Andrew L. Koenig

RATIONALE: Recent advancements have brought to light the origins, complexity, and functions of tissue-resident macrophages. However, in the context of tissue injury or disease, large numbers of monocytes infiltrate the heart and are thought to contribute to adverse remodeling and heart failure pathogenesis. Little is understood about the diversity of monocytes and monocyte-derived macrophages recruited to the heart after myocardial injury, including the mechanisms that regulate monocyte recruitment and fate specification. OBJECTIVE: We sought to test the hypothesis that distinct subsets of tissue-resident CCR2- (C-C chemokine receptor 2) and CCR2+ macrophages orchestrate monocyte recruitment and fate specification after myocardial injury. METHODS AND RESULTS: We reveal that in numerous mouse models of cardiomyocyte cell death (permanent myocardial infarction, reperfused myocardial infarction, and diphtheria toxin cardiomyocyte ablation), there is a shift in macrophage ontogeny whereby tissue-resident macrophages are predominately replaced by infiltrating monocytes and monocyte-derived macrophages. Using syngeneic cardiac transplantation to model ischemia-reperfusion injury and distinguish tissue-resident from recruited cell populations in combination with intravital 2-photon microscopy, we demonstrate that monocyte recruitment is differentially orchestrated by distinct subsets of tissue-resident cardiac macrophages. Tissue-resident CCR2+ macrophages promote monocyte recruitment through an MYD88 (myeloid differentiation primary response 88)-dependent mechanism that results in release of MCPs (monocyte chemoattractant proteins) and monocyte mobilization. In contrast, tissue-resident CCR2- macrophages inhibit monocyte recruitment. Using CD (cluster of differentiation) 169-DTR (diphtheria toxin receptor) and CCR2-DTR mice, we further show that selective depletion of either tissue-resident CCR2- or CCR2+ macrophages before myocardial infarction results in divergent effects on left ventricular function, myocardial remodeling, and monocyte recruitment. Finally, using single-cell RNA sequencing, we show that tissue-resident cardiac macrophages differentially instruct monocyte fate specification. CONCLUSIONS: Collectively, these observations establish the mechanistic basis by which monocytes are initially recruited to the injured heart and provide new insights into the heterogeneity of monocyte-derived macrophages.

Heart failure represents a major cause of morbidity and mortality worldwide. Single-cell transcriptomics have revolutionized our understanding of cell composition and associated gene expression. Through integrated analysis of single-cell and single-nucleus RNA-sequencing data generated from 27 healthy donors and 18 individuals with dilated cardiomyopathy, here we define the cell composition of the healthy and failing human heart. We identify cell-specific transcriptional signatures associated with age and heart failure and reveal the emergence of disease-associated cell states. Notably, cardiomyocytes converge toward common disease-associated cell states, whereas fibroblasts and myeloid cells undergo dramatic diversification. Endothelial cells and pericytes display global transcriptional shifts without changes in cell complexity. Collectively, our findings provide a comprehensive analysis of the cellular and transcriptomic landscape of human heart failure, identify cell type-specific transcriptional programs and disease-associated cell states and establish a valuable resource for the investigation of human heart failure.

Background: Recent studies have established that CCR2 (C-C chemokine receptor type 2) marks proinflammatory subsets of monocytes, macrophages, and dendritic cells that contribute to adverse left ventricle (LV) remodeling and heart failure progression. Elucidation of the effector mechanisms that mediate adverse effects of CCR2 + monocytes, macrophages, and dendritic cells will yield important insights into therapeutic strategies to suppress myocardial inflammation. Methods: We used mouse models of reperfused myocardial infarction, angiotensin II and phenylephrine infusion, and diphtheria toxin cardiomyocyte ablation to investigate CCL17 (C-C chemokine ligand 17). We used Ccl17 knockout mice, flow cytometry, RNA sequencing, biochemical assays, cell trafficking studies, and in vivo cell depletion to identify the cell types that generate CCL17, define signaling pathways that controlled its expression, delineate the functional importance of CCL17 in adverse LV remodeling and heart failure progression, and determine the mechanistic basis by which CCL17 exerts its effects. Results: We demonstrated that CCL17 is expressed in CCR2 + macrophages and cluster of differentiation 11b + conventional dendritic cells after myocardial infarction, angiotensin II and phenylephrine infusion, and diphtheria toxin cardiomyocyte ablation. We clarified the transcriptional signature of CCL17 + macrophages and dendritic cells and identified granulocyte-macrophage colony-stimulating factor (GM-CSF) signaling as a key regulator of CCL17 expression through cooperative activation of STAT5 (signal transducer and activator of transcription 5) and canonical NF-κB (nuclear factor κ-light-chain-enhancer of activated B cells) signaling. Ccl17 deletion resulted in reduced LV remodeling, decreased myocardial fibrosis and cardiomyocyte hypertrophy, and improved LV systolic function after myocardial infarction and angiotensin II and phenylephrine infusion. We observed increased abundance of regulatory T cells (Tregs) in the myocardium of injured Ccl17 knockout mice. CCL17 inhibited Treg recruitment through biased activation of CCR4. CCL17 activated Gq signaling and CCL22 (C-C chemokine ligand 22) activated both Gq and ARRB (β-arrestin) signaling downstream of CCR4. CCL17 competitively inhibited CCL22 stimulated ARRB signaling and Treg migration. We provide evidence that Tregs mediated the protective effects of Ccl17 deletion on myocardial inflammation and adverse LV remodeling. Conclusions: These findings identify CCL17 as a proinflammatory mediator of CCR2 + macrophages and dendritic cells and suggest that inhibition of CCL17 may serve as an effective strategy to promote Treg recruitment and suppress myocardial inflammation.

Effective tissue repair after myocardial infarction entails a vigorous angiogenic response, guided by incompletely defined immune cell–endothelial cell interactions. We identify the monocyte- and macrophage-derived cytokine METRNL (meteorin-like) as a driver of postinfarction angiogenesis and high-affinity ligand for the stem cell factor receptor KIT (KIT receptor tyrosine kinase). METRNL mediated angiogenic effects in cultured human endothelial cells through KIT-dependent signaling pathways. In a mouse model of myocardial infarction, METRNL promoted infarct repair by selectively expanding the KIT-expressing endothelial cell population in the infarct border zone. Metrnl -deficient mice failed to mount this KIT-dependent angiogenic response and developed severe postinfarction heart failure. Our data establish METRNL as a KIT receptor ligand in the context of ischemic tissue repair.