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Investigations of Rodent Urinary Bladder Carcinogens: Collection, Processing, and Evaluation of Urine and Bladders

Department of Pathology and Microbiology, and the Eppley Institute for Cancer Research, University of Nebraska Medical Center, Omaha, NE 68198-3135, USA

Correspondence: Address correspondence to: Samuel M. Cohen, University of Nebraska Medical Center, 983135 Nebraska Medical Center, Omaha, NE 68198-3135, USA; e-mail:scohen{at}unmc.edu

Examination of the urine and the bladder epithelium are essential to the investigation of mechanisms of urinary bladder carcinogens in rodents. However, urine and bladder tissue specimens must be collected and processed properly if accurate data are to be derived. The optimum specimen for analysis of urinary constituents is fresh void urine collected from nonfasting animals. Fasting the animal prior to urine collection changes the normal composition, including pH. Many of the normal urinary constituents can influence the mode of action of bladder carcinogens, especially for non-genotoxic agents. Light microscopy is routinely used to examine the bladder epithelium. However, it is often necessary to use more sensitive techniques, such as scanning electron microscopy (SEM) to detect subtle cytotoxic changes in the superficial cell layer of the urothelium, and bromodeoxyuridine (BrdU) incorporation, PCNA, or Ki-67 immunohistochemistry to determine the labeling index for cell proliferation. The urinary bladder must be handled gently and inflated with fixative in situ before the animal dies to avoid artifactual autolytic damage to the bladder epithelium that is visible by SEM and may be mistaken for treatment-related changes. The purpose of this paper is to provide information for the proper collection and examination of urine and the urinary bladder.

Key Words: Urinary bladder • urothelium • urine • scanning electron microscopy • cell proliferation • hyperplasia • urine collection

Abbreviations: BrdU, bromodeoxyuridine • DAB, diaminobenzidine tetrahydrochloride • DNA, deoxyribonucleic acid • ED₀₁, 1% effective dose • EDS, energy dispersive X-ray spectroscopy • HCl, hydrochloric acid • MgNH₄PO₄, magnesium ammonium phosphate • PCNA, proliferating cell nuclear antigen • SEM, scanning electron microscopy

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