Cardiomyocytes express most known Toll-like receptors. Of these, TLR2, TLR4 and TLR5 signal via NF-kappaB, resulting in decreased contractility and a concerted inflammatory response.

Background: Toll-like receptors (TLRs) are expressed on cardiomyocytes and recognize pathogen-associated molecular patterns. Whether endogenous molecules produced by tissue injury (damage associated molecular patterns, DAMPs) can induce cardiomyocyte inflammation via TLR signalling pathways and/or reduce cardiomyocyte contractility is unknown. Methods and Results:Primary cardiomyocytes isolated from nuclear factor κ B (NFκB)-luciferase knock-in mice were used to assess NFκB signalling. DAMPs, HSP60, HSP70 and HMGB1, increased NFκB transcriptional activity compared to controls. HSP70 stood out compared to other DAMPs and even lipopolysaccharide (LPS). Subsequent experiments focused on HSP70. Cardiomyocytes exposed to HSP70 had a 58% decrease in contractility without a decrease in calcium flux. Exposure of cultured HL-1 cardiomyocytes to HSP70 resulted in increased expression of intercellular adhesion molecule 1 (ICAM-1), interleukin 6 (IL-6) and keratinocyte-derived chemokine (KC) compared to controls. Knock-out mice for TLR2, TLR4 and MyD88, plus background strain controls (C57BL/6) were used to assess induction of cardiomyocyte inflammation by HSP70. The cardiomyocyte expression of ICAM-1 induced by HSP70 was significantly reduced in TLR2 and MyD88 knock-out mice but not TLR4 knock-out mice; implicating the TLR2 signalling pathway. Furthermore, blocking antibodies to TLR2 were able to abrogate HSP70-induced contractile dysfunction and cell death.Conclusions: Extracellular HSP70 acting via TLR2 and its obligate downstream adaptor molecule, MyD88, activate NFκB. This causes cardiomyocyte inflammation and decreased contractility. (Circ J 2011; 75: 2445 - 2452

Stimulation of toll-like receptor 9 (TLR9) by CpG-C containing oligonucleotides attenuates ischemic injury in the brain and liver. In this study, we investigate whether any of the three classes of CpG (A, B, or C) mitigate ischemia-induced cardiac dysfunction. We measured left ventricular ejection fraction (LVEF) in C57BL/6 mice using transthoracic echocardiography. Using LPS as an inflammatory stimulus, CpG-C was uniquely able to prevent cardiac dysfunction; its activity was confirmed through nuclear factor κB transcriptional activity assay in HL-1 cardiomyocytes. We went on to investigate CpG-C's efficacy and mechanism in the treatment of ischemia-reperfusion. Compared with baseline, no class of CpG significantly altered LVEF at 6 or 24 h; 40 mg/kg LPS induced a rapid, profound suppression of LVEF compared with baseline (26% ± 1.4% vs. 65% ± 1.4%), whereas pretreatment with CpG demonstrated that of the three classes, only CpG-C prevented the LPS -induced decrease in LVEF (51% ± 5.8%). In separate mice, 1-h ischemia followed by reperfusion of the left anterior descending artery resulted in a 7-day suppression of the LVEF (66% ± 5.2% at baseline; 46% ± 4.7% at day 1, and 46% ± 4.0% at day 7), whereas mice either pretreated with or begun on an infusion of CpG-C during the ischemia had no significant decline in LVEF. Gene expression microarray of CpG-C-stimulated cells revealed upregulation of the nuclear factor κB pathway inhibitors TNFAIP3, NFKBIA, TRIM30, and TNIP1. These may play a role in attenuation of cardiac inflammation. The TLR9 ligand CpG-C attenuates the acute inflammatory cardiac dysfunction induced by both LPS and ischemia-reperfusion of the left anterior descending artery.