Publication: Large-scale identification of calcium oxalate monohydrate crystal-binding proteins on apical membrane of distal renal tubular epithelial cells
Issued Date
2011-10-07
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ISSN
15353907
15353893
15353893
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2-s2.0-80053914224
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Mahidol University
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SCOPUS
Bibliographic Citation
Journal of Proteome Research. Vol.10, No.10 (2011), 4463-4477
Suggested Citation
Kedsarin Fong-Ngern, Paleerath Peerapen, Supachok Sinchaikul, Shui Tein Chen, Visith Thongboonkerd Large-scale identification of calcium oxalate monohydrate crystal-binding proteins on apical membrane of distal renal tubular epithelial cells. Journal of Proteome Research. Vol.10, No.10 (2011), 4463-4477. doi:10.1021/pr2006878 Retrieved from: https://repository.li.mahidol.ac.th/handle/20.500.14594/11454
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Title
Large-scale identification of calcium oxalate monohydrate crystal-binding proteins on apical membrane of distal renal tubular epithelial cells
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Abstract
Adhesion of calcium oxalate monohydrate (COM) crystals onto apical surface of renal tubular epithelial cells is a crucial mechanism for crystal retention, leading to kidney stone formation. Various proteins on apical membrane may bind to COM crystals; however, these crystal-binding proteins remained unidentified. The present study therefore aimed to identify COM crystal-binding proteins on apical membrane of distal renal tubular epithelial cells. Madin-Darby Canine Kidney (MDCK) cells were cultivated to be polarized epithelial cells and apical membrane was isolated from these cells using a peeling method established recently. Enrichment and purity of isolated apical membrane were confirmed by Western blot analysis for specific markers of apical (gp135) and basolateral (Na + /K + -ATPase) membranes. Proteins derived from the isolated apical membrane were then resuspended in artificial urine and incubated with COM crystals. The bound proteins were eluted, resolved by SDS-PAGE, and analyzed by Q-TOF MS and MS/MS, which identified 96 proteins. Among these, expression and localization of annexin II on apical surface of MDCK cells were confirmed by Western blot analysis, immunofluorescence staining, and laser-scanning confocal microscopic examination. Finally, the function of annexin II as the COM crystal-binding protein was successfully validated by COM crystal-binding assay. This large data set offers many opportunities for further investigations of kidney stone disease and may lead to the development of new therapeutic targets. © 2011 American Chemical Society.