title: Proteasome Inhibition Diminishes the Formation of Neutrophil Extracellular Traps and Prevents the Death of Cardiomyocytes in Coculture with Activated Neutrophils during Anoxia-Reoxygenation.
abstract: OBJECTIVE
  Polymorphic mononuclear neutrophils (PMN) are very important cells participating in nonspecific defense of the organism. Among their well-known functions, the formation of neutrophil extracellular traps (NET) is interesting and potentially dangerous for the mechanisms of other cells. Ubiquitin-dependent proteasomal proteolysis is a very important regulator of all cellular activities, but the role of proteasomal proteolysis in NET formation has not been investigated.
  
  
  METHODS
  We performed experiments with PMN activated to form NET with phorbol 12-myristate 13-acetate (PMA) and the application of a proteasome inhibitor. We also added activated neutrophils to primary culture of isolated rat neonatal cardiomyocytes with or without anoxia-reoxygenation modeling.
  
  
  RESULTS
  The data obtained show that proteasomes participate in NET formation and proteasome inhibitors facilitate the blocking of the NET program. The percentage of NET after PMA application was 70.8 ± 7.2 and the proteasome inhibitor decreased this amount to 4.7 ± 0.9%. In coculture with cardiomyocytes during anoxia-reoxygenation, this effect prevented cardiac cell death induced by activated PMN. The stimulation of NET formation by PMA in coculture with isolated cardiomyocytes led to an increase in the number of necrotic cardiomyocytes of up to 33.1 ± 12.9% and a corresponding decrease in living cardiomyocytes to 66.9 ± 12.9%. The number of living cardiomyocytes in coculture after incubation with both PMA and proteasome inhibitor was 76.6 ± 13.3% (p < 0.05), and the number of necrotic cardiomyocytes was 23.4 ± 13.3% (p < 0.05).
  
  
  CONCLUSIONS
  Proteasome inhibition blocks NET formation and prevents cardiomyocyte necrosis in coculture with activated neutrophils.
authors.affiliation: Department of General and Molecular Pathophysiology, Bogomoletz Institute of Physiology, National Academy of Science of Ukraine, Kiev, Ukraine.
authors.forename: Denis O
authors.initials: DO
authors.lastname: Pashevin
authors.forename: Vasyl S
authors.initials: VS
authors.lastname: Nagibin
authors.forename: Lesya V
authors.initials: LV
authors.lastname: Tumanovska
authors.forename: Alex A
authors.initials: AA
authors.lastname: Moibenko
authors.forename: Victor E
authors.initials: VE
authors.lastname: Dosenko
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D015687:Cell Hypoxia; D002478:Cells, Cultured; D018920:Coculture Techniques; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D065206:Extracellular Traps / Q000276:immunology / Q000502:physiology*; D007976:Leupeptins / Q000494:pharmacology; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000473:pathology / Q000502:physiology*; D018375:Neutrophil Activation; D009504:Neutrophils / Q000187:drug effects / Q000502:physiology* / Q000648:ultrastructure; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D051381:Rats; D013755:Tetradecanoylphorbol Acetate / Q000031:analogs & derivatives / Q000494:pharmacology
chemical_list: D015853:Cysteine Proteinase Inhibitors; D007976:Leupeptins; C016658:phorbolol myristate acetate; D046988:Proteasome Endopeptidase Complex; D013755:Tetradecanoylphorbol Acetate; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde