title: Mechanisms of Toll-like receptor 4 (TLR4)-mediated inflammation after cold ischemia/reperfusion in the heart.
abstract: BACKGROUND
  Toll-Like Receptor 4 (TLR4) signaling mediates early inflammation after cold ischemia-reperfusion (I/R). We hypothesized that the TLR4 coreceptor CD14, the intracellular adaptor proteins myeloid differentiation factor 88 (MyD88) and TIR domain-containing-adaptor inducing IFNbeta (TRIF) would be required for cold I/R induced inflammation. High mobility group box 1 (HMGB1) is a putative endogenous activator of TLR4. Therefore, we also assessed the contribution of HMGB1 in cold I/R induced inflammation.
  
  
  METHODS
  Syngeneic heart transplants were performed in mice deficient in CD14, MyD88, TRIF, or wild-type mice. In other experiments, anti-HMGB1 neutralizing antibody or control IgG was administered at reperfusion. Donor hearts were subjected to 2 hr of cold ischemia and retrieved after 3 hr of reperfusion.
  
  
  RESULTS
  After cold I/R, grafts revealed striking translocation of HMGB1 out of the nucleus in cardiac myocytes. Administration of an anti-HMGB1 neutralizing antibody resulted in reduced systemic interleukin (IL)-6, tumor necrosis factor alpha (TNFalpha), and intercellular adhesion molecule-1 (ICAM-1) messenger RNA (mRNA) levels (P< or =0.05). Compared with controls, CD14 knock-out (KO) mice exhibited significantly lower (P< or =0.05) systemic IL-6 and JE/monocyte chemotractant protein-1 levels after cold I/R. Intragraft TNFalpha and IL-1beta mRNA levels were also significantly lower (P< or =0.05) in CD14 KO grafts. MyD88 KO mice exhibited significantly lower (P< or=0.05) systemic IL-6 levels compared with control mice after cold I/R. Intragraft TNFalpha, IL-6, and ICAM-1 mRNA levels were also significantly lower (P< or =0.05) in MyD88 KO grafts. Significantly lower levels (P< or =0.05) of serum IL-6, monocyte chemotractant protein-1 as well as intragraft TNFalpha, IL-6, IL-1beta, and ICAM-1 were observed after cold I/R in TRIF deficient animals compared with controls.
  
  
  CONCLUSIONS
  CD14, MyD88, TRIF, and HMGB1 contribute to the inflammatory response that occurs after cold I/R. These results provide insight into the mechanisms of TLR4-mediated inflammation after cold I/R.
authors.affiliation: Department of Surgery, University of Pittsburgh Medical Center, Pittsburgh, PA 15213, USA.
authors.forename: David J
authors.initials: DJ
authors.lastname: Kaczorowski
authors.forename: Atsunori
authors.initials: A
authors.lastname: Nakao
authors.forename: Raghuveer
authors.initials: R
authors.lastname: Vallabhaneni
authors.forename: Kevin P
authors.initials: KP
authors.lastname: Mollen
authors.forename: Ryujiro
authors.initials: R
authors.lastname: Sugimoto
authors.forename: Junichi
authors.initials: J
authors.lastname: Kohmoto
authors.forename: Brian S
authors.initials: BS
authors.lastname: Zuckerbraun
authors.forename: Kenneth R
authors.initials: KR
authors.lastname: McCurry
authors.forename: Timothy R
authors.initials: TR
authors.lastname: Billiar
mesh_terms: D033942:Adaptor Proteins, Vesicular Transport / Q000172:deficiency; D000818:Animals; D050377:Cold Ischemia; D006321:Heart / Q000503:physiopathology*; D016027:Heart Transplantation / Q000276:immunology / Q000502:physiology*; D007249:Inflammation / Q000276:immunology*; D018950:Lipopolysaccharide Receptors / Q000235:genetics; D051379:Mice; D018345:Mice, Knockout; D053594:Myeloid Differentiation Factor 88 / Q000172:deficiency; D009206:Myocardium / Q000276:immunology; D032383:Myocytes, Cardiac / Q000276:immunology / Q000502:physiology; D015427:Reperfusion Injury / Q000276:immunology*; D015398:Signal Transduction; D051197:Toll-Like Receptor 4 / Q000502:physiology*; D014185:Transplantation, Isogeneic
chemical_list: D033942:Adaptor Proteins, Vesicular Transport; D018950:Lipopolysaccharide Receptors; D053594:Myeloid Differentiation Factor 88; D051197:Toll-Like Receptor 4; C507780:Myd88 protein, mouse; C476286:Ticam1 protein, mouse; C493487:Tlr4 protein, mouse