine model of cGVHD. All of these results suggest that the cGVHD murine SLE models were successfully established.
出处
《北京大学学报（医学版）》
CAS
CSCD
北大核心
2008年第4期419-424,共6页
Journal of Peking University:Health Sciences
基金
北京市自然科学基金(7072085)资助~~
关键词
移植物抗宿主病
红斑狼疮
系统性
模型
动物
Graft vs host disease
Lupus erythematosus, systemic
Models, animal
