Showing papers on "Myofibroblast published in 2019"
TL;DR: This work performed single-cell RNA-sequencing of the total non-CM fraction and enriched fibroblast lineage cells from murine hearts at days 3 and 7 post-sham or myocardial infarction (MI) surgery identified >30 populations representing nine cell lineages, including a previously undescribed fibro Blast lineage trajectory present in both sham and MI hearts leading to a uniquely activated cell state.
Abstract: Besides cardiomyocytes (CM), the heart contains numerous interstitial cell types which play key roles in heart repair, regeneration and disease, including fibroblast, vascular and immune cells. However, a comprehensive understanding of this interactive cell community is lacking. We performed single-cell RNA-sequencing of the total non-CM fraction and enriched (Pdgfra-GFP+) fibroblast lineage cells from murine hearts at days 3 and 7 post-sham or myocardial infarction (MI) surgery. Clustering of >30,000 single cells identified >30 populations representing nine cell lineages, including a previously undescribed fibroblast lineage trajectory present in both sham and MI hearts leading to a uniquely activated cell state defined in part by a strong anti-WNT transcriptome signature. We also uncovered novel myofibroblast subtypes expressing either pro-fibrotic or anti-fibrotic signatures. Our data highlight non-linear dynamics in myeloid and fibroblast lineages after cardiac injury, and provide an entry point for deeper analysis of cardiac homeostasis, inflammation, fibrosis, repair and regeneration.
371 citations
TL;DR: The authors use single-cell RNA-sequencing to identify heterogeneity among murine fibroblasts in the wound and find that recruited myeloid cells contribute to adipocyte regeneration during healing.
Abstract: During wound healing in adult mouse skin, hair follicles and then adipocytes regenerate. Adipocytes regenerate from myofibroblasts, a specialized contractile wound fibroblast. Here we study wound fibroblast diversity using single-cell RNA-sequencing. On analysis, wound fibroblasts group into twelve clusters. Pseudotime and RNA velocity analyses reveal that some clusters likely represent consecutive differentiation states toward a contractile phenotype, while others appear to represent distinct fibroblast lineages. One subset of fibroblasts expresses hematopoietic markers, suggesting their myeloid origin. We validate this finding using single-cell western blot and single-cell RNA-sequencing on genetically labeled myofibroblasts. Using bone marrow transplantation and Cre recombinase-based lineage tracing experiments, we rule out cell fusion events and confirm that hematopoietic lineage cells give rise to a subset of myofibroblasts and rare regenerated adipocytes. In conclusion, our study reveals that wounding induces a high degree of heterogeneity among fibroblasts and recruits highly plastic myeloid cells that contribute to adipocyte regeneration.
318 citations
TL;DR: Specific mechanistic principles of fibrosis regression involve the resolution of chronic tissue injury, the shift of inflammatory processes towards recovery, deactivation of myofibroblasts and finally fibrolysis of excess matrix scaffold.
317 citations
TL;DR: The data indicate that myofibroblast differentiation and proliferation are key pathological mechanisms driving fibrosis in SSc-ILD, and define multimodal transcriptome-phenotypes associated with these populations.
Abstract: Objectives Myofibroblasts are key effector cells in the extracellular matrix remodelling of systemic sclerosis-associated interstitial lung disease (SSc-ILD); however, the diversity of fibroblast populations present in the healthy and SSc-ILD lung is unknown and has prevented the specific study of the myofibroblast transcriptome. We sought to identify and define the transcriptomes of myofibroblasts and other mesenchymal cell populations in human healthy and SSc-ILD lungs to understand how alterations in fibroblast phenotypes lead to SSc-ILD fibrosis. Methods We performed droplet-based, single-cell RNA-sequencing with integrated canonical correlation analysis of 13 explanted lung tissue specimens (56 196 cells) from four healthy control and four patients with SSc-ILD, with findings confirmed by cellular indexing of transcriptomes and epitopes by sequencing in additional samples. Results Examination of gene expression in mesenchymal cells identified two major, SPINT2hi and MFAP5hi, and one minor, WIF1hi, fibroblast populations in the healthy control lung. Combined analysis of control and SSc-ILD mesenchymal cells identified SPINT2hi, MFAP5hi, few WIF1hi fibroblasts and a new large myofibroblast population with evidence of actively proliferating myofibroblasts. We compared differential gene expression between all SSc-ILD and control mesenchymal cell populations, as well as among the fibroblast subpopulations, showing that myofibroblasts undergo the greatest phenotypic changes in SSc-ILD and strongly upregulate expression of collagens and other profibrotic genes. Conclusions Our results demonstrate previously unrecognised fibroblast heterogeneity in SSc-ILD and healthy lungs, and define multimodal transcriptome-phenotypes associated with these populations. Our data indicate that myofibroblast differentiation and proliferation are key pathological mechanisms driving fibrosis in SSc-ILD.
159 citations
TL;DR: It is shown that interleukin-11 (IL11) is up-regulated in the lung of patients with IPF, associated with disease severity, and IL-11 is secreted from IPF fibroblasts, and these data prioritize IL- 11 as a drug target for lung fibrosis and IPF.
