Repair of sulfur mustard-induced DNA damage in mammalian cells measured by a host cell reactivation assay
Department of Biochemistry and Molecular Pharmacology; Department of Cell Biology
Animals; Beetles; CHO Cells; Cell Survival; Cricetinae; DNA; DNA Damage; *DNA Repair; *Dermatologic Agents; Dose-Response Relationship, Drug; *Genetic Techniques; Luciferases; Luminescent Measurements; Models, Chemical; *Mustard Gas; Plasmids; Transfection
Life Sciences | Medicine and Health Sciences
DNA damage is thought to be the initial event that causes sulfur mustard (SM) toxicity, while the ability of cells to repair this damage is thought to provide a degree of natural protection. To investigate the repair process, we have damaged plasmids containing the firefly luciferase gene with either SM or its monofunctional analog, 2-chloroethyl ethyl sulfide (CEES). Damaged plasmids were transfected into wild-type and nucleotide excision repair (NER) deficient Chinese hamster ovary cells; these cells were also transfected with a second reporter plasmid containing RENILLA: luciferase as an internal control on the efficiency of transfection. Transfected cells were incubated at 37 degrees C for 27 h and then both firefly and RENILLA: luciferase intensities were measured on the same samples with the dual luciferase reporter assay. Bioluminescence in lysates from cells transfected with damaged plasmid, expressed as a percentage of the bioluminescence from cells transfected with undamaged plasmid, is increased by host cell repair activity. The results show that NER-competent cells have a higher reactivation capacity than NER-deficient cells for plasmids damaged by either SM or CEES. Significantly, NER-competent cells are also more resistant to the toxic effects of SM and CEES, indicating that NER is not only proficient in repairing DNA damage caused by either agent but also in decreasing their toxicity. This host cell repair assay can now be used to determine what other cellular mechanisms protect cells from mustard toxicity and under what conditions these mechanisms are most effective.
Carcinogenesis. 2001 Apr;22(4):661-4.
Matijasevic, Zdenka; Precopio, Melissa Lynn; Snyder, Jennifer E.; and Ludlum, David B., "Repair of sulfur mustard-induced DNA damage in mammalian cells measured by a host cell reactivation assay" (2001). Open Access Articles. 382.