Assessment of the Developmental Toxicity of Epidermal Growth Factor using Embryonic Stem Cell Test

Purpose: To determine whether epidermal growth factor (EGF) is involved in reproductive developmental toxicity, using the embryonic stem cell test (EST), as well as ascertain how EGF influences embryonic development.
Methods: To predict developmental toxicity on the basis of reducing cell viability and inhibition of differentiation of embryonic stem cells, EST was used to assess changes in different blastodermic genes and expression of proteins including ectodermal-specific genes Pax6, NF-H and glial fibrillary acidic protein (GFAP), mesodermal-specific genes BMP4, GATA4, and MyoD, and endodermal-specific genes, viz, α-fetoprotein, transthyretin (TTR), and albumin, as well as undifferentiated genes, Nanog and Oct4.
Results: The results indicate that EGF was weakly embryotoxic with IC₅₀ESC (i.e., the concentration that reduced ESC viability by 50 %), IC₅₀3T3 (the concentration that reduced 3T3 cell viability by 50 %), and ID₅₀ESC (the concentration that inhibited differentiation of ESC by 50 %) of 6.773, 10.531, and 1.793 μg/mL, respectively. The expression levels of tissue-specific genes of the three germ layers were mainly promoted by 0.01 - 1 μg/mL EGF. Distinctively, relatively high concentrations of EGF caused a discordant effect on the three germ layers. High concentrations of EGF promoted differentiation of the ectoderm and mesoderm, and either inhibited or had mostly no impact on the endoderm.
Conclusion: The imbalance of the three layer-specific genes and expression of proteins, as a result of EGF, might be responsible for its weak level of developmental toxicity. The sensitivity of TTR means that further investigation is required to determine whether it can be used as an embryotoxicity biomarker for growth factors.

Keywords
Embryonic stem cell test; Epidermal growth factor; Developmental toxicity; Germ layers; Blastodermic genes; Protein expression