Simvastatin-induced heme oxygenase-1 increases apoptosis of neuro 2A cells in response to glucose deprivation

Heme oxygenase-1 (HO-1) has been suggested as an important mediator of the cholesterol-independent cytoprotection actions of statins, which may be of benefit for the treatment of degenerative neurological diseases and for reduction of infarct volume after cerebral ischemia. Overexpression of HO-1, however, has dual effects under oxidative stress, and the release of ferric iron from heme under these conditions may result in detrimental rather than cytoprotective effects. This study was designed to investigate the effect of simvastatin-induced HO-1 on Neuro 2A cells in response to glucose deprivation. We demonstrated that simvastatin induced a dose- and time-dependent upregulation of HO-1 protein expression in Neuro 2A cells. The induction of HO-1 after simvastatin treatment was mediated by nuclear factor erythroid 2-related factor 2 (Nrf2), which was expressed by Western blots of nuclear fractions and retarded complex formation in the electrophoretic mobility shift assay reaction. In addition, simvastatin activated the extracellular signal-regulated kinase and p38, but not the phosphorylation of c-Jun N-terminal kinase and Akt. Glucose deprivation in the cells pretreated with simvastatin induced more HO-1 expression, and the transcript could be decreased by small interfering RNA for Nrf2. This upregulation of HO-1 was significantly associated with increased apoptosis, manifested as expression at the protein level of 17-kDa cleaved caspase-3 and increased percentage of apoptotic cells shown by flow cytometry. The increased cleaved caspase-3 expression and percentage of apoptotic cells was significantly reduced by the HO inhibitor zinc protoporphyrin. Addition of the iron chelator desferrioxamine also resulted in blockade of the aggravated apoptosis, which implies that iron production from HO-1 activity may play an important role in the increased apoptosis in response to glucose deprivation in neuronal cells pretreated with simvastatin.