Withaferin-A Induces Apoptosis in Osteosarcoma U2OS Cell Line via Generation of ROS and Disruption of Mitochondrial Membrane Potential

OBJECTIVE Withaferin-A (WF-A) is a well-known dietary compound isolated from Withania somnifera. It has marked pharmacological potential and has been shown to exhibit antiproliferative activity against several types of cancerous cells. Currently, the main focus of anti-cancer therapeutic development is to identify apoptosis-inducing drug-like molecules. Osteosarcoma is a rare type of bone cancer affecting humans. The objective of the present study was therefore to evaluate the antitumor potential of WF-A against several osteosarcoma cell lines. MATERIALS AND METHODS MTT assay was used to evaluate WF-A against osteosarcoma cell lines and to calculate the IC50. DAPI staining was used to confirm the apoptosis-inducing potential of WF-A. Mitochondrial membrane potential, reactive oxygen species (ROS) assay, and Western blotting were used to confirm the basis of apoptosis. RESULTS The results of the present study revealed that WF-A exhibited strong antiproliferative activity against all the cells lines, with IC50 ranging from 0.32 to 7.6 µM. The lowest IC50 (0.32 µM) was observed against U2OS cell line and, therefore, it was selected for further analysis. DAPI staining indicated that WF-A exhibited antiproliferative activity via induction of apoptosis. Moreover, WF-A induced a ROS-mediated reduction in mitochondrial membrane potential in a dose-dependent manner and activation of caspase-3 in osteosarcoma cells. CONCLUSIONS We suggest that WF-A may prove a potent therapeutic agent for inducing apoptosis in osteosarcoma cell lines via generation of ROS and disruption of mitochondrial membrane potential.