DNA Synthesis and Genotoxicity

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Abstract
In a recent report, Burlinson et al. [Carcinogenesis 1989;10:1425–1428] published the results of an experiment which they claim shows that omeprazole induces unscheduled DNA synthesis (UDS) in rat gastric mucosa (i.e. DNA repair). The Burlinson method measures the incorporation of tritiated thymidine into cells undergoing DNA synthesis. Rat stomachs are exposed to test substances for 14 h in vivo, tritiated thymidine is injected and the animals killed 1 h later. Superficial cells are isolated by a selective digestion with pronase for 45 min, and the level of UDS in the cell isolate is measured by scintillation counting. However, differentiation of UDS in the gastric mucosa from the scheduled DNA synthesis which accompanies cell division is technically difficult. Scheduled DNA synthesis is maximal around the isthmus neck junction of the gastric glands, but is detectable (by labelling with tritiated thymidine) up to five cell positions from the surface. Any method which relies on selective separation of non-dividing surface cells to identify UDS requires stringent controls to exclude contamination with deeper dividing cells. This is particularly important in situations where there is a stimulus to cell proliferation, as occurs when testing compounds such as 1-methyl-3-nitro-1-nitrosoguanidine (MNNG) and omeprazole. The proliferation results in cells that are undergoing scheduled DNA synthesis appearing closer to the surface, where they are even more likely to contaminate the non-dividing surface layer. Any such contamination invalidates claims to selective demonstration of UDS. The need for stringent controls in experiments measuring UDS is therefore great. The method of Burlinson et al. did not inhibit scheduled DNA synthesis with hydroxyurea, nor did they perform controls to prove that their digestion was selective for surface cells. As a consequence, it is concluded that the Burlinson method is technically flawed and without scientific validity.

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