title: Calcium oxalate dihydrate crystal induced changes in glycoproteome of distal renal tubular epithelial cells.
abstract: Calcium oxalate dihydrate (COD) crystals can adhere onto the apical surface of renal tubular epithelial cells. This process is associated with crystal growth and aggregation, resulting in kidney stone formation. Glycoproteins have been thought to play roles in response to crystal adhesion. However, components of the glycoproteome that are involved in this cellular response remain largely unknown. Our present study therefore aimed to identify altered glycoproteins upon COD crystal adhesion onto tubular epithelial cells representing distal nephron, the initiating site of kidney stone formation. Madin-Darby Canine Kidney (MDCK) cells were maintained in culture medium with or without COD crystals for 48 h (n = 5 flasks per group). Cellular proteins were extracted, resolved by 2-DE and visualized by SYPRO Ruby total protein stain, whereas glycoproteins were detected by Pro-Q Emerald glycoprotein dye. Spot matching and quantitative intensity analysis revealed 16 differentially expressed glycoprotein spots, whose corresponding total protein levels were not changed by COD crystal adhesion. These altered glycoproteins were successfully identified by Q-TOF MS and/or MS/MS analyses, and potential glycosylation sites were identified by the GlycoMod tool. For example, glycoforms of three proteasome subunits (which have a major role in regulating cell-cell dissociation) were up-regulated, whereas a glycoform of actin-related protein 3 (ARP3) (which plays an important role in cellular integrity) was down-regulated. These coordinated changes implicate that COD crystal adhesion induced cell dissociation and declined cellular integrity in the distal nephron. Our findings provide some novel insights into the pathogenic mechanisms of kidney stone disease at the molecular level, particularly cell-crystal interactions.
authors.affiliation: Medical Proteomics Unit, Office for Research and Development, Faculty of Medicine Siriraj Hospital, Mahidol University, 12th Fl. Adulyadej Vikrom Bldg., 2 Prannok Rd., Bangkoknoi, Bangkok 10700, Thailand.
authors.forename: Wararat
authors.initials: W
authors.lastname: Chiangjong
authors.forename: Supachok
authors.initials: S
authors.lastname: Sinchaikul
authors.forename: Shui-Tein
authors.initials: ST
authors.lastname: Chen
authors.forename: Visith
authors.initials: V
authors.lastname: Thongboonkerd
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002129:Calcium Oxalate / Q000378:metabolism* / Q000494:pharmacology; D002460:Cell Line; D003460:Crystallization; D004285:Dogs; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism*; D006023:Glycoproteins / Q000378:metabolism*; D007669:Kidney Calculi / Q000378:metabolism; D007686:Kidney Tubules, Distal / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D020543:Proteome / Q000378:metabolism*; D020539:Sequence Analysis, Protein; D053719:Tandem Mass Spectrometry
chemical_list: D006023:Glycoproteins; D020543:Proteome; D002129:Calcium Oxalate