Unified nexus of macrophages and maresins in cardiac reparative mechanisms

Abstract

Macrophages are immune-sensing “big eater” phagocytic cells responsible for an innate, adaptive, and regenerative response. After myocardial infarction, macrophages predominantly clear the deceased cardiomyocyte apoptotic or necrotic neutrophils to develop a regenerative and reparative program with the activation of the lipoxygenase-mediated maresin (MaR) metabolome at the site of ischemic injury. The specialized proresolving molecule and macrophage mediator in resolving inflammation, MaR-1, produced by human macrophages, has potent defining effects that limit polymorphonuclear neutrophil infiltration, enhance uptake of apoptotic PMNs, regulate inflammation resolution and tissue regeneration, and reduce pain. In addition to proresolving and anti-inflammatory actions, MaR-1 displays potent tissue regenerative effects in stroke and is an antinociceptive. Macrophages actively participate in the biosynthesis of bioactive MaR-2, which exhibits anti-inflammatory, proresolving, and atherosclerotic effects. A new class of macrophage-derived molecules, MaR conjugates in tissue regeneration, is identified that regulates phagocytosis and the repair and regeneration of damaged tissue. The presented review provides a current summary of the effect of MaR in resolution pathophysiology, with relevance to a cardiac repair program.—Jadapalli, J. K., Halade, G. V. Unified nexus of macrophages and maresins in cardiac reparative mechanisms.

Prolonged ischemia in cardiac tissue, triggering heart attack or myocardial infarction (MI), is one of the major causes of myocardium injury and thereby morbidity and mortality worldwide. In the United States, more than 7 million people have cardiac infarctions each year (1, 2). With the advancement of medical technology in the past few decades, the survival after MI has significantly improved because of early percutaneous revascularization therapy and drug treatment; however, more than 45% reperfused or nonreperfused patients develop heart failure related to collateral injury. Therefore, the current focus is the discovery of new therapeutic strategies to treat the conditions that can delay advanced heart failure (3–5). This paradigm provokes immunologists to focus on ischemic heart disease, as cardiologists are more interested in the intricacies of cardiomyocyte interaction with the immune system. Recent reports indicate that atherogenesis, atheroprogression, and atherosclerosis are not merely a lipid storage disorder, but is also driven by immune system dysfunction that initiates the convergence of cardiology and immunology (6).

Overactivation of the stress-related neurohormonal axis or narrowing of a coronary vessel results in an acutely diminished oxygen supply to the myocardium, which leads to necrosis, micro or macro MI, and acute inflammation, widely known as a heart attack or MI (3). As the human heart has limited regeneration capacity, restoration of the oxygen supply, in addition to infarct healing, is much dependent on the inflammatory and resolving responses, which are sterile (7). After MI, the innate inflammatory response induced by MI instigates cardiac repair processes toward either a resolving or nonresolving pathway (8). This inflammatory response results in predominant infiltration of peripheral neutrophils and monocytes that eventually differentiate into macrophages. These are involved at different stages of infarct healing, ultimately leading to scar formation, and harmonize remodeling to preserve cardiac function (9–11). On-time leukocyte entry failure (delayed “get-in” signal) at the site of injury or prolonged residual time at the site of infarction (delayed “get-out” signal) causes development of nonresolving, uncontrolled chronic inflammation in the cardiac repair program (12–15).

