AIM: To investigate the effects of the essential oil of Curcuma wenyujin (CWO) on growth inhibition and on the induction of apoptosis in human HepG2 cancer cells. METHODS: The cytotoxic effect of drugs on HepG2 cells ...AIM: To investigate the effects of the essential oil of Curcuma wenyujin (CWO) on growth inhibition and on the induction of apoptosis in human HepG2 cancer cells. METHODS: The cytotoxic effect of drugs on HepG2 cells was measured by 3-(4,5-dimethylthiazol-2- yl)-2,5-diphenyltetra-zolium bromide (MTT) assay. DNA fragmentation was visualized by agarose gel electrophoresis. Cell cycle and mitochondrial transmembrane potential (△Ψm) were determined by flow cytometry (FCM). Cytochrome C immunostaining was evaluated by fluorescence microscopy. Caspase-3 enzymatic activity was assayed by the cleavage of Ac-DEVD-R110. Cleaved PARP and active caspase-3 protein levels were measured by FCM using BD? CBA Human Apoptosis Kit. RESULTS: Treatment with CWO inhibited the growth of HepG2 cells in a dose-dependent manner, and the IC50 of CWO was approximately 70 μg/mL. CWO was found to inhibit the growth of HepG2 cells by inducing a cell cycle arrest at S/G2. DNA fragmentation was evidentlyobserved at 70 μg/mL after 72 h of treatment. During the process, cytosolic HepG2 cytochrome C staining showed a markedly stronger green fluorescence than in control cells in a dose-dependent fashion, and CWO also caused mitochondrial transmembrane depolarization. Furthermore, the results clearly demonstrated that both, activity of caspase-3 enzyme and protein levels of cleaved PARP, significantly increased in a dose- dependent manner after treatment with CWO. CONCLUSION: CWO exhibits an antiproliferative effect in HepG2 cells by inducing apoptosis. This growth inhibition is associated with cell cycle arrest, cytochrome C translocation, caspase 3 activation, Poly- ADP-ribose polymerase (PARP) degradation, and loss of mitochondrial membrane potential. This process involves a mitochondria-caspase dependent apoptosis pathway. As apoptosis is an important anti-cancer therapeutic target, these results suggest a potential of CWO as a chemotherapeutic agent.展开更多
基金Grants from the Research Committee, Universityof Macao, Macao SAR, No RG054/05-06S and RG058/05-06Sgrants from the Science and Technology Development Fund, Macao SAR, No 012/2006/A and 045/2007/A3
文摘AIM: To investigate the effects of the essential oil of Curcuma wenyujin (CWO) on growth inhibition and on the induction of apoptosis in human HepG2 cancer cells. METHODS: The cytotoxic effect of drugs on HepG2 cells was measured by 3-(4,5-dimethylthiazol-2- yl)-2,5-diphenyltetra-zolium bromide (MTT) assay. DNA fragmentation was visualized by agarose gel electrophoresis. Cell cycle and mitochondrial transmembrane potential (△Ψm) were determined by flow cytometry (FCM). Cytochrome C immunostaining was evaluated by fluorescence microscopy. Caspase-3 enzymatic activity was assayed by the cleavage of Ac-DEVD-R110. Cleaved PARP and active caspase-3 protein levels were measured by FCM using BD? CBA Human Apoptosis Kit. RESULTS: Treatment with CWO inhibited the growth of HepG2 cells in a dose-dependent manner, and the IC50 of CWO was approximately 70 μg/mL. CWO was found to inhibit the growth of HepG2 cells by inducing a cell cycle arrest at S/G2. DNA fragmentation was evidentlyobserved at 70 μg/mL after 72 h of treatment. During the process, cytosolic HepG2 cytochrome C staining showed a markedly stronger green fluorescence than in control cells in a dose-dependent fashion, and CWO also caused mitochondrial transmembrane depolarization. Furthermore, the results clearly demonstrated that both, activity of caspase-3 enzyme and protein levels of cleaved PARP, significantly increased in a dose- dependent manner after treatment with CWO. CONCLUSION: CWO exhibits an antiproliferative effect in HepG2 cells by inducing apoptosis. This growth inhibition is associated with cell cycle arrest, cytochrome C translocation, caspase 3 activation, Poly- ADP-ribose polymerase (PARP) degradation, and loss of mitochondrial membrane potential. This process involves a mitochondria-caspase dependent apoptosis pathway. As apoptosis is an important anti-cancer therapeutic target, these results suggest a potential of CWO as a chemotherapeutic agent.
