NE-AUA 2006 Annual Meeting, September 28 - 30, 2006, The Westin Hotel & Rhode Island Convention Center Providence, Rhode Island
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The proteomic analysis of rat urine as a discovery method of urinary biomarkers of postnatal maturation and renal obstruction
Richard S. Lee, MD, Flavio Monigatti, PhD, Mohini Lutchman, PhD, Judith A.J. Steen, PhD, Michael R. Freeman, PhD, Hanno Steen, PhD.
Children's Hospital Boston, Boston, MA,

Introduction: Proteomic technologies are playing an essential role in mechanistic studies and in the search for useful biomarkers of disease. Currently, proteomic methods have the ability to define the protein composition of complex mixtures, and to determine the qualitative and quantitative differences between disease states. Here we demonstrate the advantages of a high-throughput proteomics approach to identify candidate urinary markers of postnatal maturation and renal obstruction in a rodent model.
Methods: Urine was collected from normal rats at a series of postnatal (P) days (P1, P3, P7, P14 and >P30) by sterile bladder aspiration. Additionally, >P30 rats underwent unilateral renal obstruction and obstructed renal pelvis urine was collected at postoperative day 3 (AC3). Each protein complement was analyzed and sequenced using the QSTARŪ XL LC/MS/MS (liquid chromatography / mass spectrometry / mass spectrometry). Data sets were submitted to MASCOT employing a Rat specific extract. For protein identification and relative protein quantitation, we used stringent search parameters and identification criteria to provide a 0.01% false positive rate at the protein level.
Results: We demonstrate that the composition of the urinary proteome changes dramatically during normal postnatal maturation. The number of proteins in common at each time interval decreased significantly as the animals matured. During early maturation (P1-P7), we detected 14 proteins involved with cellular adhesion, structure, or proliferation and differentiation that were not present in adults. In addition, we identified 30 proteins specific to adults, of which 13 originated from the prostate or seminal vesicle. When comparing obstructed AC3 animals to the protein composition of >P30 animals, we identified 66 new proteins that were only present in AC3 urine.
Conclusion: This is the first characterization by mass spectrometry of the normal complement of urine proteins in early postnatal rodent development and during urinary tract obstruction. In addition, we have demonstrated the ability to identify candidate markers of normal development and obstruction. Future time course experiments of urinary obstruction in both adult and neonatal rats will provide the necessary data to narrow the list of potential markers of obstructive nephropathy, and to perform a directed urinary proteomic analysis in infants with hydronephrosis.


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