MACROPHAGES AND COMPLEX TERMINOLOGY

Ilya Ilyich Mechnikov, a 1908 Nobel Laureate, first discovered the phagocytic role of macrophages (“big eaters”) that are termed phagocytes (16). Macrophages are large myeloid cells that display stellate morphology and are characterized by the presence of pseudopodia, nonspecific esterase, and phagocytic granules, which give them a foamy appearance. Resident and monocyte-derived macrophages have different roles in almost every aspect of an organism’s pathobiology (Fig. 1, cardiac pathophysiology), from embryonic development, homeostasis, electrical conduction, cardiac repair, and regeneration to immune responses against pathogens (17). In adult mammals, macrophages are found in all tissues, with the highest presence in the microenvironment of solid tumors, where they display a comprehensive anatomic and functional diversity of surface markers, depending on the microenvironment (18). Macrophages play a vibrant and multitasking role in the innate immune response to injury, and, with the integral role of phagocytosis, macrophages also regulate several wound-healing pathways, including dismantling and removing cell debris, development of mature infarct scar, and angiogenesis (9, 19). In vitro, macrophage phenotypes are the classically activated M1 macrophages (proinflammatory type) and the alternatively activated M2 macrophages (anti-inflammatory type or reparative) (20–24). In a petri dish, macrophage exposure to IFN-γ and LPS leads to M1 (inflammatory) polarization, with cytotoxic and antitumoral properties, whereas M2 (reparative or resolving) macrophages display immunoregulatory effects. The M2 phenotype is subdivided into M2a, M2b, and M2c subtypes. These M2 macrophages polarize and acquire a different functional role in response to environment-derived signals. In particular, M2a is formed on exposure to IL-4 and -13 and M2b on combined exposure to TLR or IL-1R agonists and immune complexes and promotes type II responses and immunoregulatory functions, whereas M2c macrophages (induced by IL-10) are more related to suppression of immune responses and tissue remodeling (20, 25–27). In vivo, this heterogeneous classification is challenging because of the temporal dynamics of resident vs. infiltrating macrophages in wound healing. The tension between the host defense (proinflammatory) and reparative (proresolving) properties of the mononuclear phagocytes in infarcted myocardium requires caution against indiscriminate targeting of macrophages (28). In cardiac healing, the reparative macrophages are classified based on cluster of differentiation (CD)206 (mannose receptor), but the degree to which CD206 is relevant to human healing in heart failure pathology remains unclear. The infiltrating macrophage shows biphasic activation after MI, with the first-responder M1 macrophages showing an exclusively proinflammatory character based on gene expression profiling. However, the late-responder M2 macrophages show a mixed character, expressing high levels of both reparative and proinflammatory genes (29). First, when macrophages expand from their native environments and ultimately lose their tissue-specific gene expression, it becomes difficult for them to monitor changes and secondary activation, with diverse stimuli relevant to mediators present in the local milieu that regulate deviation in the M1/M2 spectrum. For instance, like macrophage colony-stimulating factor or granulocyte macrophage colony-stimulating factor, dependent macrophage activation network revealed considerable deviation from the M1/M2 axis making it even more complicated to differentiate the overlapping phenotypes and macrophage functional capacity. In the peritoneal domain, the macrophages comprise small and large cells. Large peritoneal macrophages disappear after infection or injury and are replaced by blood monocyte–derived small macrophages that express lower levels of CD11b (surface marker for monocytes, neutrophils, and macrophages) and F4/80 (macrophage marker) but express high levels of major histocompatibility complex (MHC)-II (derived from extracellular proteins) (30, 31). Thus, macrophages are multitasking, diversified cell types with a precise residential phenotype and diversified postinjury or -infection markers.

Figure 1.

Unified roles of macrophage chemistry in cardiac pathology. Macrophages facilitate the conduction, plaque progression, and regenerative mechanisms in cardiovascular disease. Likewise, macrophages and allied cells biosynthesize maresin (MaR) and tissue-regenerating factors in cardiac pathobiology to facilitate the resolution program.

Defining the clear margin between the M1/M2 macrophages by measuring their transcripts is also not always reliable because of the diversified local milieu of each organ—for instance, during augmentation or attenuation certain cell markers consider M1 or M2 as equivalent. In some cases, markers may break from the rule entirely. Macrophages of the same tissue have differential origins—that is, from the yolk sac or hematopoietic stem cells (HSCs) or both. For instance, microglia (brain, yolk sac), Kupffer cells (liver, yolk sac+HSCs), alveolar macrophages (lungs, HSCs), red pulp macrophages (spleen, HSCs+marginal), Langerhans cells (skin, yolk sac+HSCs), dermal macrophages (skin, HSCs), C-C chemokine receptor (CCR)2⁻ macrophages (heart, yolk sac+HSCs), CCR2⁺ macrophages (heart, HSCs), kidney macrophages (HSCs), peritoneal macrophages (HSCs), and LyC6⁺, Ly6C⁻ monocytes (blood, HSCs). They do not rely on specific marking, and that it makes them hard to categorize; also, macrophages have a tendency to switch from M1 to M2, making them even more difficult to distinguish (32–38). Another recent classification based on macrophage origins (monocyte vs. locally derived tissue residents), environmental stimuli (different organs and different stimuli during steady state and inflammation), and time (development, stages of inflammation, microenvironment, and aging). Murray et al. (39) suggested stimuli dependent terminology, although such methodology goes beyond the simple dichotomy suggested using M1/M2 model. Still, this futuristic approach has its inherent limitations.

