Shikonin suppresses cell proliferation by inducing apoptosis in oral cancer

Abstract

Background: Phytotherapy has provided a new direction to the treatment of many cancers, including oral cancer, over conventional chemotherapeutic agents as it has fewer side effects. Shikonin (Shk) is a natural biologically active alkaloid found in the roots of Lithospermum erythrorhizon plant. It exhibits potent cytotoxic and anti-migratory effects in many cancer cell lines. However, the cytotoxic potential of shikonin in oral squamous cell carcinoma (OSCC) is not clearly understood. In this study, the anticancer potential of Shk has been demonstrated in SCC9 oral cancer cells. Methods: Anticancer potential of Shk in SCC9 cell line was demonstrated by investigating the anti-proliferative, anti-migratory and apoptosis promoting activity through performing MTT assay, Acridine orange/Ethidium bromide (AO/EtBr) staining, DAPI staining, reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) measurement assays, comet assay, qRT PCR, in vitro scratch assay and western blotting. Results: We observed that Shk reduced the viability of SCC9 cell in a time and concentration depending manner. It also promotes apoptosis via overexpressing pro-apoptotic proteins like Bax and Caspase 3. Additionally, Shk treatment increases the generation of intracellular ROS levels. Meanwhile, this ROS production was reversed with pre-treatment of NAC (N‐acetyl cysteine, ROS scavenger). Furthermore, production of excess ROS due to Shk treatment depletes the MMP and damages the DNA. Shk also inhibits SCC9 cells migration and proliferation by altering the expression of epithelial-mesenchymal transition (EMT) markers like E‐cadherin, ZO‐1, N‐cadherin, and Vimentin. Overall these results concluded that Shk could be an excellent phytotherapeutic agent for treatment of oral cancer.