301 Molecular and Cellular Biochemistry 234/235: 301–308, 2002. © 2002 Kluwer Academic Publishers. Printed in the Netherlands. Arsenic induces oxidative DNA damage in mammalian cells Maris Kessel, 1 Su Xian Liu, 1 An Xu, 1 Regina Santella 2 and Tom K. Hei 1,2 1 Center for Radiological Research, College of Physicians and Surgeons; 2 Department of Environmental Health Sciences, Joseph Mailman School of Public Health, Columbia University, NY, USA Abstract Although arsenic is a well-established human carcinogen, the underlying carcinogenic mechanism(s) is not known. Using the human-hamster hybrid (A L ) cell mutagenic assay that is sensitive in detecting mutagens that induce predominately multilocus deletions, we showed previously that arsenite is indeed a potent gene and chromosomal mutagen and that oxyradicals may be involved in the mutagenic process. In the present study, the effects of free radical scavenging enzymes on the cytotoxic and mutagenic potential of arsenic were examined using the A L cells. Concurrent treatment of cells with either superoxide dis- mutase or catalase reduced both the cytotoxicity and mutagenicity of arsenite by an average of 2–3 fold, respectively. Using immunoperoxidase staining with a monoclonal antibody specific for 8-hydroxy-2′-deoxyguanosine (8-OHdG), we demonstrated that arsenic induced oxidative DNA damage in A L cells. This induction was significantly reduced in the presence of the anti- oxidant enzymes. Furthermore, reducing the intracellular levels of non-protein sulfhydryls (mainly glutathione) using buthionine S-R-Sulfoximine increased the total mutant yield by more than 3-fold as well as the proportion of mutants with multilocus deletions. Taken together, our data provide clear evidence that reactive oxygen species play an important causal role in the genotoxicity of arsenic in mammalian cells. (Mol Cell Biochem 234/235: 301–308, 2002) Key words: arsenic, mutagenicity, oxidative stress, antioxidant enzymes, 8-OHdG Introduction Arsenic, as trivalent arsenite (AS 3+ ) or pentavalent arsenate (AS 5+ ), is naturally occurring and ubiquitously present in the environment. Epidemiological data have shown that chronic exposure of humans to inorganic arsenical compounds is associated with liver injury, peripheral neuropathy, and an increased incidence of cancer of the lung, skin, bladder, and liver [1, 2]. However, the mechanism(s) underlying its car- cinogenicity remains unknown. The United States Environ- mental Protection Agency has placed arsenic at the top of its Superfund contamination list [3]. Biologically, the triva- lent sodium arsenite is significantly more active than the pentavalent sodium arsenate [4]. Arsenic contamination of drinking water is a serious en- vironmental problem worldwide because of the large number of contaminated sites that have been identified and the large number of people at risk [5]. The risk of developing arsenic- induced human diseases from environmental exposure is particularly high in many developing countries. For exam- ple, it is estimated that as many as 50 million people are at risk in Bangladesh alone, where both acute and chronic ar- senic poisoning as well as increased cancer incidence have been reported [6]. Although the water supplies in the United States are generally low in arsenic, there have been reports of arsenic contamination of ground water in the Southwest with levels in the hundreds, and in few cases, more than 1,000 μg/l [7, 8], a level that is 20 times higher than the current U.S. maximum contaminant level of 50 μg/l. Occupational expo- sure occurs mainly through inhalation via nonferrous ore smelting, semiconductor and glass manufacturing, or power generation by the burning of arsenic-contaminated coal [7, Address for offprints: T.K. Hei, Center for Radiological Research, Columbia University, VC 11-205, 630 West 168th Street, NY 10032, USA (E-mail: tkhl@columbia.edu)