Abstract
Oxidative stress appears to be implicated in the pathogenesis of various diseases including alcoholic liver injury. In this study we investigated the mechanism of apoptosis induced by tert-butyl hydroperoxide (TBHP) in HepG2 human hepatoblastoma cells. Treatment with TBHP significantly reduced glutathione content and glutathione reductase activity, and increased glutathione peroxidase activity, indicating that TBHP induced oxidative stress in the HepG2 cells. TBHP also induced reduction of cell viability and DNA fragmentation, a hallmark of apoptosis, in a dose-dependent manner. In addition, TBHP induced a sustained increase in intracellular Ca2+ concentration, which was completely prevented by the extracellular Ca2+ chelation with EGTA. TBHP also induced Mn2+ influx. These results indicate that the intracellular Ca2+ increase by TBHP is exclusively due to Ca2+ influx from the extracellular site. Treatment with either an extracellular (EGTA) or an intracellular Ca2+ chelator (BAPTA/AM) significantly suppressed the TBHP-induced apoptosis. Taken together, these results suggest that TBHP induced the apoptotic cell death in the HepG2 cells and that Ca2+ influx may play an important role in the apoptosis induced by TBHP.
Highlights
Liver produces large amounts of oxygen free radicals in the course of detoxifying xenobiotic and toxic substances (Stohs, 1995)
To confirm that tert-butyl hydroperoxide (TBHP) induces oxidative stress in HepG2 human hepatoblastoma cells, we examined the effects of TBHP on the levels of GSH and the activities of GSH peroxidase and reductase
These results suggest that TBHP induced oxidative stress in the HepG2 cells
Summary
Liver produces large amounts of oxygen free radicals in the course of detoxifying xenobiotic and toxic substances (Stohs, 1995). Liver injury can occur when large acute doses or chronic exposure to toxic substances overpower the hepatic antioxidant defense system (Ishii et al, 1997). It is well known that these cellular changes induced by OFRs lead to apoptotic cell death in a variety of cell types (Slater et al, 1995; Jenner and Olanow, 1996; Stoian et al , 1996). In this study we investigated the apoptosis induced by oxidative stress in HepG2 human hepatoblastoma cell line as a model human liver cell. We tested the possible involvement of intracellular Ca2+ signal in the action mechanism of TBHP, since intracellular Ca2+ acts as a mediator of apoptosis in many cell types (McConkey and Orrenius, 1996)
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