title: Molecular mechanisms of apoptosis induced by Scorpio water extract in human hepatoma HepG2 cells.
abstract: OBJECTIVE
  To clarify the mechanism underlying the anti-mutagenic and anti-cancer activities of Scorpio water extract (SWE).
  
  
  METHODS
  Human hepatoma HepG2 cells were incubated with various concentrations of SWE. After 24-h incubation, cytotoxicity and apoptosis evaluations were determined by MTT and DNA fragmentation assay, respectively. After treatment with SWE, mitochondrial membrane potential (MMP) was determined by measuring the retention of the dye 3,3'-dihexyloxacarbocyanine (DiOC(6)(3)) and the protein expression including cytochrome C and poly-(ADP-ribose) polymerase (PARP) were measured by Western blotting. Caspase-3 and -9 enzyme activities were measured using specific fluorescence dyes such as Ac-DEVD-AFC and Ac-LEHD-AFC.
  
  
  RESULTS
  We found that treatment with SWE induced apoptosis as confirmed by discontinuous DNA fragmentation in cultured human hepatoma HepG2 cells. Our investigation also showed that SWE-induced apoptosis of HepG2 cells were associated with intracellular events including disruption of MMP, increased translocation of cytochrome C from mitochondria to cytosol, activation of caspase-3, and PARP. Pre-treatment of N-acetyl-Asp-Glu-Val-Asp-CHO (Ac-DEVD-CHO), a caspase-3 specific inhibitor, or cyclosporin A (CsA), an inhibitor of MMP disruption, completely abolished SWE-induced DNA fragmentation.
  
  
  CONCLUSIONS
  These results suggest that SWE possibly causes mitochondrial damage, leading to cytochrome C release into cytosol and activation of caspases resulting in PARP cleavage and execution of apoptotic cell death in HepG2 cells. These results further suggest that Scorpio may be a valuable agent of therapeutic intervention of human hepatomas.
authors.affiliation: Department of Physiology, School of Oriental Medicine, Wonkwang University, Iksan, 570-749 Republic of Korea.
authors.forename: Kang-Beom
authors.initials: KB
authors.lastname: Kwon
authors.forename: Eun-Kyung
authors.initials: EK
authors.lastname: Kim
authors.forename: Jung-Gook
authors.initials: JG
authors.lastname: Lim
authors.forename: Eun-Sil
authors.initials: ES
authors.lastname: Jeong
authors.forename: Byung-Cheul
authors.initials: BC
authors.lastname: Shin
authors.forename: Young-Se
authors.initials: YS
authors.lastname: Jeon
authors.forename: Kang-San
authors.initials: KS
authors.lastname: Kim
authors.forename: Eun-A
authors.initials: EA
authors.lastname: Seo
authors.forename: Do-Gon
authors.initials: DG
authors.lastname: Ryu
mesh_terms: D017209:Apoptosis / Q000187:drug effects*; D006528:Carcinoma, Hepatocellular; D053148:Caspase 3; D053453:Caspase 9; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism; D045744:Cell Line, Tumor; D016572:Cyclosporine / Q000494:pharmacology; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D045304:Cytochromes c / Q000378:metabolism; D004365:Drugs, Chinese Herbal / Q000494:pharmacology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D008113:Liver Neoplasms; D008564:Membrane Potentials / Q000187:drug effects; D008928:Mitochondria / Q000187:drug effects / Q000502:physiology; D009842:Oligopeptides / Q000494:pharmacology; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism
chemical_list: D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; D004365:Drugs, Chinese Herbal; D004791:Enzyme Inhibitors; D009842:Oligopeptides; C099158:acetyl-aspartyl-glutamyl-valyl-aspartal; D016572:Cyclosporine; D045304:Cytochromes c; D011065:Poly(ADP-ribose) Polymerases; C505407:CASP3 protein, human; C506433:CASP9 protein, human; D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases
references.citation: J Exp Med. 1996 Sep 1;184(3):1155-60
references.pmid: 9064332
references.citation: Annu Rev Physiol. 1998;60:533-73
references.pmid: 9558476
references.citation: Mol Cell. 2002 Mar;9(3):459-70
references.pmid: 11931755
references.citation: Cancer Lett. 2000 Jan 1;148(1):19-25
references.pmid: 10680588
references.citation: Science. 1998 Aug 28;281(5381):1309-12
references.pmid: 9721092
references.citation: J Biol Chem. 1998 Mar 27;273(13):7770-5
references.pmid: 9516487
references.citation: Cell. 1997 Feb 7;88(3):355-65
references.pmid: 9039262
references.citation: Cell. 1997 Nov 14;91(4):479-89
references.pmid: 9390557
references.citation: Proc Natl Acad Sci U S A. 1996 Aug 6;93(16):8395-400
references.pmid: 8710882
references.citation: Proc Natl Acad Sci U S A. 1994 Nov 22;91(24):11752-6
references.pmid: 7972136
references.citation: Nat Med. 2000 May;6(5):513-9
references.pmid: 10802706
references.citation: Cell. 1996 Oct 18;87(2):171
references.pmid: 8861900
references.citation: Cancer Res. 1999 Apr 1;59(7 Suppl):1701s-1706s
references.pmid: 10197583
references.citation: J Immunol Methods. 2002 Jul 1;265(1-2):97-110
references.pmid: 12072181
references.citation: Life Sci. 2001 Aug 17;69(13):1485-96
references.pmid: 11554610
references.citation: Leukemia. 1998 Sep;12 Suppl 1:S41-6
references.pmid: 9777895
references.citation: Blut. 1990 Feb;60(2):97-102
references.pmid: 2302467
references.citation: Cancer Res. 2002 Sep 15;62(18):5204-9
references.pmid: 12234985
references.citation: Cell. 1997 Aug 8;90(3):405-13
references.pmid: 9267021
references.citation: Biochim Biophys Acta. 1998 Aug 10;1366(1-2):151-65
references.pmid: 9714783
references.citation: J Biol Chem. 1996 Aug 30;271(35):21629-36
references.pmid: 8702951
references.citation: Anal Biochem. 1976 May 7;72:248-54
references.pmid: 942051
references.citation: J Biol Chem. 2001 Dec 14;276(50):47518-23
references.pmid: 11591724
references.citation: Cell. 1997 Feb 7;88(3):347-54
references.pmid: 9039261
references.citation: EMBO J. 1999 Apr 15;18(8):2040-8
references.pmid: 10205158
references.citation: Proc Natl Acad Sci U S A. 1996 Dec 10;93(25):14559-63
references.pmid: 8962091