Studies on mutagenesis by ethylene oxide and styrene oxide in the hprt gene in human cells

Abstract

Environmental, occupational and life style-related exposures to mutagenic agents may contribute to cancer risk in humans. To prevent the potentially hazardous effects of such agents it is important to understand their mechanisms of action. In this thesis, the mutagenic effects of two important industrial epoxides, ethylene oxide and styrene oxide, have been studied.

Ethylene oxide is used as a chemical intermediate in the production of many industrial chemicals and as a disinfectant and sterilising agent. It is a direct acting alkylating agent which has recently been classified as carcinogenic to humans. Styrene-7,8-oxide is the primary in vivo metabolite of styrene, which is used in the production of polymers and copolymers, and as an important part of unsaturated polyester resins in the production of glass-reinforced plastics products. Styrene and styrene-7,8-oxide have been classified as possibly and probably carcinogenic to humans.

The frequency and nature of ethylene oxide and styrene oxide-induced mutations were studied in the hypoxanthine guanine phosphoribosyl transferase (hprt) gene inhuman fibroblast and T-lymphocyte cultures, respectively. Polymerase chain reaction-based techniques, Southern blotting and direct DNA sequencing were used to study the molecular nature of mutations induced by the two compounds. Ethyleneoxide caused a 5-19-fold increase of hprt mutant frequency relative to the background frequency in fibroblasts. Base substitutions and large hprt gene alterations were the major types of mutation induced by ethylene oxide. Among base substitutions, both transitions and transversions were identified, the most frequent being AT to TA transversions. Among the large alterations, both total and partial gene deletions were found. The results suggest that several different target sites might be involved in ethylene oxide mutagenesis. Styrene-7,8-oxide caused a 4-fold increase of the hprt mutant frequency in cultured human T-lymphocytes.

Base substitutions were the major type of mutation and themost frequent were AT to GC transitions. Approximately 60% of the base substitutions in the coding region and the splice site sequences were at AT base pairs, suggesting that DNA adducts at these sites may be important in the mutagenesis of styrene oxide. The frequency of hprt mutant peripheral blood T-lymphocytes was studied in a group of styrene-exposed hand lamination workers. The mutant frequency was higher in the laminators than in a group of office workers from the same factory at three of four sampling times within a seven-month period. Another control group, working outside the factory and not exposed to styrene, showed a significantly lower hprt mutant frequency than the styrene exposed laminators.

Taken together, these results have contributed to the knowledge on mutagenesis by ethylene oxide and styrene oxide in human cells in vitro and support the possible mutagenic effect of human styrene exposure in vivo.