Abstract: Idiopathic pulmonary fibrosis (IPF) is a progressive fibrotic lung disease where invasive pulmonary myofibroblasts secrete collagen and destroy lung integrity. Here, we show that interleukin-11 (IL11) is up-regulated in the lung of patients with IPF, associated with disease severity, and IL-11 is secreted from IPF fibroblasts. In vitro, IL-11 stimulates lung fibroblasts to become invasive actin alpha 2, smooth muscle-positive (ACTA2+), collagen-secreting myofibroblasts in an extracellular signal-regulated kinase (ERK)-dependent, posttranscriptional manner. In mice, fibroblast-specific transgenic expression or administration of murine IL-11 induces lung myofibroblasts and causes lung fibrosis. IL-11 receptor subunit alpha-1 (Il11ra1)-deleted mice, whose lung fibroblasts are unresponsive to profibrotic stimulation, are protected from fibrosis in the bleomycin mouse model of pulmonary fibrosis. We generated an IL-11-neutralizing antibody that blocks lung fibroblast activation downstream of multiple stimuli and reverses myofibroblast activation. In therapeutic studies, anti-IL-11 treatment diminished lung inflammation and reversed lung fibrosis while inhibiting ERK and SMAD activation in mice. These data prioritize IL-11 as a drug target for lung fibrosis and IPF.
157 citations
TL;DR: This work provides a comprehensive catalogue of the aberrant cellular transcriptional programs in IPF, demonstrates a new framework for analyzing complex disease with scRNAseq, and provides the largest lung disease single-cell atlas to date.
Abstract: We provide a single cell atlas of Idiopathic Pulmonary Fibrosis (IPF), a fatal interstitial lung disease, focusing on resident lung cell populations. By profiling 312,928 cells from 32 IPF, 29 healthy control and 18 chronic obstructive pulmonary disease (COPD) lungs, we demonstrate that IPF is characterized by changes in discrete subpopulations of cells in the three major parenchymal compartments: the epithelium, endothelium and stroma. Among epithelial cells, we identify a novel population of IPF enriched aberrant basaloid cells that co-express basal epithelial markers, mesenchymal markers, senescence markers, developmental transcription factors and are located at the edge of myofibroblast foci in the IPF lung. Among vascular endothelial cells in the in IPF lung parenchyma we identify an expanded cell population transcriptomically identical to vascular endothelial cells normally restricted to the bronchial circulation. We confirm the presence of both populations by immunohistochemistry and independent datasets. Among stromal cells we identify fibroblasts and myofibroblasts in both control and IPF lungs and leverage manifold-based algorithms diffusion maps and diffusion pseudotime to infer the origins of the activated IPF myofibroblast. Our work provides a comprehensive catalogue of the aberrant cellular transcriptional programs in IPF, demonstrates a new framework for analyzing complex disease with scRNAseq, and provides the largest lung disease single-cell atlas to date.
139 citations
TL;DR: Clinical treatments that inhibit or reduce the actions of TGF-β1 and its contribution to cardiac fibrosis and heart failure are discussed.
Abstract: Experimental research has recognized the importance of cardiac fibroblast and myofibroblast cells in heart repair and function In a normal healthy heart, the cardiac fibroblast plays a central role in the structural, electrical, and chemical aspects within the heart Interestingly, the transformation of cardiac fibroblast cells to cardiac myofibroblast cells is suspected to play a vital part in the development of heart failure The ability to differentiate between the two cells types has been a challenge Myofibroblast cells are only expressed in the stressed or failing heart, so a better understanding of cell function may identify therapies that aid repair of the damaged heart This paper will provide an outline of what is currently known about cardiac fibroblasts and myofibroblasts, the physiological and pathological roles within the heart, and causes for the transition of fibroblasts into myoblasts We also reviewed the potential markers available for characterizing these cells and found that there is no single-cell specific marker that delineates fibroblast or myofibroblast cells To characterize the cells of fibroblast origin, vimentin is commonly used Cardiac fibroblasts can be identified using discoidin domain receptor 2 (DDR2) while α-smooth muscle actin is used to distinguish myofibroblasts A known cytokine TGF-β1 is well established to cause the transformation of cardiac fibroblasts to myofibroblasts This review will also discuss clinical treatments that inhibit or reduce the actions of TGF-β1 and its contribution to cardiac fibrosis and heart failure
118 citations
TL;DR: An unbiased single cell RNA-sequencing analysis of a bleomycin-induced pulmonary fibrosis model is used to characterize molecular responses to fibrotic injury and finds that activated fibroblasts are not uniquely defined but exhibit a similar, yet amplified, gene expression pattern to control cells.
