In vitro evidence of the promoting effect of testosterone in kidney stone disease: A proteomics approach and functional validation.
Incidence of kidney stone disease in males is 2- to 4-fold greater than in females. This study aimed to determine effects of testosterone on kidney stone disease using a proteomics approach. MDCK renal tubular cells were treated with or without 20nM testosterone for 7days. Cellular proteins were extracted, resolved by 2-DE, and stained with Deep Purple fluorescence dye (n=5 gels derived from 5 independent samples/group). Spot matching, quantitative intensity analysis, and statistics revealed significant changes in levels of nine protein spots after testosterone treatment. These proteins were then identified by nanoLC-ESI-Qq-TOF MS/MS. Global protein network analysis using STRING software revealed α-enolase as the central node of protein-protein interactions. The increased level of α-enolase was then confirmed by Western blotting analysis, whereas immunofluorescence study revealed the increased α-enolase on cell surface and intracellularly. Functional analysis confirmed the potential role of the increased α-enolase in enhanced calcium oxalate monohydrate (COM) crystal-cell adhesion induced by testosterone. Finally, neutralization of surface α-enolase using anti-α-enolase antibody successfully reduced the enhanced COM crystal-cell adhesion to the basal level. Our data provided in vitro evidence of promoting effect of testosterone on kidney stone disease via enhanced COM crystal-cell adhesion by the increased surface α-enolase.



The incidence of kidney stone disease in male is 2- to 4-fold greater than in female. One of the possible factors of the male preference is the higher testosterone hormone level. However, precise molecular mechanisms that testosterone plays in kidney stone disease remained unclear. Our present study is the first exploratory investigation on such aspect using a proteomics approach. Our data also provide a novel mechanistic aspect of how testosterone can impact the risk of kidney stone formation (i.e. the discovery that testosterone increases alpha-enolase expression on the surface of renal tubular cells that is responsible, at least in part, for crystal-cell adhesion).
Medical Proteomics Unit, Office for Research and Development, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok 10700, Thailand; Graduate Program in Immunology, Department of Immunology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok 10700, Thailand; Center for Research in Complex Systems Science, Mahidol University, Bangkok 10700, Thailand.
Channarong
C
Changtong
Medical Proteomics Unit, Office for Research and Development, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok 10700, Thailand; Center for Research in Complex Systems Science, Mahidol University, Bangkok 10700, Thailand.
Paleerath
P
Peerapen
Medical Proteomics Unit, Office for Research and Development, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok 10700, Thailand; Graduate Program in Immunology, Department of Immunology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok 10700, Thailand; Center for Research in Complex Systems Science, Mahidol University, Bangkok 10700, Thailand.
Supaporn
S
Khamchun
Medical Proteomics Unit, Office for Research and Development, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok 10700, Thailand; Center for Research in Complex Systems Science, Mahidol University, Bangkok 10700, Thailand.
Kedsarin
K
Fong-Ngern
Medical Proteomics Unit, Office for Research and Development, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok 10700, Thailand; Center for Research in Complex Systems Science, Mahidol University, Bangkok 10700, Thailand.
Somchai
S
Chutipongtanate
Medical Proteomics Unit, Office for Research and Development, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok 10700, Thailand; Center for Research in Complex Systems Science, Mahidol University, Bangkok 10700, Thailand. Electronic address: thongboonkerd@dr.com.
Visith
V
Thongboonkerd
D000818:Animals; D002129:Calcium Oxalate / Q000378:metabolism; D002448:Cell Adhesion; D004285:Dogs; D007669:Kidney Calculi / Q000139:chemically induced* / Q000473:pathology; D007684:Kidney Tubules / Q000737:chemistry / Q000473:pathology; D061985:Madin Darby Canine Kidney Cells; D008565:Membrane Proteins / Q000032:analysis / Q000378:metabolism; D010751:Phosphopyruvate Hydratase / Q000032:analysis / Q000378:metabolism; D060066:Protein Interaction Maps; D020543:Proteome / Q000032:analysis / Q000187:drug effects*; D040901:Proteomics / Q000379:methods*; D013739:Testosterone / Q000494:pharmacology*
D008565:Membrane Proteins; D020543:Proteome; D002129:Calcium Oxalate; D013739:Testosterone; D010751:Phosphopyruvate Hydratase
Calcium oxalate; Crystal adhesion; Enolase; Kidney stone; Nephrolithiasis; Testosterone