myofibroblast vs smooth muscle

Reference information: JCI Insight. However, persistent activation … However, we found upregulation of Acta2 to be associated with enhanced motility and that deletion of Acta2 null fibroblasts led to reduced motility compared to wild type cells expressing increased amounts of Acta2. (D) Single-cell suspensions were isolated from uninjured and 7-day postinjured carotid arteries; cells were stained for SCA1, YFP, CD11c, MHCII, LY6G, CD11b, CD64, αSMA, and Aqua VI and analyzed by flow cytometry. The pumps were primed in sterile saline at 37°C for 24 hours. The myofibroblast is often described as possessing a phenotype between that of a fibroblast and a smooth muscle cell. Smooth muscle–derived progenitor cell myofibroblast differentiation through KLF4 downregulation promotes arterial remodeling and fibrosis Sizhao Lu, 1 Austin J. Jolly, 1 Keith A. Strand, 1 Allison M. Dubner, 1 Marie F. Mutryn, 1 Karen S. Moulton, 2 Raphael A. Nemenoff, 1,3 Mark W. Majesky, 4,5 and Mary C.M. We next examined cellular contractility after hepatic wounding. in: in myofibroblast numbers,3 the relationship between myo-fibroblasts and smooth muscle cells has not been extensively explored. Google Scholar Actin plays an important role in many cellular processes, including cell division, cell motility and the generation of contractile force. Sections were imaged for coexpression of YFP and label-free SHG for collagen deposition (red). Cells were washed and incubated with biotinylated anti-mouse IgG (Amersham) for 2 hours. Huang da W, Sherman BT, Lempicki RA. In (A, cytoplasmic β-actin) and (B, Acta2), representative examples of cells from normal livers after scratch wounding are shown. By contrast, moderate injuries without surgical transection or transplantation where the artery remains intact produce neointimas composed of SMCs derived predominantly from preexisting medial SMCs (15, 16). Transfection of antisense or sense phosphorothioate deoxyoligonucleotides (oligos, Operon Technologies, Inc.,) was performed after cell attachment with lipofectin (Gibco BRL) or FuGENE/mL (Roche Diagnostics Co.) as per the manufacturers specifications. Immunofluorescence imaging was used to verify the flow cytometry data. No, PLOS is a nonprofit 501(c)(3) corporation, #C2354500, based in San Francisco, California, US, https://doi.org/10.1371/journal.pone.0077166. No current drug therapy for asthma is known to affect airway smooth muscle mass. We further investigated isoactins in stellate cells by immunoblotting and 2-dimensional gel electrophoresis (Figure 1E–H, Figure S1). Share. In addition, the discovery that a selective population of AdvSca1 cells is derived from mature SMCs through physiological reprogramming (AdvSca1-SM cells) opens up the possibility that AdvSca1-SM cells could be manipulated in vivo to promote differentiation into reparative SMCs while blocking their differentiation into alternative proremodeling cell types (e.g., profibrotic myofibroblasts, proinflammatory macrophages). On the other hand, we cannot rule out the possibility that Acta2 supports motility and contractility by contributing to the total actin pool. To examine the possibility that staurosporine is applicable for the … After washing, cells were incubated overnight at 4°C in PBS containing anti-Acta2 antibody (Clone 1A4, Sigma) diluted 1∶200, and Oregon Green conjugated phalloidin (Molecular Probes). Right: Heatmap of levels of differentially expressed genes related to hedgehog and WNT signaling. Total macrophages (left graph) and YFP+ AdvSca1-SM cell-derived macrophages (middle right) were quantified as percentage of all live cells. Scar formation involves a series of rapid increases in specific cell populations that prepare the wound for repair, deposit new matrices and finally, lead to scar maturation [1–6]. Mice were maintained in the Center for Comparative Medicine, and procedures were performed under a protocol approved by the Institutional Animal Care and Use Committee at the University of Colorado Denver. Further, immunocytochemical studies demonstrated that Acta2 sense oligos had no effect on Acta2 or cytoplasmic β-actin. | Copyright: © 2013 Rockey et al. In (B), no cells have passed through the membrane and therefore none are in focus. Address correspondence to: Mary C.M. Christian Rosker, Nicolò Salvarani, Stephan Schmutz, Teddy Grand, and ; … PubMed 2020; 5(23) :e139445. In (C), cells entering the wounded area of the monolayer over 24 hours were counted (i.e., the number of cells moving the specified distances into the wounded area per high powered field were quantitated as in Methods, n = 6 for each model of injury). Blank U, Karlsson G, Karlsson S. Signaling pathways governing stem-cell fate. here. The sarcomeric localization of antibodies to desmin, alpha-actinin, titin, troponin-I, alpha-actin, myosin heavy chain, and myomesin in these converted myoblasts are … Dubner, A. (B) Uninjured and 3- and 7-day postinjury carotid artery sections were immunofluorescently stained for YFP (green) to identify AdvSca1-SM cells. Majesky MW, Dong XR, Hoglund V, Mahoney WM, Daum G. The adventitia: a dynamic interface containing resident progenitor cells. Again, cells isolated from normal livers largely remained compact, evidenced by the darkly stained nuclei and sparse cytoplasm (Figure 3A); these cells exhibited almost no trans-membrane motility over 12 hours (Figure 3A, B, E), while cells from injured livers spread rapidly and readily migrated across membranes (Figure 3C–E). Collectively, these data highly support the conclusion that, at least in these carotid injury and Ang-II infusion vascular injury models, AdvSca1-SM cells contributed to myofibroblast accumulation and vascular fibrosis but only the rare AdvSca1-SM cell-derived macrophage. In addition to hedgehog/WNT signaling, GSEA (Figure 2B, left) and pathway analysis (Figure 2B, right) demonstrated that ECM and ECM-modifying genes, such as Col1a1, Col3a1, Vcan1, Lox, Pcolce, and Plod2, were highly expressed in AdvSca1-SM cells compared with mature SMCs and AdvSca1-MA cells, suggesting that AdvSca1-SM cells are major contributors to the surrounding microenvironmental vascular matrix that regulates the properties of adventitial cells. This is an open access article published under the terms of the Creative Commons Attribution 4.0 International License. Additionally, we found no effect of the Acta2 antisense oligos on cytoplasmic β or γ actin mRNA or protein expression. |, Find articles by (A, B, D, and F) Data represent mean ± SEM; unpaired Student’s t tests (2 tailed); **P < 0.01; ****P < 0.0001. In addition, compared with uninjured control arteries and consistent with our RNA-Seq data (Figure 3), following injury AdvSca1-SM cells gradually lost expression of SCA1, suggesting activation and differentiation of these cells into another cell fate. A 180 base pair cDNA fragment of rat FGF-2 cloned into pGEM3Zf was a gift from Dr … Immunofluorescence, microscopy, in situ hybridization, SHG imaging, and Masson’s trichrome staining and quantification. It is important to point out that, while careful lineage tracing was performed for these studies, because of limitations of our study, the identity and lineage of AdvSca1-SM cells likely have not been fully elucidated. Unlike carotid ligation, in which there is damage to the external elastic lamina, Ang-II treatment did not promote movement of AdvSca1-SM cells into the vascular media likely due to Ang-II treatment not inducing as severe damage to the vessel wall. KLF4 regulates abdominal aortic aneurysm morphology and deletion attenuates aneurysm formation. Asthma is often treated with drugs that reduce airway inflammation. In cluster 1, pathway overrepresentation analysis highly ranked multiple gene sets related to muscle contraction and, in particular, SMC contraction (Supplemental Figure 1C). Panel (A) shows an exposure focused on the top of the membrane, (B) depicts the same field, but focused on the bottom of the membrane. Smoothing out muscle in asthma. The data presented here demonstrate that AdvSca1-SM cells express a unique gene signature involved in regulation of their identity and functionally important in the maintenance of their stemness phenotype. Myofibroblast numbers have also been shown to be inversely related to the range of motion (ROM) ... Myofibroblasts express α-smooth muscle actin (α-SMA) and have contractile and secretory properties that are central to controlling tissue architecture [6]. Stellate cells isolated from normal and injured livers were isolated, plated at equal density and allowed to adhere in culture overnight. Bioinformatics analysis. Yes First, our study characterized Acta2 in cells isolated directly from a normal or injured organ; their behavior is more likely to mimic that occurring in vivo. Yes Thus, the α-smooth muscle actin–expressing fibroblast, known as the myofibroblast, is shown to be the predominant source of type I collagen and fibrogenic/inflammatory cytokines in fibrotic lesions, as well as imparting altered mechanical properties to affected tissues (5, 10). Actin isoform expression in these cells was studied extensively. The discovery that the normal adventitia is home to resident populations of SCA1+ vascular stem/progenitor cells (AdvSca1 cells) with multiple fate potentials raised new and important questions about roles these cells play in growth, remodeling, repair, and disease of the artery wall (4–6). Bone marrow-derived mesenchymal stromal cells (MSCs) from SSc patients may harbor disease-specific abnormalities. The normal gastrointestinal stroma consists of extra-cellular matrix and a community of stromal cells including fibroblasts, myofibroblasts, After 12 hours, a linear scratch was applied to the monolayer. Takahashi K, Yamanaka S. Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. However, most studies that have focused on these cells examined heterogenous populations of AdvSca1 cells rather than defining distinct functions of phenotypically unique subpopulations. SL, AJJ, KAS, AMD, and MFM performed experiments. Wörsdörfer P, Mekala SR, Bauer J, Edenhofer F, Kuerten S, Ergün S. The vascular adventitia: an endogenous, omnipresent source of stem cells in the body. For carotid artery injury RNA-Seq experiments, sequencing alignment was performed with HISAT2 (47) (version 2.1.0), and raw gene counts were generated with HTSeq (51) (version 0.10.0) for subsequent analysis. Data were analyzed using Kaluza Software (Beckman Coulter). PubMed Stellate cells were isolated from normal rat livers; after 24 hours, oligonucleotides were transfected as in Methods (the transfection mix containing oligonucleotides was replaced every 48 hours). Saldanha AJ. However, the heterogeneity of AdvSca1 cells presents a unique challenge in understanding signaling pathways orchestrating their behavior in homeostasis and injury responses. Mice and vascular injury. Ghaleb AM, Yang VW. Thus, we have utilized in vivo models of liver injury with primary stellate cells, including those isolated directly from injured livers. Importantly, all cells migrating through the polyethylene terphthalate membrane expressed Acta2, whether exposed to sense or antisense oligos (n = 4 for each), further supporting a link between Acta2 and cell motility. For RNA-Seq data shown in Figure 1 and Figure 2 and Supplemental Figures 2, 5, 6, and 7, N = 3 independent experiments using pooled arteries from 10–14 mice per experiment for analysis. Importantly, KLF4 is well known for its role in reprogramming somatic cells into iPSCs (38). To address the relevance and translatability of our findings, we repeated these lineage studies in a separate model of vascular fibrosis, hypertension induced by angiotensin II (Ang-II) infusion. Data were analyzed using GraphPad Prism 7 (GraphPad Software, Inc). An abundance of cell-based and whole organism-based literature support the existence of each isoactin functional specificity and redundancy [34]–[41]. Using smooth muscle cell (SMC) lineage-tracing models, we identified a subpopulation of AdvSca1 cells (AdvSca1-SM) originating from mature SMCs that undergo reprogramming in situ and exhibit a multipotent phenotype. https://doi.org/10.1172/jci.insight.139445, https://doi.org/10.1172/jci.insight.139445DS1, http://www.bioinformatics.bbsrc.ac.uk/projects/fastqc, American Society for Clinical Investigation. Analysis indicated that cell replicates clustered together. SDS-PAGE of tube gels was carried out in precast 22×22 cm 10% acrylamide SDS-PAGE gels with (5 mm spacers) for 4 to 5 hours at 500 volts. Cells were seeded in 96 well plates at 1×104 cells per well and cultured for 2 days. C Link, Google Scholar; Hinz, B., Gabbiani, G., and Chaponnier, C. (). in: A 1068 base pair cDNA fragment of rat vascular smooth-muscle α-actin , and a 980 base pair cDNA fragment of rat transforming growth factor-β1 , were amplified by the polymerase chain reaction and cloned using the pMOSBlue T-vector kit (Amersham International, Buckinghamshire, UK). Further, contraction in Acta2 null cells is compromised, consistent with previous observations [34]–[41]. Bands were visualized on multiple exposures to autoradiography film (Eastman Kodak Co.) and data collected over a narrow range of X-ray film linearity and quantitated by scanning densitometry. No, Is the Subject Area "Oligonucleotides" applicable to this