Abstract: Idiopathic pulmonary fibrosis (IPF) is a progressive and fatal lung disorder driven by unrelenting extracellular matrix deposition. Fibroblasts are recognized as the central mediators of extracellular matrix production in IPF; however, the characteristics of the underlying fibroblast cell populations in IPF remain poorly understood. Here, we use an unbiased single-cell RNA sequencing analysis of a bleomycin-induced pulmonary fibrosis model to characterize molecular responses to fibrotic injury. Lung cells were isolated on Day 11 to capture emerging fibrosis and gene expression was analyzed by three complementary techniques, which, together, generated a 49-gene signature that defined an activated subpopulation of fibroblasts. However, none of the identified genes were specific to the activated cells or to the disease setting, implying that the activated fibroblasts are not uniquely defined, but exhibit a similar, yet amplified, gene expression pattern to control cells. Our findings have important implications for fibrosis research, including: 1) defining myofibroblasts with any single marker will fail to capture much of the underlying biology; 2) fibroblast activation is poorly correlated with expression of transforming growth factor-β pathway genes; 3) single-cell analysis provides insight into the mechanism of action of effective therapies (nintedanib); 4) early events in lung fibrosis need not involve significant changes in fibroblast number; populations that do increase in number, such as macrophages, dendritic cells, and proliferating myeloid cells, may merit closer examination for their role in pathogenesis.
109 citations
TL;DR: The authors efficiently differentiate human pluripotent stem cells through second heart field progenitors to CFs that exhibit features and functional properties similar to native CFs.
Abstract: Cardiac fibroblasts (CFs) play critical roles in heart development, homeostasis, and disease. The limited availability of human CFs from native heart impedes investigations of CF biology and their role in disease. Human pluripotent stem cells (hPSCs) provide a highly renewable and genetically defined cell source, but efficient methods to generate CFs from hPSCs have not been described. Here, we show differentiation of hPSCs using sequential modulation of Wnt and FGF signaling to generate second heart field progenitors that efficiently give rise to hPSC-CFs. The hPSC-CFs resemble native heart CFs in cell morphology, proliferation, gene expression, fibroblast marker expression, production of extracellular matrix and myofibroblast transformation induced by TGFβ1 and angiotensin II. Furthermore, hPSC-CFs exhibit a more embryonic phenotype when compared to fetal and adult primary human CFs. Co-culture of hPSC-CFs with hPSC-derived cardiomyocytes distinctly alters the electrophysiological properties of the cardiomyocytes compared to co-culture with dermal fibroblasts. The hPSC-CFs provide a powerful cell source for research, drug discovery, precision medicine, and therapeutic applications in cardiac regeneration.
108 citations
TL;DR: MyoFb in end-stage HF have a variable degree of differentiation and retain the capacity to return to a less activated state, validating the potential for developing antifibrotic therapy targeting MyOFb.
105 citations
TL;DR: Cdh11-mediated adhesion prolongs and targets the delivery of macrophage-produced TGF-β to myofibroblasts, creating a self-sustaining profibrotic niche and investigating the key processes that turn acute beneficial repair into destructive progressive fibrosis.
Abstract: Macrophages contribute to the activation of fibroblastic cells into myofibroblasts, which secrete collagen and contract the collagen matrix to acutely repair injured tissue. Persistent myofibroblast activation leads to the accumulation of fibrotic scar tissue that impairs organ function. We investigated the key processes that turn acute beneficial repair into destructive progressive fibrosis. We showed that homotypic cadherin-11 interactions promoted the specific binding of macrophages to and persistent activation of profibrotic myofibroblasts. Cadherin-11 was highly abundant at contacts between macrophages and myofibroblasts in mouse and human fibrotic lung tissues. In attachment assays, cadherin-11 junctions mediated specific recognition and strong adhesion between macrophages and myofibroblasts. One functional outcome of cadherin-11–mediated adhesion was locally restricted activation of latent transforming growth factor–β (TGF-β) between macrophage-myofibroblast pairs that was not observed in cocultures of macrophages and myofibroblasts that were not in contact with one another. Our data suggest that cadherin-11 junctions maintain latent TGF-β–producing macrophages and TGF-β–activating myofibroblasts in close proximity to one another. Inhibition of homotypic cadherin-11 interactions could be used to cause macrophage-myofibroblast separation, thereby destabilizing the profibrotic niche.
TL;DR: Novel mechanisms for fibroblasts in expressing pro-inflammatory molecules and regulating angiogenesis following myocardial infarct are revealed.
Abstract: Cardiac fibroblasts are the major producers of extracellular matrix (ECM) to form infarct scar. We hypothesized that fibroblasts undergo a spectrum of phenotype states over the course of myocardial infarction (MI) from early onset to scar formation. Fibroblasts were isolated from the infarct region of C57BL/6J male mice (3–6 months old, n = 60) at days 0 (no MI control) and 1, 3, or 7 after MI. Whole transcriptome analysis was performed by RNA-sequencing. Of the genes sequenced, 3371 were differentially expressed after MI. Enrichment analysis revealed that MI day 1 fibroblasts displayed pro-inflammatory, leukocyte-recruiting, pro-survival, and anti-migratory phenotype through Tnfrsf9 and CD137 signaling. MI day 3 fibroblasts had a proliferative, pro-fibrotic, and pro-angiogenic profile with elevated Il4ra signaling. MI day 7 fibroblasts showed an anti-angiogenic homeostatic-like myofibroblast profile and with a step-wise increase in Acta2 expression. MI day 7 fibroblasts relied on Pik3r3 signaling to mediate Tgfb1 effects and Fgfr2 to regulate PI3K signaling. In vitro, the day 3 MI fibroblast secretome stimulated angiogenesis, while day 7 MI fibroblast secretome repressed angiogenesis through Thbs1 signaling. Our results reveal novel mechanisms for fibroblasts in expressing pro-inflammatory molecules and regulating angiogenesis following MI.
