Curcumin Enhances Low-intensity Ultrasound Killing Nasopharyngeal Carcinoma Cells

Background/Objective: Nasopharyngeal carcinoma is one of the most common malignant tumors in Southern China. Radiotherapy and chemotherapy are currently main treatments, whereas the long-term efficacy is still unsatisfactory. Curcumin, an active ingredient of Turmeric, shows promising as a naturally ocurring anti-cancer drug. Growing evidences have shown that visible light can activate curcumin and enhance the cytotoxicity of curcumin. These demonstrated that curcumin was a herbal photosensitizer. Recently, sonodynamic therapy has become a new modality for treating tumor because of its advantages in tissue penetration in comparison with photodynamic therapy. However, whether curcumin is a sensitizer to enhance ultrasound therapy on tumor remains to be clarified. This study has investigated the cell death of nasopharyngeal carcinoma cell line CNE2 cells induced by low-intensity ultrasound in the presence of curcumin. Our aims are to explore whether curcumin can enhance ultrasound therapy on tumor and provide the evidence for the combined treatment of curcumin and low-intensity ultrasound on tumor. Materials/Methods: Cytotoxicity was detected using MTT assay after the treatment with curcumin and low-intensity ultrasound together. Cell morphology was observed by inverted light microscopy. Furthermore, the temperature changes of solution were recorded as well. Apoptosis was evaluated using flow cytometry with AnnexinⅤ-FITC Apoptosis Detection Kit and nuclear staining with Hoechst 33258. Ultrastructure was observed by transmission electron microscopy (TEM). Mitochondrial membrane potential was evaluated using Rhodamine123 assay. Finally, intracellular reaction oxygen species were determined by DCFH-DA staining.Results: The cytotoxicity was the most significant when the cells were subject to ultrasound exposure for 8 s at an intensity of 0.46 W/cm2 after being sensitized for 1h with the curcumin concentration at 10μM. And the above finding was also confirmed by inverted light microscopy. No significant temperature changes were revealed. Flow cytometry showed that the combination of curcumin and ultrasound significantly increased the necrotic or late apoptotic rate up to 31.37% compared to the controls. Nuclear condensation was observed in the nuclear staining and collapse of mitochondrial membrane potential and ROS increase were found in the CNE2 cells after the treatment of curcumin and ultrasound together. Transmission electron microscopy showed the ultrastructure of the CNE2 cells after the treatment of curcumin and ultrasound together, that apoptotic bodies, necrosis and mitochondrial swelling was displayed. Conclusion: Curcumin significantly enhanced the cytotoxicity of ultrasound treatment in the CNE2 cells. Our finding also highlighted that apoptosis induction, ROS increase and mitochondrial damage might play important role in the cell death induced by the treatment of curcumin and low-intenstiy ultrasound together and indicated that the combined treatment of curcumin and ultrasound might be a potential therapeutic treatment of malignant tumor.