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Although some studies have suggested that macrophages may secrete structural collagens, and convert to fibroblast-like cells, macrophage to fibroblast transdifferentiation in infarcted and remodeling hearts remains controversial. Our study uses linage tracing approaches and single cell transcriptomics to examine whether macrophages undergo fibroblast conversion, and to characterize the extracellular matrix expression profile of myeloid cells in myocardial infarction. To examine whether infarct macrophages undergo fibroblast conversion, we identified macrophage-derived progeny using the inducible CX3CR1CreER mice crossed with the PDGFRαEGFP reporter line for reliable fibroblast identification. The abundant fibroblasts that infiltrated the infarcted myocardium after 7 and 28 days of coronary occlusion were not derived from CX3CR1+ macrophages. Infarct macrophages retained myeloid cell characteristics and did not undergo conversion to myofibroblasts, endothelial or vascular mural cells. Single cell RNA-seq of CSF1R+ myeloid cells harvested from control and infarcted hearts showed no significant expression of fibroblast identity genes by myeloid cell clusters. Moreover, infarct macrophages did not express significant levels of genes encoding structural collagens. However, infarct macrophage and monocyte clusters were the predominant source of the fibrogenic growth factors Tgfb1 and Pdgfb, and of the matricellular proteins Spp1/Osteopontin, Thbs1/Thrombospondin-1, Emilin2, and Fn1/fibronectin, while expressing significant amounts of several other matrix genes, including Vcan/versican, Ecm1 and Sparc. ScRNA-seq data suggested similar patterns of matrix gene expression in human myocardial infarction. In conclusion, infarct macrophages do not undergo fibroblast or myofibroblast conversion and do not exhibit upregulation of structural collagens but may contribute to fibrotic remodeling by producing several fibrogenic matricellular proteins.
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BACKGROUND: Cardiac fibroblast activation contributes to adverse remodeling, fibrosis, and dysfunction in the pressure-overloaded heart. Although early fibroblast TGF-ß (transforming growth factor-ß)/Smad (small mother against decapentaplegic)-3 activation protects the pressure-overloaded heart by preserving the matrix, sustained TGF-ß activation is deleterious, accentuating fibrosis and dysfunction. Thus, endogenous mechanisms that negatively regulate the TGF-ß response in fibroblasts may be required to protect from progressive fibrosis and adverse remodeling. We hypothesized that Smad7, an inhibitory Smad that restrains TGF-ß signaling, may be induced in the pressure-overloaded myocardium and may regulate fibrosis, remodeling, and dysfunction. METHODS: The effects of myofibroblast-specific Smad7 loss were studied in a mouse model of transverse aortic constriction, using echocardiography, histological analysis, and molecular analysis. Proteomic studies in S7KO (Smad7 knockout) and overexpressing cells were used to identify fibroblast-derived mediators modulated by Smad7. In vitro experiments using cultured cardiac fibroblasts, fibroblasts populating collagen lattices, and isolated macrophages were used to dissect the molecular signals responsible for the effects of Smad7. RESULTS: Following pressure overload, Smad7 was upregulated in cardiac myofibroblasts. TGF-ß and angiotensin II stimulated fibroblast Smad7 upregulation via Smad3, whereas GDF15 (growth differentiation factor 15) induced Smad7 through GFRAL (glial cell line-derived neurotrophic factor family receptor α-like). MFS7KO (myofibroblast-specific S7KO) mice had increased mortality, accentuated systolic dysfunction and dilative remodeling, and accelerated diastolic dysfunction in response to transverse aortic constriction. Increased dysfunction in MFS7KO hearts was associated with accentuated fibrosis and increased MMP (matrix metalloproteinase)-2 activity and collagen denaturation. Secretomic analysis showed that Smad7 loss accentuates secretion of structural collagens and matricellular proteins and markedly increases MMP2 secretion. In contrast, Smad7 overexpression reduced MMP2 levels. In fibroblasts populating collagen lattices, the effects of Smad7 on fibroblast-induced collagen denaturation and pad contraction were partly mediated via MMP2 downregulation. Surprisingly, MFS7KO mice also exhibited significant macrophage expansion caused by paracrine actions of Smad7 null fibroblasts that stimulate macrophage proliferation and fibrogenic activation. Macrophage activation involved the combined effects of the fibroblast-derived matricellular proteins CD5L (CD5 antigen-like), SPARC (secreted protein acidic and rich in cysteine), CTGF (connective tissue growth factor), ECM1 (extracellular matrix protein 1), and TGFBI (TGFB induced). CONCLUSIONS: The antifibrotic effects of Smad7 in the pressure-overloaded heart protect from dysfunction and involve not only reduction in collagen deposition but also suppression of MMP2-mediated matrix denaturation and paracrine effects that suppress macrophage activation through inhibition of matricellular proteins.
