scholarly journals The histone demethylase Jumonji domain-containing protein 3 (JMJD3) regulates fibroblast activation in systemic sclerosis

2017 ◽  
Vol 77 (1) ◽  
pp. 150-158 ◽  
Author(s):  
Christina Bergmann ◽  
Amelie Brandt ◽  
Benita Merlevede ◽  
Ludwig Hallenberger ◽  
Clara Dees ◽  
...  
Keyword(s):  
Systemic Sclerosis ◽  
Transforming Growth Factor Beta ◽  
Topoisomerase I ◽  
Transforming Growth Factor ◽  
Dermal Fibroblasts ◽  
Dependent Manner ◽  
Murine Models ◽  
Cultured Fibroblasts ◽  
Pharmacological Inhibition ◽  
Fibroblast Activation

ObjectivesSystemic sclerosis (SSc) fibroblasts remain activated even in the absence of exogenous stimuli. Epigenetic alterations are thought to play a role for this endogenous activation. Trimethylation of histone H3 on lysine 27 (H3K27me3) is regulated by Jumonji domain-containing protein 3 (JMJD3) and ubiquitously transcribed tetratricopeptide repeat on chromosome X (UTX) in a therapeutically targetable manner. The aim of this study was to explore H3K27me3 demethylases as potential targets for the treatment of fibrosis.MethodsJMJD3 was inactivated by small interfering RNA-mediated knockdown and by pharmacological inhibition with GSKJ4. The effects of targeted inactivation of JMJD3 were analysed in cultured fibroblasts and in the murine models of bleomycin-induced and topoisomerase-I (topoI)-induced fibrosis. H3K27me3 at the FRA2 promoter was analysed by ChIP.ResultsThe expression of JMJD3, but not of UTX, was increased in fibroblasts in SSc skin and in experimental fibrosis in a transforming growth factor beta (TGFβ)-dependent manner. Inactivation of JMJD3 reversed the activated fibroblast phenotype in SSc fibroblasts and prevented the activation of healthy dermal fibroblasts by TGFβ. Pharmacological inhibition of JMJD3 ameliorated bleomycin-induced and topoI-induced fibrosis in well-tolerated doses. JMJD3 regulated fibroblast activation in a FRA2-dependent manner: Inactivation of JMJD3 reduced the expression of FRA2 by inducing accumulation of H3K27me3 at the FRA2 promoter. Moreover, the antifibrotic effects of JMJD3 inhibition were reduced on knockdown of FRA2.ConclusionWe present first evidence for a deregulation of JMJD3 in SSc. JMJD3 modulates fibroblast activation by regulating the levels of H3K27me3 at the promoter of FRA2. Targeted inhibition of JMJD3 limits the aberrant activation of SSc fibroblasts and exerts antifibrotic effects in two murine models.

X-linked inhibitor of apoptosis protein (XIAP) inhibition in systemic sclerosis (SSc)

2021 ◽  
pp. annrheumdis-2020-219822
Author(s):  
Christina Bergmann ◽  
Ludwig Hallenberger ◽  
Sara Chenguiti Fakhouri ◽  
Benita Merlevede ◽  
Amelie Brandt ◽  
...  
Keyword(s):  
Systemic Sclerosis ◽  
Transforming Growth Factor Beta ◽  
Topoisomerase I ◽  
Transforming Growth Factor ◽  
Inhibitor Of Apoptosis ◽  
Dependent Manner ◽  
Cultured Fibroblasts ◽  
Multifunctional Protein ◽  
Inhibitor Of Apoptosis Protein ◽  
Apoptosis Protein

