scholarly journals Assessment of Brd4 Inhibition in Idiopathic Pulmonary Fibrosis Lung Fibroblasts and in Vivo Models of Lung Fibrosis

2013 ◽  
Vol 183 (2) ◽  
pp. 470-479 ◽  
Author(s):  
Xiaoyan Tang ◽  
Ruoqi Peng ◽  
Jonathan E. Phillips ◽  
Jeremy Deguzman ◽  
Yonglin Ren ◽  
...  
Keyword(s):  
Idiopathic Pulmonary Fibrosis ◽  
Pulmonary Fibrosis ◽  
Lung Fibrosis ◽  
Lung Fibroblasts ◽  
In Vivo Models ◽  
Idiopathic Pulmonary Fibrosis Lung ◽  
Brd4 Inhibition

scholarly journals Itaconate controls the severity of pulmonary fibrosis

2020 ◽  
Vol 5 (52) ◽  
pp. eabc1884 ◽  
Author(s):  
Patricia P. Ogger ◽  
Gesa J. Albers ◽  
Richard J. Hewitt ◽  
Brendan J. O’Sullivan ◽  
Joseph E. Powell ◽  
...  
Keyword(s):  
Gene Expression ◽  
Wound Healing ◽  
Idiopathic Pulmonary Fibrosis ◽  
Pulmonary Fibrosis ◽  
Lung Fibrosis ◽  
Therapeutic Strategy ◽  
Lung Fibroblasts ◽  
Disease Phenotype ◽  
Wild Type

Idiopathic pulmonary fibrosis (IPF) is a fatal lung disease in which airway macrophages (AMs) play a key role. Itaconate has emerged as a mediator of macrophage function, but its role during fibrosis is unknown. Here, we reveal that itaconate is an endogenous antifibrotic factor in the lung. Itaconate levels are reduced in bronchoalveolar lavage, and itaconate-synthesizing cis-aconitate decarboxylase expression (ACOD1) is reduced in AMs from patients with IPF compared with controls. In the murine bleomycin model of pulmonary fibrosis, Acod1−/− mice develop persistent fibrosis, unlike wild-type (WT) littermates. Profibrotic gene expression is increased in Acod1−/− tissue-resident AMs compared with WT, and adoptive transfer of WT monocyte-recruited AMs rescued mice from disease phenotype. Culture of lung fibroblasts with itaconate decreased proliferation and wound healing capacity, and inhaled itaconate was protective in mice in vivo. Collectively, these data identify itaconate as critical for controlling the severity of lung fibrosis, and targeting this pathway may be a viable therapeutic strategy.

ID: 112: ROLE OF PHOSPHOLIPASE D IN IDIOPATHIC PULMONARY FIBROSIS

2016 ◽  
Vol 64 (4) ◽  
pp. 964.1-964
Author(s):  
V Suryadevara ◽  
T Royston ◽  
E Berdyshev ◽  
L Huang ◽  
V Natarajan ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Idiopathic Pulmonary Fibrosis ◽  
Pulmonary Fibrosis ◽  
Phospholipase D ◽  
Human Lung ◽  
Lung Fibroblasts ◽  
Human Lung Fibroblasts

