Withania somnifera triggers cytotoxicity and apoptosis in cancer cells through mitochondrial dysfunction and oxidative stress

Cancer remains one of the top deadly diseases, largely due to lack of definitive and effective therapeutic agent. Withania somnifera, a renowned remedy, widely been used in ancient medication to help with several diseases. This study aims to investigate the potential anticancer effect in HepG2, MCF-7 and A-549 cells and identify the molecular mechanisms involved in cell death. All cells were exposed to W. somnifera ethanolic root extract (WEE), and cytotoxic effects were evaluated by MTT, and neutral red uptake (NRU) assays. Moreover, WEE treatment on cellular morphology, oxidative damage glutathione (GSH) and; lipid peroxidation (LPO), mitochondrial membrane stability, reactive oxygen species (ROS) production, caspase enzyme activities, and apoptosis induction in cancer cells were evaluated. The MTT and NRU assay demonstrated a strong dose-dependent cytotoxic effects of WEE on HepG2, MCF-7 and A-549 cells at 100-500 μg/mL for 24 h, with an IC 50 of 230 μg/mL, 384 μg/mL and 595 μg/mL, correspondingly. These cytotoxic effects were associated with altered cellular morphology. Furthermore, exposure of HepG2 cells to the cytotoxic concentrations of WEE led to oxidative stress, excessive ROS generation, activation of caspase enzymes, and loss of mitochondrial membrane potential. These effects observed were linked to the dysregulation of key apoptotic genes, including the upregulation of p53, caspase-3, Bax, and caspase-9, with the downregulation of antiapoptotic gene Bcl-2, which jointly triggered mitochondrial mediated apoptosis in HepG2 cells. Taken together, this study highlights the strong differential cytotoxic effects of WEE on HepG2 cells, mainly driven by oxidative stress and activation of mitochondrial apoptotic pathway.