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Cell Reports Feb 2022Intrahepatic cholangiocarcinoma (ICC) contains abundant myofibroblasts derived from hepatic stellate cells (HSCs) through an activation process mediated by TGF-β. To...
Intrahepatic cholangiocarcinoma (ICC) contains abundant myofibroblasts derived from hepatic stellate cells (HSCs) through an activation process mediated by TGF-β. To determine the role of programmed death-ligand 1 (PD-L1) in myofibroblastic activation of HSCs, we disrupted PD-L1 of HSCs by shRNA or anti-PD-L1 antibody. We find that PD-L1, produced by HSCs, is required for HSC activation by stabilizing TGF-β receptors I (TβRI) and II (TβRII). While the extracellular domain of PD-L1 (amino acids 19-238) targets TβRII protein to the plasma membrane and protects it from lysosomal degradation, a C-terminal 260-RLRKGR-265 motif on PD-L1 protects TβRI mRNA from degradation by the RNA exosome complex. PD-L1 is required for HSC expression of tumor-promoting factors, and targeting HSC PD-L1 by shRNA or Cre/loxP recombination suppresses HSC activation and ICC growth in mice. Thus, myofibroblast PD-L1 can modulate the tumor microenvironment and tumor growth by a mechanism independent of immune suppression.
Topics: Animals; B7-H1 Antigen; Cell Movement; Cell Proliferation; Hepatic Stellate Cells; Humans; Liver Neoplasms; Mice; Myofibroblasts; Receptor, Transforming Growth Factor-beta Type I; Receptors, Transforming Growth Factor beta; Signal Transduction; Tumor Microenvironment
PubMed: 35139382
DOI: 10.1016/j.celrep.2022.110349 -
Scientific Reports Aug 2018Abnormal nerve regeneration often follows corneal injury, predisposing patients to pain, dry eye and vision loss. Yet, we lack a mechanistic understanding of this...
Abnormal nerve regeneration often follows corneal injury, predisposing patients to pain, dry eye and vision loss. Yet, we lack a mechanistic understanding of this process. A key event in corneal wounds is the differentiation of keratocytes into fibroblasts and scar-forming myofibroblasts. Here, we show for the first time that regenerating nerves avoid corneal regions populated by myofibroblasts in vivo. Recreating this interaction in vitro, we find neurite outgrowth delayed when myofibroblasts but not fibroblasts, are co-cultured with sensory neurons. After neurites elongated sufficiently, contact inhibition was observed with myofibroblasts, but not fibroblasts. Reduced neurite outgrowth in vitro appeared mediated by transforming growth factor beta 1 (TGF-β1) secreted by myofibroblasts, which increased phosphorylation of collapsin response mediating protein 2 (CRMP2) in neurons. The significance of this mechanism was further tested by applying Mitomycin C after photorefractive keratectomy to decrease myofibroblast differentiation. This generated earlier repopulation of the ablation zone by intra-epithelial and sub-basal nerves. Our findings suggest that attaining proper, rapid corneal nerve regeneration after injury may require blocking myofibroblast differentiation and/or TGF-β during wound healing. They also highlight hitherto undefined myofibroblast-neuron signaling processes capable of restricting neurite outgrowth in the cornea and other tissues where scars and nerves co-exist.
Topics: Animals; Cats; Cell Differentiation; Cornea; Corneal Injuries; Eye Proteins; Myofibroblasts; Nerve Regeneration; Sensory Receptor Cells; Transforming Growth Factor beta; Wound Healing
PubMed: 30154512
DOI: 10.1038/s41598-018-30964-y -
Methods in Molecular Biology (Clifton,... 2021The stroma constitutes the structural framework of an organ and plays crucial roles in health and following organ damage. The major player of the stroma with respect to...
