Chimaphilin Induces Apoptosis In Human Breast Cancer MCF-7 Cells Through A ROS-mediated Mitochondrial Pathway

Chimaphilin,2,7-dimethyl-1,4-naphthoquinone, is extracted from pyrola [Passiflora incarnata Fisch.]. In this study, the anticancer activity and underlying mechanisms of chimaphilin toward human breast cancer MCF-7 cells are investigated. The results are as follows:1. Chimaphilin could inhibit the viability of MCF-7 cells in a concentration-dependent not time-dependent manner. Inhibition of cell proliferation by chimaphilin was measured by the MTT assay, and the IC50 values were 43.30±2.47 μM for 24 h,43.26±2.06μM for 48 h and 44.10±1.84 μM for 24 h.2. Chimaphilin markedly induced apoptosis of MCF-7 cells. The acridine orange stain assay, DNA Ladder, Annexin V-FITC/propidium iodide (PI) double stain assay were applied to measure apoptosis induced by chimaphilin. The results showed that Crescent-shaped nuclei, generations of DNA fragmentation and phosphatidylserine translocated from the inner to the outer leaflet of the plasma membrane in MCF-7 cells were obsearved.3. Chimaphilin induced apoptosis of MCF-7 cells through a Mitochondrial/Caspases pathway. The rhodamin 123 stain assay and western blotting assay were used in MCF-7 cells treated with chimaphilin. The results revealed mitochondrial membrane potential in chimaphilin-treated MCF-7 cells was disrupted. Besides, western blotting assay showed that chimaphilin suppressed Bcl-2 level and enhanced Bad level, then activated caspase-9 and caspase-3, and further activated the poly ADP-ribose polymerase (PARP), finally induced cell apoptosis involving the mitochondrial pathway.4. Generation of ROS was one cause of apoptosis induced by chimaphilin. It were measured by the DCFH-DA stain assay and western blotting assay. Chimaphilin could trigger a significant generation of ROS. Furthermore, free radical scavengers N-acetyl-L-cysteine (NAC) pretreatment removed chimaphilin-induced apoptosis of MCF-7 cells.In general, the present results demonstrated that chimaphilin induced apoptosis in human breast cancer MCF-7 cells via a ROS-mediated mitochondrial pathway. And generation of ROS was one cause of apoptosis induced by chimaphilin in MCF-7 cells. The mechanisms of chimaphilin toward human breast cancer MCF-7 cells through a ROS/Mitochondrial pathway are firstly investigated. It laid a foundation of the underlying anticancer mechanisms of chimaphilin towards MCF-7 cells and development of chimaphilin as new anticancer drugs.