Myocardial infarction remodeling that progresses to heart failure: a signaling misunderstanding

Abstract

After myocardial infarction, remodeling of the left ventricle involves a wound-healing orchestra involving a variety of cell types. In order for wound healing to be optimal, appropriate communication must occur; these cells all need to come in at the right time, be activated at the right time in the right amount, and know when to exit at the right time. When this occurs, a new homeostasis is obtained within the infarct, such that infarct scar size and quality are sufficient to maintain left ventricular size and shape. The ideal scenario does not always occur in reality. Often, miscommunication can occur between infarct and remote spaces, across the temporal wound-healing spectrum, and across organs. When miscommunication occurs, adverse remodeling can progress to heart failure. This review discusses current knowledge gaps and recent development of the roles of inflammation and the extracellular matrix in myocardial infarction remodeling. In particular, the macrophage is one cell type that provides direct and indirect regulation of both the inflammatory and scar-forming responses. We summarize current research efforts focused on identifying biomarker indicators that reflect the status of each component of the wound-healing process to better predict outcomes.

INTRODUCTION

Myocardial infarction (MI) occurs when ischemia is of sufficient duration to induce myocyte necrosis. If the time of ischemia is short and area of damage is small, the result is minimal damage to the myocardium. When the time of ischemia or the injury area is extensive, there is a variation in remodeling response across individuals in both humans and animal models that ranges from adequate wound healing to adverse remodeling that progresses to heart failure.

Once myocyte ischemia is past the point of no return and necrosis has initiated, the death of myocytes triggers an inflammatory response. The myocardium undergoes a wound-healing paradigm that involves first a proinflammatory and extracellular matrix (ECM)-degrading component to clear away the necrotic debris. The second step is anti-inflammatory and proreparative to turn off the inflammation and stimulate ECM deposition to form the new scar. Cells involved in this coordinated response include cardiac myocytes, neutrophils, macrophages, fibroblasts, endothelial cells, and nerve cells, with cross talk occurring across all cell types. Of these cell types, the macrophage and fibroblast have central roles in post-MI wound healing by providing mediators that span the whole period of the response.

When MI occurs, the best-case scenario is an optimized wound-healing response to generate an appropriate scar and minimize extension of damage to surrounding regions. Clinically, the current best treatment option is timely reperfusion, which will allow salvage of vulnerable myocytes in the border region of the infarct. In permanent occlusion coronary artery ligation animal models, which mimic patients who are not reperfused or who advance to heart failure, interventions that increase or decrease one component of the wound-healing process have consistently stimulated an adverse remodeling profile. This indicates that disruption of the spatial and temporal coordination of individual cell responses, even if beneficial to a certain cell type, can have a negative impact at the whole organ level.

There are numerous review articles summarizing the inflammatory and ECM components to post-MI remodeling (30, 99). Here, we highlight the current knowledge and focus on the key gaps that remain in our knowledge base.

INFLAMMATION

Miscommunication in inflammation includes signaling at the molecular and cellular levels, as shown in Table 1. The inflammatory response to MI has a molecular and cellular component, with a multitude of cytokines and chemokines involved (5, 20, 23, 28, 92, 124). Several groups have reported adverse MI remodeling in the setting of systemic inflammation (14, 23, 112). De Jesus et al. (11) have shown that preexisting atherosclerosis, either in apolipoprotein E (apoE)-null mice fed an atherogenic diet or simulated by elevated systemic LPS administration, exacerbates post-MI electrophysiological uncoupling and arrhythmias. Dutta et al. (19) reported the converse, that MI stimulates the progression of atherosclerosis. When chemokine (C-C motif) receptor (CCR)2 was silenced in apoE-null mice after MI, Ly-6C^high monocyte recruitment was limited, and the resulting attenuation in infarct inflammation curbed post-MI remodeling (87). In a periodontal model of chronic systemic inflammation, a baseline elevation in the inflammatory status worsens MI remodeling characterized by earlier rupture as a result of elevated matrix metalloproteinase (MMP)-9 levels (12). Chronic inflammation stimulates macrophage secretion of chemokine (C-C motif) ligand (CCL)12, which impairs post-MI reparative fibroblast activation (14). The connection between inflammatory status and the MI response, therefore, is very strong.

