Mitochondrial antioxidants decrease oxidative stress and inflammation induced bronchial epithelial cell line

Reactive oxygen species (ROS) and inflammation are main key factors driving chronic lung diseases and both can cause mitochondrial dysfunction. We hypothesized that mitochondrial function, inflammation and proliferation in bronchial epithelial line cell line (BEAS-2B) may be reduced by treatment with mitochondrial targeting antioxidants. BEAS-2B was pretreated with different concentrations of the antioxidants N-acetyl cysteine (NAC), mitoTEMPO, SS31, AP39 and AP219 for 3h, and stimulated with 200μM H₂O₂ and 1ng/ml IL-1β. Mitochondrial membrane potential (Δψ_m) and mitochondrial ROS (mROS) were measured using MitoProbe^TM JC-1 and MitoSOX^TM Red respectively, using flow cytometry. Inflammatory gene expression (<i>IL8</i>) and CXCL8 were measured using RT-PCR and ELISA, respectively. Cell proliferation was measured using BrdU assay. Results show that H₂O₂ decreased Δψ_m and increased mROS compared to control. Δψ_m is restored in mitoTEMPO, SS31 and AP39 pretreated groups compared to H₂O₂ treated alone. mROS was decreased when pretreating with 10mM NAC whereas mitochondrial targeted antioxidants did not lower mROS compared to H₂O₂ treated alone. The cells treated with IL-1β, but not H₂O₂, had significantly increased proliferation, which was decreased when treated with NAC and AP39. Proliferation was decreased in the IL-1β treated group when pretreated with NAC, mitoTEMPO and AP39 compared to IL-1β treated alone. NAC and AP39 also reduced CXCL8 release and <i>IL8</i> gene expression in response to IL-1β treatment. In conclusion, mitochondrial targeted antioxidants reduce mitochondrial oxidative stress and reduce inflammation and proliferation in BEAS-2B stimulated with H₂O₂ and IL-1β.