ROLE OF MACROPHAGES IN MYOCARDIUM HEALING

Macrophages can be of embryonic origin or can be resident in many organ tissues in the steady state condition. Resident macrophages contribute to homeostasis and development, whereas in the setting of injury or infection, they coordinate tissue repair and regeneration processes, making them prominent candidates for therapeutic intervention (26). The disruption of homeostasis associated with ageing and the absence of inflammation or injury, within the myocardium and other organs, results not only in the recruitment of blood monocyte–derived macrophages but also in the permanent replacement of embryonically established resident macrophage populations (32, 40). This replacement is reflected by dynamic changes in the resident cardiac macrophage markers and decreasing cardiac macrophage self-renewal over time (39). However, during post-MI inflammation or after macrophage depletion, Ly6C^hi monocytes contribute to all 4 macrophage populations, whereas resident macrophages numerically expand through proliferation (32).

In infarcted hearts, the chemokine expression profile is modulated over time; thus, change in the edematous chemical gradient sequentially and actively recruits Ly6C^hi and Ly6C^lo monocytes via CCR2 and CX3C chemokine receptor (CX3CR)-1, respectively. Ly6C^hi monocytes control early (phase I) and exhibit phagocytic, proteolytic, and inflammatory functions. Ly6C^lo monocytes govern later (phase II), and have attenuated inflammatory properties, and express VEGF. Therefore Ly6C^hi monocytes digest damaged tissue, whereas Ly6C^lo monocytes promote healing via myofibroblast accumulation, angiogenesis, and deposition of collagen (28). In the heart, tissue-resident cardiac macrophages are marked as CX3CR1 cells that can be found throughout the myocardium (26). Transcriptional analysis of residential cardiac macrophages revealed that monocyte-derived macrophages coordinate cardiac inflammation while playing outmoded but minor roles in antigen sampling and efferocytosis (32). Recently, it was reported that CX3CR1 residential macrophages couple electrically to cardiomyocytes in the distal AV node via connexin-43 containing gap junctions. This process aids in depolarizing resting cardiomyocytes via connexin-43 by reducing their action potential upstroke velocity and overshoot, and aids early repolarization, altered conditions showed the irregular heartbeat condition may be one of the considerable factors leading to heart failure (17). The relative balance between pathologic inflammatory pathways and on time tissue reparative processes (physiologic inflammation) defines the trajectory of heart failure development (41). The questions remain unanswered of how macrophages respond to the organ-specific milieu and what the secretome of macrophages is that contribute to the feed-forward resolving and nonresolving inflammation in heart failure pathology. The age-related immunosenescence leads to alterations in their composition, gene expression, and function precede increased cardiac fibrosis and impairments in the inflammatory response typical of the aging heart (42). From a translational perspective, whether other metabolic disease pathologies alter macrophage function, secretome, and phenotype in aging, needs to be investigated (14, 43). In a recent study, Halade et al. (44) quantitated leukocyte populations in the splenic reservoir and infarcted LV and further determined the concentrations of specialized proresolving mediators that limit cardiac inflammation post-MI. This study clearly suggests that leukocytes mobilize from the spleen to the infarcted heart to produce resolvins, lipoxins, maresins, and protectins in the infarcted heart during the acute inflammatory response, suggesting that acute inflammatory response coincides with resolving response in cardiac healing. Furthermore, the depletion of macrophages correlated with the imperceptible levels of proresolving mediators. Thus, macrophages and the macrophage interactome with innate leukocyte players are essential for the biosynthesis of bioactive mediators and cardiac healing (Fig. 2).

Figure 2.

Biosynthesis of MaR and its metabolites coordinated by the macrophage–neutrophil alliance. Shown is the overall representative sequence of biosynthesis of both MaR-1 and -2, the MCTRs produced in macrophages, and the macrophage–neutrophil interactome formed from MaR-1. DPEP, dipeptidase; GGT, γ-glutamyltransferase; GSTM4, GST-μ4; LTC4S, leukotriene C4 synthase.