Table 1.

Post-MI effect of perturbing inflammation components

Perturbation	Effect	Mechanism
Reducing inflammation:	↑LV physiology	IL-1α null: ↓innate immune response in cardiac fibroblasts
IL-1α null (79)TREM1 null (2)NF-κB null (60)TLR4 null (97)TRPV2 null (21)PS-presenting liposomes (39)	↓LV remodeling	TREM1 null: ↓myeloid cell recruitment from the spleen and bone marrow; ↓CCR2+ proinflammatory but not proreparative CX3CR1+ monocyte infiltrationTLR4 null: ↓neutrophil infiltration/activationTRPV2 null: ↓macrophage migrationPS-liposomes: ↑macrophage secretion of IL-10 and TGF-β
Total neutrophil depletion (43, 44, 85, 86)	↓LV physiology	↓Necrotic myocyte removal
	↑Fibrosis	↑Macrophage inflammatory response
		↑Myofibroblast activation
Total macrophage depletion (120)	↓LV physiology	Total: ↓necrotic/apoptotic cell removal
	↓Collagen deposition	↓Fibroblast and endothelial cell activation
M2-specific depletion (66)	↓Neovascularization	M2: prolonged N1 neutrophil/M1 macrophage activation
Macrophage modulation:	↑LV physiology	Wntless: ↑reparative M2 macrophages, neovascularization
Wntless null (98)Macrophage EP3R null (114)Macrophage EP3R transgenic (114)CD5L null (96)	↓LV remodeling	EP3R null: monocyte TGF-β signaling/↓CX3CR1 and VEGF signaling, ↓Ly6Clow reparative monocyte activationEP3R transgenic: ↑angiogenesisCD5L null: ↓neutrophils, ↓collagen accumulation, ↓IL-1 receptor-associated kinase 4, NF-κB, myeloperoxidase, and inducible nitric oxide synthase

Reference numbers are listed in parentheses. MI, myocardial infarction; TLR, Toll-like receptor; TREM1, triggering receptor expressed on myeloid cells 1; PS, phosphatidylserine; LV, left ventricular; CCR, chemokine (C-C motif) receptor; CX3CR, chemokine (C-X3-C motif) receptor; TGF-β, transforming growth factor-β; EP3R, E-prostanoid 3 receptor; ↑, increase; ↓, decrease.

For the cellular component, by numbers, neutrophils and macrophages are the predominant leukocytes that infiltrate into the infarcted left ventricle (LV) and are known participants in LV remodeling (109). While monocyte infiltration begins first, neutrophil infiltration quickly becomes the predominant leukocyte in the infarct zone by 24 h after MI (33, 44, 102). Neutrophils at day 1 post-MI release granule components (e.g., serine elastase and MMP-9) aimed at breaking down necrotic myocytes and clearing tissue debris (67, 86). Anti-inflammatory strategies that limit neutrophil infiltration reduce acute injury but exacerbate cardiac physiology and enhance the fibrotic response to promote progression to heart failure (43, 45). By day 7 post-MI, neutrophil numbers have begun to return toward baseline day 0 values. While neutrophils are primarily considered proinflammatory, we have reported that they undergo polarization, and anti-inflammatory (N2) neutrophils account for 20% of the neutrophils in the day 7 infarct (85). The N2 neutrophil is characterized by high expression of macrophage mannose receptor CD206 and IL-10 (85). Neutrophils, therefore, contribute directly to inflammation and directly and indirectly to cardiac repair after MI.