Assuntos
Fibrose , Camundongos Knockout , Miofibroblastos , Proteína Smad7 , Remodelação Ventricular , Animais , Proteína Smad7/metabolismo , Proteína Smad7/genética , Camundongos , Miofibroblastos/metabolismo , Miofibroblastos/patologia , Células Cultivadas , Camundongos Endogâmicos C57BL , Fator de Crescimento Transformador beta/metabolismo , Masculino , Fibroblastos/metabolismo , Fibroblastos/patologia , Transdução de Sinais , Miocárdio/metabolismo , Miocárdio/patologiaRESUMO
The adult mammalian heart has limited endogenous regenerative capacity and heals through the activation of inflammatory and fibrogenic cascades that ultimately result in the formation of a scar. After infarction, massive cardiomyocyte death releases a broad range of damage-associated molecular patterns that initiate both myocardial and systemic inflammatory responses. TLRs (toll-like receptors) and NLRs (NOD-like receptors) recognize damage-associated molecular patterns (DAMPs) and transduce downstream proinflammatory signals, leading to upregulation of cytokines (such as interleukin-1, TNF-α [tumor necrosis factor-α], and interleukin-6) and chemokines (such as CCL2 [CC chemokine ligand 2]) and recruitment of neutrophils, monocytes, and lymphocytes. Expansion and diversification of cardiac macrophages in the infarcted heart play a major role in the clearance of the infarct from dead cells and the subsequent stimulation of reparative pathways. Efferocytosis triggers the induction and release of anti-inflammatory mediators that restrain the inflammatory reaction and set the stage for the activation of reparative fibroblasts and vascular cells. Growth factor-mediated pathways, neurohumoral cascades, and matricellular proteins deposited in the provisional matrix stimulate fibroblast activation and proliferation and myofibroblast conversion. Deposition of a well-organized collagen-based extracellular matrix network protects the heart from catastrophic rupture and attenuates ventricular dilation. Scar maturation requires stimulation of endogenous signals that inhibit fibroblast activity and prevent excessive fibrosis. Moreover, in the mature scar, infarct neovessels acquire a mural cell coat that contributes to the stabilization of the microvascular network. Excessive, prolonged, or dysregulated inflammatory or fibrogenic cascades accentuate adverse remodeling and dysfunction. Moreover, inflammatory leukocytes and fibroblasts can contribute to arrhythmogenesis. Inflammatory and fibrogenic pathways may be promising therapeutic targets to attenuate heart failure progression and inhibit arrhythmia generation in patients surviving myocardial infarction.
