Mechanisms of toll-like receptor 4 (TLR4)-mediated inflammation after cold ischemia/reperfusion in the heart

David J. Kaczorowski, Atsunori Nakao, Raghuveer Vallabhaneni, Kevin P. Mollen, Ryujiro Sugimoto, Junichi Kohmoto, Brian S. Zuckerbraun, Kenneth R. McCurry, Timothy R. Billiar

Research output: Contribution to journalArticlepeer-review

94 Scopus citations


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 IFNβ (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 (TNFα), and intercellular adhesion molecule-1 (ICAM-1) messenger RNA (mRNA) levels (Pĝ‰Currency sign0.05). Compared with controls, CD14 knock-out (KO) mice exhibited significantly lower (Pĝ‰Currency sign0.05) systemic IL-6 and JE/monocyte chemotractant protein-1 levels after cold I/R. Intragraft TNFα and IL-1β mRNA levels were also significantly lower (Pĝ‰Currency sign0.05) in CD14 KO grafts. MyD88 KO mice exhibited significantly lower (P≥0.05) systemic IL-6 levels compared with control mice after cold I/R. Intragraft TNFα, IL-6, and ICAM-1 mRNA levels were also significantly lower (Pĝ‰Currency sign0.05) in MyD88 KO grafts. Significantly lower levels (Pĝ‰Currency sign0.05) of serum IL-6, monocyte chemotractant protein-1 as well as intragraft TNFβ, IL-6, IL-1β, 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.

Original languageEnglish
Pages (from-to)1455-1463
Number of pages9
Issue number10
StatePublished - 27 May 2009
Externally publishedYes


  • Cold storage
  • Inflammation
  • Ischemia-reperfusion
  • Toll-like receptors
  • Transplant


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