title: Monocyte activation by necrotic cells is promoted by mitochondrial proteins and formyl peptide receptors.
abstract: OBJECTIVE
  Necrotic cells evoke potent innate immune responses through unclear mechanisms. The mitochondrial fraction of the cell retains constituents of its bacterial ancestors, including N-formyl peptides, which are potentially immunogenic. Thus, we hypothesized that the mitochondrial fraction of the cell, particularly N-formyl peptides, contributes significantly to the activation of monocytes by necrotic cells.
  
  
  METHODS
  Human peripheral blood monocytes were incubated with necrotic cell fractions and mitochondrial proteins to investigate their potential for immune cell activation.
  
  
  METHODS
  University Medical Center Research Laboratory.
  
  
  METHODS
  Healthy human adults served as blood donors.
  
  
  RESULTS
  Human blood monocyte activation was measured after treatment with cytosolic, nuclear and mitochondrial fractions of necrotic HepG2 cells or necrotic HepG2 cells depleted of N-formyl peptides [Rho(0) cells]. The specific role of the high affinity formyl peptide receptor (FPR) was then tested using specific pharmacologic inhibitors and RNA silencing. The capacity of mitochondrial N-formyl peptides to activate monocytes was confirmed using a synthetic peptide conforming to the N-terminus of mitochondrial nicotinamide adenine dinucleotide subunit 6. The results demonstrated that mitochondrial cell fractions most potently activated monocytes, and interleukin (IL)-8 was selectively released at low-protein concentrations. Mitochondria from Rho(0) cells induced minimal monocyte IL-8 release, and specific pharmacologic inhibitors and RNA-silencing confirmed that FPR contributes significantly to monocyte IL-8 responses to both necrotic cells and mitochondrial proteins. N-formyl peptides alone did not induce monocyte IL-8 release; whereas, the combination of mitochondrial N-formyl peptides and mitochondrial transcription factor A (TFAM) dramatically increased IL-8 release from monocytes. Likewise, high mobility group box 1, the nuclear homolog of TFAM, did not induce monocyte IL-8 release unless combined with mitochondrial N-formyl peptides.
  
  
  CONCLUSIONS
  Interactions between mitochondrial N-formyl peptides and FPR in the presence of other mitochondrial antigens (e.g., TFAM) contributes significantly to the activation of monocytes by necrotic cells.
authors.affiliation: Division of Pulmonary, Allergy, Critical Care and Sleep Medicine, Dorothy M Davis Heart and Lung Research Institute, The Ohio State University Medical Center, Columbus, OH, USA. elliott.crouser@osumc.edu
authors.forename: Elliott D
authors.initials: ED
authors.lastname: Crouser
authors.forename: Guohong
authors.initials: G
authors.lastname: Shao
authors.forename: Mark W
authors.initials: MW
authors.lastname: Julian
authors.forename: Jennifer E
authors.initials: JE
authors.lastname: Macre
authors.forename: Gerald S
authors.initials: GS
authors.lastname: Shadel
authors.forename: Susheela
authors.initials: S
authors.lastname: Tridandapani
authors.forename: Qin
authors.initials: Q
authors.lastname: Huang
authors.forename: Mark D
authors.initials: MD
authors.lastname: Wewers
mesh_terms: D002478:Cells, Cultured; D004272:DNA, Mitochondrial / Q000502:physiology; D004268:DNA-Binding Proteins / Q000502:physiology; D006801:Humans; D024101:Mitochondrial Proteins / Q000502:physiology*; D009000:Monocytes / Q000502:physiology*; D009336:Necrosis / Q000473:pathology*; D044042:Receptors, Formyl Peptide / Q000502:physiology*; D014157:Transcription Factors / Q000502:physiology
chemical_list: D004272:DNA, Mitochondrial; D004268:DNA-Binding Proteins; D024101:Mitochondrial Proteins; D044042:Receptors, Formyl Peptide; C499631:TFAM protein, human; D014157:Transcription Factors