ObjectiveX-linked inhibitor of apoptosis protein (XIAP) is a multifunctional protein with important functions in apoptosis, cellular differentiation and cytoskeletal organisation and is emerging as potential target for the treatment of various cancers. The aim of the current study was to investigate the role of XIAP in the pathogenesis of systemic sclerosis (SSc).MethodsThe expression of XIAP in human skin samples of patients with SSc and chronic graft versus host disease (cGvHD) and healthy individuals was analysed by quantitative PCR, immunofluorescence (IF) and western blot. XIAP was inactivated by siRNA-mediated knockdown and pharmacological inhibition. The effects of XIAP inactivation were analysed in cultured fibroblasts and in the fibrosis models bleomycin-induced and topoisomerase-I-(topoI)-induced fibrosis and in Wnt10b-transgenic mice.ResultsThe expression of XIAP, but not of other inhibitor of apoptosis protein family members, was increased in fibroblasts in SSc and sclerodermatous cGvHD. Transforming growth factor beta (TGF-β) induced the expression of XIAP in a SMAD3-dependent manner. Inactivation of XIAP reduced WNT-induced fibroblast activation and collagen release. Inhibition of XIAP also ameliorated fibrosis induced by bleomycin, topoI and overexpression of Wnt10b in well-tolerated doses. The profibrotic effects of XIAP were mediated via WNT/β-catenin signalling. Inactivation of XIAP reduces binding of β-catenin to TCF to in a TLE-dependent manner to block WNT/β-catenin-dependent transcription.ConclusionsOur data characterise XIAP as a novel link between two core pathways of fibrosis. XIAP is overexpressed in SSc and cGvHD in a TGF-β/SMAD3-dependent manner and in turn amplifies the profibrotic effects of WNT/β-catenin signalling on fibroblasts via transducin-like enhancer of split 3. Targeted inactivation of XIAP inhibits the aberrant activation of fibroblasts in murine models of SSc.

Cartilage oligomeric matrix protein expression in systemic sclerosis reveals heterogeneity of dermal fibroblast responses to transforming growth factor β

2008 ◽  
Vol 68 (3) ◽  
pp. 435-441 ◽  
Author(s):  
G Farina ◽  
R Lemaire ◽  
P Pancari ◽  
J Bayle ◽  
R L Widom ◽  
...  
Keyword(s):  
Systemic Sclerosis ◽  
Growth Factor ◽  
Mrna Expression ◽  
Cartilage Oligomeric Matrix Protein ◽  
Matrix Protein ◽  
Transforming Growth Factor ◽  
Dermal Fibroblast ◽  
Smooth Muscle Actin ◽  
Dermal Fibroblasts ◽  
Cultured Fibroblasts

Objective:Cartilage oligomeric matrix protein (COMP) accumulates in systemic sclerosis (SSc) skin and is upregulated by transforming growth factor (TGF)β. To further characterise the response to TGFβ in SSc, we investigated TGFβ1 and COMP expression and myofibroblast staining in SSc skin.Methods:Skin biopsies from patients with diffuse cutaneous SSc (dSSc), limited cutaneous SSc (lSSc) and healthy controls were evaluated for COMP mRNA expression using real-time PCR. COMP, α-smooth muscle actin (SMA) and TGFβ were assessed in skin sections and in cultured fibroblasts by immunohistochemistry. Clinical disease status was assessed by the modified Rodnan skin score (mRSS).Results:Myofibroblasts expressing SMA and COMP were found coexpressed in many cells in dSSc dermis, but each also stained distinct cells in the dermis. Cultured SSc dermal fibroblasts also showed heterogeneity for COMP and SMA expression, with cells expressing SMA, COMP, both or neither. TGFβ treatment increased COMP and SMA-expressing cells. COMP mRNA expression in lesional skin from patients with dSSc correlated with the mRSS and TGFβ1 staining.Conclusion:These findings suggest that TGFβ upregulation of COMP and/or SMA expression in subpopulations of fibroblasts contributes to different pathways of fibrosis and that multiple TGFβ regulated genes may serve as biomarkers for the degree of SSc skin involvement.

scholarly journals The Cell-Permeable Derivative of the Immunoregulatory Metabolite Itaconate, 4-Octyl Itaconate, Is Anti-Fibrotic in Systemic Sclerosis

