Identification of a Direct Hemolytic Effect Dependent on the Catalytic Activity Induced by Phospholipase-D (Dermonecrotic Toxin) From Brown Spider Venom

Identification of a Direct Hemolytic Effect Dependent on the Catalytic Activity Induced by Phospholipase-D (Dermonecrotic Toxin) From Brown Spider Venom

Author Chaves-Moreira, Daniele Google Scholar
Chaim, Olga M. Autor UNIFESP Google Scholar
Sade, Youssef B. Google Scholar
Paludo, Katia S. Autor UNIFESP Google Scholar
Gremski, Luiza H. Autor UNIFESP Google Scholar
Donatti, Lucelia Google Scholar
Moura, Juliana de Google Scholar
Mangili, Oldemir C. Google Scholar
Gremski, WaIdemiro Google Scholar
Silveira, Rafael B. da Google Scholar
Senff-Ribeiro, Andrea Google Scholar
Veiga, Silvio S. Google Scholar
Institution Univ Fed Parana
Universidade Federal de São Paulo (UNIFESP)
Catholic Univ Parana
Univ Estadual Ponta Grossa
Abstract Brown spiders have world-wide distribution and are the cause of health problems known as loxoscelism. Necrotic cutaneous lesions surrounding the bites and less intense systemic signs like renal failure, DIC, and hemolysis were observed. We studied molecular mechanism by which recombinant toxin, biochemically characterized as phospholipase-D, causes direct hemolysis (complement independent). Human erythrocytes treated with toxin showed direct hemolysis in a dose-dependent and time-dependent manner, as well as morphological changes in cell size and shape. Erythrocytes from human, rabbit, and sheep were more susceptible than those from horse. Hemolysis was not dependent on ABO group or Rhesus system. Confocal and FACS analyses using antibodies or GFP-phospholipase-D protein showed direct toxin binding to erythrocytes membrane. Moreover, toxin-treated erythrocytes reacted with annexin-V and showed alterations in their lipid raft profile. Divalent ion chelators significantly inhibited hemolysis evoked by phospholipase-D, which has magnesium at the catalytic domain. Chelators were more effective than PMSF (serine-protease inhibitor) that had no effect on hemolysis. By site-directed mutation at catalytic domain (histidine 12 by alanine), hemolysis and morphologic changes of erythrocytes (but not the toxin's ability of membrane binding) were inhibited, supporting that catalytic activity is involved in hemolysis and cellular alterations but not toxin cell binding. the results provide evidence that L. intermedia venom phospholipase-D triggers direct human blood cell hemolysis in a catalytic-dependent manner. J. Cell. Biochem. 107: 655-666, 2009. (C) 2009 Wiley-Liss, Inc.
Keywords BROWN SPIDER
VENOM
PHOSPHOLIPASE-D
CATALYSIS
HEMOLYSIS
Language English
Sponsor Secretaria de Estado de Ciencia, Tecnologia e Ensino Superior (SETI) do Parana
Fundacao Araucaria-PR
Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)
Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)
Date 2009-07-01
Published in Journal of Cellular Biochemistry. Hoboken: Wiley-liss, v. 107, n. 4, p. 655-666, 2009.
ISSN 0730-2312 (Sherpa/Romeo, impact factor)
Publisher Wiley-Blackwell
Extent 655-666
Origin http://dx.doi.org/10.1002/jcb.22148
Access rights Closed access
Type Article
Web of Science ID WOS:000267813600011
URI http://repositorio.unifesp.br/handle/11600/31627

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