Withaferin A induces apoptosis and inhibits adipogenesis in 3T3‐L1 adipocytes

Withaferin A (WA), a highly oxygenated steroidal lactone that is found in the medicinal plant Withania somnifera (ashwagandha) has been reported to have anti‐tumor, anti‐angiogenesis, and pro‐apoptotic activity. We investigated the effects of WA on viability, apoptosis and adipogenesis using 3T3‐L1 adipocytes. Mature adipocytes were treated with WA (1μM to 25μM) for 6, 12 and 24h. Viability and apoptosis were measured by CellTiter‐Blue® Cell Viability Assay and single strand DNA ELISA assay, respectively. WA decreased viability in a dose‐ and time‐dependent manner. WA 10μM and WA 25μM increased apoptosis by 73.2 ± 8.1 % and 104.1 ± 19.7% (p<0.001) after 24 hours. WA's effect on adipogenesis was examined by AdipoRed™ assay after treating with WA (0.1μM – 5μM) during differentiation. WA decreased lipid accumulation in a dose‐dependent manner without decreasing viability up to 1μM. WA 0.1μM, 0.5μM and 1μM decreased lipid accumulation by 6.5 ± 5.1%, 47.0 ± 2.7% and 86.1 ± 0.6% (p<0.001). Treatment with WA during the first 2 days seems critical in lipid accumulation. The effect on apoptosis and lipid accumulation were also confirmed with Hoechst staining and Oil Red O staining, respectively. These results show that WA has a potential to act directly on adipocytes to reduce cell viability and adipogenesis and also induce apoptosis. The outcome of this study may have applications in the treatment of obesity.