TL;DR: This work focuses on mechanical and chemical regulation of collagen contraction by fibroblast and the involvement of these factors in their phenotypic conversion to myofibroblasts.
TL;DR: It is reported here that Hippo signaling cell-autonomously regulates CF fate transitions and proliferation, and non-cell-autonomicously regulates both myeloid and CF activation in the heart.
Abstract: Cardiac fibroblasts (CFs) respond to injury by transitioning through multiple cell states, including resting CFs, activated CFs, and myofibroblasts. We report here that Hippo signaling cell-autonomously regulates CF fate transitions and proliferation, and non-cell-autonomously regulates both myeloid and CF activation in the heart. Conditional deletion of Hippo pathway kinases, Lats1 and Lats2, in uninjured CFs initiated a self-perpetuating fibrotic response in the adult heart that was exacerbated by myocardial infarction (MI). Single cell transcriptomics showed that uninjured Lats1/2 mutant CFs spontaneously transitioned to a myofibroblast cell state. Through gene regulatory network reconstruction, we found that Hippo-deficient myofibroblasts deployed a network of transcriptional regulators of endoplasmic reticulum (ER) stress, and the unfolded protein response (UPR) consistent with elevated secretory activity. We observed an expansion of myeloid cell heterogeneity in uninjured Lats1/2 CKO hearts with similarity to cells recovered from control hearts post-MI. Integrated genome-wide analysis of Yap chromatin occupancy revealed that Yap directly activates myofibroblast cell identity genes, the proto-oncogene Myc, and an array of genes encoding pro-inflammatory factors through enhancer-promoter looping. Our data indicate that Lats1/2 maintain the resting CF cell state through restricting the Yap-induced injury response.
TL;DR: The data suggest that reduced autophagy induces EMT of alveolar epithelial cells and can contribute to fibrosis via aberrant epithelial–fibroblast crosstalk.
Abstract: Idiopathic pulmonary fibrosis (IPF), the prototypic progressive fibrotic interstitial lung disease, is thought to be a consequence of repetitive micro-injuries to an ageing, susceptible alveolar epithelium. Ageing is a risk factor for IPF and incidence has been demonstrated to increase with age. Decreased (macro)autophagy with age has been reported extensively in a variety of systems and diseases, including IPF. However, it is undetermined whether the role of faulty autophagy is causal or coincidental in the context of IPF. Here, we report that in alveolar epithelial cells inhibition of autophagy promotes epithelial–mesenchymal transition (EMT), a process implicated in embryonic development, wound healing, cancer metastasis and fibrosis. We further demonstrate that this is attained, at least in part, by increased p62/SQSTM1 expression that promotes p65/RELA mediated-transactivation of an EMT transcription factor, Snail2 (SNAI2), which not only controls EMT but also regulates the production of locally acting profibrogenic mediators. Our data suggest that reduced autophagy induces EMT of alveolar epithelial cells and can contribute to fibrosis via aberrant epithelial–fibroblast crosstalk.
TL;DR: The current understanding of how CAFs fuel immune escape, as well as their potential clinical applications are outlined, and whether these therapeutics really have clinically significant activity remains to be seen, but the outlook is positive.
Abstract: Immune escape is central to the persistence of most, if not all, solid tumors and poses a critical obstacle to successful cancer (immuno)therapy. Cancer-associated fibroblasts (CAFs) constitute the most prevalent, yet heterogeneous, component of the tumor stroma, where they 'cool down' the immune microenvironment. The central role played by CAFs, both as a physical barrier and source of immunosuppressive molecules, sets them as a target to enhance immunotherapy of cancer. We outline the current understanding of how CAFs fuel immune escape, as well as their potential clinical applications. Whether these therapeutics really have clinically significant activity remains to be seen, but the outlook is positive.
TL;DR: This chapter discusses the origins, heterogeneity, and activation of myofibroblasts in diseased kidneys, and highlights novel strategies for the treatment of patients with fibrotic kidney diseases.