ORCHESTRATION OF MONOCYTES AND MACROPHAGES INTO THE ISCHEMIC HEART

In the normal steady state, most cardiac macrophages are embryonically derived, and a smaller contribution is derived from HSCs. At the time of cardiac ischemic injury or inflammation, Ly6C^hi monocytes migrate into tissue and undergo proliferation, and compete with resident cardiac macrophages proliferation to re-establish the steady state in the macrophage pool. The pool of resident cardiac macrophages has undergone ‘‘ontological reprogramming’’ as a result and now is primarily composed of adult-derived macrophages. In conditions such as the absence of circulating blood monocytes (as in CCR2⁻mice), resident cardiac macrophages are fully capable of rebuilding steady-state levels solely through in-situ proliferation (45–48). At baseline, the adult heart contained 2 immunosurveillance resident macrophages (MHC-II^loCCR2⁻ and MHC−II^hiCCR2⁻) subsets, 2 monocyte-derived macrophages (MHC-II^hiCCR2⁺), and (MHC-II^loCR2⁺) subsets. The neonatal heart included one resident macrophage (MHC-II^loCR2⁻) and 1 monocyte-derived (MHC-II^loCCR2⁺) subset. In response to injury, the neonatal heart selectively expanded the number of MHC-II^loCCR2⁻ macrophages and did not recruit additional CCR2⁺ monocytes. In contrast, the injured adult heart selectively recruited monocytes and MHC-II^hiCCR2⁺ monocyte-derived macrophages. An additional surface marker such as tyrosine-protein kinase Mer and cluster of differentiation-64 confirms that MHC-II^loCR2⁻, MHC-II^hiCCR2⁻, and MHC-II^hiCCR2⁺ cells represent macrophages, whereas MHC-II^loCCR2⁺ cells are monocytes in both the neonatal and adult heart (49). A flow cytometry–based quantitative approach or qualitative immunostaining revealed the rapid expansion of macrophages in the neonatal heart that resolved within 5 d after injury. In contrast, monocytes infiltrated the adult heart after injury and transitioned to macrophages progressively (50, 51). Together, these data indicate that, in response to injury, the population of resident CCR2⁻ reparative cardiac macrophages expands in the neonatal heart, whereas the adult heart recruits CCR2⁺ monocyte-derived macrophages (51). The consequences of this difference are regeneration vs. repair. The neonatal heart has a unique and transient ability for myocardial regeneration after mild-to-moderate cardiac injury by surgical apical resection, cryoablation, or MI (50–57). The amplified regenerative response can be supported by a resolution of inflammation with the biosynthesis of resolving lipids and reactive cardiomyocyte dedifferentiation and redifferentiation (56). The regenerative response in newborn mice is marked with angiogenesis, which is scar formation in adults. Our knowledge is still limited, but initial research in this model has led to the understanding that myocardial regeneration in the neonatal heart is dependent on diversified macrophage phenotypes and transcription profiles (32, 54, 57). When a myocardial resection was performed, fibrin clots sealed the resection site and created a matrix for recruiting myeloid cells, and macrophages secreted growth factors and paracrine factors that promote regeneration (58–61). Inhibition of myocardial regeneration by anti-inflammatory glucocorticoids was seen in zebrafish (60). Thus, the mechanism of immune system control in myocardial regeneration and repair is an important area of research that has therapeutic implications in human cardiac healing.

MACROPHAGES AND REPARATIVE MECHANISMS

Aurora and colleagues (54) were the first to show with the quantitative approach of flow cytometry that macrophages are essential for neonatal heart regeneration. In the intact heart, without MI, macrophages are significantly more abundant at d 1 after birth than on d 14. Seven days after birth, post-MI d 1 macrophages appear more abundantly overall in the heart and are distributed throughout the myocardium, both at the infarct and in remote areas, whereas on d 14 after birth, there were fewer post-MI d 1 macrophages, and they were almost exclusively localized to the infarct zone (54). The regenerative capacity of the neonatal heart was eliminated by macrophage depletion (62). Macrophage depletion did not reduce cardiomyocyte proliferation but was associated with impaired myocardial vascularization and regeneration (54). These findings indicate the importance of macrophages in the regeneration of the neonatal heart and suggest that newborn (d 1) mice are rich in M2 macrophages, with high regenerative angiogenic potential, distinctive from the profibrotic and scar-forming activities of adult monocytes/macrophages (54). These fundamental insights have been gained from cardiac regeneration in fish, salamander, and neonatal mouse; are translatable; and have paved the way to study cardiac repair in humans (54, 63). As in newborn mice, newborn humans with signs of ischemic injury or heart attack heal faster than adult humans; the mechanism that facilitates this healing is unclear (64).