Macrophages have polarization profiles similar to neutrophils, with day 1 post-MI macrophages exhibiting a proinflammatory M1 profile (6, 25, 42). Over the first 7 days of MI, there is a transition from primarily M1 macrophages to primarily anti-inflammatory and then reparative M2 cells (75, 84). While the M1 and M2 classification system is frequently used to define pro- and anti-inflammatory macrophages, there is a growing effort to replace the M1/M2 nomenclature, as this naming was set up to describe the in vitro situation and does not apply to the MI scenario where there is a heterogeneous set of polarization stimuli (93).

In addition to known roles as phagocytic cells that remove necrotic debris and apoptotic neutrophils, macrophages have been assigned roles in facilitating electrical conduction in the atria and regulating diastolic pathophysiology in the setting of hypertension (46, 47, 57). In skin wound healing, macrophage-depleted mice showed a severely impaired wound morphology and delayed healing due to the persistence of large numbers of neutrophils, proinflammatory mediators (macrophage inflammatory protein-3, macrophage chemoattractant protein-1, IL-1b, and cycyooxygenase-2), impaired neovascularization, and fibroblast activation (37). Post-MI, macrophage-depleted mice show impaired wound healing and exacerbated LV remodeling, leading to high mortality and excessive LV dilation (120). Deletion of macrophage polarization components show similar negative phenotypes. For example, CCR5 deletion does not limit macrophage infiltration into the infarct region; rather, it attenuates both pro- and anti-inflammatory proteins, resulting in poor healing and increased rupture rates (131). Periodontal-induced chronic inflammation triggers macrophage secretion of CCL12 to inhibit fibroblast-mediated cardiac wound healing (14). Obesity superimposed on aging magnifies inflammation and delays the resolving response after MI (78). While total depletion of macrophages is detrimental to post-MI wound healing, in vivo silencing of the interferon regulatory factor 5 transcription factor reprograms the macrophage away from an inflammatory polarization status to attenuate inflammation and remodeling after MI (9). Likewise, infusion of anti-inflammatory IL-10 early post-MI shifts macrophage polarization and improves LV remodeling (56).

These studies highlight the role of the macrophage in cross talk with neutrophils, fibroblasts, and endothelial cells (33, 84). Neutrophil depletion is also detrimental to MI wound healing, in part because secretion of neutrophil gelatinase-associated lipocalin (NGAL; lipocalin 2) by neutrophils stimulates macrophage polarization toward the reparative phenotype (43). A number of reviews have summarized current efforts to target post-MI inflammation (5, 23, 28, 45, 103, 124). A major take-home message here is that modulation of some aspects of inflammation feedforward to modify other aspects. Caution should be given to strategies that totally block the entry of any one cell type, as this approach is likely overly simplistic. To date, inhibition of IL-1β is a target that shows promise in the post-MI setting in both animal models and humans (29, 94, 100). The consistency across species indicates that the inflammatory component is a common denominator and translational research focused on inflammation is likely to inform clinical practice.

SCAR FORMATION

Miscommunication in ECM signaling also occurs at the molecular and cellular levels, as shown in Table 2 and Fig. 1. Major ECM components include structural proteins (collagens and fibronectin), accessory proteins [osteopontin, secreted protein acidic and rich in cysteine (SPARC), and thrombospondin-1 (TSP-1)], and MMPs. MMPs are a family of enzymes that degrade ECM constituents as well as many of the inflammatory mediators listed above (15). Tissue inhibitors of metalloproteinase (TIMPs) are the endogenous inhibitors of MMPs.

Table 2.

