Mutant frequency in comparison to oxidative DNA damage induced by ochratoxin A in L5178Y tk^+/− (3.7.2C) mouse lymphoma cells

Abstract

Ochratoxin A (OTA) is a naturally occurring mycotoxin that contaminates animal feed and human food. OTA is nephrotoxic, hepatotoxic, immunosuppressive and a potent renal carcinogen in rodents. In the present study, we evaluated the genotoxicity of OTA in L5178Y tk^+/− (3.7.2C) mouse lymphoma cells using the microwell version of the mouse lymphoma gene mutation assay (MLA) and the comet assay modified to detect oxidative DNA damage. Cells were treated for 4 hours with 0, 5, 10, 25, 50 or 100 µM of OTA in the presence and absence of exogenous metabolic activation (S9). Benzo[a]pyrene (1 µg/mL) and 4-nitroquinoline-1-oxide (0.1 µg/mL) were used as positive control with and without S9, respectively. OTA treatment produced dose-dependent increases in cytotoxicity and tk mutant frequency, with significant increases in mutant frequency detected at concentrations ≥25 µM with and without S9. Similarly treated cells were used for the comet assay conducted with and without formamidopyrimidine-DNA glycosylase for the determination of oxidative DNA damage. OTA exposure resulted in a significant increase in both direct and oxidative DNA damage, with induction of oxidative damage being greater. The results indicate that OTA is mutagenic in mouse lymphoma assay; and that OTA-generated oxidative DNA damage is, at least partially, responsible for its mutagenicity in the assay.

• Comet assay
• mouse lymphoma assay
• mutant frequency
• ochratoxin A
• oxidative DNA damage

Acknowledgements

The authors are thankful to Dr. Robert H. Heflich for supporting Rahat Ali at NCTR to conduct a part of his Ph.D. research work, which resulted in this manuscript. The authors are also thankful to the Higher Education Commission (HEC), Pakistan, for providing the financial support under the project “International Research Support Initiative Program (IRSIP)” and to the Oak Ridge Institute for Scientific Education (ORISE), for hosting the scientific interaction that resulted in this study. The views presented in this article do not necessarily reflect those of the U.S. Food and Drug Administration.