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运用逆转录聚合酶链反应、免疫印迹法和免疫细胞化学法对水通道蛋白-7在大鼠和小鼠肾脏中的定位进行研究。

Localization of aquaporin-7 in rat and mouse kidney using RT-PCR, immunoblotting, and immunocytochemistry.

作者信息

Nejsum L N, Elkjaer M, Hager H, Frokiaer J, Kwon T H, Nielsen S

机构信息

Department of Cell Biology, Institute of Anatomy, Aarhus, DK-8000, Denmark.

出版信息

Biochem Biophys Res Commun. 2000 Oct 14;277(1):164-70. doi: 10.1006/bbrc.2000.3638.

Abstract

To establish the segmental, cellular, and subcellular localization of AQP7 in rat and mouse kidney, we used RT-PCR, immunocytochemical, and immunoblotting approaches. RT-PCR of rat and mouse kidney zones revealed AQP7 mRNA in cortex and outer stripe of the outer medulla. RT-PCR on microdissected nephron segments revealed AQP7 mRNA in proximal convoluted and straight tubules. Immunoblotting using peptide-derived rabbit antibodies to either rat or mouse AQP7 revealed a 28-kDa band in kidney and testes from rat and mouse, respectively. Immunocytochemistry revealed strong AQP7 labeling of segment 3 proximal tubules and weaker labeling of proximal convoluted tubules in both rat and mouse kidneys. The labeling was almost exclusively confined to the brush border with no basolateral labeling. No labeling was observed of thin descending limbs or collecting duct. Immunolabeling controls were negative. The presence of AQP7 in the proximal tubule brush border indicates a role of AQP7 in proximal tubule water reabsorption.

摘要

为确定水通道蛋白7(AQP7)在大鼠和小鼠肾脏中的节段、细胞及亚细胞定位,我们采用了逆转录聚合酶链反应(RT-PCR)、免疫细胞化学和免疫印迹法。对大鼠和小鼠肾脏不同区域进行RT-PCR检测,结果显示皮质和外髓质外带存在AQP7信使核糖核酸(mRNA)。对显微解剖的肾单位节段进行RT-PCR检测,结果显示近曲小管和近端直小管存在AQP7 mRNA。使用针对大鼠或小鼠AQP7的肽段衍生兔抗体进行免疫印迹检测,结果分别在大鼠和小鼠的肾脏及睾丸中显示出一条28 kDa的条带。免疫细胞化学检测显示,大鼠和小鼠肾脏的近端小管S3段有强AQP7标记,近曲小管标记较弱。标记几乎完全局限于刷状缘,无基底外侧标记。细降支或集合管未观察到标记。免疫标记对照为阴性。近端小管刷状缘存在AQP7表明AQP7在近端小管水重吸收中发挥作用。

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