The Role of Oxidative Stress and Epigenetic Modifications in Valproic Acid-Induced Teratogenesis in the Mouse

Abstract

Exposure to the anticonvulsant valproic acid (VPA) is associated with a 7.5% rate of major malformations and a 1-2% incidence of neural tube defects (NTDs). Although the teratogenic outcomes resulting from VPA use during pregnancy were first identified in the 1980s, the mechanisms by which VPA induces birth defects are not fully elucidated. Based on evidence in the literature, the studies in this thesis examined the role of in utero VPA exposure on oxidative stress and epigenetic alterations in the developing embryo to provide further mechanistic insight into VPA’s teratogenic pathway. The first study investigated the role of oxidative stress in VPA-induced teratogenesis. Using CD-1 mice, catalase was shown to protect against VPA-induced effects on developmental and morphological parameters in both whole embryo culture and in vivo models. Studies in whole embryo culture demonstrated that markers of oxidative damage were not altered by VPA; however, VPA increased apoptosis in the neuroepithelium, which was attenuated by the addition of catalase. The second objective addressed epigenetic modifications induced by VPA in an in vivo mouse model. Maternal administration of VPA resulted in increased acetylation of histones H3 and H4, increased methylation of histone H3K4, and decreased methylation of histone H3K9. Furthermore, these changes were localized to VPA target tissues including the neuroepithelium, heart, and somites. Global DNA methylation in the embryo was not altered by VPA. The final objective was to determine VPA’s effect on a marker of DNA damage, markers of cell cycle proteins, and a marker of apotosis in vivo. Maternal administration of VPA resulted in a rapid increase of γH2A.X, a marker of DNA double strand breaks (DSBs). Increased expression of p27KIP1, a cyclin-dependent kinase inhibitor, and activated caspase-3, a marker of apoptosis, were observed and these changes were localized to the neuroepithelium of developing embryos. In conclusion, this thesis supports the hypothesis that VPA-induced increases in ROS production and HDAC inhibition may lead to altered gene expression patterns and consequently teratogenic effects, namely NTDs.