上海交通大学学报（医学版）

上海交通大学学报（医学版）

• 论著（基础研究） • 上一篇    下一篇

转基因IgA肾病小鼠模型的建立与鉴定

谷裕，刘爽，朱逸，林芙君，蒋更如   

  1. 上海交通大学 医学院附属新华医院肾脏科， 上海 200092
  • 出版日期:2016-01-28 发布日期:2016-02-26
  • 通讯作者: 蒋更如, 电子信箱: jianggeng-ru@hotmail.com。
  • 作者简介:谷裕（1989—）， 女， 硕士生; 电子信箱: guyu198901@163.com。
  • 基金资助:

    上海市科学技术委员会基金（12140903002）； 上海市卫生和计划生育委员会科研课题（20144Y0145）

Establishment and identification of a mice model of transgenic IgA nephropathy

GU Yu, LIU Shuang, ZHU Yi, LIN Fu-jun, JIANG Geng-ru   

  1. Department of Nephrology, Xinhua Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200092， China
  • Online:2016-01-28 Published:2016-02-26
  • Supported by:

    Science and Technology Commission of Shanghai Municipality, 12140903002； Municipal Commission of Health and Family Planning Projects of Shanghai， 20144Y0145。

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摘要：

目的 利用转基因方法构建一种新的自发性IgA肾病小鼠模型，模拟人体IgA1分子铰链区结构的改变在IgA肾病发病过程中的作用。方法 利用转基因方法将野生型（转基因A组）及糖基化位点突变（转基因B组）的人IgA1铰链区序列转入C57BL/6J小鼠体内，置换小鼠原有的铰链区序列。依据遗传学规则进行繁殖，所有小鼠均经过PCR检测筛选出基因型阳性。阳性亲代鼠于24周龄，阳性F1代小鼠于8周龄采集尿样进行性状观察、尿沉渣镜检和尿蛋白/肌酐检测，取肾脏组织进行IgA免疫荧光染色和H-E染色以评估IgA沉积情况和肾脏组织学变化；心脏采血分离血清，检测IgA、IgG水平。 结果 转基因A组和B组分别得到阳性亲代鼠3只和8只，分别获得阳性F1代小鼠17只和52只。2个转基因组的阳性亲代鼠和F1代小鼠肾脏IgA免疫荧光染色显示，IgA在肾小球系膜区沉积；H-E染色显示肾小球系膜细胞增生，系膜基质增多，间质有炎症细胞浸润；新鲜尿沉渣镜检均未见红细胞。尿蛋白/肌酐、血清IgA和IgG水平均显著高于同周龄野生型小鼠。结论 采用转入人IgA1铰链区的方法可以构建出IgA肾病小鼠模型，IgA1铰链区结构的改变在IgA肾病发生中具有重要的作用。

关键词: IgA肾病, 转基因, 小鼠, 铰链区, 糖基化

Abstract:

Objective To mimic the effects of structural changes of the hinge area of human IgA1 molecules on the pathogenesis of IgA nephropathy via a novel mouse model of spontaneous IgA nephropathy established by the transgenic method. Methods Sequences of the hinge area of wild type (transgenic group A) and O-glycan sites-mutated (transgenic group B) human IgA1 were inserted into the fertilized eggs of C57BL/6J mice by the transgenic method to replace the original sequences. Mice propagated based on genetic rules and genotype-positive mice were identified by PCR. The urine samples of positive parental mice at the age of 24 weeks and positive F1 mice at the age of 8 weeks were collected to conduct the property observation, microscopic examination of urinary sediment, and detection of the ratio of urine protein to urine creatinine. Renal tissues were harvested and IgA immunofluorescence staining and H-E staining were performed to assess the IgA deposition and histological changes of renal tissues. The blood was drawn from the heart of mice and the serum was isolated. Then IgA and IgG levels were detected. Results A total of 3 and 8 positive parental mice were obtained in transgenic group A and B, from which 17 and 52 positive F1 mice were obtained. The results of immunofluorescence staining indicated that IgA deposited in the glomerular mesangial area of kidneys of positive parental mice and positive F1 mice. The results of H-E staining showed mesenteric cell proliferation, increase of mesangial matrix, and inflammatory cell infiltration in mesenchyma. No erythrocytes were found in fresh urinary sediment by microscopic examination. The ratio of urine protein to urine creatinine and serum IgA and IgG levels of transgenic mice were significantly higher than those of wild type mice with the same age. Conclusion A mice model of IgA nephropathy is established by inserting sequences of the hinge area of human IgA1. Structural changes of the hinge area of IgA1 play a crucial role in the pathogenesis of IgA nephropathy.

Key words: IgA nephropathy, transgene, mouse, hinge area, glycosylation