The stroma constitutes the structural framework of an organ and plays crucial roles in health and following organ damage. The major player of the stroma with respect to extracellular matrix deposition, maintenance, and remodeling is the fibroblast and its activated derivative, the myofibroblast. It has long been recognized that there is considerable variability to the fibroblast phenotype. The recent advent of new single cell "omics" technologies has revolutionized our understanding and appreciation of cellular heterogeneity of fibroblasts been revolutionized. With these tools, the nature and defining characteristics of the cells comprising the stroma is finally being defined not just through a few markers, but by taking a wholistic look at transcriptional programs. It is now apparent that stromal cells are not only transcriptionally diverse, but also functionally, epigenetically, and spatially heterogeneous. Studying populations at single cell resolution has enabled identification of new clusters of cells with unique transcriptional signatures. Whether these clusters truly represent distinct subpopulations or different states of the same population remains to be clarified. In this chapter, we first describe a procedure for purification and preparation of a single cell suspension from tissue samples (in this case the heart) for single cell RNA sequencing. We also describe preparation of high-quality tissue sections for spatial transcriptomics. Secondly, we outline a workflow for computational analysis of single cell RNA sequencing and spatial transcriptomics data, as well as integrating them together, to explore the heterogeneity within fibroblasts/myofibroblasts and identify different subtypes and their locations in the heart.
Topics: Animals; Cell Differentiation; Cells, Cultured; Computational Biology; Extracellular Matrix; Fibroblasts; Gene Expression Profiling; Gene Regulatory Networks; Mice; Myofibroblasts; Sequence Analysis, RNA; Single-Cell Analysis; Workflow
PubMed: 34028734
DOI: 10.1007/978-1-0716-1382-5_4 -
Cells Dec 2020We identify the focal adhesion protein kindlin-2 as player in a novel mechanotransduction pathway that controls profibrotic cardiac fibroblast to myofibroblast...
We identify the focal adhesion protein kindlin-2 as player in a novel mechanotransduction pathway that controls profibrotic cardiac fibroblast to myofibroblast activation. Kindlin-2 is co-upregulated with the myofibroblast marker α-smooth muscle actin (α-SMA) in fibrotic rat hearts and in human cardiac fibroblasts exposed to fibrosis-stiff culture substrates and pro-fibrotic TGF-β1. Stressing fibroblasts using ferromagnetic microbeads, stretchable silicone membranes, and cell contraction agonists all result in kindlin-2 translocation to the nucleus. Overexpression of full-length kindlin-2 but not of kindlin-2 missing a putative nuclear localization sequence (∆NLS kindlin-2) results in increased α-SMA promoter activity. Downregulating kindlin-2 with siRNA leads to decreased myofibroblast contraction and reduced α-SMA expression, which is dependent on CC(A/T)-rich GG(CArG) box elements in the α-SMA promoter. Lost myofibroblast features under kindlin-2 knockdown are rescued with wild-type but not ∆NLS kindlin-2, indicating that myofibroblast control by kindlin-2 requires its nuclear translocation. Because kindlin-2 can act as a mechanotransducer regulating the transcription of α-SMA, it is a potential target to interfere with myofibroblast activation in tissue fibrosis.
Topics: Actins; Adult; Animals; Cell Nucleus; Cells, Cultured; Fibroblasts; Humans; Male; Mechanotransduction, Cellular; Membrane Proteins; Microscopy, Fluorescence; Myofibroblasts; Neoplasm Proteins; Promoter Regions, Genetic; RNA Interference; RNA, Small Interfering; Rats; Transforming Growth Factor beta1
PubMed: 33348602
DOI: 10.3390/cells9122702 -
The Journal of Pathology Nov 2013Fibrosis and scar formation results from chronic progressive injury in virtually every tissue and affects a growing number of people around the world. Myofibroblasts... (Review)
Review
Fibrosis and scar formation results from chronic progressive injury in virtually every tissue and affects a growing number of people around the world. Myofibroblasts drive fibrosis, and recent work has demonstrated that mesenchymal cells, including pericytes and perivascular fibroblasts, are their main progenitors. Understanding the cellular mechanisms of pericyte/fibroblast-to-myofibroblast transition, myofibroblast proliferation and the key signalling pathways that regulate these processes is essential to develop novel targeted therapeutics for the growing patient population suffering from solid organ fibrosis. In this review, we summarize the current knowledge about different progenitor cells of myofibroblasts, discuss major pathways that regulate their transdifferentiation and discuss the current status of novel targeted anti-fibrotic therapeutics in development.