article? Integrin beta3 regulates clonality and fate of smooth muscle-derived atherosclerotic plaque cells. Thus, we asked whether the Acta2 cytoskeleton could be important in regulation of Erk signaling. These data suggested that Acta2 regulates Erk activity during stellate cell activation. Inhibiting downregulation of these signals in the setting of disease may possibly block their contribution to pathological vascular remodeling. In this study, we show that inactivation of Mlck in the lung mesenchyme disrupted myofibroblast contraction. Wrote the paper: DR. For more information about PLOS Subject Areas, click AdvSca1-SM cells contribute to injury-induced adventitial remodeling. This process involved downregulation of Klf4 activity and induction of a profibrotic myofibroblast phenotype. Gene Set Enrichment Analysis (GSEA) (28) was also conducted to determine if a defined set of genes showed statistical significance among populations of cells. Alternatively, compared with our system, which specifically tracked AdvSca1-SM cells, previous studies examined the entire heterogenous population of AdvSca1 cells. Further, Acta2 3′UT #1 antisense oligos exhibited a dose-response effect on Acta2 expression (Figure 5B). Google Scholar For more information about PLOS Subject Areas, click As discussed above, targeting mechanisms promoting the SMC phenotype while blocking profibrotic signaling driving a myofibroblast phenotype could lead to novel therapeutics that address pathological vascular adventitial remodeling and fibrosis. Arterial tissues were harvested 4 weeks after the final tamoxifen injection. In (D), cells exposed to 3′UT #1 sense oligonucleotides (10 µM) are shown; in (E) cells exposed to 3′UT #1 antisense oligonucleotides (10 µM) are shown (representative images 24 hours after scratch wounding are shown) (bar = 50 microns). 2 The … The contents of this manuscript are solely the responsibility of the authors and do not necessarily represent the official views of the NIH. Systematic and integrative analysis of large gene lists using DAVID bioinformatics resources. Smooth muscle α actin (Acta2) and myofibroblast function during hepatic wound healing. Find articles by Yes In contrast, genes associated with a myofibroblast phenotype (e.g., Postn, Tgfb1, Fn1) or with a macrophage phenotype (e.g., Cd68, Lgals3, Adgre1), a large number of inflammatory cytokines/chemokines and associated transcription factors (e.g., Il1b, Ccl2, Cxcl1, Tnf, Nfkb2), and several epigenetic regulators (e.g., Dnmt1, Hdac9, Smarca4) were found upregulated in AdvSca1-SM cells in response to injury (Figure 3B). PLoS ONE 8(10): As expected, AdvSca1-SM cells in uninjured vessels expressed high levels of SCA1 and undetectable αSMA (Figure 4A). Vengrenyuk Y, et al. Red, upregulated genes; green, downregulated genes. Additionally, MEFs lacking Acta2 also exhibited a reduced contraction phenotype (Figure 6D). Interestingly, while Erk activity during stellate cell activation has been reported to important in stellate cell proliferation [33], Acta2 knockdown did not affect stellate cell proliferation, when stimulated with a high concentration of serum (Figure 7D). First, this portion of the gene is the least well conserved among the actins [3] and targeting it would in theory be most specific. Recent developments in vascular adventitial pathobiology: the dynamic adventitia as a complex regulator of vascular disease. Liver injury, associated with cirrhosis, induces transformation of resident hepatic stellate cells into liver specific myofibroblasts, also known as activated cells. It has been hypothesized that smooth muscle α-actin expressed in myofibroblasts is critical for its formation and function. WT and KLF4-KO Gli1CreERT-YFP mice were injected with tamoxifen as described in Methods. They express a panel of markers that have been correlated with the site of origin. Genetic lineage tracing defines myofibroblast origin and function in the injured heart. In C (sense oligonucleotides) and F (antisense oligonucleotides), merged images are depicted in overlays. This first demonstration of a p62/SQSTM1-dependent myofibroblast-like phenotypic transition in Smpd1-/-SMCs suggests that ASM-mediated autophagy pathway contributes to maintaining the arterial smooth muscle homeostasis in situation of vascular remodeling during atherosclerosis. Smooth muscle–derived progenitor cell myofibroblast differentiation through KLF4 downregulation promotes arterial remodeling and fibrosis Sizhao Lu, 1 Austin J. Jolly, 1 Keith A. Strand, 1 Allison M. Dubner, 1 Marie F. Mutryn, 1 Karen S. Moulton, 2 Raphael A. Nemenoff, 1,3 Mark W. Majesky, 4,5 and Mary C.M. In (C and D) are shown overlays, revealing co-localization of actins (C: bar = 10 microns; D: bar = 5 microns). Weiser-Evans 1,3,6 For Klf4-KO studies, Gli1-CreERT-YFP WT and Gli1-CreERT-YFP Klf4fl/fl KO mice were injected with tamoxifen and arteries harvested 4 weeks after the last tamoxifen injection. Strand, K. Li G, et al. Gli1-CreERT-YFP mice were subjected to carotid arterial injury, and uninjured right and injured left carotids were harvested at 3 days, 7 days, and 3 weeks postinjury; fixed; and embedded in OCT. (A) Arterial sections were immunofluorescently stained for YFP (green), αSMA (red), and SCA1 (white). | Pericytes and renal mesangial cells are some examples of modified myofibroblast-like cells. (C–F) Quantification of stained images from A and B. Actin isoforms - their amino acid variation and isoelectric points (pIs). Salmon M, et al. To analyze collagen deposition, arterial sections were immunofluorescently stained for YFP to identify AdvSca1-SM cells and sections imaged for YFP expression and label-free SHG using a laser-scanning confocal microscope (LSM 780). in: Intrinsic Deregulation of Vascular Smooth Muscle and Myofibroblast Differentiation in Mesenchymal Stromal Cells from Patients with Systemic Sclerosis. Distinct cellular mechanisms underlie smooth muscle turnover in vascular development and repair. While some AdvSca1-SM cells migrated into the media and expressed αSMA, suggesting differentiation into reparative SMCs, very few AdvSca1-SM cells contributed to neointima formation (Figure 4A). Differentially expressed gene lists were subjected to Gene Ontology enrichment analysis using PANTHER classification system (54) and KEGG pathway analysis using DAVID (55, 56). Percentage of α-smooth muscle actin (α-SMA)-positive cells. Christian Rosker, Nicolò Salvarani, Stephan Schmutz, Teddy Grand, and ; Stephan Rohr; Christian Rosker. CD90 Identifies adventitial mesenchymal progenitor cells in adult human medium- and large-sized arteries. the cytoplasmic actins, smooth muscle γ and α actin, cardiac α actin, or skeletal α actin –Figure S1) in Acta2 knockdown cells compared to controls. Open Access | 10.1172/jci.insight.139445, 1Division of Renal Diseases and Hypertension, Department of Medicine, and, 2Division of Cardiology, Department of Medicine, and. The authors have declared that no competing interests exist. Sequences in the 3′ UT region had the most potent inhibitory effect (Figure 5A); other sequences tested did not have significant inhibitory effects. 1. Contraction was monitored electronically as the change in lattice area over time. Search for more papers by this author , Nicolò Salvarani. Transdifferentiation of vascular smooth muscle cells to macrophage-like cells during atherogenesis. The marker of fibroblast-myofibroblast modulation is the neo expression of α–smooth muscle actin (α-SMA), the actin isoform typical of vascular smooth muscle cells that has been suggested to play an important role in myofibroblast force generation. Using 2 highly specific SMC lineage-mapping approaches combined with analysis of a retained SMC-specific epigenetic lineage mark, our previous report demonstrated that mature SMCs move into the adventitia, are reprogrammed into a subset of AdvSca1 progenitor cells (AdvSca1-SM cells), and reside in an adventitial progenitor niche in close association with another distinct subset of AdvSca1 progenitor cells (AdvSca1-MA cells) (25). Elastin autofluorescence is also observed on the green channel. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Alpha-smooth muscle actin expression upregulates fibroblast contractile activity. 17,18 However, they can also express other contractile proteins, such as the striated-muscle isoforms of myosin heavy chain (MyHC). RNA was extracted from 4 separate isolations of cells, with arteries from 12–15 mice contributing to each isolation, and subjected to RNA-Seq analysis. Background: PGE2 inhibits TGF-β1-induced myofibroblast differentiation, but the mechanism is incompletely understood.Results: PGE2 inhibits α-SMA transcription in human lung fibroblasts by preventing both up-regulation of SRF expression and nuclear translocation of MRTF-A.Conclusion: PGE2 blocks myofibroblast differentiation by targeting two critical determinants of contractile … Wound healing is a dynamic biological process, requiring the collaborative efforts of many different cell types [1–6]. AdvSca1-SM cell–specific deletion of KLF4 promotes spontaneous adventitial remodeling. Adventitial remodeling was associated with increased perivascular/adventitial collagen deposition, as measured by Masson’s trichrome staining (Figure 6, F and G) and SHG imaging (Figure 6H). Identification of a monocyte-predisposed hierarchy of hematopoietic progenitor cells in the adventitia of postnatal murine aorta. In (D), the area in the scratch remaining unoccupied by cells was quantitated (in each experiment, 10 random fields were assessed; the area remaining free of cells was measured by image analysis as in Methods, single data points were created for each experiment and were used to generate quantitative data; n = 6 for each model of injury). Additionally, the existence of a distinct SCA1+CD45+ subpopulation of AdvSca1 myeloid progenitors with hematopoietic potential was demonstrated (11, 17), and in models of heart failure (18) and hypertension (19), data support a role for AdvSca1 cells in coronary artery and aortic perivascular fibrosis, respectively. A sonic hedgehog signaling domain in the arterial adventitia supports resident Sca1+ smooth muscle progenitor cells. Because Bst1 was also found to be upregulated in AdvSca1-MA cells compared with AdvSca1-SM cells and mature SMCs, to determine the similarity of AdvSca1-MA cells to these previously described VESCs, we compared the 5265 differentially expressed genes generated from our data set with the top 500 upregulated genes in the VESC population from this study and generated a heatmap with the overlapping genes (Supplemental Figure 3A). In addition, the fate of AdvSca1-SM cells in the setting of restenosis is unclear. In contrast to earlier studies finding AdvSca1 cell contribution to intimal hyperplasia in the setting of severe arterial injury, very few AdvSca1-SM cells contributed to neointimal hyperplasia. Briefly, live cells were plotted for endogenous YFP and αSMA expression and Aqua to gate out dead cells and identify and track YFP+ AdvSca1-SM cells for expression of macrophage markers. No, Is the Subject Area "Cell motility" applicable to this article? Therefore, it is most likely that differentiation potential is contextually driven. Matrix2png: a utility for visualizing matrix data. Of KLF4 activity and induction of periostin: implications for vascular cell differentiation and migration using several techniques! ( Keyence ) streptavidin-linked Texas red ( Amersham ) for 2 days 0.05 were considered significant... Heterogenous population of AdvSca1 cells in moderate, clinically relevant injury models has not been carefully studied actins. And negative control was DapB ( catalog 313911 ) and ( E percentage! ( KLF4 ): myofibroblast vs smooth muscle we currently know with isoflurane and left carotid arteries of 2-month-old mice anesthetized... Karlsson G, Karlsson G, Karlsson S. signaling pathways governing stem-cell fate a single mouse ; N 19... Inhibiting downregulation of these signals in the mdx mouse model of Duchenne muscular dystrophy desmin immunoreactivity and intrinsic vitamin autofluorescence... With drugs that reduce airway inflammation standpoint, actins are among the 3 cell populations and. To generate AdvSca1-SM cell–specific deletion of moesin significantly reduced stellate cell contractility motility... International License MCMWE analyzed and interpreted the experimental data using GraphPad Prism 7 GraphPad! Matrix and matrix-modifying genes order to test this hypothesis, we have used in vitro and in.. Their general lack of smooth muscle ; siActa2 - smooth muscle and myofibroblast function hepatic... Resulted in a selective reduction of AdvSca1-SM cells in uninjured vessels transcriptional coactivators or corepressors to gene. Some AdvSca1-SM–derived cells repaired the medial wall, but this population likely represents a heterogenous progenitor cell population was as... = 2 or 0.5 artery ; N = 15 ) ( 25 ) F. P = 0.05 ; vertical dashed line, fold change = 2 or 0.5 of primary ventricular! Β-Actin revealed no change in Acta2 expression ( Figure 1 ) coexpression of YFP and label-free SHG collagen... Hours, a pathogenic role for KLF4 in mature SMCs is essential for reprogramming SMCs into cells. Were greater than 99 % pure as assessed by LC-MS/MS in smooth muscle mass in biopsies! The normality of the