The neonatal heart is responsive to cardiac inflammation with the progression of MHC-II^loCCR2⁻ embryo-derived macrophages (51). These macrophages have superior phagocytic, angiogenic, and reparative properties that enhance the regenerative capacity of the neonatal heart (65). In contrast, adult mice respond with the recruitment of inflammatory MHC-II^hiCCR2⁺ monocytes that replace the reparative resident CCR2⁻ macrophages and lead to adverse cardiac remodeling and dysfunction. Inhibiting the accumulation of CCR2⁺ monocytes in the injured myocardium preserves reparative cardiac MHC-II^lo macrophages in the adult heart and improves coronary angiogenesis and reduced inflammation (51).

Han et al. (52) recently showed that cardiomyocyte proliferation induced by acute inflammation after apical resection results in infiltration of Ly6G inflammatory leukocytes. When the regenerative response in neonatal mouse heart was blocked after immunosuppression with dexamethasone, it showed that IL-6 is essential for cardiomyocyte proliferation. The same results were observed when deletion of the cardiomyocyte-specific signal transducer and activation of transcription-3 caused the downregulation of IL-6 signaling and decreased reactive cardiomyocyte proliferation after apical resection (52). These results are in harmony with recent profiling studies that indicated that cardiomyocyte signal transducer and activation of transcription-3 mediate injury-induced myocyte proliferation and heart regeneration in the adult zebrafish (60, 66) and the neonatal mouse heart (67). This study suggests that the immune system stimulates the regenerative response in the mouse heart.

Aging is also a primary factor in MI-induced heart failure pathology superimposed on additional risk factors, such as obesity, insulin resistance, and hypertension (8, 43). A recent study revealed that aging impairs the phagocytotic capacity of brain macrophages (microglia) (68). In addition to aging, macrophages exhibit diminished ability to phagocytose apoptotic cells, indicating systemic metabolic dysfunction, which is often seen in old obese mice (69). This effect shows that diminished macrophage phagocytotic activity with aging results in failed resolution and accumulation of damage-associated molecular patterns in aged animals (70) and confirms that aging-related immunosenescence leads to a decreased immunosurveillance capacity of macrophages and thereby ultimately affects immunoregulatory and tolerogenic mechanisms of the immune system in aging (71, 72).

MARESINS

Macrophages also participate in the biosynthesis of specialized proresolving mediators (SPMs), which are bioactive autacoids with both anti-inflammatory and proresolving properties derived from fatty acid immunometabolism. SPMs are enzymatically biosynthesized from essential fatty acids, with different stereochemistries. Recently, a new family of macrophage-derived mediators was identified and named maresins (MaRs) for macrophage mediator resolving inflammation (73). These newly identified bioactive lipid mediators display potent anti-inflammatory and proresolving actions inhibiting neutrophil infiltration in vivo and stereoselectively stimulating macrophage phagocytosis and efferocytosis to enhance the clearance of inflammation without affecting the innate response (74, 75). MaRs are a family of bioactive lipid mediators biosynthesized by activated macrophages from docosahexaenoic acid (DHA) via human 12-lipoxygenase (LOX) (75). Chemically, MaR-1 is 7R,14S, dihydroxy-docosa-4Z,8E,10E,12Z,16Z,19Z-hexaenoic acid, and is the first member of the MaR family to be identified. The complete stereochemistry of MaR-1 has been established in several in vitro and in vivo models. MaR-1 also possesses potent tissue regenerative and antinociceptive effects (74). Recently a new bioactive MaR metabolome was identified as 13R,14S dihydroxy-4Z,7Z,9E,11E,16Z,19Z-hexaenoic acid (Mar-2), which displays anti-inflammatory, proresolving, and antiatherosclerotic effects (Table 1) (76, 77).

TABLE 1.

