中国组织工程研究

中国组织工程研究 ›› 2010, Vol. 14 ›› Issue (40): 7524-7528.doi: 10.3969/j.issn.1673-8225.2010.40.025

• 干细胞基础实验 basic experiments of stem cells • 上一篇    下一篇

建立人源β-地中海贫血小鼠模型

赖颖晖，赖永榕，杨高晖，罗  琳   

  1. 广西医科大学第一附属医院血液科，广西壮族自治区南宁市 530021
  • 出版日期:2010-10-01 发布日期:2010-10-01
  • 通讯作者: 赖永榕，博士，教授，广西医科大学第一附属医院血液科，广西壮族自治区南宁市 530021
  • 作者简介:赖永榕，博士，教授，广西医科大学第一附属医院血液科，广西壮族自治区南宁市 530021
  • 基金资助:

    国家自然科学基金项目(30860307)，课题名称：反义寡核苷酸对人源β地中海贫血鼠模型α珠蛋白基因的调控作用。广西研究生教育创新计划资助项目(2009105981002D60)，课题名称：建立人源β-地中海贫血小鼠模型的实验研究。

Establishment of humanized beta-thalassemia mouse models

Lai Ying-hui, Lai Yong-rong, Yang Gao-hui, Luo Lin   

  1. Department of Hematology, First Affiliated Hospital, Guangxi Medical University, Nanning  530021, Guangxi Zhuang Autonomous Region, China
  • Online:2010-10-01 Published:2010-10-01
  • Contact: Lai Yong-rong, Doctor, Professor, Department of Hematology, First Affiliated Hospital, Guangxi Medical University, Nanning 530021, Guangxi Zhuang Autonomous Region, China
  • About author:Lai Ying-hui☆, Doctor, Attending physician, Department of Hematology, First Affiliated Hospital, Guangxi Medical University, Nanning 530021, Guangxi Zhuang Autonomous Region, China yinghuilai@sina.com
  • Supported by:

     the National Natural Science Foundation of China, No. 30860307*; the Postgraduate Education Innovation Project of Guangxi Zhuang Autonomous Region, No. 2009105981002D60*

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摘要：

背景：没有适合的β-地中海贫血动物模型的发展，就没有β-地中海贫血基因治疗的进步，而小鼠模型是最有用的且通常用于第一线的体内试验。
目的：拟建立人源β-地中海贫血小鼠模型，为β-地中海贫血的基因治疗提供与人类疾病相似、可靠的动物模型。
方法：非肥胖糖尿病/重症联合免疫缺陷(NOD/SCID)小鼠15只，随机均分为3组：β-地贫/HbE骨髓移植组、β-地贫骨髓移植组、空白对照组。取β-地中海贫血/HbE和β-地中海贫血患者骨髓的单个核细胞，分别从尾静脉输注给经60Co γ射线350 cGy全身照射预处理后的β-地贫/HbE骨髓移植组、β-地贫骨髓移植组小鼠，空白对照组输注RPMI-1640培养液。移植后小鼠腹腔注射重组人粒-巨噬细胞集落刺激因子及重组人促红细胞生成素。观察小鼠一般状况和存活情况，移植5周后取小鼠骨髓及外周血，检测人CD45+细胞比例、人β-地贫基因，观察红系细胞内α-珠蛋白链的沉积情况、小鼠的肝脏和脾脏病理及铁染色检查。
结果与结论：移植5周后，小鼠骨髓和外周血人CD45+细胞比例结果分别为：β-地贫/HbE骨髓移植组为7.21%、4.08%(+47 d)，7.37%、6.03%(+61 d)；β-地贫骨髓移植组为8.17%、3.43%(+36 d)，16.82%、8.89%(+45 d)，空白对照组均为0。β-地贫基因检测结果与供者相符，部分红系细胞内出现电子致密物(过剩α-珠蛋白肽链)沉积。移植组小鼠脾脏长度与其体质量的比值比空白对照组的大(P < 0.05)，肝脾出现较明显的铁沉积。提示，所建立的小鼠模型具有人源化β-地中海贫血的表现，基本符合人源β-地中海贫血小鼠模型的要求。

关键词: 动物模型, &beta, -地中海贫血, NOD/SCID小鼠, 60Co &gamma, 射线, 人源化

Abstract:

BACKGROUND: Progress in the gene therapy of β-thalassemia would not have been possible without the development of suitable animal models of the disease. Mouse models are the most useful and are generally adopted as first line in vivo studies.
OBJECTIVE: To generate humanized β-thalassemia mice models, and to provide the similar reliable humanized animal models towards the genetic treatment of β-thalassemia.
METHODS: Totally 15 nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice were randomly divided into three groups: β- thalassemia/HbE bone marrow transplantation group, β-thalassemia bone marrow transplantation group and blank control group. The bone marrow mononuclear cells of β-thalassemia/HbE and β-thalassemia patients were transplanted into the sublethally irradiated (60Coγ ray 350cGy) NOD/SCID mice of the transplantation group by tail vein injection. RPMI-1640 medium was infused into blank control group. After transplantation, human recombinant growth factors consisting of recombinant human erythropoietin (rhEPO) and recombinant human granulocyte-macrophage colony stimulating factor (rhGM-CSF) were injected into the mice intraperitoneally. The survival status of the mice was recorded. Five weeks after transplantation, bone marrow and peripheral blood were collected to detect human CD45+cell proportion, β-thalassemia gene, to observe α-globin chain precipitates in erythroid cells.The spleen and liver were observed, and iron staining was performed.
RESULTS AND CONCLUSION: Five weeks after transplantation, the percentages of human CD45+ cells of the β-thalassemia/HbE bone marrow transplantation group in bone marrow and peripheral blood were 7.21% and 4.08% in +47 day, 7.37% and 6.03% in +61 day, respectively; those of the β-thalassemia bone marrow transplantation group were 8.17% and 3.43% in +36 day, 16.82% and 8.89% in +45 day, respectively. While in blank control group, the percentage of that of each mouse was 0, respectively. The results of β-thalassemia gene mutations corresponded with the donor. There were α-globin chain precipitates in some erythroid cells. The ratios of the spleen length to body weight of the transplantation group were greater than those of blank control group (P < 0.05). The iron deposition was observed in liver and spleen of mice from the transplantation group. Results suggest that there are humanized cell and β-thalassemia features in the mice of β-thalassemia or β-thalassemia/HbE bone marrow transplantation group, and humanized β-thalassemia mice models are generated successfully.

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