Valproic acid (VPA), an important drug for treatment of epilepsy, has severe teratogenic effects on 1-2 % of fetuses of treated pregnant women. The effects of VPA are termed "fetal valproate syndrome", which describes predominantly neural tube defects, i.e. Spina bifida). The teratogenic mechanisms of VPA is unknown, but different from the anticonvulsive action of VPA, as distuingished by the effects of a number of VPA analogues.

Mouse F9 teratocarcinoma cells were identified as an in vitro system to study the teratogenic potential of VPA and it's analogues. Treatment of F9 cells with VPA induced F9 cell differentiation, as observed by an altered cell morphology, inhibition of cell proliferation, and induced expression of differentiation marker genes (Keratin 18, but not Laminin b1 and Kollagen(a1)IV. Exogenously administered retinoic acid causes similar teratogenic effects in the mouse embryo (Spina bifida) as VPA, which is reflected in F9 cells. Nevertheless, VPA-induced F9 cell differentiation is different from retinoic acid-induced F9 cell differentiation (cell morphology, differentiation marker expression).
Another indication of F9 cell differentiation was the induction of transiently transfected reporter constructs containing a viral promotor (RSV, CMV, SV40 promotors). This was reporter gene-independent, and a reporter with a non-viral promotor was not induced. Viral promotors are repressed in undifferentiated embryonal carcinoma cells as e.g. F9 cells, but they are activated upon induction of cell differentiation. The VPA-induced transcription of a viral reporter was completely inhibited by cycloheximide, i.e. VPA-induction of viral reporter is dependent on de-novo protein synthesis.

The induction of transiently transfected viral reporter by VPA is a useful system to study the possible regulation of the VPA-induced F9 cell differentiation by peroxisome proliferator-activated receptors (PPARs). PPAR receptors are nuclear receptors of the steroid hormone receptor superfamily, which are activated by fatty acids and derivatives. VPA activates PPAR receptors: in CHO cells, a hybrid receptor containing the ligand binding domain of rat PPARa and the DNA binding domain of the human glucocorticoid receptor was stereospecifically activated by VPA and derivatives, as measured by induction of a GRE-driven reporter gene. The teratogenic potential of VPA derivatives in mice correlates with their potential to activate PPAR a expressed in CHO cells. Only the PPARd isoform of the mammalian PPAR receptors (PPARa, d, g1/2) was detected in F9 cells, i.e. only PPARd could be responsible for mediation of the VPA effects. The teratogenic action of VPA and analogues in the embryo correlates with the induction of viral reporters in F9 cells; therefore, it is possible to measure indirectly the teratogenic potential of VPA analogues in F9 cells. F9 cells are derived from an early phase of mouse embryonal development (comparable to embryonal stem cells), and represent therefore a cell model to analyse the molecular mechanisms of mouse embryonal differentiation (expression of teratogenic target genes, mechanisms of target gene regulation during cell differentiation, and the role of receptors and their subtype-specific function in the regulation of cell differentiation).

The teratogenicity of VPA could be explained by the model of VPA-induced activation of PPARd in the embryo, and hence interference with embryonal differentiation processes by VPA. PPARd is the only subtype which is expressed during the VPA-sensitive phase of the mouse embyonal development, and in situ hybridisation revealed expression in the neural tube of mouse embryos (the teratogenic target tissue of VPA). F9 cells might allow an initial screening of VPA analogues in search of a drug, which possesses the anticonvulsive action of VPA, but no teratogenic side effects.