DNA adduct formation and induction of micronuclei and mutations in B6C3F₁/Tk mice treated neonatally with acrylamide or glycidamide

Abstract

Acrylamide, a food contaminant, is carcinogenic in experimental animals, with both genotoxic and nongenotoxic pathways being proposed. To obtain information regarding mechanisms of acrylamide tumorigenesis, we compared the extent of DNA adduct formation and induction of micronuclei and mutations in mice treated neonatally with acrylamide and its electrophilic metabolite glycidamide. Male and female B6C3F 1 /Tk mice were treated intraperitoneally on postnatal days (PNDs) 1, 8 and 15 or PNDs 1-8 with 0.14 or 0.70 mmol acrylamide or glycidamide per kg body weight per day. One day after the final dose, B6C3F 1 /Tk 1/1 mice were killed to measure DNA adduct levels and peripheral blood micronuclei. Three weeks after the last treatment, B6C3F 1 /Tk 1/2 mice were killed to assess the Hprt and Tk mutant frequencies in spleen lymphocytes. The levels of N7-(2-carbamoyl-2-hydroxyethyl)guanine, the major glycidamide-DNA adduct, decreased in the order 0.70 mmol glycidamide > 0.70 mmol acrylamide > 0.14 mmol glycidamide 0.14 mmol acrylamide. Only glycidamide increased the frequency of micronucleated reticulocytes and normochromatic erythrocytes. In mice treated on PNDs 1, 8 and 15, the Hprt mutant frequency was increased by 0.70 mmol glycidamide. In mice dosed on PNDs 1-8, 0.70 mmol glycidamide caused extensive mortality; each of the other treatments increased the Tk mutant frequency, whereas acrylamide increased the Hprt mutant frequency. These data suggest that the mutagenic response in neonatal mice treated on PNDs 1, 8 and 15 is due to glycidamide, whereas mutations resulting from dosing on PNDs 1-8 are due to another mechanism.

Scientific Opinion on acrylamide in food

“…The i.p. exposure to AA on PND1-8 also increased mutations at the TK and hprt loci in spleen lymphocytes of mice (von Tungeln et al, 2009) and at lac Z loci in transgenic mice (Muta®Mouse) (Hoorn et al, 1993).…”

Genotoxicity of Acrylamide and Glycidamide: A Review of the Studies by HPRT Gene and TK Gene Mutation Assays

“…In the assessment of mutagenic eŠects induced by AA and its metabolite GA, the cell subclones employed in TK and HPRT gene mutation assays include L5178Y TK ＋/-mouse lymphoma cell (18,(28)(29)(30), human lymphoblastoid cell TK6 (31), Chinese hamster ovary (CHO) cell (32), Chinese hamster V79 cell (33)(34)(35) and human promyelocytic leukaemia cells HL-60 and NB4 (36). More recently, the development of the mammalian cell gene assays employing rodent lymphocytes has also contributed signiˆcantly to the understanding of in vivo somatic cell mutagenesis induced by AA, including the assays performed in Big Blue mice, B6C3F1/TK mice and Big Blue rats (37)(38)(39).…”

“…Mutagenicity of AA and GA at the HPRT Locus in DiŠerent Test Systems GA is consistently mutagenic at HPRT mutation assays in various test systems involving mammalian cell lines (32,34,35) and rodents (37)(38)(39). On the contrary, some studies suggested that AA may not induce gene mutation in HPRT test.…”

“…Although a linear increase in the mutant frequency (MF) with the increasing concentration of AA was found, the signiˆcant diŠerence occurred only at the highest concentration of AA (700 mg/L) in HL-60 cells (pº0.01) and NB4 cells ( pº0.01), respectively, where the MF was aboutˆve times higher than that of the control cultures in both cell lines. More recently, there was increasing evidence which suggested that AA as well as GA, produced increased lymphocyte Hprt mutant frequencies (MFs) in rodent animals, including in Big Blue mice, B6C3F1/TK mice and Big Blue rats (37)(38)(39). Manjanatha et al investigated the mutagenicity of AA and GA in Big Blue mice (37).…”

“…AA in vitro induces chromosomal aberrations, micronuclei, sister chromatid exchanges and mitotic disturbance with or without the metabolic activation. It also induces positive responses in the TK locus in vivo and in vitro (18,(28)(29)(30)(31)38). Further analysis of Tk mutants in mouse lymphoma cells showed AA induced almost exclusively small-colony mutants, which appear to represent chromosome alterations to that chromosome 11 carring the Tk locus (28).…”