Abstract: Renal fibrosis is characterized by excessive deposition of extracellular matrix (ECM), leading to destruction of normal kidney architecture and loss of renal function. The activation of α-smooth muscle actin-positive myofibroblasts plays a key role in this process. After kidney injury, profibrotic factors are secreted by injured tubular epithelia and infiltrated inflammatory cells to promote complex cascades of signaling events leading to myofibroblastic activation, proliferation, and ECM production. The origins of myofibroblasts remain controversial, and possibilities include resident fibroblasts, pericytes, bone marrow-derived cells, and endothelial cells. Recent evidence supports the existence of localized fibrogenic niches, which provides a specialized tissue microenvironment for myofibroblastic activation and expansion. Myofibroblasts often undergo epigenetic modifications, leading to their sustained activation and resistance to apoptosis. In this chapter, we discuss the origins, heterogeneity, and activation of myofibroblasts in diseased kidneys. We also highlight novel strategies for the treatment of patients with fibrotic kidney diseases.
TL;DR: Transforming growth factor–β1 (TGF-β1), the key pro-fibrotic cytokine implicated in multiple fibrotic conditions, increased the production of activating transcription factor 4 (ATF4), the transcriptional master regulator of amino acid metabolism, to supply glucose-derived glycine to meet the amino acid requirements associated with enhanced collagen production in response to myofibroblast differentiation.
Abstract: The differentiation of fibroblasts into a transient population of highly activated, extracellular matrix (ECM)-producing myofibroblasts at sites of tissue injury is critical for normal tissue repair. Excessive myofibroblast accumulation and persistence, often as a result of a failure to undergo apoptosis when tissue repair is complete, lead to pathological fibrosis and are also features of the stromal response in cancer. Myofibroblast differentiation is accompanied by changes in cellular metabolism, including increased glycolysis, to meet the biosynthetic demands of enhanced ECM production. Here, we showed that transforming growth factor-β1 (TGF-β1), the key pro-fibrotic cytokine implicated in multiple fibrotic conditions, increased the production of activating transcription factor 4 (ATF4), the transcriptional master regulator of amino acid metabolism, to supply glucose-derived glycine to meet the amino acid requirements associated with enhanced collagen production in response to myofibroblast differentiation. We further delineated the signaling pathways involved and showed that TGF-β1-induced ATF4 production depended on cooperation between canonical TGF-β1 signaling through Smad3 and activation of mechanistic target of rapamycin complex 1 (mTORC1) and its downstream target eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1). ATF4, in turn, promoted the transcription of genes encoding enzymes of the de novo serine-glycine biosynthetic pathway and glucose transporter 1 (GLUT1). Our findings suggest that targeting the TGF-β1-mTORC1-ATF4 axis may represent a novel therapeutic strategy for interfering with myofibroblast function in fibrosis and potentially in other conditions, including cancer.
TL;DR: Evidence is provided that ZEB1-mediated EMT in human alveolar epithelial type II (ATII) cells contributes to the development of lung fibrosis by paracrine signalling to underlying fibroblasts by controlling the expression of tissue plasminogen activator.
Abstract: The contribution of epithelial–mesenchymal transition (EMT) to human lung fibrogenesis is controversial. Here we provide evidence that ZEB1-mediated EMT in human alveolar epithelial type II (ATII) cells contributes to the development of lung fibrosis by paracrine signalling to underlying fibroblasts. Activation of EGFR–RAS–ERK signalling in ATII cells induced EMT via ZEB1. ATII cells had extremely low extracellular matrix gene expression even after induction of EMT, however conditioned media from ATII cells undergoing RAS-induced EMT augmented TGFβ-induced profibrogenic responses in lung fibroblasts. This epithelial–mesenchymal crosstalk was controlled by ZEB1 via the expression of tissue plasminogen activator (tPA). In human fibrotic lung tissue, nuclear ZEB1 expression was detected in alveolar epithelium adjacent to sites of extracellular matrix (ECM) deposition, suggesting that ZEB1-mediated paracrine signalling has the potential to contribute to early fibrotic changes in the lung interstitium. Targeting this novel ZEB1 regulatory axis may be a viable strategy for the treatment of pulmonary fibrosis.
TL;DR: This review focuses on the contractile properties of myofibroblasts and the conductor, TGF-β1, which together control the opposing interplay between PPARγ and the canonical WNT/β-catenin pathway.
Abstract: Myofibroblasts are non-muscle contractile cells that play a key physiologically role in organs such as the stem villi of the human placenta during physiological pregnancy. They are able to contract and relax in response to changes in the volume of the intervillous chamber. Myofibroblasts have also been observed in several diseases and are involved in wound healing and the fibrotic processes affecting several organs, such as the liver, lungs, kidneys and heart. During the fibrotic process, tissue retraction rather than contraction is correlated with collagen synthesis in the extracellular matrix, leading to irreversible fibrosis and, finally, apoptosis of myofibroblasts. The molecular motor of myofibroblasts is the non-muscle type IIA and B myosin (NMMIIA and NMMIIB). Fibroblast differentiation into myofibroblasts is largely governed by the transforming growth factor-β1 (TGF-β1). This system controls the canonical WNT/β-catenin pathway in a positive manner, and PPARγ in a negative manner. The WNT/β-catenin pathway promotes fibrosis, while PPARγ prevents it. This review focuses on the contractile properties of myofibroblasts and the conductor, TGF-β1, which together control the opposing interplay between PPARγ and the canonical WNT/β-catenin pathway.