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Infarto do Miocárdio , Humanos , Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/patologia , Animais , Transdução de Sinais , Regeneração , Mediadores da Inflamação/metabolismo , Miocárdio/metabolismo , Miocárdio/patologiaRESUMO
The adult mammalian heart contains a large population of pericytes that play important roles in homeostasis and disease. In the normal heart, pericytes regulate microvascular permeability and flow. Myocardial diseases are associated with marked alterations in pericyte phenotype and function. This review manuscript discusses the role of pericytes in cardiac homeostasis and disease. Following myocardial infarction (MI), cardiac pericytes participate in all phases of cardiac repair. During the inflammatory phase, pericytes may secrete cytokines and chemokines and may regulate leukocyte trafficking, through formation of intercellular gaps that serve as exit points for inflammatory cells. Moreover, pericyte contraction induces microvascular constriction, contributing to the pathogenesis of 'no-reflow' in ischemia and reperfusion. During the proliferative phase, pericytes are activated by growth factors, such as transforming growth factor (TGF)-ß and contribute to fibrosis, predominantly through secretion of fibrogenic mediators. A fraction of pericytes acquires fibroblast identity but contributes only to a small percentage of infarct fibroblasts and myofibroblasts. As the scar matures, pericytes form a coat around infarct neovessels, promoting stabilization of the vasculature. Pericytes may also be involved in the pathogenesis of chronic heart failure, by regulating inflammation, fibrosis, angiogenesis and myocardial perfusion. Pericytes are also important targets of viral infections (such as SARS-CoV2) and may be implicated in the pathogenesis of cardiac complications of COVID19. Considering their role in myocardial inflammation, fibrosis and angiogenesis, pericytes may be promising therapeutic targets in myocardial disease.
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Infarto do Miocárdio , Pericitos , Pericitos/fisiologia , Humanos , Infarto do Miocárdio/fisiopatologia , Fibrose , COVID-19 , Miocárdio/patologia , Insuficiência Cardíaca/fisiopatologia , Cardiomiopatias/fisiopatologia , Animais , Miofibroblastos/fisiologiaRESUMO
INTRODUCTION: Myocardial fibrosis accompanies most cardiac conditions and can be reparative or maladaptive. Transforming Growth Factor (TGF)-ß is a potent fibrogenic mediator, involved in repair, remodeling, and fibrosis of the injured heart. AREAS COVERED: This review manuscript discusses the role of TGF-ß in heart failure focusing on cellular mechanisms and therapeutic implications. TGF-ß is activated in infarcted, remodeling and failing hearts. In addition to its fibrogenic actions, TGF-ß has a broad range of effects on cardiomyocytes, immune, and vascular cells that may have both protective and detrimental consequences. TGF-ß-mediated effects on macrophages promote anti-inflammatory transition, whereas actions on fibroblasts mediate reparative scar formation and effects on pericytes are involved in maturation of infarct neovessels. On the other hand, TGF-ß actions on cardiomyocytes promote adverse remodeling, and prolonged activation of TGF-ß signaling in fibroblasts stimulates progression of fibrosis and heart failure. EXPERT OPINION: Understanding of the cell-specific actions of TGF-ß is necessary to design therapeutic strategies in patients with myocardial disease. Moreover, to implement therapeutic interventions in the heterogeneous population of heart failure patients, mechanism-driven classification of both HFrEF and HFpEF patients is needed. Heart failure patients with prolonged or overactive fibrogenic TGF-ß responses may benefit from cautious TGF-ß inhibition.
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Insuficiência Cardíaca , Infarto do Miocárdio , Humanos , Insuficiência Cardíaca/tratamento farmacológico , Fator de Crescimento Transformador beta/metabolismo , Volume Sistólico , Infarto do Miocárdio/terapia , Fibroblastos/metabolismo , Fibrose , Miocárdio/patologiaRESUMO
Macrophages sense changes in the extracellular matrix environment through the integrins and play a central role in regulation of the reparative response after myocardial infarction. Here we show that macrophage integrin α5 protects the infarcted heart from adverse remodeling and that the protective actions are associated with acquisition of an angiogenic macrophage phenotype. We demonstrate that myeloid cell- and macrophage-specific integrin α5 knockout mice have accentuated adverse post-infarction remodeling, accompanied by reduced angiogenesis in the infarct and border zone. Single cell RNA-sequencing identifies an angiogenic infarct macrophage population with high Itga5 expression. The angiogenic effects of integrin α5 in macrophages involve upregulation of Vascular Endothelial Growth Factor A. RNA-sequencing of the macrophage transcriptome in vivo and in vitro followed by bioinformatic analysis identifies several intracellular kinases as potential downstream targets of integrin α5. Neutralization assays demonstrate that the angiogenic actions of integrin α5-stimulated macrophages involve activation of Focal Adhesion Kinase and Phosphoinositide 3 Kinase cascades.
