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Analysis of Biomarkers of DNA Damage and Mutagenicity in Mice Exposed to Acrylonitrile
Vernon E Walker1,2,3, Dale M Walker3,4, Burhan I Ghanayem5
1Wadsworth Center, New York State Department of Health, Albany, New York 12201, United States.
Abstract:
Acrylonitrile (ACN), which is a widely used industrial chemical, induces cancers in the mouse via unresolved mechanisms. For this report, complementary and previously described methods were used to assess in vivo genotoxicity and/or mutagenicity of ACN in several mouse models, including (i) female mice devoid of cytochrome P450 2E1 (CYP2E1), which yields the epoxide intermediate cyanoethylene oxide (CEO), (ii) male lacZ transgenic mice, and (iii) female (wild-type) B6C3F1 mice. Exposures of wild-type mice and CYP2E1-null mice to ACN at 0, 2.5 (wild-type mice only), 10, 20, or 60 (CYP2E1-null mice only) mg/kg body weight by gavage for 6 weeks (5 days/week) produced no elevations in the frequencies of micronucleated erythrocytes, but induced significant dose-dependent increases in DNA damage, detected by the alkaline (pH >13) Comet assay, in one target tissue (forestomach) and one nontarget tissue (liver) of wild-type mice only. ACN exposures by gavage also caused significant dose-related elevations in the frequencies of mutations in the hypoxanthine-guanine phosphoribosyltransferase (Hprt) reporter gene of T-lymphocytes from spleens of wild-type mice; however, Hprt mutant frequencies were significantly increased in CYP2E1-null mice only at a high dose of ACN (60 mg/kg) that is lethal to wild-type mice. Similarly, drinking water exposures of lacZ transgenic mice to 0, 100, 500, or 750 ppm ACN for 4 weeks caused significant dose-dependent elevations in Hprt mutant frequencies in splenic T-cells; however, these ACN exposures did not increase the frequency of lacZ transgene mutations above spontaneous background levels in several tissues from the same animals. Together, the Comet assay and Hprt mutant frequency data from these studies indicate that oxidative metabolism of ACN by CYP2E1 to CEO is central to the induction of the majority of DNA damage and mutations in ACN-exposed mice, but ACN itself also may contribute to the carcinogenic modes of action via mechanisms involving direct and/or indirect DNA reactivity.
Insights
Acrylonitrile (ACN) causes DNA damage and mutations in mice, primarily through its metabolite cyanoethylene oxide (CEO) produced by CYP2E1. This research clarifies ACN
Area of Science:
- Toxicology
- Genetics
- Biochemistry
Background:
- Acrylonitrile (ACN) is a widely used industrial chemical linked to cancer in mice through poorly understood mechanisms.
- Cytochrome P450 2E1 (CYP2E1) metabolizes ACN into the epoxide intermediate cyanoethylene oxide (CEO), a suspected genotoxic agent.
- Understanding the genotoxic and mutagenic pathways of ACN is crucial for assessing its carcinogenic risk.
Purpose of the Study:
- To investigate the in vivo genotoxicity and mutagenicity of Acrylonitrile (ACN) in various mouse models.
- To determine the role of CYP2E1-mediated metabolism in ACN-induced DNA damage and mutations.
- To elucidate the mechanisms underlying ACN's carcinogenicity.
Main Methods:
- Assessment of micronucleated erythrocytes and DNA damage via the alkaline Comet assay in wild-type and CYP2E1-null mice exposed to ACN via gavage.
- Measurement of hypoxanthine-guanine phosphoribosyltransferase (Hprt) gene mutations in T-lymphocytes from ACN-exposed wild-type and CYP2E1-null mice.
- Evaluation of Hprt and lacZ transgene mutations in T-lymphocytes and various tissues of lacZ transgenic mice exposed to ACN in drinking water.
Main Results:
- ACN exposure induced significant dose-dependent DNA damage in the forestomach and liver of wild-type mice, but not CYP2E1-null mice.
- Hprt mutant frequencies increased in wild-type mice, with a more pronounced effect in CYP2E1-null mice only at a lethal ACN dose.
- ACN increased Hprt mutations in lacZ mice but did not elevate lacZ transgene mutations, suggesting CYP2E1-dependent genotoxicity.
Conclusions:
- Oxidative metabolism of ACN by CYP2E1 to CEO is the primary driver of ACN-induced DNA damage and mutations in mice.
- While CEO is the main genotoxic metabolite, ACN itself may contribute to carcinogenicity through direct or indirect DNA reactivity.
- These findings highlight the critical role of metabolic activation in ACN toxicity and cancer risk.
