title: Effect of the silencing of the Ehcp112 gene on the in vitro virulence of Entamoeba histolytica.
abstract: BACKGROUND
  Entamoeba histolytica is an intestinal protozoan parasite that causes amoebiasis in humans, affecting up to 50 million people worldwide each year and causing 40,000 to 100,000 deaths annually. EhCP112 is a cysteine proteinase of E. histolytica able to disrupt cell monolayers and digest extracellular matrix proteins, it is secreted by trophozoites and it can be active in a wide range of temperature and pH. These characteristics have encouraged the use of EhCP112 in the design and production of possible vaccines against amoebiasis, obtaining promising results. Nevertheless, we have no conclusive information about the role of EhCP112 in the E. histolytica pathogenesis.
  
  
  METHODS
  A set of three specific siRNA sequences were used to silence the Ehcp112 gene via the soaking system. Silencing was evaluated by Western blot using an antibody against the EhCP112 recombinant protein. Finally, we analyzed the protease activity, the phagocytosis rate and the ability to destroy MDCK cells of the EhCP112-silenced trophozoites.
  
  
  RESULTS
  The highest silencing effect on EhCP112 was detected at 16 h of treatment; time enough to perform the in vitro virulence assays, which showed that EhCP112 silencing produces a significant reduction in cytolysis and phagocytosis of target cells, indicating the participation of this proteinase in these events.
  
  
  CONCLUSIONS
  EhCP112 is involved in the in vitro virulence of E. histolytica.
authors: Departamento de Infectómica y Patogénesis Molecular, CINVESTAV-IPN, A,P, 14-740, México, D,F,, México. marodri@cinvestav.mx.
authors: Ramón
authors: R
authors: Ocádiz-Ruiz
authors: Wendy
authors: W
authors: Fonseca
authors: Máximo B
authors: MB
authors: Martínez
authors: Ramón
authors: R
authors: Ocádiz-Quintanar
authors: Esther
authors: E
authors: Orozco
authors: Mario A
authors: MA
authors: Rodríguez
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D015153:Blotting, Western; D016923:Cell Death; D002460:Cell Line; D003546:Cysteine Endopeptidases / Q000235:genetics / Q000378:metabolism*; D004285:Dogs; D004748:Entamoeba histolytica / Q000235:genetics / Q000472:pathogenicity*; D004847:Epithelial Cells / Q000469:parasitology; D020868:Gene Silencing; D010587:Phagocytosis; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D037521:Virulence Factors / Q000235:genetics / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D034741:RNA, Small Interfering; D037521:Virulence Factors; D003546:Cysteine Endopeptidases; C122004:EhCP112 protein, Entamoeba histolytica
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