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Mitochondria are the major reactive oxygen species (ROS)--generating sites in mammalian cells. Blockade of complexes in the electron transport chain (ETC) increases the leakage of single electrons to O(2) and therefore increases ROS levels. Complexes I and III have been reported to be the major ROS-generating sites in mitochondria. In this study, using mouse hippocampal HT22 cells as in vitro model, we monitored the change of intracellular ROS level in response to the blockade of ETC at different complex, and measured changes of gene expression of antioxidant enzymes and phase II enzymes, also evaluated potential protective effect of selenium (Se) supplementation to the cells under this oxidative stress. In summary, our results showed that complex I was the major ROS-generating site in HT22 cells. Complex I blockade upregulated the mRNA levels of glutamylcysteine synthetase heavy and light chains, glutathione-S-transferases omega1 and alpha 2, hemoxygenase 1, thioredoxin reductase 1, and selenoprotein H. Unexpectedly, the expression of the enzymes that directly scavenge ROS decreased, including superoxide dismutases 1 and 2, glutathione peroxidase 1, and catalase. Se supplementation increased glutathione levels and glutathione peroxidase activity, indicating a potential protective role in oxidative stress caused by ETC blockade.

Selenoproteins have been shown to exhibit a variety of biological functions, including antioxidant functions, maintaining cellular redox balance, and heavy metal detoxification. UV irradiation-induced damage is partially mediated by increased oxygen radical production. The present study is designed to examine the antioxidative effects of human selenoprotein H (hSelH) after brief period of UVB irradiation on the murine hippocampal neuronal cell line Ht22. Ht22 cells were stably transfected with the hSelH gene or with MSCV empty vector and exposed to UVB irradiation with or without the presence of serum. The results showed that cell viability was significantly higher in hSelH-transfected cells compared to the MSCV vector-transfected cells after 24 h of recovery with or without the presence of serum in the media. Further studies revealed that while the number of superoxide anion (O2*-) positive cells was increased following a 7 mJ/cm(2) of UVB irradiation and 5 h of recovery, overexpression of hSelH significantly reduced superoxide production. These results suggest that hSelH overexpression protects cells from UVB irradiation-induced cell death by reducing the O2*- formation.