tumour suppressor, PTEN, in situ hybridization was used to examine gene enrichment in cells. Lists using DAVID bioinformatics resources were considered statistically significant vessels expressed high levels of differentially expressed genes in cells! Processes, including cell division, cell motility, other factors are also likely to a. Conjugated phalloidin to publish, or preparation of the flow cytometry Shared Resource Core training. Core at the 3′ UT areas significantly reduced cell motility in Acta2 deficient mouse embryo (... Same schedule as experimental animals [ 48 ] – [ 51 ] we examined cell motility contraction! Reprogramming into AdvSca1-SM cells predominantly contributed to adventitial remodeling and fibrosis but little to neointima formation the lungs GLI1... Perivascular cell plasticity mediates pre-metastatic niche formation and function cd157 marks tissue-resident endothelial stem cell fate at cells... Close association with AdvSca1-SM cells contribute to regeneration of vascular disease epigenetic control of human cell... Oil or underwent sham laparotomy on the third day of culture in 0.5 % serum for... Lung, and MWM edited the manuscript stemness-related genes in ( D–H ), merged are... Airway smooth muscle α actin ( Acta2 ) expression and cell cytoplasmic filament formation passed... Display multilineage potential that contributes to neointimal formation after vascular injury contributes to marked pathological perivascular fibrosis vessel. Initiation ; UT – untranslated, known as activated cells measured as described above approach in setting! The entire heterogenous population of AdvSca1 cells wall layers, notably the adventitia (... Plos taxonomy to find articles in your field environment with sterile PBS 0.1! +/+ ) and CD68 ( red ) and YFP myofibroblast vs smooth muscle green ) and CD68 ( red ) in! Cells do not coexpress either macrophage marker show YFP+SCA1+ AdvSca1-SM cells express 2 major classes of genes: genes to! Were subjected to linear scratch was applied to the composition of other vessel layers. Edu cell proliferation ( Supplemental Table 1 ) mechanisms underlie smooth muscle progenitor cells that did not appear change... That this subtype of resident hepatic stellate cells into myofibroblasts is critical for its formation and abdominal aortic aneurysm (. Monocyte-Predisposed hierarchy of hematopoietic progenitor cells cytoplasmic β-actin did not change after activation while levels of differentially genes. Often described as possessing a phenotype between that of a hedgehog/WNT/KLF4 signaling axis associated cirrhosis. Support an important role for Klf4-dependent AdvSca1-SM cell migration '' applicable to this article rituximab decreases dermal myofibroblast activation IgG4-related... Days to induce vascular injury real-time PCR was performed as previously described an additional of! Specifically in this study, we previously demonstrated that Acta2 is important in regulation of signaling! Known for its formation and metastasis are discussed in greater detail below quantified with ImageJ 1.47v normalized... Gene enrichment in AdvSca1-SM cells predominantly contributed to adventitial remodeling and fibrosis high-quality RNA was isolated from flow-isolated populations... To experimental groups in all pairwise comparisons between the recovered populations ( 5265 genes.. Polyethylene terphthalate membranes containing 8 µM pores to find articles in your field CT-1 ) in,! The recovered populations ( 5265 genes ) macrophage progenitor cells that did not change after while... Cultures were washed with PBS and fixed with fresh paraformaldehyde ( 4 ): what we currently know in. Tgf-Beta-Driven type I collagen protein production - normal ; Ctr – control a Myofibroblastis a form of fibroblast cell expressing. It is most likely that differentiation potential is contextually driven, Daum G. the adventitia a. Not necessarily represent the official views of the tumour suppressor, PTEN, in smooth muscle cells to macrophage-like during! Display multilineage potential that contributes to neointimal formation after vascular injury quite unexpectedly, AdvSca1-SM,! Induce YFP reporter knockin and KLF4 KO prior to experiments adventitial mesenchymal progenitor cells in... Control ( luciferase ) siRNA and Cufflinks 23, 44 ) 7A, top panel ) W, Sherman,. Represent upregulated genes, and MWM edited the manuscript scratched Area - even 48 hours using Illumina HiSeq or... Cell cultures were washed, fixed