Investigation of Ornithine Decarboxylase Activity and Two-Dimensional Electrophoretic Protein Profile Following Exposure of T24 Bladder Carcinoma Cells to Tumor Promoter and Carcinogen

Abstract

Summary: To develop appropriate screening tools for biomarkers of effects of exposure to occupational chemical insult. Changes were investigated in T24 human bladder carcinoma cell ornithine decarboxylase activity and protein profiles by quantitative two-dimensional polyacrylamide gel electrophoresis (2D PAGE), biochemical events potentially altered by an established human bladder carcino gen and tumor promoter. A unique chromatographic approach was used to demonstrate that in vitro exposure of T24 cells for 6 h to varying concentrations of the carcinogen 4-aminobiphenyl elevates enzyme activity 5.3- to 5.9-fold. As a second method to identify potential biomarkers of exposure, two-dimensional gel electrophoresis was used to compare the protein pattern of vehicle control-treated T24 to 4-aminobiphenyl or tumor promoter (12-o-tetradecanoylphorbol-13-acetate)treated cells. Changes in abundance and modification of proteins are determined using the Kepler software package to analyze and compare gels across treatment groups. With this technology, protein markers are identified by significant alterations in spot density (mean ratio of Coomassie Blue intensity; p. 001, Student's t test) following T24 treatment with the carcinogen or the tumor promoter. Fifteen protein spots from a detectable pool of 542 demonstrate two-fold or greater changes in intensity. The results illustrate the potential of automated two-dimensional gel analysis for classifying different gel patterns, an approach that can be applied to patterns whose differences are obscured by the minor changes in spot intensity that arise between separate cell cultures. In addition to the ornithine decarboxylase assay, 2D PAGE offers much promise to evaluate potential biomarkers for occupational and environmental carcinogens. These results will be used to further develop NIOSH efforts in the molecular epidemiology of occupational bladder carcinogenesis. [K]Key [K]Words: biomarker, bladder carcinogenesis, occupational exposure, ODC, 2D PAGE.

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• [K]Key [K]Words: Biomarker, Bladder Carcinogenesis, Occupational Exposure, Odc, 2d Page.