title: Role of P2X7 receptor in Clostridium perfringens beta-toxin-mediated cellular injury.
abstract: BACKGROUND
  Clostridium perfringens beta-toxin is a pore-forming toxin (PFT) and an important agent of necrotic enteritis and enterotoxemia. We recently reported that beta-toxin strongly induced cell death in THP-1 cells via the formation of oligomers. We here describe that the P2X(7) receptor, which is an ATP receptor, interacts with beta-toxin.
  
  
  METHODS
  We tested the role of P2X(7) receptor in beta-toxin-induced toxicity using specific inhibitors, knockdown of receptor, expression of the receptor and interaction by dot-blot assay. The potency of P2X(7) receptor was further determined using an in vivo mouse model.
  
  
  RESULTS
  Selective P2X(7) receptor antagonists (oxidized ATP (o-ATP), oxidized ADP, and Brilliant Blue G (BBG)) inhibited beta-toxin-induced cytotoxicity in THP-1 cells. o-ATP also blocked the binding of beta-toxin to cells. The P2X(7) receptor and beta-toxin oligomer were localized in the lipid rafts of THP-1 cells. siRNA for the P2X(7) receptor inhibited toxin-induced cytotoxicity and binding of the toxin. In contrast, the siRNA knockdown of P2Y(2) or P2Y(6) had no effect on beta-toxin-induced cytotoxicity. The addition of beta-toxin to P2X(7)-transfected HEK-293 cells resulted in binding of beta-toxin oligomer. Moreover, beta-toxin specifically bound to immobilized P2X(7) receptors in vitro and colocalized with the P2X(7) receptor on the THP-1 cell surface. Furthermore, beta-toxin-induced lethality in mice was blocked by the preadministration of BBG.
  
  
  CONCLUSIONS
  The results of this study indicate that the P2X(7) receptor plays a role in beta-toxin-mediated cellular injury.
  
  
  CONCLUSIONS
  P2X(7) receptor is a potential target for the treatment of C. perfringens type C infection.
authors: Department of Microbiology, Faculty of Pharmaceutical Sciences, Tokushima Bunri University, Yamashiro-cho, Tokushima 770-8514, Japan. Electronic address: nagahama@ph.bunri-u.ac.jp.
authors: Masahiro
authors: M
authors: Nagahama
authors: Department of Microbiology, Faculty of Pharmaceutical Sciences, Tokushima Bunri University, Yamashiro-cho, Tokushima 770-8514, Japan.
authors: Soshi
authors: S
authors: Seike
authors: Department of Microbiology, Faculty of Pharmaceutical Sciences, Tokushima Bunri University, Yamashiro-cho, Tokushima 770-8514, Japan.
authors: Hidenori
authors: H
authors: Shirai
authors: Department of Microbiology, Faculty of Pharmaceutical Sciences, Tokushima Bunri University, Yamashiro-cho, Tokushima 770-8514, Japan.
authors: Teruhisa
authors: T
authors: Takagishi
authors: Department of Microbiology, Faculty of Pharmaceutical Sciences, Tokushima Bunri University, Yamashiro-cho, Tokushima 770-8514, Japan.
authors: Keiko
authors: K
authors: Kobayashi
authors: Department of Microbiology, Faculty of Pharmaceutical Sciences, Tokushima Bunri University, Yamashiro-cho, Tokushima 770-8514, Japan.
authors: Masaya
authors: M
authors: Takehara
authors: Department of Microbiology, Faculty of Pharmaceutical Sciences, Tokushima Bunri University, Yamashiro-cho, Tokushima 770-8514, Japan.
authors: Jun
authors: J
authors: Sakurai
mesh_terms: D051722:ADAM Proteins / Q000502:physiology; D000072197:ADAM10 Protein; D053829:Amyloid Precursor Protein Secretases / Q000502:physiology; D000818:Animals; D001427:Bacterial Toxins / Q000633:toxicity*; D002118:Calcium / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D008565:Membrane Proteins / Q000502:physiology; D051379:Mice; D008813:Mice, Inbred ICR; D034741:RNA, Small Interfering / Q000494:pharmacology; D058486:Receptors, Purinergic P2X7 / Q000502:physiology*; D012394:Rosaniline Dyes / Q000494:pharmacology
chemical_list: D001427:Bacterial Toxins; C031884:CPB protein, Clostridium perfringens; D008565:Membrane Proteins; D034741:RNA, Small Interfering; D058486:Receptors, Purinergic P2X7; D012394:Rosaniline Dyes; D053829:Amyloid Precursor Protein Secretases; D051722:ADAM Proteins; D000072197:ADAM10 Protein; C491236:ADAM10 protein, human; C004692:coomassie Brilliant Blue; D002118:Calcium
keywords: C. perfringens beta-toxin; Oligomer formation; P2X(7) receptor; Pore-forming toxin