Topics: Animals; Cell Lineage; Cell Proliferation; Cell Transdifferentiation; Cicatrix; Extracellular Matrix; Fibrosis; Humans; Myofibroblasts; Pericytes; Signal Transduction
PubMed: 24006178
DOI: 10.1002/path.4253 -
Experimental Cell Research Jul 2013Cancers become significantly more dangerous when the tumor progresses from in situ, or contained, to an invasive state, in which the cancer cells acquire the ability to... (Review)
Review
Cancers become significantly more dangerous when the tumor progresses from in situ, or contained, to an invasive state, in which the cancer cells acquire the ability to pass through the surrounding basement membrane (BM), a specialized extracellular matrix (ECM) that provides structure and contextual information to the underlying tissue. While the majority of tumors are carcinomas, derived from epithelial cells, it is the stromal cells surrounding the epithelial-derived tumor cells, including fibroblasts and myofibroblasts, vasculature, and immune cells, that are largely responsible for the production and remodeling of the ECM. Here, we will discuss myofibroblasts as key effectors of tumor progression, focusing on recent advances in breast and pancreatic carcinoma, showing how myofibroblasts may function properly in normal tissue remodeling and wound-healing processes, how in the tumor context they can drive cancer invasion and metastasis, and how the pathogenic functions of myofibroblasts may be targeted therapeutically.
Topics: Animals; Cell Transformation, Neoplastic; Humans; Myofibroblasts; Neoplasm Invasiveness; Neoplasms
PubMed: 23528452
DOI: 10.1016/j.yexcr.2013.03.015 -
Oral Diseases Jul 2020Myofibroblasts constitute a specific cell phenotype involved in connective tissue healing. Diabetes alters the wound healing response. However, it is not clear whether...
OBJECTIVES
Myofibroblasts constitute a specific cell phenotype involved in connective tissue healing. Diabetes alters the wound healing response. However, it is not clear whether diabetes modifies the involvement of myofibroblasts in periodontal wounds.
MATERIALS AND METHODS
Type I diabetes was induced in rats through streptozotocin injection, and periodontal wounds were performed. Wound healing was evaluated histologically at 2, 5, 7, and 15 days by measuring epithelial migration, neutrophil infiltration, and collagen and biofilm formation. Distribution of myofibroblasts was evaluated through immunofluorescence for α-smooth muscle actin. Data analyses were performed using the Shapiro-Wilk, ANOVA, or Kruskal-Wallis tests.
RESULTS
Diabetic wounds were characterized by delayed epithelial closure, increased neutrophil infiltration, biofilm formation, and reduced collagen formation. Quantification of the myofibroblasts showed a significant reduction at 5 and 7 days in wounds of diabetic rats and an increase at 15 days when compared to wounds of non-diabetic rats.
CONCLUSIONS
Diabetic wound healing was associated with decreased epithelial and connective tissue healing, increased levels of inflammation, and biofilm formation. Myofibroblast differentiation was delayed in diabetic periodontal wounds at early time points. However, myofibroblasts persisted at later time points of healing. The present study suggests that diabetes alters the involvement of myofibroblasts during periodontal wound healing.