Cells ◽  
2021 ◽  
Vol 10 (8) ◽  
pp. 2053
Author(s):  
John Henderson ◽  
Sharadha Dayalan Naidu ◽  
Albena T. Dinkova-Kostova ◽  
Stefan Przyborski ◽  
Richard Stratton ◽  
...  
Keyword(s):  
Gene Expression ◽  
Systemic Sclerosis ◽  
Transforming Growth Factor Beta ◽  
Target Genes ◽  
Transforming Growth Factor ◽  
Dermal Fibroblasts ◽  
Heme Oxygenase 1 ◽  
Quinone Oxidoreductase ◽  
Tgf Β1

Systemic sclerosis (SSc) is an autoimmune connective tissue disease that leads to skin fibrosis. Altered metabolism has recently been described in autoimmune diseases and SSc. Itaconate is a product of the Krebs cycle intermediate cis-aconitate and is an immunomodulator. This work examines the role of the cell-permeable derivative of itaconate, 4-octyl itaconate (4-OI), in SSc. SSc and healthy dermal fibroblasts were exposed to 4-OI. The levels of collagen Nrf2-target genes and pro-inflammatory cytokines interleukin 6 (IL-6) and monocyte chemotactic protein 1 (MCP-1) were determined. Levels of reactive oxygen species (ROS) as well as the gene expression of collagen and Cellular Communication Network Factor 2 (CCN2) were measured after transforming growth factor beta 1 (TGF-β1) stimulation in the presence or absence of 4-OI. Wild-type or Nrf2-knockout (Nrf2-KO) mouse embryonic fibroblasts (MEFs) were also treated with 4-OI to determine the role of Nrf2 in 4-OI-mediated effects. 4-OI reduced the levels of collagen in SSc dermal fibroblasts. Incubation with 4-OI led to activation of Nrf2 and its target genes heme oxygenase 1 (HO-1) and NAD(P)H quinone oxidoreductase 1 (NQO1). 4-OI activated antioxidant response element (ARE)-dependent gene expression, reduced inflammatory cytokine release and reduced TGF-β1-induced collagen and ROS production in dermal fibroblasts. The effects of 4-OI are dependent on Nrf2. The cell-permeable derivative of itaconate 4-OI is anti-fibrotic through upregulation of Nrf2 and could be a potential therapeutic option in an intractable disease.

scholarly journals Therapeutic interleukin-6 blockade reverses transforming growth factor-beta pathway activation in dermal fibroblasts: insights from the faSScinate clinical trial in systemic sclerosis

2018 ◽  
Vol 77 (9) ◽  
pp. 1362-1371 ◽  
Author(s):  
Christopher P Denton ◽  
Voon H Ong ◽  
Shiwen Xu ◽  
Haiyin Chen-Harris ◽  
Zora Modrusan ◽  
...  
Keyword(s):  
Systemic Sclerosis ◽  
Growth Factor ◽  
Transforming Growth Factor Beta ◽  
Interleukin 6 ◽  
Transforming Growth Factor ◽  
Expression Profiles ◽  
Dermal Fibroblasts ◽  
Sequencing Analysis ◽  
Growth Factor Beta

ObjectivesSkin fibrosis mediated by activated dermal fibroblasts is a hallmark of systemic sclerosis (SSc), especially in the subset of patients with diffuse disease. Transforming growth factor-beta (TGFβ) and interleukin-6 (IL-6) are key candidate mediators in SSc. Our aim was to elucidate the specific effect of IL-6 pathway blockade on the biology of SSc fibroblasts in vivo by using samples from a unique clinical experiment—the faSScinate study—in which patients with SSc were treated for 24 weeks with tocilizumab (TCZ), an IL-6 receptor-α inhibitor.MethodsWe analysed the molecular, functional and genomic characteristics of explant fibroblasts cultured from matched skin biopsy samples collected at baseline and at week 24 from 12 patients receiving placebo (n=6) or TCZ (n=6) and compared these with matched healthy control fibroblast strains.ResultsThe hallmark functional and molecular-activated phenotype was defined in SSc samples and was stable over 24 weeks in placebo-treated cases. RNA sequencing analysis robustly defined key dysregulated pathways likely to drive SSc fibroblast activation in vivo. Treatment with TCZ for 24 weeks profoundly altered the biological characteristics of explant dermal fibroblasts by normalising functional properties and reversing gene expression profiles dominated by TGFβ-regulated genes and molecular pathways.ConclusionsWe demonstrated the exceptional value of using explant dermal fibroblast cultures from a well-designed trial in SSc to provide a molecular framework linking IL-6 to key profibrotic pathways. The profound impact of IL-6R blockade on the activated fibroblast phenotype highlights the potential of IL-6 as a therapeutic target in SSc and other fibrotic diseases.Trial registration numberNCT01532869; Post-results.

