It has been reported that the release of cyto c appears to

be dependent on the induction of mitochondrial permeability transition, which is associated with a decrease in Δφm; therefore, the loss of Δφm and the release of apoptogenic factors, such as cyto c, from the mitochondria into the cytosol are associated with apoptosis induced by chemotherapeutic drugs[25–27]. In the present study, loss of Δφm and release of Cyto c were observed in NCTD-treated cells, resulting in caspase-9 and caspase-3 find more activation and PARP cleavage and, finally, apoptosis. Moreover, the loss of Δφm may, in fact, be a consequence of massive cytochrome c release from the mitochondria. Thus, a mitochondrial damage-dependent pathway may be involved in NCTD-induced apoptosis in HepG2 cells. Some studies have MLN4924 mouse reported that ROS act as secondary messengers in apoptosis induced by anti-cancer and chemopreventive agents[28, 29]. The generation of ROS can cause the loss of Δφm, and induce apoptosis by releasing pro-apoptotic proteins such as AIF and Cyto c from mitochondria to the cytosol.The generation of ROS may contribute to mitochondrial

damage and lead to cell death by acting as an apoptotic signaling molecule[30, 31]. To reveal if NCTD influenced the level of ROS, we stained drug treated cells with DCFH-DA. We found that, in addition to its effect on Δφm, NCTD caused an increase in ROS production in HepG2 cells. The NCTD -induced increase in ROS and antiproliferation in HepG2 cells are apparently dependent on ROS generation, because the NCTD -induced increase in ROS can be abolished or Selleck MAPK inhibitor attenuated by antioxidants, such as NAC. In addition, we found that NCTD -induced antiproliferation in HepG2 cells was also abolished by the antioxidant NAC. Conclusions In conclusion, our data indicate that NCTD induced apoptosis in HepG2 cells via ROS generation and mitochondrial pathway (Figure 7)[32]. These findings suggest that NCTD

may one day be used in the prevention and treatment of cancer. Figure 7 A proposed model showing the mechanism of NCTD anti-proliferative and apoptosis effects in HepG2 cells. ROS, reactive oxygen species; PARP, poly (ADP ribose)polymerase; Δφm, mitochondrial membrane potential; selleck Apaf-1, apoptotic protease activating factor-1. Acknowledgements We thank Yan Wan, Department of Immunology, Wuhan University, for exceptional technical assistance in flow cytometry analysis. References 1. El-Serag HB, Rudolph KL: Hepatocellular carcinoma:epidemiology and molecular carcinogenesis. Gastroenterology 2007, 132: 2557–2576.PubMedCrossRef 2. Wang GS: Medical uses of mylabris in ancient China and recent studies. J Ethnopharmacol 1989, 26: 147–162.PubMedCrossRef 3. Peng F, Wei YQ, Tian L, Yang L, Zhao X, Lu Y: Induction of apoptosis by norcantharidin in human colorectal carcinoma cell lines: involvement of the CD95 receptor/ligand. J Cancer Res Clin Oncol 2002, 128: 223–230.PubMedCrossRef 4.