The Mechanism of t-Butylhydroperoxide-Induced Apoptosis in IMR- 32 Human Neuroblastoma Cells

The Mechanism of t-Butylhydroperoxide-Induced Apoptosis in IMR- 32 Human Neuroblastoma Cells

<P> Apoptosis has been implicated in the pathophysiological mechanisms of various neurodegenerative diseases. In a variety of cell types, oxidative stress has been demonstrated to play an important role in the apoptotic cell death. However, the exact mechanism of oxidative stress-induced apoptosis in neuronal cells is not known. In this study, we induced oxidative stress in IMR-32 human neuroblastoma cells with <I>tert</I>- butylhydroperoxide (TBHP), which was confirmed by significantly reduced glutathione content and glutathione reductase activity, and increased glutathione peroxidase activity. TBHP induced decrease in cell viability and increase in DNA fragmentation, a hallmark of apoptosis, in a dose-dependent manner. TBHP also induced a sustained increase in intracellular Ca<SUP>2&#8290;</SUP> concentration, which was completely prevented either by EGTA, an extracellular Ca<SUP>2&#8290;</SUP> chelator or by flufenamic acid (FA), a non-selective cation channel (NSCC) blocker. These results indicate that the TBHP-induced intracellular Ca<SUP>2&#8290;</SUP> increase may be due to Ca<SUP>2&#8290;</SUP> influx through the activation of NSCCs. In addition, treatment with either an intracellular Ca<SUP>2&#8290;</SUP> chelator (BAPTA/AM) or FA significantly suppressed the TBHP-induced apoptosis. Moreover, TBHP increased the expression of p53 gene but decreased c-myc gene expression. Taken together, these results suggest that the oxidative stress-induced apoptosis in neuronal cells may be mediated through the activation of intracellular Ca<SUP>2&#8290;</SUP> signals and altered expression of <I>p53</I> and <I>c-myc</I>.