Each siRNA was applied separately for 6 h at 50 nM using INTERFERin. After transfection, the culture media was changed, and the cells were incubated for an additional 24 h.

Dichlorodiphenyldichloroethylene (DDE) is a primary environmental and metabolic degradation product of the pesticide dichlorodiphenyltrichloroethane (DDT). It is one of the most toxic compounds belonging to organochlorines. DDE has never been commercially produced; however, the parent pesticide DDT is still used in some developing countries for disease-vector control of malaria. DDT and DDE remain in the environment because these chemicals are resistant to degradation and bioaccumulate in the food chain. Little is known, however, about DDE toxicity during the early stages of neural development. The results of the present study demonstrate that DDE induced a caspase-3-dependent apoptosis and caused the global DNA hypomethylation in mouse embryonic neuronal cells. This study also provided evidence for DDE-isomer-non-specific alterations of retinoid X receptor alpha (RXRalpha)- and retinoid X receptor beta (RXRbeta)-mediated intracellular signaling, including changes in the levels of the receptor mRNAs and changes in the protein levels of the receptors. DDE-induced stimulation of RXRalpha and RXRbeta was verified using selective antagonist and specific siRNAs. Co-localization of RXRalpha and RXRbeta was demonstrated using confocal microscopy. The apoptotic action of DDE was supported at the cellular level through Hoechst 33342 and calcein AM staining experiments. In conclusion, the results of the present study demonstrated that the stimulation of RXRalpha- and RXRbeta-mediated intracellular signaling plays an important role in the propagation of DDE-induced apoptosis during early stages of neural development.