Apoptosis assay

Cell apoptosis was detected by the Annexin V-FITC/PI Kit (4A Biotech Co. Ltd., Beijing, China; Cat. No.: FXP018-100), according to the manufacturer’s protocol. Cells were seeded into 6-well plates at a density 1 × 10⁵ cells/well and treated with 0–400 μg/mL tanshinol for 48 h. Cells were then collected and re-suspended in 200 μL binding buffer containing 10 μL Annexin V-FITC and 5 μL PI with an incubation in the dark for 30 min. Finally, the apoptotic cells were distinguished by a FACS Calibur flow cytometer (Becton-Dickinson) immediately.