Various roles of MaRs

Bioactive resolution	Action	Model system/cell type	Reference
MaR-1 (7R,14S-dihydroxy-docosa-4Z,8E,10E,12Z,16Z,19Z-hexaenoic acid)	↓ PMN infiltration, ↑ phagocyte clearance	Murine peritonitis/lung	110
	Antinociceptive	Mouse/pain	111
	↓ PMN infiltration and cytokine expression as well as NF-κB activation	Mouse/colitis	112
	↑ Tissue regeneration in Planaria	Planaria/regeneration	90
	Phenotype switch	Human macrophages	75
	↑ Apoptotic PMNs	Human macrophages	90
	↑ Phagocytosis and efferocytosis	Human macrophages	73, 113
	↓ Cytokine release with organic dust	Human bronchial epithelial cells	114
	Identified in synovial fluid extracts	Human/rheumatoid arthritis	115
MaR-2 (13R,14S-dihydroxy-4Z,7Z,9E,11Z,16Z,19Z-DHA)	↓ PMN infiltration	Human macrophages	76
	↑ Phagocytosis and efferocytosis	Human macrophages	76, 116

In macrophages, MaR-1 biosynthesis (Fig. 2) is initiated by 12-LOX from DHA, producing 14S-hydroperoxydocosa-4Z,7Z,10Z,12E,16Z,19Z-hexaenoic acid, the hydroperoxy intermediate, which undergoes further conversion via enzymatic 13(14)-epoxidation. This epoxide intermediate is hydrolyzed enzymatically via an acid-catalyzed nucleophilic attack by water at carbon 7, resulting in the introduction of a hydroxyl group at that position and a double bond rearrangement to form the stereochemistry of bioactive MaR1 (7R,14S-dihydroxy-docosa-4Z,8E,10E,12Z,16Z,19Z-hexaenoic acid) with potent proresolution properties (74, 75). The 13S,14S-epoxy-MaR the intermediate in MaR1 synthesis is also the precursor of MaR2. This is the product of DHA biosynthesize by 12-LOX to produce the 14S-hydroperoxide that is converted to the 13S,14S-epoxy-MaR and finally converted by a soluble epoxide hydrolase into MaR2 (78).

MARESINS ROLE IN THE CLEARANCE OF ATHEROSCLEROTIC INFLAMMATION

Atherogenesis and atheroprogression have long been regarded as signs of chronic low-grade inflammation. However, modern concepts also view atherosclerosis as a low-grade chronic consequence of failed resolution (79, 80). Resolution is crucially orchestrated by a switch from inflammatory to resolving by specialized lipid mediators in the affected tissue (81), an event that may be impaired during atheroprogression (82, 83). In contrast to acute inflammatory situations, atherogenesis and atheroprogression are characterized by the persistence of the initiating stimulus, including hypertension, hyperlipidemia, and oxidative stress.

Macrophages in atherosclerotic lesions actively participate in lipoprotein ingestion, and their accumulation gives rise to the formation of foam cells filled with lipid droplets (Table 1) (84). Accumulation of foam cells in the lesions results in lipid storage and growth of atherosclerotic plaque. This effect was determined by the increase in several M1 macrophages in progressing plaques, and a greater number of M2 macrophages present in regressing plaques, where they were involved in tissue repair and remodeling (85). Macrophages are highly heterogeneous, plastic cells that can rapidly adapt their phenotype/function and change the chemical milieu to facilitate resolution. During the inflammatory phase, they phagocytose necrotic cells and intensify inflammation by releasing proteases, proinflammatory cytokines/chemokines, and reactive oxygen species. Later these macrophages, in the reparative phase, undergo transformation and exert profibrotic functions by activating collagen production and neovascularization. A timely resolution of each phase is crucial because spontaneous, prolonged inflammation and fibrosis may be detrimental and determine whether progression to heart failure or repair of cardiac tissue occurs (86).

The milieu of progressive atherosclerotic lesions increases the synthesis of leukotriene (LT)B₄ driven by 5-LOX (79, 80). Also, the local cytokine environment favors the accumulation of inflammatory over reparative macrophages (87). LTB₄ and prostaglandin (PG)E₂ are produced in abundance at sites of inflammation (88–90), suggesting that their production by classic inflammatory macrophages favors the synthesis of inflammatory lipid mediators, in comparison to their reparative macrophage counterparts (91).