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Integrina alfa5 , Infarto do Miocárdio , Camundongos , Animais , Integrina alfa5/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Infarto do Miocárdio/genética , Infarto do Miocárdio/metabolismo , Macrófagos/metabolismo , Camundongos Knockout , RNA/metabolismoRESUMO
At least seven cell death programs are activated during myocardial infarction (MI), but which are most important in causing heart damage is not understood. Two of these programs are mitochondrial-dependent necrosis and apoptosis. The canonical function of the pro-cell death BCL-2 family proteins BAX and BAK is to mediate permeabilization of the outer mitochondrial membrane during apoptosis allowing apoptogen release. BAX has also been shown to sensitize cells to mitochondrial-dependent necrosis, although the underlying mechanisms remain ill-defined. Genetic deletion of Bax or both Bax and Bak in mice reduces infarct size following reperfused myocardial infarction (MI/R), but the contribution of BAK itself to cardiomyocyte apoptosis and necrosis and infarction has not been investigated. In this study, we use Bak-deficient mice and isolated adult cardiomyocytes to delineate the role of BAK in the pathogenesis of infarct generation and post-infarct remodeling during MI/R and non-reperfused MI. Generalized homozygous deletion of Bak reduced infarct size â¼50% in MI/R in vivo, which was attributable primarily to decreases in necrosis. Protection from necrosis was also observed in BAK-deficient isolated cardiomyocytes suggesting that the cardioprotection from BAK loss in vivo is at least partially cardiomyocyte-autonomous. Interestingly, heterozygous Bak deletion, in which the heart still retains â¼28% of wild type BAK levels, reduced infarct size to a similar extent as complete BAK absence. In contrast to MI/R, homozygous Bak deletion did not attenuate acute infarct size or long-term scar size, post-infarct remodeling, cardiac dysfunction, or mortality in non-reperfused MI. We conclude that BAK contributes significantly to cardiomyocyte necrosis and infarct generation during MI/R, while its absence does not appear to impact the pathogenesis of non-reperfused MI. These observations suggest BAK may be a therapeutic target for MI/R and that even partial pharmacological antagonism may provide benefit.
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Infarto do Miocárdio , Proteína Killer-Antagonista Homóloga a bcl-2 , Animais , Camundongos , Apoptose/fisiologia , Proteína X Associada a bcl-2/genética , Proteína X Associada a bcl-2/metabolismo , Homozigoto , Infarto do Miocárdio/genética , Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/patologia , Necrose/genética , Deleção de Sequência , Proteína Killer-Antagonista Homóloga a bcl-2/metabolismoRESUMO
BACKGROUND: Pericytes have been implicated in tissue repair, remodeling, and fibrosis. Although the mammalian heart contains abundant pericytes, their fate and involvement in myocardial disease remains unknown. METHODS: We used NG2Dsred;PDGFRαEGFP pericyte:fibroblast dual reporter mice and inducible NG2CreER mice to study the fate and phenotypic modulation of pericytes in myocardial infarction. The transcriptomic profile of pericyte-derived cells was studied using polymerase chain reaction arrays and single-cell RNA sequencing. The role of transforming growth factor-ß (TGF-ß) signaling in regulation of pericyte phenotype was investigated in vivo using pericyte-specific TGF-ß receptor 2 knockout mice and in vitro using cultured human placental pericytes. RESULTS: In normal hearts, neuron/glial antigen 2 (NG2) and platelet-derived growth factor receptor α (PDGFRα) identified distinct nonoverlapping populations of pericytes and fibroblasts, respectively. After infarction, a population of cells expressing both pericyte and fibroblast markers emerged. Lineage tracing demonstrated that in the infarcted region, a subpopulation of pericytes exhibited transient expression of fibroblast markers. Pericyte-derived cells accounted for ~4% of PDGFRα+ infarct fibroblasts during the proliferative