How do endosomal Toll-like receptors sense and extend ischaemic myocardial injury?

This editorial refers to ‘Toll-like receptor 7 deficiency promotes survival and reduces adverse left ventricular remodelling after myocardial infarction’ by D.P.V. de Kleijn et al., pp. 1791–1803.

Following myocardial infarction, dying cells and fragmented extracellular matrix release damage-associated molecular patterns, which activate a family of pattern recognition receptors, the Toll-like receptors (TLRs), triggering innate immune signalling pathways.¹ The TLR family is comprised of plasma membrane receptors (including TLR1, TLR2, TLR4, TLR6, and TLR10), and nucleic acid-sensing endosomal receptors, such as TLR3, TLR7, TLR8, and TLR9.^2,3 Although TLRs have evolved as a warning system against infectious agents that elicits a protective response, inappropriate or excessive activation of TLRs following tissue injury may have catastrophic consequences. In the infarcted myocardium, the cell surface TLRs, TLR2, and TLR4, have been implicated in extension of ischaemic injury, and in the pathogenesis of adverse remodelling.^4–8 In contrast, the potential role of endosomal TLRs that may respond to nucleic acids released by dying cells to activate innate immune signalling, remains poorly understood. TLR7 recognizes purine-rich single-stranded RNA (ssRNA) and is known to elicit a protective immune response to viral infection. In vitro, RNA released by injured cardiomyocytes has been demonstrated to induce TLR7-dependent inflammatory leucocyte activation.⁹ The potential role of TLR7-dependent signalling in the pathogenesis of myocardial injury in vivo remains unknown.

de Kleijn et al.¹⁰ studied the role of TLR7 in repair and remodelling of the infarcted heart. They found that TLR7 expression is up-regulated in the infarcted mouse heart, and in human myocardium from patients with ischaemic heart failure. In a model of non-reperfused myocardial infarction, mice with global loss of TLR7 had a decreased incidence of cardiac rupture and exhibited attenuated adverse remodelling, associated with increased recruitment of myofibroblasts and a favourable protease/antiprotease balance. Experiments using bone marrow chimaeras demonstrated that TLR7-dependent dysfunction and remodelling of the infarcted heart were mediated predominantly through effects on bone marrow-derived cells. The findings add TLR7 to the growing list of pattern recognition receptors involved in post-infarction remodelling, and suggest a potential role for extracellular ssRNA, released by dying cells, in mediating rupture, dilation and dysfunction of the infarcted heart. Moreover, the observations raise several intriguing questions regarding the identity of the ligands activating TLR7 in the infarcted heart, the cellular targets of TLR7-mediated responses, the mechanism of delivery of infarct-derived TLR7 ligands in the endosomal compartment of leucocytes, and the downstream signalling pathways responsible for the injurious actions.

How do ssRNAs access endosomal TLR7 in leucocytes infiltrating the infarct?

Due to its endosomal localization, TLR7 requires an interaction with endocytosed ssRNA to activate an innate immune signalling response.¹¹ Following viral infection, viral ssRNA enters the endosome via receptor-mediated endocytosis and interacts with TLR7 initiating signal transduction. One can only speculate regarding the mechanism responsible for delivery of ssRNA released by dying cardiomyocytes to immune cells infiltrating the infarcted heart (Figure 1). In autoimmune diseases, such as systemic lupus erythematosus, autophagic responses have been implicated in TLR7-dependent activation of inflammatory signalling.¹² Binding of nucleic acids to high-mobility group box proteins (HMGBs) has been implicated in selective activation of endosomal nucleic-acid-sensing TLRs. In healing infarcts, early release of HMGB1¹³ may prime immune cells for endocytosis of ssRNAs, thus setting the stage for activation of TLR7-mediated signalling.^14,15

Figure 1.