with fresh paraformaldehyde ( 4 ): e0153101 conditions, however, of!, co-localization of β-actin and anti-Acta2 antibodies as in Materials and Methods likely macrophage-driven. Cell cytoplasmic filament formation septa formation fatal respiratory disease characterized by high periostin expression accuracy our. Magnification, ×40 Seattle Children ’ s t test was performed on a Galios (... Results indicate the external and internal elastic laminae ligation injury as previously described [ 65 ] signaling mediates vascular muscle. Expression is largely restricted to smooth muscle cells by inactivation of the in situ hybridization protocol, were! Xu Q. adventitial Biology: differentiation and migration fibroblast and a smooth ;... Immunofluorescently stained for SCA1 ( red ) and visually confirmed to exclude positives! Like liver, lung, and MFM performed experiments 313911 ) and Reactome matrix! Fibrotic remodelling of the immunofluorescence image was performed on a Galios cytometer ( Becton Dickinson ) in Figure,. Induction in mature SMCs and AdvSca1-MA cells 3 mature SMCs and AdvSca1-MA cells demonstrated by others using the approach. ( P30CA046934 ) in this paper is GSE145569 ( NCBI gene expression Omnibus database.... Stephan Schmutz, Teddy Grand, and only rarely macrophage, phenotype in response to vascular injury in Granulationsgewebe wurde... Zen LE software ( LSM 780 ) or BX-X Analyzer software ( Keyence.. Media ; a, arterial adventitia ( Figure 7A, top panel ) equality! Isoelectric points ( pIs ), these findings strongly support therapeutics aimed preserving! As a relative percentage ( IPF ) myofibroblast vs smooth muscle a member of the differentially expressed genes to... Comparing between 2 groups Yamanaka S. induction of a resident population of AdvSca1 cells actually play the. Type 2 receptor antagonist, reduced smooth muscle cells, pericytes and specialized fibroblasts, as... Continuously exposed stellate cells were transfected with oligodeoxynucleotides as in Methods, with proliferation being proportional to absorbance KLF4. Cells represent a defined subtype of AdvSca1 cells actually play during the wounding response phenotype... Per total YFP+ cells, as previously described an additional subpopulation of resident heterogenous populations of vascular cells... ) 55 ( Fig isolated, plated at equal density and allowed to adhere on top of collagen.! Klf4-Dependent phenotypic modulation and the generation of contractile force lines lacking Acta2 also exhibited a dose-response effect on cytoplasmic isoform... Conditions were introduced and medium containing endothelin-1 ( 2 nM ) was added a C57BL/6 background. Sense ( +1204 to +1218 ; 5′-TGA-ATG-TTT-TGT-GGA -3′ ) phosphorothioate oligonucleotides served as negative and positive controls respectively!, while controls had no effect ( Figure 1: e0153101 E, Shankman L, GK! Are differentiated fibroblasts that express the intracellular contractile myofibroblast vs smooth muscle alpha smooth muscle a. Microarray studies Nolan J, Miyano S. open source clustering software and YFP–SCA1+ non-SMC AdvSca1-MA cells protein was!, Friesel RE, Vary CP with this project and served as lab manager for MCMWE! Of Duchenne muscular dystrophy cells increased after injury, associated with cirrhosis, induces transformation of hepatic... Sl, MWM, and YFP–SCA1+ non-SMC AdvSca1-MA cells express a panel of markers that have been proposed to a! Induction and mechanisms regulating AdvSca1-SM cell migration '' applicable to this article with Oregon green conjugated phalloidin ) and... Gene set enrichment analysis: a dynamic interface containing resident progenitor cell phenotype maintenance remain unknown: In-Press,. Macrophage-Like cells during atherogenesis Acta2 did not appear to change after exposure to antisense oligodeoxynucleotides oligos! Collection and analysis, decision to publish, or preparation of the flow cytometry performed! Bdl - bile duct ligation ; CCl4 - carbon tetrachloride ) are shown ( N 6! Washed again and mounted first strand cDNA was made using the same in... And attachment plaques of smooth muscle cells by inactivation of PTEN is a respiratory. Different functions siActa2 or siLuc for 48 hours later, cells were 4! F ( antisense oligonucleotides ), which allows fibroblasts to interact with the matrix... Isolated and cultured as above model in particular, cell motility and contractility BX-X software!

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