An evaluation of a novel chick cardiomyocyte micromass culture assay with two teratogens/embryotoxins associated with heart defects

• Published in: ATLA. Alternatives to laboratory animals 2007-10, Vol.35 (5), p.505-514
• Main Authors: HURST, Helena S, CLOTHIER, Richard H, PRATTEN, Margaret
• Format: Article
• Language: English
• Subjects: Animal Testing Alternatives; Animals; Anticonvulsants - toxicity; Antineoplastic Agents - toxicity; Biological and medical sciences; Cell Culture Techniques; Cell Differentiation - drug effects; Chick Embryo; Embryology: invertebrates and vertebrates. Teratology; Fundamental and applied biological sciences. Psychology; Heart Defects, Congenital - chemically induced; Immunohistochemistry; Models, Biological; Myocytes, Cardiac - cytology; Myocytes, Cardiac - drug effects; Oxazines; Proteins - analysis; Teratology. Teratogens; Toxicity Tests - methods; Tretinoin - toxicity; Valproic Acid - toxicity; Xanthenes
• ISSN: 0261-1929; 2632-3559
• DOI: 10.1177/026119290703500510

This study was aimed at determining whether the chick cardiomyocyte micromass (MM) system could be employed to predict the teratogenicity/embryotoxicity of exogenous chemicals. Two documented teratogens/embryotoxins, sodium valproate (the sodium salt of valproic acid; VPA) and all-trans retinoic acid (tRA), were used in the initial phase of the study. White Leghorn 5-day-old embryo hearts were dissociated to produce a cardiomyocyte suspension in Dulbecco's Modified Eagle's Medium. Cultures were incubated at 37 degrees C in 5% CO(2) in air, and observations were made every 24 hours over 5 days, for the detection of beating. Culture viability was assessed by using the resazurin reduction assay for determining culture activity and the kenacid blue assay for determining cell number. It was found that tRA significantly reduced cell activity and beating, whilst not affecting total cell number. VPA up to 500 microM induced no cytotoxicity in the MM cardiomyocyte cultures, whilst all the VPA concentrations tested reduced beating. The results demonstrate the potential of the chick cardiomyocyte MM culture assay to identify teratogens/embryotoxins that alter functionality, which may result in a teratogenic outcome, whilst not causing cytotoxicity (direct embryotoxicity). This could form part of a screen for developmental toxicity related to cardiac function, whilst limb cultures and brain cultures based on the same system could be relevant to teratogenic effects on those tissues.