Post-MI effects of perturbing extracellular matrix components

Perturbation	Effect	Mechanism
Collagen type VI null (80)	↓Infarct size	Accelerated acute apoptosis, ↓chronic apoptosis
	↑LV physiology
MMP and TIMP null	Improved LV remodeling	MMP-2: ↓M2 macrophage and myofibroblast activation
MMP-2 (40)		MMP-7: ↓connexin-43 cleavage, ↑conduction velocity
MMP-7 (8, 70)		MMP-9: ↓macrophage accumulation (late phase)
MMP-9 (18, 69)		MMP28: ↓inflammation and ↓M2 macrophage and fibroblast activation
MMP-28 (82)		TIMP null: ↑MMP activity
TIMP-1 (49)
TIMP-2 (59)
TIMP-3 (38, 115)
TIMP-4 (64)
Macrophage-specific MMP-9 transgenic (90)	↑LV physiology	↓Expression of other MMPs, ↑expression of profibrotic genes
	↓LV inflammation
Cardiac-specific MMP-1 transgenic (61)	↑LV remodeling	↑Collagen degradation
Osteopontin null (117)	↓LV physiology, collagen deposition	↑MMP-2 and ↓MMP-9 activity
Secreted protein acidic and rich in cysteine null (88, 104)	↑LV physiology early; ↑rupture later	↑Connective tissue growth factor activity and collagen synthesis
Thrombospondin-1 null (24)	↑LV dilation, inflammation	↓Transforming growth factor-β and Smad-2 signaling

Reference numbers are listed in parentheses. MI, myocardial infarction; LV, left ventricular; MMP, matrix metalloproteinase; TIMP, tissue inhibitor of metalloproteinase; ↑, increase; ↓, decrease.

Fig. 1.

After myocardial infarction (MI), there is a sequential wound-healing response comprised of inflammation followed by scar formation. When components are aligned ( = ), a stable scar forms and a new homeostatic-like environment ensues. When miscommunication among the components occurs (+/−), the scar is unstable and adverse remodeling occurs. ECM, extracellular matrix; MMP, matrix metalloproteinase. [Modified from Ref. 14.]

While collagen- and fibronectin-null mice are both embryonically lethal, mice with cleavage-resistant collagen showed no effect on post-MI LV remodeling and mice that lack the fibronectin EDA domain showed improved survival and better remodeling compared with wild-type control mice (1, 76). Osteopontin-null mice show exaggerated LV dilation through reduced collagen accumulation in the scar region, which can be ameliorated with MMP inhibition (65, 117). Deletion of SPARC reduced early rupture and improved LV remodeling through day 3 post-MI but enhanced rupture and impaired LV remodeling through day 14 post-MI, indicating biphasic roles for SPARC in early macrophage polarization and later fibroblast entry (88, 104, 125). TSP-1 limits infarct expansion and plays an important role in the transition from MI to heart failure (24, 63).

A number of MMPs and all four TIMPs have been evaluated in the post-MI LV, including MMP-2, MMP-3, MMP-7, MMP-9, MMP-10, MMP-12, MMP-13, MMP-14, and MMP-28 and all four TIMPs (15, 51, 54, 55, 72, 77, 126). Deletion of MMP-2, MMP-7, MMP-9, MMP-14, and MMP-28 overall improved LV remodeling through mechanisms ranging from effects on myocytes to inflammatory cells to endothelial cells (8, 17, 69, 70, 82, 132). MMP inhibition is complex for two reasons. First, the specific MMP that is inhibited and the time of its inhibition have a large effect on reparative outcomes. We will use MMP-9 as an example to illustrate this point (53). When MMP-9 is globally deleted from birth or is overexpressed in macrophages only, the net effect is improved LV remodeling (albeit through different mechanisms) (16, 18, 69, 90, 130). When MMP-9 is inhibited beginning at 3 h post-MI, inflammation resolution is delayed and the net effect is detrimental (52). The baseline and post-MI environments are also altered with age (7, 89, 116, 127, 128). To optimize scar formation, understanding how MMPs, MMP-9 in particular, coordinate inflammation with ECM deposition and cross linking is needed (122, 123).

