title: Inhibition of connexin 43 hemichannel-mediated ATP release attenuates early inflammation during the foreign body response.
abstract: BACKGROUND
  In the last 50 years, the use of medical implants has increased dramatically. Failure of implanted devices and biomaterials is a significant source of morbidity and increasing healthcare expenditures. An important cause of implant failure is the host inflammatory response. Recent evidence implicates extracellular ATP as an important inflammatory signaling molecule. A major pathway for release of cytoplasmic ATP into the extracellular space is through connexin hemichannels, which are the unpaired constituents of gap junction intercellular channels. Blockade of hemichannels of the connexin 43 (Cx43) isoform has been shown to reduce inflammation and improve healing. We have developed a Cx43 mimetic peptide (JM2) that targets the microtubule-binding domain of Cx43. The following report investigates the role of the Cx43 microtubule-binding domain in extracellular ATP release by Cx43 hemichannels and how this impacts early inflammatory events of the foreign body reaction.
  
  
  METHODS
  In vitro Cx43 hemichannel-mediated ATP release by cultured human microvascular endothelial cells subjected to hypocalcemic and normocalcemic conditions was measured after application of JM2 and the known hemichannel blocker, flufenamic acid. A submuscular silicone implant model was used to investigate in vivo ATP signaling during the early foreign body response. Implants were coated with control pluronic vehicle or pluronic carrying JM2, ATP, JM2+ATP, or known hemichannel blockers and harvested at 24 h for analysis.
  
  
  RESULTS
  JM2 significantly inhibited connexin hemichannel-mediated ATP release from cultured endothelial cells. Importantly, the early inflammatory response to submuscular silicone implants was inhibited by JM2. The reduction in inflammation by JM2 was reversed by the addition of exogenous ATP to the pluronic vehicle.
  
  
  CONCLUSIONS
  These data indicate that ATP released through Cx43 hemichannels into the vasculature is an important signal driving the early inflammatory response to implanted devices. A vital aspect of this work is that it demonstrates that targeted molecular therapeutics, such as JM2, provide the capacity to regulate inflammation in a clinically relevant system.
authors.affiliation: 1Department of Surgery, Medical University of South Carolina, Charleston, South Carolina.
authors.forename: Bennett W
authors.initials: BW
authors.lastname: Calder
authors.affiliation: 1Department of Surgery, Medical University of South Carolina, Charleston, South Carolina.
authors.forename: Joshua
authors.initials: J
authors.lastname: Matthew Rhett
authors.affiliation: 1Department of Surgery, Medical University of South Carolina, Charleston, South Carolina.
authors.forename: Heather
authors.initials: H
authors.lastname: Bainbridge
authors.affiliation: 1Department of Surgery, Medical University of South Carolina, Charleston, South Carolina.
authors.forename: Stephen A
authors.initials: SA
authors.lastname: Fann
authors.affiliation: 2Center for Heart and Regenerative Medicine Research, Virginia Tech Carilion School of Medicine and Research Institute, Roanoke, Virginia.
authors.forename: Robert G
authors.initials: RG
authors.lastname: Gourdie
authors.affiliation: 1Department of Surgery, Medical University of South Carolina, Charleston, South Carolina.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Yost
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism* / Q000494:pharmacology; D000595:Amino Acid Sequence; D000818:Animals; D019921:Biotinylation; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D018031:Connexin 43 / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000502:physiology*; D004353:Drug Evaluation, Preclinical; D042783:Endothelial Cells / Q000378:metabolism; D005439:Flufenamic Acid / Q000378:metabolism; D005549:Foreign-Body Reaction / Q000276:immunology / Q000378:metabolism*; D006801:Humans; D007249:Inflammation; D008264:Macrophages / Q000276:immunology; D008297:Male; D008870:Microtubules / Q000378:metabolism; D008969:Molecular Sequence Data; D009504:Neutrophils / Q000276:immunology; D010446:Peptide Fragments / Q000378:metabolism / Q000494:pharmacology; D017434:Protein Structure, Tertiary; D051381:Rats; D017207:Rats, Sprague-Dawley; D012828:Silicones
chemical_list: D018031:Connexin 43; C500216:GJA1 protein, human; D010446:Peptide Fragments; D012828:Silicones; D005439:Flufenamic Acid; D000255:Adenosine Triphosphate; D002118:Calcium