Topics: Animals; Collagen; Diabetes Mellitus, Experimental; Myofibroblasts; Periodontics; Rats; Streptozocin; Wound Healing
PubMed: 32147898
DOI: 10.1111/odi.13325 -
Journal of Cell Science Jul 2020In 1971, Gabbiani and co-workers discovered and characterized the "" (contraction) in rat wound granulation tissue and, accordingly, named these cells 'myofibroblasts'.... (Review)
Review
In 1971, Gabbiani and co-workers discovered and characterized the "" (contraction) in rat wound granulation tissue and, accordingly, named these cells 'myofibroblasts'. Now, myofibroblasts are not only recognized for their physiological role in tissue repair but also as cells that are key in promoting the development of fibrosis in all organs. In this Cell Science at a Glance and the accompanying poster, we provide an overview of the current understanding of central aspects of myofibroblast biology, such as their definition, activation from different precursors, the involved signaling pathways and most widely used models to study their function. Myofibroblasts will be placed into context with their extracellular matrix and with other cell types communicating in the fibrotic environment. Furthermore, the challenges and strategies to target myofibroblasts in anti-fibrotic therapies are summarized to emphasize their crucial role in disease progression.
Topics: Animals; Cell Differentiation; Extracellular Matrix; Fibroblasts; Fibrosis; Myofibroblasts; Rats; Wound Healing
PubMed: 32651236
DOI: 10.1242/jcs.227900 -
Journal of the College of Physicians... Jan 2017Myofibroblast-mediated contraction is viewed as a cycle of four steps. The first step is stimulation of myofibroblasts by lysophospholipids leading to the activation of... (Review)
Review
Myofibroblast-mediated contraction is viewed as a cycle of four steps. The first step is stimulation of myofibroblasts by lysophospholipids leading to the activation of G proteins and ending with contraction of the actin-myosin complex. The next step is the transmission of the intracellular contractile force at the focal adhesions of myofibroblasts; a step that involves talin, vinculin, paxillin, Hic-5, and the integrin receptors. In the third step, fibronectin will act as the extracellular link between the integrin receptors and the extracellular collagen. Finally, "sensing" tension and the maintenance of myofibroblast activity represent the fourth step. The clinical relevance of each step is then discussed in the form of modalities to prevent excessive scarring/fibrosis.
Topics: Animals; Fibroblasts; Fibronectins; Focal Adhesions; Humans; Muscle Contraction; Muscle, Skeletal; Muscle, Smooth; Myofibroblasts; Vinculin; Wound Healing
PubMed: 28292367
DOI: No ID Found -
The Journal of Biological Chemistry Mar 2015Myofibroblasts have increased expression of contractile proteins and display augmented contractility. It is not known if the augmented contractile gene expression...
Myofibroblasts have increased expression of contractile proteins and display augmented contractility. It is not known if the augmented contractile gene expression characterizing the myofibroblast phenotype impacts its intrinsic ability to assemble fibronectin (FN) and extracellular matrix. In this study we investigated whether myofibroblasts displayed increased rates of FN fibril assembly when compared with their undifferentiated counterparts. Freshly plated myofibroblasts assemble exogenous FN (488-FN) into a fibrillar matrix more rapidly than fibroblasts that have not undergone myofibroblast differentiation. The augmented rate of FN matrix formation by myofibroblasts was dependent on intact Rho/Rho kinase (ROCK) and myosin signals inasmuch as treatment with Y27632 or blebbistatin attenuated 488-FN assembly. Inhibiting contractile gene expression by pharmacologic disruption of the transcription factors megakaryoblastic leukemia-1 (MKL1)/serum response factor (SRF) during myofibroblast differentiation resulted in decreased contractile force generation and attenuated 488-FN incorporation although not FN expression. Furthermore, disruption of the MKL1/SRF target gene, smooth muscle α-actin (α-SMA) via siRNA knockdown resulted in attenuation of 488-FN assembly. In conclusion, this study demonstrates a linkage between increased contractile gene expression, most importantly α-SMA, and the intrinsic capacity of myofibroblasts to assemble exogenous FN into fibrillar extracellular matrix.
Topics: Actins; Cell Differentiation; Cells, Cultured; Extracellular Matrix; Fibroblasts; Fibronectins; Humans; Myofibroblasts; Pulmonary Fibrosis; Serum Response Factor; Trans-Activators; Transforming Growth Factor beta
PubMed: 25627685
DOI: 10.1074/jbc.M114.606186