实验动物与比较医学 ›› 2023, Vol. 43 ›› Issue (5): 531-540.DOI: 10.12300/j.issn.1674-5817.2023.070

• 研究报告 • 上一篇    下一篇

成年斑马鱼体肾的微细结构及其外泌体的分布鉴定

林金杏1()(), 王新栋2,3, 白雪兵2, 冯丽萍1, 谢淑武1, 陈秋生2()()   

  1. 1.上海实验动物研究中心, 上海 201203
    2.南京农业大学动物医学院, 南京 210095
    3.枝江市畜牧兽医服务中心, 宜昌 443200
  • 收稿日期:2023-06-05 修回日期:2023-10-12 出版日期:2023-10-25 发布日期:2023-11-01
  • 通讯作者: 陈秋生(1963—),男,博士,教授,研究方向:动物组织细胞形态及功能研究。E-mail: chenqsh305@njau.edu.cn。ORCID: 0000-0002-6087-7029
    林金杏(1983—),女,博士,研究员,研究方向:水生实验动物生理功能及标准化研究。E-mail: linjinxing@slarc.org.cn。ORCID: 0000-0002-7649-9552;
  • 作者简介:林金杏(1983—),女,博士,研究员,研究方向:水生实验动物生理功能及标准化研究。E-mail: linjinxing@slarc.org.cn。ORCID:0000-0002-7649-9552;
  • 基金资助:
    上海市科技计划项目“上海市水生实验动物专业技术服务平台”(22DZ2291200);上海市科研计划项目“斑马鱼器官形态参数完善与立体重构整合”(16140904300)

Fine Structure of the Trunk Kidney and Distribution of Its Secreted Exosomes in the Adult Zebrafish

Jinxing LIN1()(), Xindong WANG2,3, Xuebing BAI2, Liping FENG1, Shuwu XIE1, Qiusheng CHEN2()()   

  1. 1.Shanghai Laboratory Animal Research Center, Shanghai 201203, China
    2.College of Veterinary Medicine, Nanjing Agricultural University, Nanjing 210095, China
    3.Zhijiang Animal Husbandry and Veterinary Service Center, Yichang 443200, China
  • Received:2023-06-05 Revised:2023-10-12 Published:2023-10-25 Online:2023-11-01
  • Contact: CHEN Qiusheng (ORCID: 0000-0002-6087-7029), E-mail:chenqsh305@njau.edu.cn
    LIN Jinxing (ORCID: 0000-0002-7649-9552), E-mail:linjinxing@slarc.org.cn;

摘要:

目的 观察和分析斑马鱼体肾的微细结构,并对其分泌的外泌体进行分离鉴定。 方法 应用光学显微镜和电子显微镜技术系统观察斑马鱼体肾的显微结构和超微结构,用纳米颗粒跟踪分析(nanoparticle tracking analysis,NTA)技术检测外泌体粒径大小。 结果 斑马鱼体肾紧贴并平行于脊椎,肾单位由肾小管和肾小体组成,其中肾小管可细分为近曲小管、远曲小管及颈段3种类型,肾小体由肾小球和肾小囊构成;经高碘酸-希夫(periodic acid-Schiff,PAS)染色后,可见近曲小管富含糖原颗粒,其游离面有明显的刷状缘结构。透射电子显微镜下,在肾小管的管腔观察到外泌体分布,上皮细胞游离面的胞质有少量多囊泡体和大量的晚期内涵体分布,肾小管的顶端和肾小球足细胞中亦观察到少量多囊泡体分布。通过CD9、CD63和TSG101免疫组织化学和免疫荧光检测显示,多囊泡体及其外泌体在肾小管管腔的游离面高表达,在肾小体和管腔内表达较弱。NTA和透射电子显微镜观察结果显示,分离获得的斑马鱼体肾外泌体呈茶托状,粒径众数为144.4 nm,符合外泌体形态和直径大小特征。 结论 斑马鱼体肾具有哺乳动物肾脏的典型结构,为体内的泌尿器官。肾小管具有分泌外泌体的能力,其形成是多囊泡体向细胞游离面释放到胞外的过程。本研究结果为水生实验动物泌尿器官的功能研究及相关模型的开发应用奠定了形态学基础。

关键词: 斑马鱼, 体肾, 外泌体, 多囊泡体

Abstract:

Objective To observe the fine structure of the trunk kidney in zebrafish, and to identify its secreted exosomes. Methods The microstructure and ultrastructure of the trunk kidney in zebrafish were observed by light microscopy and electron microscopy, and the particle size of exosomes was detected by nanoparticle tracking analysis (NTA). Results The trunk kidney was close and parallel to the spine in adult zebrafish. The nephron consisted of renal tubules and renal corpuscles. The renal tubules could be further divided into three types: proximal convoluted tubules, distal convoluted tubules, and cervical segments. The renal corpuscles were composed of glomerulus and renal capsules. The periodic acid-Schiff (PAS) staining results revealed that there were abundant glycogen granules in the proximal convoluted tubules, with brush-like outline in the apical surface of epithelial cells. Under transmission electron microscopy (TEM), there were exosomes distributed in the lumen of renal tubules, with numerous late endosomes and few number of multivesicular bodies (MVBs) in the cytoplasm of the epithelial cells concentrating on the apical side. Meanwhile, MVBs were also distributed in the apical regions of the renal tubules and the podocytes of the renal glomeruli. Immunohistochemical staining results showed that CD9, CD63 and TSG101 were strongly expressed in the lumen surface of the renal tubules, but weakly expressed in the corpuscles and lumen. NTA and TEM results showed that the exosomes isolated from zebrafish trunk kidney were saucer-like outline, and the particle size mode was 144.4 nm, which was consistent with the characteristics of morphological futures of exosome. Conclusion The zebrafish somatic kidney has the typical structure of the mammalian kidney and is the urinary organ in the body. The renal tubules have the ability to secrete exosomes, and their formation is a process of releasing poly-vesicles to the free surface of epithelial cells into the extracellular space. This study laid a morphological foundation for further study of exosomes in urinary function in aquatic experimental animals as well as the development and application of related models.

Key words: Zebrafish, Trunk kidney, Exosome, Multivesicular body

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