Activation of genotoxins to DNA-damaging species in exfoliated breast milk cells.
ABSTRACT Exfoliated cells, isolated from breast milk samples donated by UK-resident women (n=15), were incubated, either immediately or after culture for 7 days, with one of a series of genotoxins, either in the presence or absence of the DNA-repair inhibitors, hydroxyurea (HU), and cytosine arabinoside (ara-C). The numbers of DNA single-strand breaks induced were then assessed as comet tail length (CTL) (microm) using the alkaline single cell-gel electrophoresis ('Comet') assay; cell viability was measured by trypan blue exclusion. The heterocyclic aromatic amines (HAAs) (2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) (0.4 mM), 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1) (1.67 mM), 3-amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2) (1.77 mM)), a polycyclic aromatic hydrocarbon (benzo[a]pyrene (B[a]P) (0.36 mM)), a nitro-polycyclic aromatic hydrocarbon (1-nitropyrene (1-NP) (1.84 mM)) and aromatic amines (o-toluidine (0.85 mM), p-chloroaniline (0. 71 mM)) each induced statistically significant (P<0.0001, Mann-Whitney test) increases in median CTLs in breast milk cells from all the donors examined when incubated (30 min, 37 degrees C) in the presence of HU/ara-C. In some cases, these compounds were also active in the absence of the repair inhibitors. There were marked variations in comet formation between donors and between genotoxins. Cell culture appeared to increase the epithelial cell proportion and cultured cells retained their ability to activate genotoxins. The results suggest that breast milk is a valuable source of human mammary cells for the study of the metabolic activation of possible carcinogens.
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ABSTRACT: Extensive research indicates that the etiology of breast cancer is complex and multifactorial and may include environmental risk factors. Breast cancer etiology and exposure to xenobiotic compounds, diet, electromagnetic fields, and lifestyle have been the subject of numerous scientific inquiries, but research has yielded inconsistent results. Biomonitoring has been used to explore associations between breast cancer and levels of environmental chemicals in the breast. Research using breast tissues and fluids to cast light on the etiology of breast cancer is, for the most part, predicated on the assumption that the tissue or fluid samples either contain measurable traces of the environmental agent(s) associated with the cancer or that they retain biological changes that are biomarkers of such exposure or precursors of carcinogenic effect. In this paper, we review breast cancer etiology research utilizing breast biomonitoring. We first provide a brief synopsis of the current state of understanding of associations between exposure to environmental chemicals and breast cancer etiology. We then describe the published breast cancer research on tissues and fluids, which have been used for biomonitoring, specifically human milk and its components, malignant and benign breast tissue, nipple aspirate fluid (NAF) and breast cyst fluid. We conclude with a discussion on recommendations for biomonitoring of breast tissues and fluids in future breast cancer etiology research. Both human milk and NAF fluids, and the cells contained therein, hold promise for future biomonitoring research into breast cancer etiology, but must be conducted with carefully delineated hypotheses and a scientifically supportable epidemiological approach.Journal of Exposure Science and Environmental Epidemiology 10/2007; 17(6):525-40. DOI:10.1038/sj.jes.7500548 · 3.05 Impact Factor
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ABSTRACT: Cumulative exposure to oestrogen has been linked to increased risk of breast cancer. Whilst oestrogens induce cancers in rodent bioassays it is unclear whether the mechanisms involved are genotoxic and/or epigenetic. The cytokinesis block micronucleus (CBMN) and the alkaline single cell-gel electrophoresis 'Comet' assays were used to examine MCF-7 cells for chromosomal damage and DNA single-strand breaks (SSBs), respectively. The comet-forming activities of oestrogens were also tested in a 72 h primary culture of cells isolated from freshly expressed breast milk. Micronuclei (MN) were scored in 500 binucleate cells per treatment and SSBs were quantified by comet tail length (CTL) (microm). Effects on mitotic rate (per cent binucleate cells) and cell viability (per cent plating efficiency) were also assessed. beta-Oestradiol, oestrone and oestriol were tested for genotoxicity in the 10(-10)-10(-4) M and 10(-10)-10(-2) M concentration ranges in the CBMN and Comet assays, respectively. Beta-Oestradiol, following 24 h treatment but not 120 h treatment, induced increases (up to 3-fold) in MN at a concentration of 10(-9) M. Oestrone induced dose-related increases in MN (up to 5-fold) following both 24 and 120 h treatment, whereas oestriol appeared not to induce MN. All three oestrogens induced dose-related increases in per cent binucleate cells suggesting that they enhance mitotic rate. In the Comet assay both beta-oestradiol and oestrone induced dose-related increases in SSBs (up to 7-fold over control CTL) and were significantly comet-forming (P < 0.0001) at concentrations as low as 10(-9) and 10(-8) M, respectively, whereas oestriol was less genotoxic. All three oestrogens were significantly comet-forming (P < 0.0001) in a primary culture of breast milk cells, suggesting that they can damage the target cells from which breast cancers may eventually arise.Mutagenesis 07/2002; 17(4):345-52. · 3.50 Impact Factor
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ABSTRACT: Epidemiological studies indicate the involvement of environmental factors in the etiology of breast cancer, but have not provided clear indications of the nature of the agents responsible. Several environmental carcinogens are known to induce mammary tumors in rodents, and the abundance of adipose tissue in the human breast suggests that the epithelial cells, from which breast tumors commonly arise, could be exposed to lipid-soluble carcinogens sequestered by the adipose tissue. In this report we review our studies in which we have examined human mammary lipid, obtained from elective reduction mammoplasties from healthy donors, and human milk from healthy mothers, for the presence of components with genotoxic activity in several in vitro assays. A significant proportion of lipid extracts induced mutations in bacteria and micronuclei in mammalian cells. They also caused DNA damage, detected as single-strand breaks in the alkaline single-cell gel electrophoresis (comet) assay, in both the MCL-5 cell line and in primary cultures of human mammary epithelial cells. Genotoxic activity was also found in a significant proportion of extracts of human breast milk. Viable cells recovered from milk samples showed evidence of DNA damage and were susceptible to comet formation by genotoxic agents in vitro. Genotoxic activity was found to be less prevalent in milk samples from countries of lower breast cancer incidence (the Far East) compared with that in samples from the UK. The agents responsible for the activity in milk appear to be moderately polar lipophilic compounds and of low molecular weight. Identification of these agents and their sources may hold clues to the origins of breast cancer. Environ. Mol. Mutagen. 39:143–149, 2002. © 2002 Wiley-Liss, Inc.Environmental and Molecular Mutagenesis 01/2002; 39(2‐3):143 - 149. DOI:10.1002/em.10049 · 2.55 Impact Factor