Eucalrobusone C suppresses cell proliferation and induces ROS-dependent mitochondrial apoptosis via the p38 MAPK pathway in hepatocellular carcinoma cells

ABSTRACT Background Eucalyptus extracts have anti-cancer activity against various cancer cells . Formyl-phloroglucinol meroterpenoids (FPMs), which are typical secondary metabolites of the genera Eucalyptus, have many important pharmacological activities . Purpose Eucalrobusone C (EC), a new bioactive phytochemical , was first isolated from the leaves of Eucalyptus robusta in our laboratory. EC is a FPM, and our previous research revealed that EC showed strongest cytotoxicity in three cancer models than other compounds isolated from the leaves of E. robusta . This study investigated its anti-tumor effects on human hepatocellular carcinoma (HCC) and its underlying mechanisms. Methods Cell viability was measured by MTT assay . Cell cycle, apoptosis and mitochondrial transmembrane potential were determined by flow cytometry. Immunofluorescence was determined by a laser scanning confocal microscope . Protein levels were analyzed by Western blotting . Results Our results showed that EC exerted strong anti-proliferative activity against HCC cells in a concentration- and time-dependent manner. EC markedly induced apoptosis through the caspase-dependent mitochondrial pathway, and the cell cycle was arrested at S phase. SB203580 , a p38 MAPK inhibitor, effectively decreased cell death caused by EC. Moreover, the ROS scavenger N-acetyl cysteine (NAC) significantly attenuated apoptosis induced by EC and reversed EC-induced p38 MAPK activation. Conclusion Our findings indicate that EC induces mitochondrial-dependent apoptosis in HCC cells through ROS generation and p38 MAPK activation, making EC a promising candidate for further development as an anticancer agent for HCC cells.