TL;DR: The authors report that alterations in mitochondrial calcium uptake is essential for metabolic reprogramming and epigenetic signaling for activation of the myofibroblast gene program and uncover an important role for the mtCU beyond metabolic regulation and cell death.
Abstract: Fibroblast to myofibroblast differentiation is crucial for the initial healing response but excessive myofibroblast activation leads to pathological fibrosis. Therefore, it is imperative to understand the mechanisms underlying myofibroblast formation. Here we report that mitochondrial calcium (mCa2+) signaling is a regulatory mechanism in myofibroblast differentiation and fibrosis. We demonstrate that fibrotic signaling alters gating of the mitochondrial calcium uniporter (mtCU) in a MICU1-dependent fashion to reduce mCa2+ uptake and induce coordinated changes in metabolism, i.e., increased glycolysis feeding anabolic pathways and glutaminolysis yielding increased α-ketoglutarate (αKG) bioavailability. mCa2+-dependent metabolic reprogramming leads to the activation of αKG-dependent histone demethylases, enhancing chromatin accessibility in loci specific to the myofibroblast gene program, resulting in differentiation. Our results uncover an important role for the mtCU beyond metabolic regulation and cell death and demonstrate that mCa2+ signaling regulates the epigenome to influence cellular differentiation. Myofibroblast differentiation contributes to extracellular matrix remodeling and fibrosis. Here, the authors report that alterations in mitochondrial calcium uptake is essential for metabolic reprogramming and epigenetic signaling for activation of the myofibroblast gene program.
TL;DR: TGF-β1-induced miR-133a functions as a feed-back negative regulator of TGF- β1 profibrogenic pathways and may provide a new therapeutic strategy to halt and perhaps even partially reverse the progression of IPF.
Abstract: Transforming growth factor (TGF)-β1, a main profibrogenic cytokine in the progression of idiopathic pulmonary fibrosis (IPF), induces differentiation of pulmonary fibroblasts to myofibroblasts that produce high levels of collagen, leading to concomitantly loss of lung elasticity and function. Recent studies implicate the importance of microRNAs (miRNAs) in IPF but their regulation and individual pathological roles remain largely unknown. We used both RNA sequencing and quantitative RT-PCR strategies to systematically study TGF-β1-induced alternations of miRNAs in human lung fibroblasts (HFL). Our data show that miR-133a was significantly upregulated by TGF-β1 in a time- and concentration-dependent manner. Surprisingly, miR-133a inhibits TGF-β1-induced myofibroblast differentiation whereas miR-133a inhibitor enhances TGF-β1-induced myofibroblast differentiation. Interestingly, quantitative proteomics analysis indicates that miR-133a attenuates myofibroblast differentiation via targeting multiple components of TGF-β1 profibrogenic pathways. Western blot analysis confirmed that miR-133a down-regulates TGF-β1-induced expression of classic myofibroblast differentiation markers such as ɑ-smooth muscle actin (ɑ-SMA), connective tissue growth factor (CTGF) and collagens. miRNA Target Searcher analysis and luciferase reporter assays indicate that TGF-β receptor 1, CTGF and collagen type 1-alpha1 (Col1a1) are direct targets of miR-133a. More importantly, miR-133a gene transferred into lung tissues ameliorated bleomycin-induced pulmonary fibrosis in mice. Together, our study identified TGF-β1-induced miR-133a as an anti-fibrotic factor. It functions as a feed-back negative regulator of TGF-β1 profibrogenic pathways. Thus, manipulations of miR-133a expression may provide a new therapeutic strategy to halt and perhaps even partially reverse the progression of IPF.
TL;DR: The findings indicate that malignant PTs recruit and repolarize TAMs through a CCL5-CCR5–driven signaling cascade, and targeting CCR5 with maraviroc represents a potential clinically available strategy to treat malignantPTs.
Abstract: Purpose: Malignant phyllodes tumor (PT) is a fast-progression neoplasm derived from periductal stromal cells of the breast, which currently still lack effective treatment strategies. Our previous studies showed that the high density of tumor-associated macrophages (TAM) plays an important role in the malignant progression of PTs. TAMs secreted large amount of CCL18 to promote myofibroblast differentiation and invasion via binding to its receptor PIPTNM3 on myofibroblasts. Herein, we investigate the mechanism of how TAMs are recruited and repolarized by PTs to drive the malignant progression. Experimental Design: The cytokines secreted by PTs were identified by the cytokine array. The clinical and pathologic correlations of the cytokine with PTs were estimated with IHC. The mechanisms of the cytokine that recruited and polarized the macrophage were explored with a coculture model of primary PT cells and macrophages in vitro and in vivo. The patient-derived xenografts (PDX) of malignant PTs were used to evaluate the therapeutic effect of CCR5 inhibitor. Results: A high level of malignant PT-secreted CCL5 correlated with poor outcome of PTs and could be an independent prognostic factor of PTs. CCL5 bound to its receptor, CCR5, on macrophages thus activated AKT signaling to recruit and repolarize TAMs. Subsequently, the TAMs released CCL18 to further promote the aggressive phenotype of malignant PTs by enhancing and maintaining the myofibroblast differentiation and invasion in vitro and in vivo. In a murine PDX model of human malignant PTs, the CCL5–CCR5 axis blocked by maraviroc, an FDA-proved CCR5 inhibitor, prevented recruitment of monocytes to the tumor and dramatically suppressed tumor growth. Conclusions: Our findings indicate that malignant PTs recruit and repolarize TAMs through a CCL5-CCR5–driven signaling cascade. Thus, a positive feedback loop of CCL5-CCR5 and CCL18-PIPTNM3 between myofibroblast and TAMs is constituted to drive the malignant progression of PTs. Furthermore, targeting CCR5 with maraviroc represents a potential clinically available strategy to treat malignant PTs.