scholarly journals IL11 is elevated in systemic sclerosis and IL11-dependent ERK signaling underlies TGFβ-mediated activation of dermal fibroblasts

Rheumatology ◽  
2021 ◽  
Author(s):  
Eleonora Adami ◽  
Sivakumar Viswanathan ◽  
Anissa A Widjaja ◽  
Benjamin Ng ◽  
Sonia Chothani ◽  
...  
Keyword(s):  
Systemic Sclerosis ◽  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Neutralizing Antibody ◽  
Dermal Fibroblasts ◽  
Lung Fibroblasts ◽  
Loss Of Function ◽  
Autocrine Loop ◽  
Erk Activation ◽  
Erk Activity

Abstract Objectives Interleukin 11 (IL11) is highly upregulated in skin and lung fibroblasts from patients with systemic sclerosis (SSc). Here we tested whether IL11 is mechanistically linked with activation of human dermal fibroblasts (HDFs) from patients with SSc or controls. Methods We measured serum IL11 levels in volunteers and patients with early diffuse SSc and manipulated IL11 signalling in HDFs using gain- and loss-of-function approaches that we combined with molecular and cellular phenotyping. Results In patients with SSc, serum IL11 levels are elevated as compared to healthy controls. All transforming growth factor beta (TGFβ) isoforms induced IL11 secretion from HDFs, which highly express IL11RA and the gp130 co-receptor, suggestive of an autocrine loop of IL11 activity in HDFs. IL11 stimulated ERK activation in HDFs and resulted in HDF-to-myofibroblast transformation and extracellular matrix secretion. The pro-fibrotic action of IL11 in HDFs appeared unrelated to STAT3 activity, independent of TGFβ upregulation and was not associated with phosphorylation of SMAD2/3. Inhibition of IL11 signaling using either a neutralizing antibody against IL11 or siRNA against IL11RA reduced TGFβ-induced HDF proliferation, matrix production and cell migration, which was phenocopied by pharmacologic inhibition of ERK. Conclusions These data reveal that autocrine IL11-dependent ERK activity alone, or downstream of TGFβ stimulation, promotes fibrosis phenotypes in dermal fibroblasts and suggest IL11 as a potential therapeutic target in SSc.

Acyltransferase skinny hedgehog regulates TGFβ-dependent fibroblast activation in SSc

2019 ◽  
Vol 78 (9) ◽  
pp. 1269-1273 ◽  
Author(s):  
Ruifang Liang ◽  
Rosebeth Kagwiria ◽  
Ariella Zehender ◽  
Clara Dees ◽  
Christina Bergmann ◽  
...  
Keyword(s):  
Long Range ◽  
Transforming Growth Factor ◽  
Human Fibroblasts ◽  
Transforming Growth Factor Β ◽  
Dermal Fibroblasts ◽  
Myofibroblast Differentiation ◽  
Dependent Manner ◽  
Hedgehog Signalling ◽  
Fibroblast Activation ◽  
Novel Approach

