title: Vibrio vulnificus cytolysin induces superoxide anion-initiated apoptotic signaling pathway in human ECV304 cells.
abstract: Previous studies showed that exposure to Vibrio vulnificus cytolysin (VVC) caused characteristic morphologic changes and dysfunction of vascular structures in lung. VVC showed cytotoxicity for mammalian cells in culture and acted as a vascular permeability factor. In this study, the underlying mechanisms of VVC-induced cytotoxicity was investigated on ECV304 cell, a human vascular endothelial cell line. When cells were exposed to 0.4 hemolytic units (HU) of VVC, consecutive apoptotic events were observed; the elevation of superoxide anion (O (-.)(2)), the release of cytochrome c, the activation of caspase-3, the cleavage of poly(ADP-ribose) polymerase, and the DNA fragmentation. The pretreatment with 4-hydroxy-2,2,6,6-tetramethylpiperidine-N-oxyl (TEMPO), O(-.) 2) scavenger, completely abolished O(-.)(2) levels and downstream apoptotic events. Moreover, pretreatment with cyclosporin A (CsA), a mitochondrial permeability transition inhibitor, was capable of attenuating O(-.)(2)-mediated cytochrome c release and caspase-3 activation, and consequent apoptosis. Apoptosis, as demonstrated by oligonucleosomal DNA fragmentation and fluorescence microscopy, was induced 24 h after VVC treatment, which was also prevented by caspase-3 inhibitor, Ac-DEVD-CHO. Caspase-1 inhibitor, Ac-YVAD-CHO, did not protect ECV 304 cells from apoptosis. These results suggest a scenario where VVC-induced apoptosis is triggered by the generation of O(-.)(2), release of cytochrome c from mitochondria, activation of caspase-3, degradation of poly(ADP-ribose) polymerase, and DNA fragmentation. The induction of apoptosis in endothelial cells by VVC may provide a pivotal mechanism for understanding the pathophysiology of septicemia.
authors.affiliation: Department of Physiology, School of Oriental Medicine, Won-Kwang University, Iksan 570-749, Republic of Korea.
authors.forename: K B
authors.initials: KB
authors.lastname: Kwon
authors.forename: J Y
authors.initials: JY
authors.lastname: Yang
authors.forename: D G
authors.initials: DG
authors.lastname: Ryu
authors.forename: H W
authors.initials: HW
authors.lastname: Rho
authors.forename: J S
authors.initials: JS
authors.lastname: Kim
authors.forename: J W
authors.initials: JW
authors.lastname: Park
authors.forename: H R
authors.initials: HR
authors.lastname: Kim
authors.forename: B H
authors.initials: BH
authors.lastname: Park
mesh_terms: D000838:Anions / Q000378:metabolism*; D000975:Antioxidants / Q000494:pharmacology; D017209:Apoptosis; D015153:Blotting, Western; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D002460:Cell Line; D002470:Cell Survival; D003497:Cyclic N-Oxides / Q000494:pharmacology; D016572:Cyclosporine / Q000494:pharmacology; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D003574:Cytochrome c Group / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D003603:Cytotoxins / Q000737:chemistry* / Q000378:metabolism*; D053938:DNA Fragmentation; D004305:Dose-Response Relationship, Drug; D004727:Endothelium / Q000378:metabolism; D004730:Endothelium, Vascular / Q000166:cytology; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D008856:Microscopy, Fluorescence; D008928:Mitochondria / Q000378:metabolism; D009842:Oligopeptides / Q000494:pharmacology; D010100:Oxygen / Q000378:metabolism; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D015398:Signal Transduction; D013481:Superoxides / Q000378:metabolism*; D013997:Time Factors; D014733:Vibrio / Q000378:metabolism*
chemical_list: D000838:Anions; D000975:Antioxidants; D003497:Cyclic N-Oxides; D015853:Cysteine Proteinase Inhibitors; D003574:Cytochrome c Group; D003603:Cytotoxins; D004791:Enzyme Inhibitors; D009842:Oligopeptides; C099158:acetyl-aspartyl-glutamyl-valyl-aspartal; D013481:Superoxides; C096429:L 709049; D016572:Cyclosporine; D011065:Poly(ADP-ribose) Polymerases; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; D010100:Oxygen; C003959:TEMPO