The role of α5 Integrin in regulating macrophage phenotype and function during cardiac remodeling after myocardial infarction
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ABSTRACT: Macrophages infiltrate the infarcted heart and play a critical role in repair, remodeling and fibrosis. Macrophages sense changes in the extracellular matrix (ECM) environment through Integrins, thus activating signaling pathways that regulate their function. Analysis of our previous RNA sequencing data identified integrin α5 (Itgα5) as one of the most upregulated integrin genes in infarct macrophages. Accordingly, we hypothesized that integrin α5 signaling in infarct macrophages transduces ECM-derived signals, regulating responses critical for repair and remodeling of the infarcted heart.
Project description:Macrophages infiltrate the infarcted heart and play a critical role in repair, remodeling and fibrosis. Macrophages sense changes in the extracellular matrix (ECM) environment through Integrins, thus activating signaling pathways that regulate their function. Analysis of our previous RNA sequencing data identified integrin α5 (Itgα5) as one of the most upregulated integrin genes in infarct macrophages. Accordingly, we hypothesized that integrin α5 signaling in infarct macrophages transduces ECM-derived signals, regulating responses critical for repair and remodeling of the infarcted heart.
Project description:Macrophages infiltrate the infarcted heart and play a critical role in repair, remodeling and fibrosis. Macrophages sense changes in the extracellular matrix (ECM) environment through Integrins, thus activating signaling pathways that regulate their function. Our data show that av integrin is highly expressed in isolated bone marrow macrophages and is markedly upregulated in response to TGF-β, a growth factor known to be activated in the infarcted heart. Accordingly, we hypothesize that aV integrin induction in infarct macrophages may regulate macrophage phenotype, function and response to key macrophage-activating signals following MI.
Project description:Conclusions: 1. ITGα5-SRC signalling regulates the CLDN1 expression and hepatic polarity. 2. ITGα5-SRC signalling regulates the TET catelyzed DNA 5hmc/5mc level. 3. Itgα5 regulates hydroxymethylation level of genes critical for polarity in vivo. 4. Excessive deposition of ECM blocks the regulatory effect of ITGα5 on CLDN1 in vitro and in vivo.
Project description:Conclusions:1、 ITGα5-SRC signalling regulates the CLDN1 expression and hepatic polarity.2、ITGα5-SRC signalling regulates the TET catelyzed DNA 5hmc/5mc level. 3、Itgα5 regulates hydroxymethylation level of genes critical for polarity in vivo. 4、Excessive deposition of ECM blocks the regulatory effect of ITGα5 on CLDN1 in vitro and in vivo.
Project description:Integrin trafficking to and from membrane adhesions is a crucial mechanism that dictates many aspects of a cell’s behaviour, including motility, polarisation, and invasion. In endothelial cells (ECs), the intracellular traffic of α5-integrin is regulated by both neuropilin 1 (NRP1) and neuropilin 2 (NRP2), yet the redundancies in function between these co-receptors remain unclear. Moreover, the endocytic complexes that participate in NRP-directed-traffic remain poorly annotated. Here we identify an important role for the GTPase-activating protein p120RasGAP in ECs, promoting the recycling of α5-integrin from early endosomes. Mechanistically, p120RasGAP enables transit of endocytosed α5-integrin-NRP1-NRP2 complexes to Rab11+ recycling endosomes, promoting cell polarisation and fibronectin (FN) fibrillogenesis. Silencing of both NRP receptors, or p120RasGAP, resulted in the accumulation of α5-integrin in early endosomes, a loss of α5-integrin from surface adhesions, and attenuated EC polarisation. Endothelial-specific deletion of both NRP1 and NRP2 in the postnatal retina recapitulated our in vitro findings, severely impairing FN fibrillogenesis and polarised sprouting. Our data assign an essential role for p120RasGAP during integrin traffic in ECs and support a hypothesis that NRP receptors co-traffic internalised cargoes. Importantly, we utilise comparative proteomics analyses to, for the first time, isolate a comprehensive map of NRP1-dependent and NRP2-dependent-α5-integrin interactions in ECs.
Project description:The airway epithelium of asthmatics is characterized by intrinsically abnormal wound repair that may contribute to disease pathobiology. In this study, we show that in asthma, the airway epithelial cells at the leading edge of a wound display aberrant migration patterns, reduced expression of α5 and β1 integrin subunits at baseline and during wound repair, resulting in dysregulated cell migration and an inability to fully repair. Transcriptional profiling identified the PI3K/Akt signaling pathway as the top upstream transcriptional regulator of integrin α5β1. Significantly, activation of Akt signaling enhanced airway epithelial repair in cultures derived from asthmatic children via upregulation of α5 and β1 integrin subunits. Conversely, inhibition of the PI3K/Akt signaling pathway in airway epithelial cultures from non-asthmatic children attenuated epithelial repair and reduced α5 and β1 integrin expression. Importantly, the FDA-approved drug celecoxib, and its non-COX2 inhibitory analogue dimethyl-celecoxib, also stimulated the PI3K/Akt/integrin α5β1 axis and restored airway epithelial repair in cells from asthmatics. Thus, targeting the PI3K/Akt pathway may represent a novel therapeutic avenue for asthma.