ObjectivesSystemic sclerosis (SSc) is characterised by aberrant hedgehog signalling in fibrotic tissues. The hedgehog acyltransferase (HHAT) skinny hedgehog catalyses the attachment of palmitate onto sonic hedgehog (SHH). Palmitoylation of SHH is required for multimerisation of SHH proteins, which is thought to promote long-range, endocrine hedgehog signalling. The aim of this study was to evaluate the role of HHAT in the pathogenesis of SSc.MethodsExpression of HHAT was analysed by real-time polymerase chain reaction(RT-PCR), immunofluorescence and histomorphometry. The effects of HHAT knockdown were analysed by reporter assays, target gene studies and quantification of collagen release and myofibroblast differentiation in cultured human fibroblasts and in two mouse models.ResultsThe expression of HHAT was upregulated in dermal fibroblasts of patients with SSc in a transforming growth factor-β (TGFβ)/SMAD-dependent manner. Knockdown of HHAT reduced TGFβ-induced hedgehog signalling as well as myofibroblast differentiation and collagen release in human dermal fibroblasts. Knockdown of HHAT in the skin of mice ameliorated bleomycin-induced and topoisomerase-induced skin fibrosis.ConclusionHHAT is regulated in SSc in a TGFβ-dependent manner and in turn stimulates TGFβ-induced long-range hedgehog signalling to promote fibroblast activation and tissue fibrosis. Targeting of HHAT might be a novel approach to more selectively interfere with the profibrotic effects of long-range hedgehog signalling.

PGC-1α regulates autophagy to promote fibroblast activation and tissue fibrosis

2020 ◽  
Vol 79 (9) ◽  
pp. 1227-1233 ◽  
Author(s):  
Yun Zhang ◽  
Lichong Shen ◽  
Honglin Zhu ◽  
Katja Dreissigacker ◽  
Diana Distler ◽  
...  
Keyword(s):  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Therapeutic Potential ◽  
Human Fibroblasts ◽  
Dermal Fibroblasts ◽  
Skin Fibrosis ◽  
Type I ◽  
Myofibroblast Differentiation ◽  
Tissue Fibrosis ◽  
Fibroblast Activation

ObjectivesCoactivators are a heterogeneous family of transcriptional regulators that are essential for modulation of transcriptional outcomes and fine-tune numerous cellular processes. The aim of the present study was to evaluate the role of the coactivator peroxisome proliferator-activated receptor gamma coactivator-1α (PGC-1α) in the pathogenesis of systemic sclerosis (SSc).MethodsExpression of PGC-1α was analysed by real-time PCR, western blot and immunofluorescence. Modulation of autophagy was analysed by reporter studies by expression of autophagy-related genes. The effects of PGC-1α knockdown on collagen production and myofibroblast differentiation were analysed in cultured human fibroblasts and in two mouse models with fibroblast-specific knockout of PGC-1α.ResultsThe expression of PGC-1α was induced in dermal fibroblasts of patients with SSc and experimental murine fibrosis. Transforming growth factor beta (TGFβ), hypoxia and epigenetic mechanisms regulate the expression of PGC-1α in fibroblasts. Knockdown of PGC-1α prevented the activation of autophagy by TGFβ and this translated into reduced fibroblast-to-myofibroblast differentiation and collagen release. Knockout of PGC-1α in fibroblasts prevented skin fibrosis induced by bleomycin and by overexpression of a constitutively active TGFβ receptor type I. Moreover, pharmacological inhibition of PGC-1α by SR18292 induced regression of pre-established, bleomycin-induced skin fibrosis.ConclusionPGC-1α is upregulated in SSc and promotes autophagy to foster TGFβ-induced fibroblast activation. Targeting of PGC-1α prevents aberrant autophagy, inhibits fibroblast activation and tissue fibrosis and may over therapeutic potential.

scholarly journals Tribbles homologue 3 stimulates canonical TGF-β signalling to regulate fibroblast activation and tissue fibrosis

2015 ◽  
Vol 75 (3) ◽  
pp. 609-616 ◽  
Author(s):  
Michal Tomcik ◽  
Katrin Palumbo-Zerr ◽  
Pawel Zerr ◽  
Barbora Sumova ◽  
Jerome Avouac ◽  
...  
Keyword(s):  
Cell Activation ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β ◽  
Dependent Manner ◽  
Cultured Fibroblasts ◽  
Fibroblast Activation ◽  
Modulation Of Gene Expression ◽  
Β Receptor ◽  
Interfering Rna

