282 - The protective effects of Nrf2 against valproic acid in differentiating P19 cells

Abstract

Valproic acid (VPA) is an effective antiepileptic drug that has been shown to cause birth defects, especifically neural tube defects (NTD), when taken by pregnant mothers. The mechanisms responsible for the embryotoxic effects of VPA remain unclear, but there is evidence to suggest that they are related to oxidative stress. Nuclear factor (erythroid-derived 2)-like 2 (Nrf2) is a transcription factor that activates the cellular antioxidant response leading to an upregulation of proteins/enzymes that combat the negative effects caused by oxidative stress. The goal of this study is to determine if VPA alters neuronal differentiation in vitro and evaluate the protective role of Nrf2. P19 cells are an embryonic carcinoma cell line that can be differentiated into neurons over the course of four days. Data show that as P19 cells progressively become more differentiated, glutathione redox potentials become increasingly oxidized, suggesting a possible level of redox regulation during neurogenesis. To evaluate the effect of VPA on differentiation, every 12 hours throughout the four day differentiation period, cells were dosed with VPA (0.5 mM) or vehicle, and were collected at the termination of differentiation (day 4). Levels of beta-III Tubulin, a neuronal marker, were measured and compared to an undosed, fully differentiated control. VPA significantly decreased beta-III Tubulin expression compared to the undosed control, suggesting decreased differentiation. This approach was repeated but 3H-1,2-dithiole-3-thione (D3T), a potent Nrf2 inducer, was added (10 uM) 6 hours prior to dosing with VPA or vehicle. Decreased differentiation was improved in cells that were co-treated with VPA and D3T compared to those that were treated with VPA only. These results suggest that prior Nrf2 induction in VPA-treated cells may be a means to promote normal neurogenesis.