Involvement of activation of the Nrf2/ARE pathway in protection against 6-OHDA-induced SH-SY5Y cell death by α-iso-cubebenol

Free radical-mediated neurodegeneration is one of the many causes of Parkinson's disease (PD). As part of our ongoing studies on the identification of biologically active Schisandra chinensis components, we have isolated and structurally elucidated alpha-iso-cubebenol. This study was carried out in an attempt to clarify the neuroprotective effect of alpha-iso-cubebenol on toxin-insulted dopaminergic neuronal death using 6-hydroxy-dopamine (6-OHDA)-induced dopaminergic SH-SY5Y cells. alpha-iso-cubebenol significantly attenuated the loss of mitochondria] function (MTT assay) and membrane integrity (lactate dehydrogenase assay) associated with 6-OHDA-induced neurotoxicity. Pretreatment of the cells with alpha-iso-cubebenol diminished the intracellular accumulation of reactive oxygen species (ROS) and calcium in esponse to 6-OHDA. Moreover, alpha-iso-cubebenol protected against 6-OHDA-induced neurotoxicity through inhibition of SH-SY5Y cell apoptosis. In addition, JC-1 staining, which is a well-established measure of mitochondria] damage, was decreased after treatment with alpha-iso-cubebenol. Notably, alpha-iso-cubebenol inhibited the release of mitochondria] flavoprotein apoptosis inducing factor (AIF) from the mitochondria to the cytosol and nucleus following 6-OHDA treatment. In addition, alpha-iso-cubebenol reduced the 6-OHDA-induced phosphorylation of ERK and induced the phosphorylation of PKA, PKB, and CREB in a dose-dependent manner. Moreover, alpha-iso-cubebenol stimulated the activation of Nrf2, a downstream target of CREB. Furthermore, alpha-iso-cubebenol stimulated the expression of multiple antioxidant response genes (NQO-1 and HO-1). Finally, CREB and Nrf2 siRNA transfection diminished alpha-iso-cubebenol-mediated neuroprotection. (C) 2014 Elsevier Inc. All rights reserved.