Methods: OKF6/TERT2 cells were treated for 12-48 h with 2.5 mM CQ alone and for 24 h also in combination with various concentrations of GSH (0.5 -5.0 mM) in the dark. Cell viability and apoptosis were assessed by flow cytometry using the Annexin V-FITC/PI assay. Bonferroni´s ANOVA (p < 0.05) was used for statistical analysis.
Results: Exposure of OKF6/TERT2 to 2.5 mM CQ resulted in a significant increase of early apoptotic cells (10.8 ± 2.3%) in comparison to untreated control cells (c: 0.5 ± 0.1%; p < 0.05) after 24h only. This increase was associated with a marked decrease in cell viability (2.5 mM CQ: 58.3 ± 18.6%;c: 95.2 ± 1.3%; p < 0.05). GSH in concentrations of 0.5-2.5 mM showed no effect on CQ-induced apoptosis, while 5.0 mM GSH caused a significant decrease of apoptotic cells to 2.7 ± 0.6% (p <0.05). Concurrently, GSH increased the amount of viable OKF6/TERT2 cells exposed to CQ to 78.6 ± 9.7% in comparison to control cells, but this elevated cell viability was not statistically significant. GSH alone showed no effects on apoptosis and viability of OKF6/TERT2 cells (5.0 mM GSH: 0.4% ± 0.0% early apoptotic cells; 94.1 ± 1.2% viable cells).
Conclusion: Co-treatment with GSH clearly inhibited CQ-triggered apoptosis in OKF6/TERT2 cells. Therefore it may be concluded that reactive oxygen species play a crucial role in apoptosis induced by CQ in human oral keratinocytes.