Systematically, LTB₄ potentiates atherosclerosis by increasing the expression of fatty acid translocase/CD36 and C-C motif chemokine ligand-2, thereby inducing a positive feedback loop to recruit leukocytes (92, 93). PGE₂ and PGD₂ have been reported to have a dual, paradoxical role, functioning as a proinflammatory and proresolving mediators, depending on their location (94). In the context of atherosclerosis, PGE₂ is associated with disease progression, mediated by activation of platelets (89) and, consequently, monocyte recruitment (95). In general, lipid mediators are mostly produced by a myeloid cells, where the same cell or combination of cells can often be a source of different classes of lipid mediators, depending on the substrate and temporal injury setting. Under inflammatory conditions, neutrophils (96, 97) and macrophages (91, 98) preferentially synthesize LTB₄ and PGE₂ over MaR-1 and resolvin (Rv)-D-2. Also, endothelial (99) and epithelial cells (100) add to the pool of LTB₄ and PGE₂ production. Upon a switch toward resolution, macrophages (reparative) and neutrophils (possibly second wave neutrophils) also synthesize MaR-1 (75) and RvD-2 (91, 101). However, during chronic inflammation, such as atherosclerosis it is likely that, overall, the inflammatory environment prevails, favoring the synthesis of inflammatory lipid mediators. MaR-1 and RvD-2 act on lesional macrophages, inducing phenotype changes toward a reparative phenotype. These reparative macrophages reduce the inflammatory environment by enhancing secretion of TGF-β and decreasing the concentrations of secreted IL-6 and TNF-α. The resolving milieu aids in reducing macrophage accumulation and smaller necrotic core sizes and instructs smooth muscle cells to produce collagen, resulting in stable thickening of the fibrous cap and increased plaque stability, eventually resulting in a halt of atheroprogression.

MACROPHAGES AND MaRs NEXUS

Recently, two new macrophage–neutrophil interactomes that are formed from MaR-1 when metabolized in macrophages and neutrophils—14-oxo-MaR-1 and 22-OH-MaR-1—were identified, respectively. Incubation of human macrophages with MaR-1 biosynthesize 14-oxo-MaR-1, reaching a maximum level at 4 h, whereas the primary product with human neutrophils was 22-OH-MaR-1, reaching a maximum at 24 h (102) (Fig. 2). Each of these newly identified molecules retained at low concentrations the bioactions of their parent SPMs—that is, MaR-1, with human macrophages. 22-OH-MaR-1 gave a biphasic dose response and up-regulated E. coli phagocytosis by human macrophages. This response is suggestive of the engagement of a high- and a low-affinity receptor by 22-OH-MaR-1 as observed for other proresolving mediators including RvD-1 (103). Also, both MaR-1 and its metabolite 22-OH-MaR-1 act at the human leukotriene-1 receptor and affect LTB₄ signaling. These results recognize heretofore unidentified molecules and establish unique functions for the macrophage-derived MaR-1 bioactive metabolome (102, 104).

Macrophages also produce a family of bioactive peptide-conjugated mediators called MaR conjugates in tissue regeneration (MCTRs). These recently identified innovative MCTRs regulate the mechanisms in inflammation resolution, as well as tissue regeneration establishing links between local inflammatory exudates and tissue regeneration via newly identified chemical signals (105). MCTR compounds are produced from the 13(S),14S-epoxide MaR intermediate during the MaR biosynthesis. This epoxide intermediate is then enzymatically converted to MCTRs. MCTR-1 (13R-glutathionyl, 14S-hydroxy-4Z,7Z,9E,11E,13R,14S,16Z,19Z-DHA) is biosynthesized in the presence of leukotriene C4 synthase and γ-glutamyltransferase-μ4 in human macrophages. γ-Glutamyl transferase aids in the conversion of MCTR-1 to MCTR2 (13R-cysteinylglycinyl, 14S-hydroxy-4Z,7Z,9E,11E,13R,14S,16Z,19Z-DHA), which is then further converted to MCTR-3 (13R-cysteinyl,14S-hydroxy-4Z,7Z,9E,11E,13R,14S,16Z,19Z-DHA) by a dipeptidase. The direct emphasis on the MCTR biosynthetic pathway and the enzymes that catalyze these reactions, along with their relation with MaRs in human macrophages are unclear (106). The simplified and overall nexus between the MaR and MCTR metabolome and macrophages is demonstrated in Fig. 2.

MaRs AS THERAPEUTIC TARGETS

The resolution of inflammation is an active process orchestrated by lipid SPMs. Uncontrolled and chronic inflammation facilitates tissue destruction and impedes healing. With the significant roles of MaR-1 in regulating phagocytic functions, tissue regeneration, and wound healing properties, it can be used as a therapeutic agent, as it impedes the rate of inflammation during atherosclerosis by limiting the preferential synthesis of LTB₄ by inhibiting the production of PGE₂. MaR-1 can also be employed as a therapeutic target as MCTR-1, -2 and -3 (Table 2) conjugates derived from MaR-1 counter both vascular and negative inotropic actions that are stimulated by LTD₄, by blocking the MK571 receptor in tissue regeneration (107).