Second, TIMP inhibition has shown beneficial effects on LV remodeling through effects on both MMPs and non-MMP roles (22). While TIMPs function through overlapping roles to endogenously inhibit MMPs and there is some compensation across their profiles, each TIMP has distinct mechanisms in LV remodeling. TIMP-1 deficiency accelerates post-MI LV remodeling, an effect that was pharmacologically modified with an MMP inhibitor (10, 49). Of note, TIMP-1 is an independent predictor of all-cause mortality, cardiac mortality, and MI (4). Deletion of TIMP-2 amplifies LV remodeling by enhancing activation of MMP-14 (59). TIMP-3-null mice show early MMP activation that leads to exacerbated systolic and diastolic dysfunction after MI through the activation of epidermal growth factor signaling, whereas TIMP-3 overexpression shows improvement by preventing ECM proteolysis (38, 58, 113). TIMP-4-null mice are susceptible to MI-induced rupture and mortality but not pressure overload pathology (64).

The ECM is an important modifier of fibroblast phenotype, with fibroblasts sensing changes to the ECM as an important cue (27). Depending on the ECM environment, fibroblasts can polarize through homeostatic, proinflammatory, anti-inflammatory, and reparative phenotypes to reset at a new homeostatic-like end point (81, 83, 106, 121). In the early post-MI proinflammatory environment, high concentrations of proinflammatory cytokines such as IL-1β and the generation of ECM fragments by MMPs induce an ECM-degrading fibroblast phenotype. Deposition of EDA-fibronectin, a common myofibroblast marker, activates myofibroblast differentiation in a feedforward manner (1, 119). In the later post-MI anti-inflammatory and reparative environments, growth factors such as transforming growth factor-β and the synthesis of new ECM induce an ECM-accumulating fibroblast phenotype (129).

A number of reviews have summarized current efforts to target post-MI ECM remodeling (26, 71, 81, 95, 110). Similar to inflammation, modulation of particular ECM proteins feeds forward to modify other aspects. To date, targeting specific MMPs has promise in the post-MI setting (15, 16, 51, 72, 111). Of interest, a collagen fragment generated post-MI by MMP-9 actually promotes wound healing by limiting dilation and promoting neovascularization when exogenously administrated after MI (73). The ECM, MMP, and TIMP environment is complex, and understanding how individual patterns fit into the entire LV effect, including cross talk among cell types, requires systems biology approaches to map the interplay (33, 34, 48, 72). More information on ECM remodeling is needed before approaches targeting ECM components can be translated.

FUTURE DIRECTIONS

There are four gaps to our understanding of how the inflammatory and ECM components interact in the MI wound-healing response. The four gaps are as follows: 1) harnessing the heterogeneity across cell subtypes to target most effective interventions, 2) linking molecular and cellular phenotypes to cell and tissue physiology to complete the knowledge maps, 3) figuring out how to best target endogenous and exogenous signaling pathways, and 4) translating findings to the clinic. In particular, sensors (marker subsets that define network status) are needed to better predict outcomes based on responses of the individual elements.

Heterogeneity Along the Response

While we have appreciated changes at the molecular and macrophage cell levels, molecular and cellular heterogeneity in polarization phenotypes along the MI time continuum is greater than previously realized. In addition to macrophage polarization (32, 35, 48, 75, 84), there is growing awareness for neutrophil and fibroblast polarization (41, 81, 83, 86, 91, 106, 107, 109, 118, 121). This is consistent with the polarization of the MI environment, which quickly transitions through phases, starting with the shift from homeostasis to inflammatory and then shifting to turn off inflammation and initiate repair. The final stage is a homeostatic-like end point, where a new homeostasis is maintained thereafter. To fill these gaps, there are three targets.

First, there is a need to map the molecular and cellular phenotypes, particularly during the early phase (i.e., first week in mice). Using omics approaches (transcriptomics, metabolomics, and proteomics) will allow the full molecular component to be defined, which will provide markers that best define tissue or cell phenotypes.

Second, there is a need for big data tools to harness the large data sets and distill the most informative details of the system (105). For example, identifying which genes, metabolites, proteins, and cell phenotypes provide the best diagnostic or prognostic information will focus attempts to provide mechanistic insights to those signaling pathways that most directly impact the system.

