MG患者胸腺移植BALB／c nu／nu裸鼠模型的建立及其功能研究

The establishment of BALB/c nu/nu nude mouse model of myasthenia gravis by grafting the MG thymus and it functional study

目的:通过建立MG患者胸腺移植BALB/c裸鼠动物模型,探讨MG患者胸腺对脐血干细胞分化的影响。方法:将4mm3无菌切除的MG患者胸腺和非自身免疫病正常胸腺组织行BALB/c裸鼠左肾包膜下移植,2周后给BALB/c裸鼠静脉输入脐血单个核细胞。胸腺移植60天后通过免疫组化试验检测CD3分子的表达,利用流式细胞仪检测BALB/c裸鼠外周血中人源T细胞、B细胞、NK细胞、调节性T细胞数量。结果:胸腺移植60天,移植的胸腺组织在BALB/c裸鼠肾包膜下存活,组织结构清晰,免疫组化显示移植块中的胸腺细胞表达CD3分子;流式细胞分析显示CD3+、CD4+、CD19+、CD161+细胞以及CD3+CD56+、CD4+CD25+细胞百分率在MG胸腺、正常胸腺和脐血MNC联合移植组明显高于对照组,正常组CD3+、CD19+比例增加更明显,MG组CD4+细胞比例高于先心组。结论:胸腺组织块植入BALB/c裸鼠肾被膜下,能有效存活,并参与脐血细胞在体内的分化;移植的MG胸腺与正常胸腺在脐血细胞中的作用存在差异。MG患者胸腺、脐血干细胞联合移植建立的人胸腺-裸鼠动物模型可作为MG胸腺功能研究、免疫细胞分化成熟研究的工具。

Objective：To determine the effect of the thymuses from patients with myasthenia gravis （MG） on differentiation of cord blood stem cells. Methods： Thymuses from MG patients were transplanted into nude mice of BALB/c to establish the animal models by the following methods. 4 mm3 of thymses from MG patients, congenital heart disease patients were transplanted respectively into the subeapsular region of left kidneys of nude mice of BALB/c. After 2 weeks, human cord blood stem cells were transplanted into the nude mice through intravenous injection. 60 days after thymus transplantation, CD3 were detected with immunohistochemical test and the amounts of humanized T cells, B cells, NK cells and Ts cells from the transplanted nude mice were detected by flow cytometry. Results：The thymus transplants surived for over 60 days in nude mice of BALB/c with evidence nf restoration of thymic structure. It is manifested with immunohistochemical test that CD3 were present in the thymus transplants. It is found by flow cytometry that the transplantation with thymus from patients with MG and congenital heart disease held a significant higher percentage of CD3^＋ , CD4^ ＋ , CD19^＋ , CD161^＋ cells and CD3^＋ CD56^＋ , CD4^＋ CD25^＋ cells compared with those of the control group. The CD3^＋ and CD19^＋ cells were more and CD4＋ cells were less when compared the two groups of normal and MG. Conclusion：The findings suggest that the thymus transplants survive in nude mice of BALB/c and the transplantation restore thymic tunction. It can promote in vivo proliferation of cord blood stem cells transfused in unde mice. There exists difference in the promotion of cord blood stem cell proliferation between the group of MG transplantation and the control group. The hmnan thymus-nude mouse animal model can be established hy transplanting the MG thymus combined with transfusion of cord blood stem cells. The animal model provides a good approach to MG thymic function study and also to the study in immune cell differentiation.