Cytotoxic potency of H₂O₂ in cell cultures: Impact of cell concentration and exposure time

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• 作者：Michael Gü ; lden ; Anne Jess ; Julia Kammann ; Edmund Maser ; Hasso Seibert
• 关键词：Hydrogen peroxide ; C6 cells ; Oxidative stress ; Cytotoxicity ; Free radicals
• 刊名：Free Radical Biology and Medicine
• 出版年：2010
• 出版时间：1 November 2010
• 年：2010
• 卷：49
• 期：8
• 页码：1298-1305
• 全文大小：688 K

文摘

Using C6 glioma cells in this study we investigated in detail how exposure time and cell concentration affect the cytotoxic potency of H₂O₂ in vitro. Median cytotoxic concentrations (EC₅₀) decreased from 500 to 30 μM with increasing incubation time from 1 to 24 h. Twenty-four hours proved to be sufficient to determine incipient cytotoxic concentrations of H₂O₂. The incipient EC₅₀ values were linearly related to the cell concentration. A cell concentration-independent median cytotoxic cell dose (ED₅₀) of 430 nmol/mg cell protein or 860 nmol/10⁷ cells was derived. Median cytotoxic H₂O₂ concentrations were completely eliminated from the culture medium at a rate proportional to both the H₂O₂ and the cell concentrations. In contrast to EC₅₀ values the corresponding areas under the concentration versus time curve (AUC) were independent of the cell concentration and amounted to 1800 μM × min. With decreasing cell concentration the H₂O₂ elimination decelerates and, thus, exposure to H₂O₂ applied as a bolus approaches a continuous exposure to a steady H₂O₂ concentration. Taken together, our results indicate that the cytotoxic potency of H₂O₂ administered to cultured cells as a bolus is characterized by the AUC, which depends on its initial concentration, the ability of the cells to eliminate H₂O₂, and the cell concentration. We recommend expressing the toxic potency of H₂O₂ in vitro by the incipient toxic cell dose (e.g., nmol H₂O₂/mg cell protein or nmol H₂O₂/10⁷ cells), in particular for comparative purposes.