Selenium deficiency in tissue culture: Implications for oxidative metabolism

Background: Selenium is located at the catalytic site of the enzyme glutathione peroxidase, and with selenium deficiency the activity of glutathione peroxidase is decreased. Cell culture is an important tool for studying oxidative processes-that is generation and metabolism of oxygen-derived metabolites in the gastrointestinal system. Cell culture is also used to understand the mechanisms of cell injury by oxygen-derived metabolites. Methods: To assess the importance of the selenium content of cell culture media, Caco-2 cells and the hepatoma-derived cell lines, Hep3B and HepG2, were grown to confluence and placed in media with various concentrations of selenium. After 7 to 14 days, cells were harvested and assayed for glutathione peroxidase, lactate dehydrogenase, and protein content. Results: Cells maintained in media unsupplemented with selenium demonstrated a progressive decrease in glutathione peroxidase activity. Cells maintained in media supplemented with various concentrations of selenium demonstrated a dose-dependent increase in glutathione peroxidase until a plateau was reached. The plateau was reached at approximately 400 times the selenium concentration routinely used in cell culture. In the Caco-2 and hepatoma cells, no toxicity was observed at selenium supplementation five times the lowest concentration needed to reach a plateau. Conclusions: Cell culture media are routinely deficient in selenium, and cells that are cultured in this medium are deficient in glutathione peroxidase activity. Studies of oxidative metabolism based on cultures deficient in selenium may yield results that could be falsely interpreted. The addition of 1 nM selenium is sufficient for these cell lines to reach a plateau for intracellular glutathione peroxidase activity. These observations may have important ramifications for the study of reactive oxygen metabolite injury in cell culture.