Proceedings of The Physiological Society
University of Oxford (2011) Proc Physiol Soc 23, C36
Urinary aminopeptidase activities as early and predictive biomarkers of renal dysfunction in cisplatin-treated rats
A. Quesada1, F. O'Valle2, M. Rodríguez-Martínez2, A. Segarra1, I. Prieto1, M. Ramírez1, F. Vargas3, R. Wangensteen1
Introduction: There is an urgent need for better biomarkers to permit more timely diagnosis of acute kidney injury (AKI), prediction of injury severity, and safety assessment during drug development (1). This study analyzes the fluorimetric determination of alanyl- (Ala), glutamyl- (Glu), leucyl-cystinyl- (Cys) and aspartyl-aminopeptidase (AspAp) urinary enzymatic activities as early and predictive biomarkers of renal dysfunction in cisplatin-treated rats. Methods: Male Wistar rats (n=5 each group) received a single subcutaneous injection of either saline or cisplatin 3.5 or 7 mg/kg, and urine samples were taken at 0, 2, 3, 7, 10 and 14 days after treatment. In urine samples we determined Ala, Glu, Cys and AspAp activities, proteinuria, N-acetyl-β-D-glucosaminidase (NAG), albumin, cistatin and β2-microglobulin. Plasma creatinine and creatinine clearance were measured at the end of the experiment. All experimental procedures were performed according to the European Union Guidelines to the Care and Use of Laboratory Animals and approved by the Ethical Committee of the University of Jaén. Blood samples were obtained from left ventricle under terminal anesthesia (pentobarbital, 50 mg/kg, i.p.) and then killed with an overdose of pentobarbital (150 mg/kg, i.p.). Kidneys were removed and fixed in formalin solution for morphological analysis. Results: AlaAp, albumin and cistatin were increased at the second day of treatment in the cisplatin 3.5 mg/kg treated group, showing their ability to detect a slight renal damage. All aminopeptidase activities and urinary biomarkers were significantly increased at the third day of treatment in the cisplatin 7 mg/kg treated group. At this point, AlaAp (r2=0.998; p=0.0001), CysAp (r2=0.992; p=0.0003), GluAp (r2=0.951; p=0.0046), NAG (r2=0.863; p=0.0225) and total protein (r2=0.787; p=0.0446) urinary excretion correlated with plasma creatinine level at the end of the experiment and could be considered as predictive biomarkers of renal injury severity. Cisplatin significantly enlarged the fibrosis area in the tubular interstitium of the kidney, as observed by other authors (2) and it was dose-dependent. ROC-Area under the curve (AUC) for aminopeptidase activities at day 3 was >0.5. Conclusion: Ala, Cys, Glu and AspAp enzymatic activities are urinary biomarkers of the AKI induced by cisplatin. AlaAp activity is an early biomarker and Ala, Cys and GluAp activities are also predictive biomarkers of renal dysfunction. These determinations can be very useful in the prognostic and diagnostic of renal dysfunction in preclinical research and clinical practice.
Where applicable, experiments conform with Society ethical requirements