phase of repair. Pericyte-derived fibroblasts were overactive, expressing higher levels of extracellular matrix genes, integrins, matricellular proteins, and growth factors, when compared with fibroblasts from other cellular sources. Another subset of pericytes contributed to infarct angiogenesis by forming a mural cell coat, stabilizing infarct neovessels. Single-cell RNA sequencing showed that NG2 lineage cells diversify after infarction and exhibit increased expression of matrix genes, and a cluster with high expression of fibroblast identity markers emerges. Trajectory analysis suggested that diversification of infarct pericytes may be driven by proliferating cells. In vitro and in vivo studies identified TGF-ß as a potentially causative mediator in fibrogenic activation of infarct pericytes. However, pericyte-specific TGF-ß receptor 2 disruption had no significant effects on infarct myofibroblast infiltration and collagen deposition. Pericyte-specific TGF-ß signaling was involved in vascular maturation, mediating formation of a mural cell coat investing infarct neovessels and protecting from dilative remodeling. CONCLUSIONS: In the healing infarct, cardiac pericytes upregulate expression of fibrosis-associated genes, exhibiting matrix-synthetic and matrix-remodeling profiles. A fraction of infarct pericytes exhibits expression of fibroblast identity markers. Pericyte-specific TGF-ß signaling plays a central role in maturation of the infarct vasculature and protects from adverse dilative remodeling, but it does not modulate fibrotic remodeling.
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Infarto do Miocárdio , Pericitos , Gravidez , Camundongos , Feminino , Humanos , Animais , Pericitos/metabolismo , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/metabolismo , Placenta/metabolismo , Infarto do Miocárdio/genética , Infarto do Miocárdio/metabolismo , Fibrose , Camundongos Knockout , Fenótipo , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/metabolismo , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , MamíferosRESUMO
Background Interstitial and perivascular fibrosis may contribute to diabetes-associated heart failure. Pericytes can convert to fibroblasts under conditions of stress and have been implicated in the pathogenesis of fibrotic diseases. We hypothesized that in diabetic hearts, pericytes may convert to fibroblasts, contributing to fibrosis and to the development of diastolic dysfunction. Methods and Results Using pericyte:fibroblast dual reporter (NG2Dsred [neuron-glial antigen 2 red fluorescent protein variant]; PDGFRαEGFP [platelet-derived growth factor receptor alpha enhanced green fluorescent protein]) mice in a type 2 diabetic db/db background, we found that diabetes does not significantly affect pericyte density but reduces the myocardial pericyte:fibroblast ratio. Lineage tracing using the inducible NG2CreER driver, along with reliable labeling of fibroblasts with the PDGFRα reporter system, showed no significant pericyte to fibroblast conversion in lean and db/db hearts. In addition, db/db mouse cardiac fibroblasts did not undergo myofibroblast conversion and had no significant induction of structural collagens but exhibited a matrix-preserving phenotype, associated with increased expression of antiproteases, matricellular genes, matrix cross-linking enzymes, and the fibrogenic transcription factor cMyc. In contrast, db/db mouse cardiac pericytes had increased expression of Timp3, without any changes in expression of other fibrosis-associated genes. The matrix-preserving phenotype of diabetic fibroblasts was associated with induction of genes encoding oxidative (Ptgs2/cycloxygenase-2, and Fmo2) and antioxidant proteins (Hmox1, Sod1). In vitro, high glucose partially recapitulated the in vivo changes in diabetic fibroblasts. Conclusions Diabetic fibrosis is not mediated through pericyte to fibroblast conversion but involves acquisition of a matrix-preserving fibroblast program, which is independent of myofibroblast conversion and is only partially explained by the effects of the hyperglycemic environment.