Idiopathic pulmonary fibrosis (IPF) is a deadly interstitial disease that leads to scarring and fibrosis of the lung tissue. In pulmonary fibrosis, there is injury and denudation of the alveolar epithelium, which further leads to activation of fibroblasts which differentiate into myofibroblasts. This includes several mechanisms including epithelial to mesenchymal transition (EMT). In this study, we investigated the role of phospholipase D (PLD) in IPF and also its underlying mechanism like EMT and fibroblast proliferation and differentiation. An in vivo murine model of bleomycin-induced pulmonary fibrosis (PF) and in vitro models of murine alveolar type-II epithelial cells (MLE-12) and human lung fibroblasts were used. C57BL/6 and genetically engineered PLD2−/− mice were intratracheally challenged with bleomycin (1.5 U/kg animal) for 14 days and markers of inflammation, EMT and fibrosis were determined. MLE-12 cells were treated with specific PLD1 or PLD2 inhibitors prior to bleomycin (10 mU/ml) challenge, and the role of PLD in EMT and apoptosis of alveolar epithelial cells was studied. Human lung fibroblasts were serum-starved (3h), pretreated with PLD1 or PLD2 inhibitors, and the effect of TGF-β (5 ng/ml) on differentiation of lung fibroblast to myofibroblast was determined. Intra-tracheal instillation of bleomycin in the mice for 14 days leads to the progression of fibrosis in the lung. The lung tissues of the bleomycin treated mice were found to have increased PLD2 protein expression, myofibroblast markers like α-SMA, fibronectin, mesenchymal markers like vimentin, inflammatory cytokines and collagen. Genetic deletion of PLD2 in mice attenuated bleomycin-induced lung inflammation and pulmonary fibrosis. In vitro, MLE-12 cells pretreated with either PLD1 or PLD2 inhibitor did not show a profound reduction either in apoptosis or the expression of transcription factors such as SNAIL, and other markers of EMT. However, MLE-12 cells pretreated with both PLD1 (250 nM) and PLD2 (500 nM) inhibitors were resistant to bleomycin-induced apoptosis, and exhibited reduced expression of SNAIL and mesenchymal markers. On the contrary, human lung fibroblasts pretreated with PLD1 and PLD2 inhibitors showed increased fibroblast to myofibroblast differentiation mediated by TGF-β. The present study suggests a role for PLD2 in bleomycin-induced PF. In vitro, inhibition of both PLD1 and PLD2 was necessary to attenuate bleomycin-induced EMT in epithelial cells and TGF-β mediated differentiation of fibroblasts to myofibroblasts. The in vivo and in vitro results identify the mechanism by which PLD regualtes PF and suggest PLD as a potential therapeutic target in pulmonary fibrosis. This work was supported by National Institutes of Health grant P01 HL98050 to VN.

scholarly journals NADPH oxidase DUOX1 sustains TGF-β1 signalling and promotes lung fibrosis

2020 ◽  
pp. 1901949
Author(s):  
Ruy Andrade Louzada ◽  
Raphaël Corre ◽  
Rabii Ameziane El Hassani ◽  
Lydia Meziani ◽  
Madeleine Jaillet ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Pulmonary Fibrosis ◽  
Nadph Oxidase ◽  
Lung Fibrosis ◽  
Transforming Growth Factor ◽  
Lung Fibroblasts ◽  
Mouse Lung ◽  
Epithelial Surface ◽  
Tgf Β1

Interstitial lung fibroblast activation coupled with extracellular matrix production is a pathological signature of pulmonary fibrosis, and is governed by transforming growth factor (TGF-β1)/Smad signalling. TGF-β1 and oxidative stress cooperate to drive fibrosis. Cells can produce reactive oxygen species (ROS) through activation and/or induction of NADPH oxidases, such as dual oxidase (DUOX1/2). Since DUOX enzymes, as extracellular H2O2-generating systems, are involved in extracellular matrix formation and in wound healing in different experimental models, we hypothesised that DUOX-based NADPH oxidase plays a role in the pathophysiology of pulmonary fibrosis.Our in vivo data (IPF patients and mouse models of lung fibrosis) showed that the NADPH oxidase DUOX1 is induced in response to lung injury. DUOX1-deficient mice (DUOX1+/- and DUOX1-/-) had an attenuated fibrotic phenotype. In addition to being highly expressed at the epithelial surface of airways, DUOX1 appears to be also well expressed in the fibroblastic foci of remodelled lungs. By using primary human and mouse lung fibroblasts, we showed that TGF-β1 upregulates DUOX1 and its maturation factor DUOXA1 and that DUOX1-derived H2O2 promoted the duration of TGF-β1-activated Smad3 phosphorylation by preventing phospho-Smad3 degradation. Analysis of the mechanism revealed that DUOX1 inhibited the interaction between phospho-Smad3 and the ubiquitin ligase NEDD4L, preventing NEDD4L-mediated ubiquitination of phospho-Smad3 and its targeting for degradation.These findings highlight a role for DUOX1-derived H2O2 in a positive feedback that amplifies the signalling output of the TGF-β1 pathway and identify DUOX1 as a new therapeutic target in pulmonary fibrosis.