ObjectivesTribbles homologue 3 (TRB3) is a pseudokinase that modifies the activation of various intracellular signalling pathways to control fundamental processes extending from mitosis and cell activation to apoptosis and modulation of gene expression. Here, we aimed to analyse the role of TRB3 in fibroblast activation in systemic sclerosis (SSc).MethodsThe expression of TRB3 was quantified by quantitative PCR, western blot and immunohistochemistry. The role of TRB3 was analysed in cultured fibroblasts and in experimental fibrosis using small interfering RNA (siRNA)-mediated knockdown and overexpression of TRB3.ResultsTRB3 expression was increased in fibroblasts of patients with SSc and in murine models of SSc in a transforming growth factor-β (TGF-β)/Smad-dependent manner. Overexpression of TRB3 stimulated canonical TGF-β signalling and induced an activated phenotype in resting fibroblasts. In contrast, knockdown of TRB3 reduced the profibrotic effects of TGF-β and decreased the collagen synthesis. Moreover, siRNA-mediated knockdown of TRB3 exerted potent antifibrotic effects and ameliorated bleomycin as well as constitutively active TGF-β receptor I-induced fibrosis with reduced dermal thickening, decreased hydroxyproline content and impaired myofibroblast differentiation.ConclusionsThe present study characterises TRB3 as a novel profibrotic mediator in SSc. TGF-β induces TRB3, which in turn activates canonical TGF-β/Smad signalling and stimulates the release of collagen, thereby inducing a positive feedback loop that may contribute to aberrant TGF-β signalling in SSc.

scholarly journals P149 The intracellular chloride channel 4 (CLIC4) plays an important role in systemic sclerosis fibroblast activation

Rheumatology ◽  
2021 ◽  
Vol 60 (Supplement_1) ◽  
Author(s):  
Christopher Wasson ◽  
Rebecca Ross ◽  
Ruth Morton ◽  
Jamel Mankouri ◽  
Francesco Del Galdo
Keyword(s):  
Systemic Sclerosis ◽  
Ion Channel ◽  
Smooth Muscle Actin ◽  
Chloride Ion ◽  
Dermal Fibroblasts ◽  
Cancer Associated Fibroblasts ◽  
Alpha Smooth Muscle Actin ◽  
Direct Role ◽  
Fibroblast Activation ◽  
Intracellular Chloride

Abstract Background/Aims  The intracellular chloride ion channel CLIC4 mediates the activation of cancer associated fibroblasts. Interestingly, systemic sclerosis (SSc) fibroblasts display a number of similar properties to cancer associated fibroblasts. Tissue fibrosis in SSc is driven by active fibroblasts (myofibroblasts). Therefore in this study we investigated the role of CLIC4 in SSc fibroblast activation. Methods  Dermal fibroblasts were obtained from full thickness skin biopsies from SSc patients (early-diffuse). RNA and protein were collected from the fibroblasts and CLIC4 transcript and protein levels were assessed by qPCR and western blot. SSc patient fibroblasts were treated with the chloride ion channel inhibitors NPPB and IAA-94. Results  CLIC4 was found to be expressed at significantly higher levels in SSc patients fibroblasts compared to healthy controls, at both the transcript (3.7 fold) and protein (1.7 fold) levels. Inhibition of the TGF-β signalling pathway led to reduced CLIC4 expression in SSc fibroblasts, confirming this pathway as the main driver of CLIC4 expression. Finally, treatment of SSc fibroblasts with small molecule inhibitors that target the channel led to reduced expression of the myofibroblast markers collagen type 1 and alpha-smooth muscle actin, suggesting a direct role for CLIC4 in SSc associated skin fibrosis. Conclusion  We have identified a novel role for CLIC4 in SSc myofibroblast activation, which further strengthen the similarities between SSc fibroblasts and cancer associated fibroblasts. Furthermore this study highlights this channel as a novel target for therapeutic intervention. Disclosure  C. Wasson: None. R. Ross: None. R. Morton: None. J. Mankouri: None. F. Del Galdo: None.

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