TABLE 2.

Different actions of MCTRs in inflammation-resolution as well as tissue regeneration

Name of the action	Order of potency between MCTRs
Stimulating tissue regeneration	MCTR3 ≈ MCTR2 > MCTR1
Macrophage phagocytic responses	MCTR3 > MCTR1 > MCTR2
Resolution of live bacterial infections	MCTR3 ≥ MCTR2 > MCTR1
Controlling exudate eicosanoids during infections	MCTR1 > > MCTR3 > MCTR2

Data from ref. 106.

MaR-1 is effective in controlling inflammation in periodontal infections with unresolved inflammation and leukocyte-mediated tissue destruction (108), and restoring the reparative function of nonhealing diabetic wounds impaired the macrophages (109). MaR-1 has also been tested in the treatment of diabetic nephropathy by inhibiting nucleotide binding and oligomerization domain-like receptor family pyrin domain–containing-3 and controls the inflammation rate in renal failure (110).

FUTURE PROSPECTIVE

What is first in cardiac healing and repair after ischemic injury? Is it inflammation, or is it a resolution of inflammation, or do both overlap in a time-dependent manner to set the stage for healing program of myocardium after MI (44)? In response to injury, infection, or stress, an acute inflammatory response primarily involves early tissue edema within seconds to minutes followed by infiltration of neutrophils within minutes to hours and later mononuclear cells, some of which transition at maturity to macrophages. During host-mediated acute inflammation that resolves and returns the infarcted cardiac tissue to homeostasis by formation of SPMs, such as lipoxins, Rvs, protectins, and MaRs (44). The main hallmark of chronic inflammation (particularly in aging marked with decreased biosynthesis of SPMs) is when acute inflammation fails to resolve, or excessive inflammation leads to bystander tissue injury. However, the defective or delayed resolution also may result in chronic inflammation, which relates to other human diseases. Resolution defects may arise because of one of the following reasons: decreased SPM formation, ineffective LOX, inhibition of SPM sites of action, and perhaps genetic variation or different pharmacologic treatment that interferes with resolution physiology. Identification of the mediators and mechanisms underlying the resolution and regulation of inflammation will provide new insights into the pathobiology of human disease and lead to the development of novel therapeutic approaches to such diseases as atheroprogression characterized by excess or chronic inflammation.

SUMMARY

Several lines of evidence point out that inflammation and resolution of inflammation are primordial and inherent components of the wound-healing mechanism, including that for cardiac ischemic injury. Now, it is time to decode the therapeutic potential of MaR biomolecules involved in the resolution process with the inbuilt endogenously enzymatic process, to amplify the resolution program, identify the factors that contribute to resolution deficit, impair MaR formation in response to injuries such as aging, and test whether restoration of these molecules improves myocardial healing in a translational prospective. Therefore, a detailed study of resolution biology with major emphasis on comprehensive molecular and cellular mechanisms is necessary in acute and chronic heart failure pathology. Furthermore, a precise focus of human macrophage plasticity and function in the steady-state condition or in a setting of injury could guide the development of macrophage-derived molecules in patients with ischemic cardiac diseases. Finally, we should test in the clinical setting whether a single MaR molecule or a combination of specialized proresolving molecules is necessary to accelerate a resolution program without affecting the innate response to control major ischemic disease pathology and to delay heart failure.

Glossary

CCR2

C-C chemokine receptor

CD

cluster of differentiation

CX3CR

CX3C chemokine receptor

DHA

docosahexaenoic acid

HSC

hemapoietic stem cell

LOX

lipoxygenase

LT

leukotriene

MaR

maresin

MCTR

maresin conjugates in tissue regeneration

MHC

major histocompatibility complex

MI

myocardial infarction

PG

prostaglandin

PMN

polymorphonuclear neutrophil

Rv

resolvin

SPM

specialized proresolving mediator

AUTHOR CONTRIBUTIONS

J. K. Jadapalli wrote the manuscript draft based on the outline and comments on a series of drafts; and G. V. Halade designed the review scheme and edited, revised, and approved the submission to the journal.