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Diabetes Mellitus , Pericitos , Camundongos , Animais , Pericitos/patologia , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/metabolismo , Fibroblastos/metabolismo , Diabetes Mellitus/metabolismo , Fenótipo , FibroseRESUMO
The heart contains a population of resident macrophages that markedly expands following injury through recruitment of monocytes and through proliferation of macrophages. In myocardial infarction, macrophages have been implicated in both injurious and reparative responses. In coronary atherosclerotic lesions, macrophages have been implicated in disease progression and in the pathogenesis of plaque rupture. Following myocardial infarction, resident macrophages contribute to initiation and regulation of the inflammatory response. Phagocytosis and efferocytosis are major functions of macrophages during the inflammatory phase of infarct healing, and mediate phenotypic changes, leading to acquisition of an anti-inflammatory macrophage phenotype. Infarct macrophages respond to changes in the cytokine content and extracellular matrix composition of their environment and secrete fibrogenic and angiogenic mediators, playing a central role in repair of the infarcted heart. Macrophages may also play a role in scar maturation and may contribute to chronic adverse remodeling of noninfarcted segments. Single cell studies have revealed a remarkable heterogeneity of macrophage populations in infarcted hearts; however, the relations between transcriptomic profiles and functional properties remain poorly defined. This review manuscript discusses the fate, mechanisms of expansion and activation, and role of macrophages in the infarcted heart. Considering their critical role in injury, repair, and remodeling, macrophages are important, but challenging, targets for therapeutic interventions in myocardial infarction.
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Infarto do Miocárdio , Citocinas , Humanos , Macrófagos/patologia , Monócitos/patologia , Infarto do Miocárdio/patologia , FagocitoseRESUMO
TGF-ßs regulate macrophage responses, by activating Smad2/3. We have previously demonstrated that macrophage-specific Smad3 stimulates phagocytosis and mediates anti-inflammatory macrophage transition in the infarcted heart. However, the role of macrophage Smad2 signaling in myocardial infarction remains unknown. We studied the role of macrophage-specific Smad2 signaling in healing mouse infarcts, and we explored the basis for the distinct effects of Smad2 and Smad3. In infarct macrophages, Smad3 activation preceded Smad2 activation. In contrast to the effects of Smad3 loss, myeloid cell-specific Smad2 disruption had no effects on mortality, ventricular dysfunction and adverse remodeling, after myocardial infarction. Macrophage Smad2 loss modestly, but transiently increased myofibroblast density in the infarct, but did not affect phagocytic removal of dead cells, macrophage infiltration, collagen deposition, and scar remodeling. In isolated macrophages, TGF-ß1, -ß2 and -ß3, activated both Smad2 and Smad3, whereas BMP6 triggered only Smad3 activation. Smad2 and Smad3 had similar patterns of nuclear translocation in response to TGF-ß1. RNA-sequencing showed that Smad3, and not Smad2, was the main mediator of transcriptional effects of TGF-ß on macrophages. Smad3 loss resulted in differential expression of genes associated with RAR/RXR signaling, cholesterol biosynthesis and lipid metabolism. In both isolated bone marrow-derived macrophages and in infarct macrophages, Smad3 mediated synthesis of Nr1d2 and Rara, two genes encoding nuclear receptors, that may be involved in regulation of their phagocytic and anti-inflammatory properties. In conclusion, the in vivo and in vitro effects of TGF-ß on macrophage function involve Smad3, and not Smad2.
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Infarto do Miocárdio , Proteína Smad2 , Proteína Smad3 , Animais , Colesterol , Colágeno/metabolismo , Macrófagos/metabolismo , Camundongos , Camundongos Knockout , Infarto do Miocárdio/metabolismo , Miocárdio/metabolismo , Fenótipo , RNA , Proteína Smad2/metabolismo , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta/metabolismoRESUMO
Cells sense mechanical stress and changes in their matrix environment through the integrins, a family of heterodimeric surface receptors that bind to extracellular matrix ligands and trigger cytoskeletal remodeling, while transducing a wide range of intracellular signals. Integrins have been extensively implicated in regulation of inflammation, repair and fibrosis in many different tissues. This review manuscript discusses the role of integrin-mediated cascades in myocardial fibrosis. In vitro studies have demonstrated that ß1 and αv integrins play an important role in fibrogenic conversion of cardiac fibroblast, acting through direct stimulation of FAK/Src cascades, or via accentuation of growth factor signaling. Fibrogenic actions of αv integrins may be mediated, at least in part, through pericellular activation of latent TGF-ß stores. In vivo evidence supporting the role of integrin heterodimers in fibrotic cardiac remodeling is limited to associative evidence, and to experiments using pharmacologic inhibitors, or global loss-of-function approaches. Studies documenting in vivo actions of integrins on fibroblasts using cell-specific strategies are lacking. Integrin effects on leukocytes may also contribute to the pathogenesis of fibrotic myocardial responses by mediating recruitment and activation of fibrogenic macrophages. The profile and role of integrins in cardiac fibrosis may be dependent on the underlying pathologic condition. Considering their cell surface localization and the availability of small molecule inhibitors, integrins may be attractive therapeutic targets for patients with heart failure associated with prominent fibrotic remodeling.
