Decreased renal expression of the putative calcium oxalate inhibitor Tamm-Horsfall protein in the ethylene glycol rat model of calcium oxalate urolithiasis.
OBJECTIVE
Tamm-Horsfall protein is believed to inhibit calcium oxalate crystallization, aggregation or adhesion to the renal epithelium. We determined whether ethylene glycol induced urolithiasis changes the expression of renal and urinary Tamm-Horsfall protein. For comparison the expression of another calcium oxalate inhibitor, osteopontin, was also analyzed.


METHODS
Male rats were treated with 0.75% ethylene glycol plus an AIN-76 diet (Dyets, Bethlehem Pennsylvania) (ethylene glycol group) or standard rat chow and water (control group) for up to 8 weeks (6 per group for 8 weeks and 3 per group for 3 days to 6 weeks). Kidneys and urine (8 weeks only) were harvested and analyzed by Northern and Western blot analysis, and immunohistochemistry.


RESULTS
Tamm-Horsfall protein message and protein (membrane bound form) were decreased, while those of osteopontin were increased in the kidneys of rats treated with ethylene glycol for 8 weeks. As judged by immunochemistry Tamm-Horsfall protein and osteopontin were consistently present in a few tubules in rats in the ethylene glycol and control groups, respectively. In urine expression of the free form of Tamm-Horsfall protein (approximately 75 kDa.) was decreased but detectable in ethylene glycol treated rats. Although readily detected in tissue, osteopontin was not detected in the urine of control or ethylene glycol treated rats. In the time course experiment Tamm-Horsfall protein did not decrease until 4 weeks, when calcium oxalate crystals were detectable in the kidneys of treated rats. In contrast, osteopontin was increased, although inconsistently, beginning at 3 days.


CONCLUSIONS
Unlike other calcium oxalate inhibitors, such as osteopontin, renal message and protein for Tamm-Horsfall protein was decreased in ethylene glycol treated rats. Tamm-Horsfall protein expression did not decrease until aggregates of crystals had been deposited in the kidneys, while osteopontin expression began to increase almost immediately. Comparisons of the data on kidneys and urine obtained by RNA or protein blot analysis and immunochemistry underscore the need to examine tissue and urine by multiple techniques to obtain the most accurate assessment of how protein expression is changed by a given treatment.
Marengo
Susan Ruth
SR
Jim and Eilleen Dicke Research Laboratory, Department of Urology and Anatomy, School of Medicine, Case Western Reserve University-University Hospitals of Cleveland, Cleveland, Ohio, USA.
Chen
Daniel H-C
DH
Kaung
Hue-Lee C
HL
Resnick
Martin I
MI
Yang
Lizhu
L
D000818:Animals; D002129:Calcium Oxalate / Q000652:urine*; D019855:Ethylene Glycol; D005260:Female; D015870:Gene Expression / Q000187:drug effects; D007668:Kidney / Q000473:pathology*; D007669:Kidney Calculi / Q000139:chemically induced / Q000235:genetics* / Q000473:pathology; D008297:Male; D009088:Mucoproteins / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D017207:Rats, Sprague-Dawley; D058949:Uromodulin
D009088:Mucoproteins; D012333:RNA, Messenger; C548614:Umod protein, rat; D058949:Uromodulin; D002129:Calcium Oxalate; D019855:Ethylene Glycol