H₂O₂ inhibits the growth of human pulmonary fibroblast cells by inducing cell death, GSH depletion and G1 phase arrest

Cultured normal human cells are invaluable biological models for mechanistic studies of oxidative stress. Exogenous hydrogen peroxide (H ₂O₂) is often utilized as a representative model of an oxidative stressor. In the present study, the effect of exogenous H ₂O₂ on cell growth and death was evaluated in normal human pulmonary fibroblast (HPF) cells with respect to reactive oxygen species (ROS) and glutathione (GSH) levels. In MTT assays, H₂O₂ inhibited the growth of HPF cells with an IC₅₀ of ~50 μM at 24 h. DNA flow cytometric analysis indicated that 50-500 μM H₂O ₂ significantly induced G1 phase arrest of the cell cycle. H ₂O₂ induced cell death in the HPF cells, which was accompanied by cleavage of caspase-3 and loss of mitochondrial membrane potential (MMP; Δψ_m). However, H₂O₂ was not observed to significantly induce sub-G1 cells. H₂O ₂ increased superoxide anion (O₂^•-) levels from 120 min and increases in ROS, including ₂^•-, were also detected at 24 h. H₂O₂ increased the activity of superoxide dismutase (SOD). H₂O₂ also induced GSH depletion in HPF cells at 24 h and decreased GSH levels after only 25 min. In conclusion, H₂O₂ inhibited the growth of HPF cells via apoptosis and/or necrosis as well as G1 phase arrest, which was accompanied by an intracellular increase in ROS levels and the depletion of GSH. The present study provides an important insight into the toxicological effects of exogenous H₂O₂ on normal HPF cells.