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Fibroblastos , Integrinas , Humanos , Integrinas/metabolismo , Fibrose , Fibroblastos/metabolismo , Matriz Extracelular/metabolismo , Fator de Crescimento Transformador beta/metabolismoRESUMO
Smad7 restrains TGF-ß responses, and has been suggested to exert both pro- and anti-inflammatory actions that may involve effects on macrophages. Myocardial infarction triggers a macrophage-driven inflammatory response that not only plays a central role in cardiac repair, but also contributes to adverse remodeling and fibrosis. We hypothesized that macrophage Smad7 expression may regulate inflammation and fibrosis in the infarcted heart through suppression of TGF-ß responses, or via TGF-independent actions. In a mouse model of myocardial infarction, infiltration with Smad7+ macrophages peaked 7 days after coronary occlusion. Myeloid cell-specific Smad7 loss in mice had no effects on homeostatic functions and did not affect baseline macrophage gene expression. RNA-seq predicted that Smad7 may promote TREM1-mediated inflammation in infarct macrophages. However, these alterations in the transcriptional profile of macrophages were associated with a modest and transient reduction in infarct myofibroblast infiltration, and did not affect dysfunction, chamber dilation, scar remodeling, collagen deposition, and macrophage recruitment. In vitro, RNA-seq and PCR arrays showed that TGF-ß has profound effects on macrophage profile, attenuating pro-inflammatory cytokine/chemokine expression, modulating synthesis of matrix remodeling genes, inducing genes associated with sphingosine-1 phosphate activation and integrin signaling, and inhibiting cholesterol biosynthesis genes. However, Smad7 loss did not significantly affect TGF-ß-mediated macrophage responses, modulating synthesis of only a small fraction of TGF-ß-induced genes, including Itga5, Olfml3, and Fabp7. Our findings suggest a limited role for macrophage Smad7 in regulation of post-infarction inflammation and repair, and demonstrate that the anti-inflammatory effects of TGF-ß in macrophages are not restrained by endogenous Smad7 induction.
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Infarto do Miocárdio , Proteína Smad7/metabolismo , Animais , Fibrose , Inflamação , Macrófagos/metabolismo , Camundongos , Infarto do Miocárdio/genética , Infarto do Miocárdio/metabolismo , Fenótipo , Proteína Smad7/genética , Fator de Crescimento Transformador beta/metabolismoRESUMO
The development of novel therapies based on understanding the pathophysiologic basis of disease is a major goal of biomedical research. Despite an explosion in new knowledge on the molecular mechanisms of disease derived from animal model investigations, translation into effective treatment for human patients has been disappointingly slow. Several fundamental problems may explain the translational failures. First, the emphasis on novel and highly significant findings selectively rewards implausible, low-probability observations and high-magnitude effects, providing a biased perspective of the pathophysiology of disease that underappreciates the complexity and redundancy of biological systems. Second, even when a sound targetable mechanism is identified, animal models cannot recapitulate the pathophysiologic heterogeneity of the human disease, and are poor predictors of therapeutic success. Third, traditional classifications of most complex diseases are based primarily on clinical criteria and do not reflect the diverse pathophysiologic mechanisms that may be involved. The development of a flexible and dynamic conceptual paradigm that takes into account the totality of the evidence on the mechanisms of disease, and pathophysiologic stratification of patients to identify subpopulations with distinct pathogenetic mechanisms, are crucial for the development of new therapeutics.