Cytogenetic Damage Induced by Acrylamide and Glycidamide in Mammalian Cells: Correlation with Specific Glycidamide-DNA Adducts

Abstract

Acrylamide (AA) is a suspected human carcinogen generated in carbohydrate-rich foodstuffs upon heating. Glycidamide (GA), formed via epoxidation, presumably mediated by cytochrome P450 2E1, is thought to be the active metabolite playing a central role in AA genotoxicity. In this work we investigated DNA damage induced by AA and GA in mammalian cells, using V79 Chinese hamster cells. For this purpose, we evaluated two cytogenetic end points, chromosomal aberrations (CAs) and sister chromatid exchanges (SCEs), as well as the levels of specific GA-DNA adducts, namely, N7-(2-carbamoyl-2-hydroxyethyl)guanine (N7-GA-Gua) and N3-(2-carbamoyl-2-hydroxyethyl)adenine (N3-GA-Ade) using high-performance liquid chromatography coupled with tandem mass spectrometry. GA was more cytotoxic and clastogenic than AA. Both AA and GA induced CAs (breaks and gaps) and decreased the mitotic index. GA induced SCEs in a dose-responsive manner; with AA, SCEs were increased at only the highest dose tested (2mM). A linear dose-response relationship was observed between the GA concentration and the levels of N7-GA-Gua. This adduct was detected for concentrations as low as 1 microM GA. N3-GA-Ade was also detected, but only at very high GA concentrations (>or= 250 microM). There was a very strong correlation between the levels of N7-GA-Gua in the GA- and AA-treated cells and the extent of SCE induction. Such correlation was not apparent for CAs. These data suggest that the induction of SCEs by AA is associated with the metabolism of AA to GA and subsequent formation of depurinating DNA adducts; however, other mechanisms must be involved in the induction of CAs.

Genotoxic effects of acrylamide and glycidamide in mouse lymphoma cells

“…Compared to AA, GA (2-4 mM) was much more mutagenic, about 5-10-fold more potent. This is consistent with several previously reported genotoxicity studies conducted using mammalian cells (Baum et al, 2005;Besaratinia and Pfeifer, 2003;Koyama et al, 2006;Martins et al, 2006). Since the genotoxic responses at higher concentration are reproducible, LOH analysis of the mutants induced by 4 mM GA and 16 mM AA treatments was conducted to determine the types of the mutations.…”

“…Those chromosomal mutations are most likely associated with GA-derived DNA adducts. A recent study shows that there is a strong correlation between the levels of N7-GA-Gua and the extent of sister-chromatid exchange induction in GA-treated V79 cells (Martins et al, 2006). It has been suggested that the clastogenicity results from formation of depurinating DNA lesions and the formation of DNA breaks when repairing the DNA adducts.…”

“…The genotoxicity of AA has been studied extensively, but there are fewer experimental data available with GA as the test substance (Baum et al, 2005;Besaratinia and Pfeifer, 2004;Koyama et al, 2006;Martins et al, 2006;Paulsson et al, 2003). AA is not mutagenic in bacterial assays, and it is mutagenic in mammalian somatic and germinal cells in vitro and in vivo (Dearfield et al, 1995).…”

“…GA is positive for micronucleus induction in vivo in both mice and rats. The micronuclei result from chromosome breakage rather than aneuploidy (Dearfield et al, 1995;Martins et al, 2006). Recently, Manjanatha et al have reported that AA and GA in drinking water increased micronucleated reticulocytes, lymphocyte Hprt mutant frequencies (MFs), and liver cII MFs in Big Blue mice (Manjanatha et al, 2006).…”

Scientific Opinion on acrylamide in food

“…c-mitosis) in the absence of metabolic activation (Baum et al, 2005;Koyama et al, 2006;Jiang et al, 2007;Martins et al, 2007;Katic et al, 2010;ATSDR, 2012;Pingarilho et al, 2013) (Table 21). GA induced also chromosomal aberrations, micronuclei or SCE (Table 21), however at lower concentrations that are not severely cytotoxic.…”