Mitochondrial dysfunction is now widely anticipated as a major underlying mechanism of aging and a range of chronic diseases, especially through its role in oxidative stress, apoptosis, and cellular senescence. Identifying the mechanisms connecting mitochondrial dysfunction to cell fate decisions is important, particularly restoring the role of reactive oxygen species (ROS) in the regulation of apoptosis and senescence. In vitro experimental settings were designed using HEK293 and HeLa Cell Lines under optimized laboratory controlled conditions. Mitochondrial dysfunction was induced by hydrogen peroxide at concentrations ranging from 50--500 µM, and then measured by fluorescence-based assays using DCFH-DA for ROS detection and JC-1 dye for mitochondrial membrane potential (ΔΨm) (Table 1). Annexin V-FITC/PI staining, caspase-3 activity assays, and β-galactosidase staining were employed in order to assess apoptosis and senescence respectively. Statistical analysis was undertaken using ANOVA, correlation and regression modeling.

Our findings showed that elevated ROS level reduced mitochondrial membrane potential (Δψm, from 1.00 to 0.55) and promoted both apoptosis (from 5% to 55%) and senescence (from 3% to 40%). There were significant correlations between ROS and apoptosis (r = 0.95) and between ROS and Δψm (r = −0.91). ROS was also identified as an important predictor of apoptosis by regression analysis (R² = 0.90). The Time-dependent studies showed that the short treatment with ODSs mainly causes apoptosis, while the long exposure promoted cellular senescence. Furthermore, this suggests that ROS-mediated mitochondrial dysfunction are decisive regulators of the balance between apoptosis and senescence. These results provide useful information on cellular ageing mechanisms and suggest that targeting of mitochondrial pathways and oxidative stress may represent a potential therapeutic option for diseases of aging.