Investigating the hepatic developmental toxicity of synthetic phenolic antioxidant 3-BHA: A comprehensive network-based approach combined with human embryonic stem cell-derived hepatocyte models.

During the production and use of plastics, rubber and other food contact materials, free radicals may be generated, which may lead to material aging. In order to alleviate this situation, antioxidants are usually added in the manufacturing process to prevent oxidation reactions, thereby extending the service life. 3-tert-butyl-4-hydroxyanisole (3-BHA) is a synthetic monophenolic antioxidant and isomer of butyl hydroxyanisole, which has been widely used in the food industry. In this study, we combined the method of network toxicology and found that 3-BHA may affect the expression of AURKB, AURKA, and CDK2, thus leading to congenital liver dysplasia. Furthermore, we further tested the developmental toxicity of 3-BHA using the human embryonic stem cells (hESCs) hepatocyte differentiation model combined with molecular biology experimental methods. The results showed that 3-BHA significantly affected the expression of the endoderm marker gene FOXA2. In addition, the expression levels of liver marker genes alpha fetoprotein (AFP) and hepatocyte nuclear factor 4α (HNF4α) were significantly suppressed at the late stage of differentiation as well as hepatocyte-specific functions. These findings suggest that 3-BHA inhibits hepatocyte differentiation and may produce potential toxicity during embryonic development and liver formation.