TL;DR: CTHRC1 can be used for predicting pirfenidone response and developing new therapeutic targets for lung fibrosis through inverse regulation of CTHRC 1-induced lung fibroblast activity.
Abstract: Pirfenidone, an antifibrotic agent used for the treatment of idiopathic pulmonary fibrosis (IPF), functions by inhibiting myofibroblast differentiation, which is involved in transforming growth factor (TGF)-β1-induced IPF pathogenesis. However, unlike normal lung fibroblasts, the relationship between pirfenidone responses of TGF-β1-induced human fibrotic lung fibroblasts and lung fibrosis has not been elucidated. The effects of pirfenidone were evaluated in lung fibroblasts isolated from fibrotic human lung tissues after TGF-β1 exposure. The ability of two new pharmacological targets of pirfenidone, collagen triple helix repeat containing protein 1(CTHRC1) and four-and-a-half LIM domain protein 2 (FHL2), to mediate contraction of collagen gels and migration toward fibronectin were assessed in vitro. Compared to control lung fibroblasts, pirfenidone significantly restored TGF-β1-stimulated fibroblast-mediated collagen gel contraction, migration, and CTHRC1 release in lung fibrotic fibroblasts. Furthermore, pirfenidone attenuated TGF-β1- and CTHRC1-induced fibroblast activity, upregulation of bone morphogenic protein-4(BMP-4)/Gremlin1, and downregulation of α-smooth muscle actin, fibronectin, and FHL2, similar to that observed post-CTHRC1 inhibition. In contrast, FHL2 inhibition suppressed migration and fibronectin expression, but did not downregulate CTHRC1. Overall, pirfenidone suppressed fibrotic fibroblast-mediated fibrotic processes via inverse regulation of CTHRC1-induced lung fibroblast activity. Thus, CTHRC1 can be used for predicting pirfenidone response and developing new therapeutic targets for lung fibrosis.
TL;DR: In vivo, it is shown that interfering with DNM3OS function not only prevents lung fibrosis but also improves established pulmonary fibrosis.
Abstract: Rationale: Given the paucity of effective treatments for idiopathic pulmonary fibrosis (IPF), new insights into the deleterious mechanisms controlling lung fibroblast activation, the key cell type driving the fibrogenic process, are essential to develop new therapeutic strategies. TGF-β (transforming growth factor-β) is the main profibrotic factor, but its inhibition is associated with severe side effects because of its pleiotropic role. Objectives: To determine if downstream noncoding effectors of TGF-β in fibroblasts may represent new effective therapeutic targets whose modulation may be well tolerated. Methods: We investigated the whole noncoding fraction of TGF-β-stimulated lung fibroblast transcriptome to identify new genomic determinants of lung fibroblast differentiation into myofibroblasts. Differential expression of the long noncoding RNA (lncRNA) DNM3OS (dynamin 3 opposite strand) and its associated microRNAs (miRNAs) was validated in a murine model of pulmonary fibrosis and in IPF tissue samples. Distinct and complementary antisense oligonucleotide-based strategies aiming at interfering with DNM3OS were used to elucidate the role of DNM3OS and its associated miRNAs in IPF pathogenesis. Measurements and Main Results: We identified DNM3OS as a fibroblast-specific critical downstream effector of TGF-β-induced lung myofibroblast activation. Mechanistically, DNM3OS regulates this process in trans by giving rise to three distinct profibrotic mature miRNAs (i.e., miR-199a-5p/3p and miR-214-3p), which influence SMAD and non-SMAD components of TGF-β signaling in a multifaceted way. In vivo, we showed that interfering with DNM3OS function not only prevents lung fibrosis but also improves established pulmonary fibrosis. Conclusions: Pharmacological approaches aiming at interfering with the lncRNA DNM3OS may represent new effective therapeutic strategies in IPF.
TL;DR: Major therapeutic antifibrotic strategies developed in the last two decades in preclinical studies, some translated to clinical conditions, designed to interfere directly or indirectly with the Ras/Raf/MEK/ERK signaling pathway in activated hepatic MFs are analyzed.
