Sister Chromatid Exchange Studies in the Chick Embryo and Neonate: Actions of Mutagens in a Developing System

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Abstract
The embryonic and neonatal periods represent times when the disease process may be initiated as a result of exposure to environ mental mutagens and teratogens. We are using the chick embryo/neo-nate as an experimental system to detect and study the genotoxicity of environmental chemicals in developing tissues and the resultant biological alterations in survivors of perinatal chemical exposure. In vivo bromodeoxyuridine (BrdUrd) labeling of replicating DNA has been employed to measure basal and induced sister chromatid exchanges (SCEs), a candidate cytogenetic endpoint in genetic toxicology testing. Additionally, SCE induction studies with model promu-tagens have permitted the detection and study of components of the developing mixed-function oxidase (MFO) system of the liver and other organs. The relationship between specific MFO enzyme induction and SCE generation by promutagens has been studied in ovo and using in vitro assays. The in vivo SCE induction potential of 53 Compounds, including known mutagens and nonmutagens, was evaluated in the early chick embryo. About 90% of the mutagens induced SCEs; all nonmutagens fail-ed to induce SCE above baseline. Clastogens such as bleomycin did not induce SCE but did cause massive chromosome damage that was easlly detected. Gentian violet (GV) is a direct-acting clastogen that did not show any SCE induction. This agrees with the findings from in vitro mutation assays that incorporate rat liver S-9 preparations. Potency for inducing SCE in the chick embryo correlates well with true mutagenic potency, DNA Inhibition, and to some extent with carcinogenic activity. Indirect-acting mutagen-carcinogens induced SCEs and also un scheduled DNA synthesis (UDS) in embryonic cells. Biochemical studies revealed that aryl hydrocarbon hydroxylase (AHH) activity develops in the early embryonic liver as it is first formed at 4-5 da of incubation. The level of AHH activity is sufficient to account for the dramatic SCE response. Enhanced SCE induction occurred in older stage embryos, correlating with the increased basal AHH level and enhanced induction capacity of the liver. Modulation of the MFO enzyme system with specific inducers resulted in altered SCE and UDS responses in vivo and in vitro using a chick microsome/chinese hamster ovary (CHO) mammalian cell assay. Viable embryos and neonates have been obtained following exposure to SCE-inducing levels of the mutagen-carcinogen aflatoxin Bl applied at either 6 da or 12 da of development. Phenotypically “normal” chicks demonstrated deficiencies in the hematolymphoid system and in postnatal growth potential. The postnatal biological outcome was correlated to some degree with the developmental/differentiation status at the time of toxicant exposure. Thus, baseline and induced SCEs (if they occur in the absence of the BrdUrd probe) in the above instance are not associated with gross disturbances of development. Rather, any induced cellular alterations are subtle and expression is delayed. The chick embryo should be useful for the further study of SCEs in a developing system, the effects of chemicals on tissue-specific SCE induction, the role of MFO enzymes in mediating the production of DNA-damaging and SCE-inducing metabolites, and for studying the cellular and developmental consequences of embryo exposure to geno-toxic environmental chemicals.