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      Assessment of the role of DNA damage and repair in the survival of primary cultures of rat cutaneous keratinocytes exposed to bis(2-chloroethyl)sulfide.

      Toxicology and Applied Pharmacology

      Animals, Cell Survival, drug effects, physiology, Cells, Cultured, Centrifugation, Density Gradient, methods, DNA, biosynthesis, DNA Damage, DNA Repair, Dose-Response Relationship, Drug, Keratinocytes, Kinetics, Mustard Gas, adverse effects, Rats, Skin, cytology, Tritium, Thymidine, metabolism

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          Toxicity manifests itself as vesication in human skin exposed topically to bis(2-chloroethyl)sulfide (BCES). The destruction of the proliferating population of epidermal cells is a major component of the pathogenic process. Available data strongly suggest that damage to cellular DNA is a critical factor in the loss of these cells. However, the influence of DNA repair on this toxic response has not been adequately studied. Therefore, a study was undertaken to ascertain the influence of DNA repair on the survival of primary monolayer cultures of rat cutaneous keratinocytes exposed to BCES. The sensitive nucleoid sedimentation assay was employed for the determination of DNA damage in cultures exposed to very low levels of BCES. Initial experiments demonstrated that within 1 hr of exposure to as little as 0.1 microM BCES the structural integrity of cellular DNA was compromised, presumably resulting from the appearance of single-strand breaks in the nucleic acid. This same effect was demonstrated in basal cells derived from a stratified, cornified culture grown at the air-liquid interface and exposed topically to the vesicant. Further studies with the monolayer culture demonstrated that the gross structural integrity of the DNA in cells exposed to as much as 5 microM BCES was completely restored within the first 22 hr following the exposure. However, this repair process appeared to be inefficient since a depression of thymidine incorporation into DNA and a significant loss of DNA were exhibited in exposed cultures as long as 72 hr after the initial exposure.

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