Abstract: Fibrogenic progression of chronic liver disease, whatever the etiology, is characterized by persistent chronic parenchymal injury, chronic activation of inflammatory response, and sustained activation of liver fibrogenesis, and of pathological wound healing response. A critical role in liver fibrogenesis is played by hepatic myofibroblasts (MFs), a heterogeneous population of α smooth-muscle actin—positive cells that originate from various precursor cells through a process of activation and transdifferentiation. In this review, we focus the attention on the role of extracellular signal-regulated kinase (ERK) signaling pathway as a critical one in modulating selected profibrogenic phenotypic responses operated by liver MFs. We will also analyze major therapeutic antifibrotic strategies developed in the last two decades in preclinical studies, some translated to clinical conditions, designed to interfere directly or indirectly with the Ras/Raf/MEK/ERK signaling pathway in activated hepatic MFs, but that also significantly increased our knowledge on the biology and pathobiology of these fascinating profibrogenic cells.
TL;DR: A functional cross-talk between TGF-β and PDGFRα signaling pathways that is involved in regulating the biology of FAPs and/or MSCs is established.
Abstract: Fibro-adipogenic progenitors (FAPs) are tissue-resident mesenchymal stromal cells (MSCs) required for proper skeletal muscle development, regeneration and maintenance. However, FAPs are also responsible for fibro-fatty scar deposition following chronic damage. We aimed to investigate the role of functional cross-talk between TGF-β and PDGFRα signaling pathways in the fate of FAPs. Here, we show that the number of FAPs correlates with TGF-β levels and with extracellular matrix deposition during regeneration and repair. Interestingly, the expression of PDGFRα changed dynamically in the fibroblast lineage after injury. Furthermore, PDGFRα-dependent immediate early gene expression changed during regeneration and repair. We also found that TGF-β signaling reduces PDGFRα expression in FAPs, mouse dermal fibroblasts and in two related mesenchymal cell lines. Moreover, TGF-β promotes myofibroblast differentiation of FAPs but inhibits their adipogenicity. Accordingly, TGF-β impairs the expression of PDGFRα-dependent immediate early genes in a TGFBR1-dependent manner. Finally, pharmacological inhibition of PDGFRα activity with AG1296 impaired TGF-β-induced extracellular matrix remodeling, Smad2 signaling, myofibroblast differentiation and migration of MSCs. Thus, our work establishes a functional cross-talk between TGF-β and PDGFRα signaling pathways that is involved in regulating the biology of FAPs and/or MSCs.This article has an associated First Person interview with the first author of the paper.
TL;DR: This study reveals that ROS, lipid peroxidation, and GPX4 play important roles in pulmonary fibrosis and ferroptosis induced by erastin.
Abstract: Ferroptosis is a new form of regulated cell death. Fibroblast-to-myofibroblast differentiation is known to be involved in the pathogenesis of idiopathic pulmonary fibrosis. Utilizing HFL1 cell line...
TL;DR: This review explores the emerging importance of the p38 MAPK pathway in cardiac fibroblasts, describes the molecular mechanisms by which it regulates the expression of key genes, and highlights its potential as a therapeutic target for reducing adverse myocardial remodeling.
Abstract: The cardiac fibroblast is a remarkably versatile cell type that coordinates inflammatory, fibrotic and hypertrophic responses in the heart through a complex array of intracellular and intercellular signaling mechanisms. One important signaling node that has been identified involves p38 MAPK; a family of kinases activated in response to stress and inflammatory stimuli that modulates multiple aspects of cardiac fibroblast function, including inflammatory responses, myofibroblast differentiation, extracellular matrix turnover and the paracrine induction of cardiomyocyte hypertrophy. This review explores the emerging importance of the p38 MAPK pathway in cardiac fibroblasts, describes the molecular mechanisms by which it regulates the expression of key genes, and highlights its potential as a therapeutic target for reducing adverse myocardial remodeling.
TL;DR: These findings indicate for the first time that a cardiomyocyte-specific miRNA, transferred to fibroblasts in form of exosomal cargo, is crucial in the activation of myofibroblasta following ischemic injury.
Abstract: The inclusion of microRNAs (miRNAs) in extracellular microvesicles/exosomes (named cardiosomes when deriving from cardiomyocytes) allows their active transportation and ensures cell-cell communication. We hypothesize that cardiosomal miRNAs play a pivotal role in the activation of myofibroblasts following ischemic injury. Using a murine model of myocardial infarction (MI), we tested our hypothesis by measuring in isolated fibroblasts and cardiosomes the expression levels of a set of miRNAs, which are upregulated in cardiomyocytes post-MI and involved in myofibroblast phenoconversion. We found that miR-195 was significantly upregulated in cardiosomes and in fibroblasts isolated after MI compared with SHAM conditions. Moreover, primary isolated cardiac fibroblasts were activated both when incubated with cardiosomes isolated from ischemic cardiomyocytes and when cultured in conditioned medium of post-MI cardiomyocytes, whereas no significant effect was observed following incubation with cardiosomes or medium from sham cardiomyocytes. Taken together, our findings indicate for the first time that a cardiomyocyte-specific miRNA, transferred to fibroblasts in form of exosomal cargo, is crucial in the activation of myofibroblasts.