The role of the endosomal Toll-like receptor 7 (TLR7) in remodelling of the infarcted heart. The study by De Kleijn et al. suggests that endosomal TLR7 may be activated in infarct leucocytes, presumably by single-stranded RNA (ssRNA) released by dying cardiomyocytes, accentuating the immune response and promoting extracellular matrix degradation through release of matrix metalloproteinases (MMPs). Several important questions remain to be answered: (i) How does ssRNA find its way to leucocyte endosomes to activate TLR7? Receptor-mediated endocytosis and autophagic pathways may be implicated. Moreover, release of high-mobility group box protein 1 (HMGB1) following infarction may prime ssRNA for endocytosis. (ii) Which cells exhibit TLR7 activation? TLR7 is constitutively expressed by dendritic cells and B lymphocytes and is markedly induced following inflammation in macrophages and in non-immune cell types. (iii) Which downstream signals mediate the effects of TLR7 in the infarcted heart? Following interaction with ssRNA in endosomes, TLR7 dimerizes and may activate nuclear factor (NF)-κB and mitogen-activated protein kinase (MAPK) through a myeloid differentiation primary response protein 88 (MyD88)-dependent pathway, thus inducing synthesis of cytokines and chemokines. TLR7-mediated MMP secretion may also play a role in mediating cardiac rupture and adverse remodelling of the infarcted heart. Additional symbols: IL, interleukin; TNF, tumour necrosis factor; IRAK, interleukin-1 receptor-associated kinase.

Which leucocyte subpopulations are activated by TLR7?

Although the effects of TLR7 in cardiac rupture and adverse remodelling appear to involve bone marrow cells, the specific leucocyte subsets that respond to ssRNAs, activating TLR7 signalling is unknown. Constitutive expression of TLR7 has been documented in dendritic cells and B cells; lower levels of TLR7 may also be expressed in non-immune cells, including cardiomyocytes.⁹ Following injury, activation of nuclear factor (NF)-κB induces TLR7 in a wide range of cell types, including macrophages.¹⁶ Thus, in the cytokine-rich environment of the infarcted heart, increased expression of TLR7 in many different cell types may account for its pro-inflammatory and matrix-degrading actions. Cell-specific loss-of-function approaches are needed to dissect the relative role of various immune cell subsets in mediating TLR7-dependent responses.

Which molecular signals are responsible for TLR7-dependent actions?

Upon interaction with ssRNA, TLR7 dimerizes and associates with myeloid differentiation primary response protein 88 (MyD88), thus initiating signal transduction, and stimulating downstream activation of mitogen-activated protein kinases (MAPKs), the NF-κB cascade, interleukin-1 receptor-associated kinases (IRAKs), and tumour necrosis factor receptor associated factors (TRAF) 3/6 (Figure 1). In human immune cells, TLR7 activation triggers synthesis of a wide range of pro-inflammatory cytokines, including tumour necrosis factor (TNF)-α, interleukin (IL)-1β, and IL-6,¹⁷ mediators with a critical role in the post-infarction inflammatory response.¹⁸ Accentuated induction of pro-inflammatory cytokines may account for TLR7-dependent cardiac rupture and may accentuate dilative remodelling following infarction. The current study suggests that TLR7 signalling mediates MMP release in the infarcted area, thus promoting a matrix-degrading environment that may cause left ventricular rupture. TLR7-mediated MMP release may be due to cytokine-stimulated protease induction, or may reflect direct effects of TLR7 on MMP synthesis by activated leucocytes.

Conclusions

Emerging evidence suggests that endosomal TLRs (such as TLR7) do not simply serve to protect from viral infections, but may be inappropriately activated following non-viral tissue injury, accentuating inflammation. In the infarcted myocardium, activation of TLR7 signalling may have catastrophic consequences, causing rupture and precipitating heart failure. Further studies are needed to dissect the cellular basis for the deleterious effects of TLR7 in the infarcted heart, and to suggest a plausible mechanism for endosomal delivery of ssRNA in infarct leucocytes. If consistent and reproducible, the impressive beneficial effects of disrupting the ssRNA: TLR7 interaction suggested by the current study, may suggest TLR7 antagonism as a promising therapeutic approach for patients with myocardial infarction.

Conflict of interest: none declared.

Funding

This work was supported by the National Institutes of Health (R01 HL76246 and R01 HL85440) and by the Department of Defense (PR151134 and PR151029) (to N.G.F.).

The opinions expressed in this article are not necessarily those of the Editors of Cardiovascular Research or of the European Society of Cardiology.