Development of enzyme-linked immunosorbent assays for urinary thiazide-sensitive Na-Cl cotransporter measurement

Kiyoshi Isobe, Takayasu Mori, Takako Asano, Hiroyuki Kawaguchi, Shigeaki Nonoyama, Naonori Kumagai, Fumiaki Kamada, Tetsuji Morimoto, Matsuhiko Hayashi, Eisei Sohara, Tatemitsu Rai, Sei Sasaki, Shinichi Uchida

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Abstract

The Na-Cl cotransporter (NCC) in the distal convoluted tubules in kidney is known to be excreted in urine. However, its clinical significance has not been established because of the lack of quantitative data on urinary NCC. We developed highly sensitive enzymelinked immunosorbent assays (ELISAs) for urinary total NCC (tNCC) and its active form, phosphorylated NCC (pNCC). We first measured the excretion of tNCC and pT55-NCC in urinary exosomes in pseudohypoaldosteronism type II (PHAII) patients since PHAII is caused by NCC activation. Highly increased excretion of tNCC and pNCC was observed in PHAII patients. In contrast, the levels of tNCC and pNCC in the urine of patients with Gitelman's syndrome were not detectable or very low, indicating that both assays could specifically detect the changes in urinary NCC excretion caused by the changes of NCC activity in the kidney. Then, to test whether these assays could be feasible for a more general patient population, we measured tNCC and pNCC in the urine of outpatients with different clinical backgrounds. Although urinary protein levels >30 mg/dl interfered with our ELISA, we could measure urinary pNCC in all patients without proteinuria. Thus we established highly sensitive and quantitative assays for urinary NCC, which could be valuable tools for estimating NCC activity in vivo.

Original languageEnglish
Pages (from-to)F1374-F1381
JournalAmerican Journal of Physiology - Renal Physiology
Volume305
Issue number9
DOIs
Publication statusPublished - 2013 Nov 1

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Keywords

  • Exosome
  • Hypertension
  • Pseudohypoaldosteronism type II
  • WNK kinase

ASJC Scopus subject areas

  • Physiology
  • Urology

Cite this

Isobe, K., Mori, T., Asano, T., Kawaguchi, H., Nonoyama, S., Kumagai, N., Kamada, F., Morimoto, T., Hayashi, M., Sohara, E., Rai, T., Sasaki, S., & Uchida, S. (2013). Development of enzyme-linked immunosorbent assays for urinary thiazide-sensitive Na-Cl cotransporter measurement. American Journal of Physiology - Renal Physiology, 305(9), F1374-F1381. https://doi.org/10.1152/ajprenal.00208.2013