The Relative Semi-quantification of mRNA Expression as a Useful Toxicological Endpoint for the Identification of Embryotoxic/Teratogenic Substances.

Embryonic stem cells, which resemble the undifferentiated cells of the epiblast in a blastocyst, are able to differentiate into derivatives of the primary germ layers, including cardiomyocytes. The effects of embryotoxic/teratogenic compounds on the differentiation of cells was examined by semi-quantification of the mRNA expression using the RT-PCR protocol. Alpha and beta myosin heavy chain (alpha-MHC and beta-MHC, respectively) mRNA expression were chosen as tissue-specific markers, characteristic of early cardiac muscle development. Nine chemicals were investigated, chosen according to their in vivo embryotoxic/teratogenic potential in mice. The teratogens all-trans retinoic acid (RA), 5-fluorouracil (5-FU), hydroxyurea (HU), diphenylhydantoin (DPH) and caffeine (Caff) caused a significant reduction in MHC mRNA expression at a dose lower than that required for cytotoxicity. Saccharin (Sacc) had a similar effect, while penicillin G (PenG), isoniazid (Iso) and cytarabine (Ara-C) only showed an effect on MHC mRNA expression when the cells had a significant loss in viability. The inability to identify the strong teratogen Ara-C is related to the inappropriate target tissue.