Third, there is a need to use computational models, which amplify the possible post-MI remodeling scenarios that can be studied and have the added benefit of limiting in vivo experiments. Computational modeling will also allow the introduction of multiple confounding factors, which more closely mimics the clinical scenario. While the possible permutations of intervention are extremely large, it is not possible to explore all using time-consuming in vivo approaches, which only evaluate a few factors at a time. Using knowledge maps to reduce the possible choices down to the most probable will focus and facilitate efforts.

Linking Molecular and Cellular Phenotypes to Cell and Tissue Physiology

Going beyond marker expression profiling is needed to build our understanding at the systems level and completely evaluate how the marker is influencing LV remodeling. To fill these gaps, there are two targets.

There is a need to understand the connections between molecular changes to cell physiology changes. Key cell physiology phenotypes that coordinate LV physiology phenotypes include activation of the inflammatory cascade for resident cardiac myocytes and fibroblasts, tissue clearance for neutrophils and macrophages, and ECM secretion for the fibroblast. Understanding cell activation and deactivation mechanisms is needed to better link molecular to cell to tissue pathophysiological responses to MI.

There is a need to understand the connections that link individual cell physiology to LV tissue physiology. Key tissue physiology phenotypes include dilation (elevated end-systolic and end-diastolic dimensions), wall thinning, and ejection fraction. While we know individual cell effects in culture, how these combine in the complex in vivo environment is still being investigated.

Targeting Endogenous and Exogenous Signaling Pathways

There is a need to know how to intervene in a way that will change the molecular, cellular, and tissue phenotypes in predictable ways and is amenable to therapeutic intervention. This will tell us if the key drivers of LV remodeling have been identified and if there is translational potential. To fill these gaps, there are two targets. For molecular phenotypes, we need to identify the triggers that stimulate particular phenotypic outputs. Presently, IL-1β is a key candidate for inflammation, whereas IL-10 is a key candidate for anti-inflammation (5, 36, 50, 56, 62, 101).

There is a need to identify the subnetworks downstream of the candidates that may be more amenable to inhibition. If the main driver has off-target actions, inhibiting it may not be the best approach. For example, MMP inhibition, even if specific for a particular MMP, may not be as specific as targeting one of its downstream substrates.

Translating From Animal Models to Humans

While most interventions target endogenous mediators, consideration should also be given to using exogenous systems as intervention targets. For example, IL-4 is not elevated post-MI, yet the LV has the capability to respond to IL-4, which is a known in vitro anti-inflammation stimulus (108). With any intervention strategy, consideration should be given to unintentional consequences, and a good experimental design will consider possible secondary effects. A number of guidelines have recently been established to increase rigor and reproducibility in ischemia and infarction animal models, with antibody use, and in cardiac physiology measurements, all of which have an application here (3, 68, 74).

CONCLUSIONS

In summary, a large amount of data has been assimilated over the past two decades to increase our knowledge of inflammation and ECM roles in post-MI remodeling. In particular, the macrophage and fibroblast are the key cell types that regulate the inflammatory and scar formation processes. With focused systems biology approaches, future efforts can effectively translate strategies to improve LV remodeling.

GRANTS

We acknowledge funding from National Institutes of Health Grants GM-104357, GM-114833, GM-115428, HL-051971, HL-075360, HL-105324, and HL-129823 and Biomedical Laboratory Research and Development Service of the Veterans Affairs Office of Research and Development Grant 5I01BX000505.

DISCLAIMERS

The content of this article is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health or the Veterans Administration.

DISCLOSURES

No conflicts of interest, financial or otherwise, are declared by the authors.

AUTHOR CONTRIBUTIONS

A.J.M., O.J.R., and M.L.L. edited and revised manuscript; A.J.M., O.J.R., and M.L.L. approved final version of manuscript; M.L.L. prepared figures; M.L.L. drafted manuscript.