Project description:Ly6Clow macrophages promote scar formation and prevent early infarct expansion after myocardial infarction (MI). Although CD4+ T cells influence the regulation of Ly6Clow macrophages after MI, the mechanism remains largely unknown. Here, we focused on IL-21 and uncovered its physiological relevance in post-MI hearts. CD4+ T cells harvested from the infarcted heart produce IL-21 upon stimulation, and IL-21 receptor was expressed on Ly6Clo macrophages in the infarcted heart. The survival rate after MI was significantly improved in IL-21-deficient mice compared with WT mice. Moreover, transcriptome analysis of infarcted heart tissue demonstrated that inflammation was persistent in WT mice compared with IL-21-deficient mice. The number of neutrophils was significantly decreased, whereas the number of Ly6Clow macrophages was significantly increased in IL-21-deficient mice. Consistently, IL-21 enhanced the apoptosis of Ly6Clow macrophages. Furthermore, RNA-seq analysis of Ly6Chi and Ly6Clo macrophages stimulated with or without IL-21 for 24 hours revealed that IL-21 induces inflammatory responses in both Ly6Chi and Ly6Clo macrophages. Finally, the treatment with IL-21 receptor Fc protein significantly increased the survival after MI. Thus, the deletion of IL-21 improves survival after MI by preventing Ly6Clo macrophage apoptosis.
Project description:Pericytes have been implicated in regulation of inflammatory, reparative, fibrogenic and angiogenic responses in several different organs and pathologic conditions. Although the adult mammalian heart contains abundant pericytes, their fate and involvement in myocardial disease remains unknown. We used NG2Dsred;PDGFRaEGFP pericyte-fibroblast dual reporter mice and inducible NG2CreER mice to study the fate and phenotypic modulation of pericytes in a model of myocardial infarction. The transcriptomic profile of pericyte-derived fibroblasts was studied using PCR arrays. The transcriptomic profile of NG2 lineage cells (pericytes) was studied in control and infarcted hearts using single cell RNA-sequencing analysis. The role of TGF-b signaling in regulation of pericyte phenotype in vivo was investigated using pericyte-specific Tgfbr2 knockout mice. In vitro, the effects of TGF-b were studied in cultured human placental pericytes.In normal mouse hearts, NG2 and PDGFRa identified distinct non-overlapping populations of pericytes and fibroblasts respectively. Following myocardial infarction, a population of cells expressing both pericyte and fibroblast markers emerged. These cells expressed large amounts of extracellular matrix (ECM) genes. Lineage tracing demonstrated that in the infarcted region, a subpopulation of pericytes underwent fibroblast conversion. Single cell RNA-seq experiments demonstrated expansion and diversification of pericyte-derived cells in the infarct, associated with emergence of subpopulations exhibiting accentuated matrix gene synthesis. In vitro studies and the profile of pericyte-derived fibroblasts identified TGF-b as a potentially causative mediator in fibrogenic activation of infarct pericytes. However, pericyte-specific Tgfbr2 disruption had no significant effects on myofibroblast infiltration and collagen deposition in the infarct. Pericyte-specific TGF-b signaling was involved in vascular maturation, mediating formation of a mural cell coat investing infarct neovessels. These reparative effects of infarct pericytes protected the infarcted heart from dilative remodeling.
Project description:Cardiac macrophages are heterogenous in phenotype and functions, which has been associated with differences in their ontogeny. Despite extensive research, our understanding of the precise role of different subsets of macrophages in ischemia/reperfusion injury remains incomplete. We here investigated macrophage lineages and ablated tissue macrophages in homeostasis and after I/R injury in a CSF1R-dependent manner. Genomic deletion of a fms-intronic regulatory element (FIRE) in the Csf1r locus resulted in specific absence of resident homeostatic and antigen-presenting macrophages, without affecting the recruitment of monocyte-derived macrophages to the infarcted heart. Specific absence of homeostatic, monocyte-independent macrophages altered the immune cell crosstalk in response to injury and induced proinflammatory neutrophil polarization, resulting in impaired cardiac remodelling without influencing infarct size. In contrast, continuous CSF1R inhibition led to depletion of both resident and recruited macrophage populations. This augmented adverse remodelling after I/R and led to an increased infarct size and deterioration of cardiac function. In summary, resident macrophages orchestrate inflammatory responses improving cardiac remodelling, while recruited macrophages determine infarct size after I/R injury. These findings attribute distinct beneficial effects to different macrophage populations in the context of myocardial infarction.
Project description:Cardiac macrophages are heterogenous in phenotype and functions, which has been associated with differences in their ontogeny. Despite extensive research, our understanding of the precise role of different subsets of macrophages in ischemia/reperfusion injury remains incomplete. We here investigated macrophage lineages and ablated tissue macrophages in homeostasis and after I/R injury in a CSF1R-dependent manner. Genomic deletion of a fms-intronic regulatory element (FIRE) in the Csf1r locus resulted in specific absence of resident homeostatic and antigen-presenting macrophages, without affecting the recruitment of monocyte-derived macrophages to the infarcted heart. Specific absence of homeostatic, monocyte-independent macrophages altered the immune cell crosstalk in response to injury and induced proinflammatory neutrophil polarization, resulting in impaired cardiac remodelling without influencing infarct size. In contrast, continuous CSF1R inhibition led to depletion of both resident and recruited macrophage populations. This augmented adverse remodelling after I/R and led to an increased infarct size and deterioration of cardiac function. In summary, resident macrophages orchestrate inflammatory responses improving cardiac remodelling, while recruited macrophages determine infarct size after I/R injury. These findings attribute distinct beneficial effects to different macrophage populations in the context of myocardial infarction.