Genotoxicity of styrene–acrylonitrile trimer in brain, liver, and blood cells of weanling F344 rats

Abstract

Styrene–acrylonitrile Trimer (SAN Trimer), a by-product in production of acrylonitrile styrene plastics, was identified at a Superfund site in Dover Township, NJ, where childhood cancer incidence rates were elevated for a period of several years. SAN Trimer was therefore tested by the National Toxicology Program in a 2-year perinatal carcinogenicity study in F344/N rats and a bacterial mutagenicity assay; both studies gave negative results. To further characterize its genotoxicity, SAN Trimer was subsequently evaluated in a combined micronucleus (MN)/Comet assay in juvenile male and female F344 rats. SAN Trimer (37.5, 75, 150, or 300 mg/kg/day) was administered by gavage once daily for 4 days. Micronucleated reticulocyte (MN-RET) frequencies in blood were determined by flow cytometry, and DNA damage in blood, liver, and brain cells was assessed using the Comet assay. Highly significant dose-related increases (P < 0.0001) in MN-RET were measured in both male and female rats administered SAN Trimer. The RET population was reduced in high dose male rats, suggesting chemical-related bone marrow toxicity. Results of the Comet assay showed significant, dose-related increases in DNA damage in brain cells of male (P < 0.0074) and female (P < 0.0001) rats; increased levels of DNA damage were also measured in liver cells and leukocytes of treated rats. Chemical-related cytotoxicity was not indicated in any of the tissues examined for DNA damage. The results of this subacute MN/Comet assay indicate induction of significant genetic damage in multiple tissues of weanling F344 male and female rats after oral exposure to SAN Trimer.

Perinatal toxicity and carcinogenicity studies of styrene-acrylonitrile trimer, a ground water contaminant

“…Interestingly, a stand-alone genetic toxicology study was conducted by the NTP with the same batch of SAN Trimer as used in the current bioassay confirmed the lack of mutagenicity seen in the earlier bacterial and human studies, but found that SAN Trimer, administered once daily for 4 days by gavage to male and female F344/N juvenile rats was associated with significantly increased levels of DNA damage in brain cells from the cerebrum and cerebellum, measured by the Comet assay, and chromosomal damage in peripheral blood reticulocytes, measured by the micronucleus assay (NTP 2012, Hobb et al, 2012). Additional evidence of DNA damage, measured by the Comet assay, was seen in liver cells and peripheral blood leukocytes of exposed male and female rats.…”

Evaluation of the genotoxicity of cell phone radiofrequency radiation in male and female rats and mice following subchronic exposure

“…Slides were prepared and analyzed as described previously (Hobbs et al ; Recio et al ) with some modifications. In a laboratory with controlled humidity (≤60%), samples were thawed on ice and a portion of the cell suspension was diluted with 0.5% low melting point agarose (Lonza, Walkersville, MD) dissolved in Dulbecco's phosphate buffer (Ca +2 , Mg +2 , and phenol free) at 37°C and layered onto each well of a 2‐well CometSlide™ (Trevigen, Gaithersburg, MD).…”

Statistical evaluation of toxicological bioassays – a review

“…As an example, the method description for Comet and MN assay data analysis is used from a recently published study on the genotoxicity of Styrene Acrylonitrile Trimer in brain, liver, and blood cells of weanling F344 rats 45 : The Shapiro-Wilk test … was used to assess normality of the vehicle control group. Data that were normally distributed were analyzed using an independent sample's t-test to compare each dose level to the concurrent control … normally distributed data were also tested for homogeneity of variances using the F test; for data of unequal variances, the Welch's approximation … was used.…”