title: B4G2 induces mitochondrial apoptosis by the ROS-mediated opening of Ca(2+)-dependent permeability transition pores.
abstract: OBJECTIVE
  Hepatocellular carcinoma (HCC) is the most common type of liver cancer. At present, only sorafenib is approved to treat HCC. In this study, we found that a 23-hydroxybetulinic acid derivative, B4G2, exhibited potent antiproliferative activity in HCC cell lines.
  
  
  METHODS
  We used four HCC cell lines (HepG2, HepG2/ADM, Hep3B and Bel-7402) to evaluate the anti-tumour activity and explore underlying mechanisms by which B4G2 induces apoptosis.
  
  
  RESULTS
  Among these cell lines, HepG2 showed the highest sensitivity to B4G2. HepG2 cells treated with B4G2 showed a depolarized mitochondrial membrane potential, released cytochrome c, activated caspase-9 and caspase-3 and cleaved poly ADP-ribose polymerase (PARP). However, Z-VAD-FMK, a pan-caspase inhibitor, did not attenuate B4G2-induced apoptosis, implying that the induction of mitochondrial apoptosis by B4G2 may be independent of caspases. Moreover, pre-treatment with MgCl2, a blocker of Ca2+-dependent permeability transition (PT) pores, attenuated the depolarization of the mitochondrial potential and decreased the population of apoptotic cells, indicating that B4G2-induced apoptosis was partly dependent on the opening of the Ca2+-dependent PT pores. B4G2 also increased the levels of intracellular calcium and reactive oxygen species (ROS). Furthermore, an ROS scavenger, N-acetyl-cysteine (NAC), markedly decreased the accumulation of intracellular calcium and apoptosis.
  
  
  CONCLUSIONS
  This is the first demonstration that B4G2 inhibits the growth of HCC cells and induces mitochondrial apoptosis in hepatocellular carcinoma cells by the ROS-mediated opening of Ca2+-dependent permeability transition pores.
authors.affiliation: Institute of Traditional Chinese Medicine &amp; Natural Products, College of Pharmacy, Jinan University, Guangzhou, China.
authors.forename: Nan
authors.initials: N
authors.lastname: Yao
authors.forename: Ying-Jie
authors.initials: YJ
authors.lastname: Li
authors.forename: Dong-Mei
authors.initials: DM
authors.lastname: Zhang
authors.forename: Dao-Lu
authors.initials: DL
authors.lastname: Liu
authors.forename: Ming-Kuen
authors.initials: MK
authors.lastname: Tang
authors.forename: Anita
authors.initials: A
authors.lastname: Yiu
authors.forename: Yong
authors.initials: Y
authors.lastname: Li
authors.forename: Wei-Min
authors.initials: WM
authors.lastname: Chen
authors.forename: Ping
authors.initials: P
authors.lastname: Lan
authors.forename: Zhe
authors.initials: Z
authors.lastname: Yao
authors.forename: Zhe-Sheng
authors.initials: ZS
authors.lastname: Chen
authors.forename: Wen-Cai
authors.initials: WC
authors.lastname: Ye
mesh_terms: D017209:Apoptosis / Q000187:drug effects; D002118:Calcium / Q000378:metabolism*; D006528:Carcinoma, Hepatocellular / Q000378:metabolism*; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D056945:Hep G2 Cells; D006801:Humans; D008113:Liver Neoplasms / Q000378:metabolism*; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D033681:Mitochondrial Membrane Transport Proteins / Q000378:metabolism*; D000083162:Mitochondrial Permeability Transition Pore; D017382:Reactive Oxygen Species / Q000378:metabolism; D014315:Triterpenes / Q000494:pharmacology*
chemical_list: C468651:23-hydroxybetulinic acid; D033681:Mitochondrial Membrane Transport Proteins; D000083162:Mitochondrial Permeability Transition Pore; D017382:Reactive Oxygen Species; D014315:Triterpenes; D002118:Calcium