scholarly journals Galectin-3 levels are elevated following nintedanib treatment

2020 ◽  
Vol 11 ◽  
pp. 204062232096841
Author(s):  
Gali Epstein Shochet ◽  
Alon Pomerantz ◽  
David Shitrit ◽  
Becky Bardenstein-Wald ◽  
Kjetil Ask ◽  
...  
Keyword(s):  
Pulmonary Fibrosis ◽  
Mouse Model ◽  
Quantitative Polymerase Chain Reaction ◽  
Lung Fibroblasts ◽  
Mrna Levels ◽  
Serum Samples ◽  
In Vivo Models ◽  
Galectin 3 ◽  
The Impact

Background and Aims: Idiopathic pulmonary fibrosis (IPF) is a common and severe form of pulmonary fibrosis. Nintedanib, a triple angiokinase inhibitor, is approved for treating IPF. Galectin 3 (Gal-3) activates a variety of profibrotic processes. Currently, the Gal-3 inhibitor TD139 is being tested in phase II clinical trials. Since this treatment is given ‘on top’ of nintedanib, it is important to estimate its effect on Gal-3 levels. Therefore, we evaluated the impact of nintedanib on Gal-3 expression using both in vitro and in vivo models, in addition to serum samples from patients with IPF. Methods: Gal-3 levels were evaluated in IPF and control tissue samples, primary human lung fibroblasts (HLFs) following nintedanib treatment (10–100 nM, quantitative polymerase chain reaction), and in a silica-induced fibrosis mouse model with/without nintedanib (0.021–0.21 mg/kg) by immunohistochemistry. In addition, Gal-3 levels were analyzed in serum samples from 41 patients with interstitial lung disease patients with/without nintedanib treatment by ELISA. Results: Nintedanib addition to HLFs resulted in significant elevations in Gal-3, phospho-signal transducer and activator of transcription 3 (pSTAT3), as well as IL-8 mRNA levels ( p < 0.05). Gal-3 expression was higher in samples from IPF patients compared with non-IPF controls at the protein and mRNA levels ( p < 0.05). In the in vivo mouse model, Gal-3 levels were increased following fibrosis induction and even further increased with the addition of nintedanib, mostly in macrophages ( p < 0.05). Patients receiving nintedanib presented with higher Gal-3 serum levels compared with those who did not receive nintedanib ( p < 0.05). Conclusion: Nintedanib elevates Gal-3 levels in both experimental models, along with patient samples. These findings highlight the possibility of using combined inhibition therapy for patients with IPF.

scholarly journals SIRT3 blocks myofibroblast differentiation and pulmonary fibrosis by preventing mitochondrial DNA damage

2017 ◽  
Vol 312 (1) ◽  
pp. L68-L78 ◽  
Author(s):  
Samik Bindu ◽  
Vinodkumar B. Pillai ◽  
Abhinav Kanwal ◽  
Sadhana Samant ◽  
Gökhan M. Mutlu ◽  
...  
Keyword(s):  
Dna Damage ◽  
Mitochondrial Dna ◽  
Pulmonary Fibrosis ◽  
Lung Fibrosis ◽  
Lung Fibroblasts ◽  
Whole Body ◽  
Myofibroblast Differentiation ◽  
Mitochondrial Dna Damage

Myofibroblast differentiation is a key process in the pathogenesis of fibrotic diseases. Transforming growth factor-β1 (TGF-β1) is a powerful inducer of myofibroblast differentiation and is implicated in pathogenesis of tissue fibrosis. This study was undertaken to determine the role of mitochondrial deacetylase SIRT3 in TGF-β1-induced myofibroblast differentiation in vitro and lung fibrosis in vivo. Treatment of human lung fibroblasts with TGF-β1 resulted in increased expression of fibrosis markers, smooth muscle α-actin (α-SMA), collagen-1, and fibronectin. TGF-β1 treatment also caused depletion of endogenous SIRT3, which paralleled with increased production of reactive oxygen species (ROS), DNA damage, and subsequent reduction in levels of 8-oxoguanine DNA glycosylase (OGG1), an enzyme that hydrolyzes oxidized guanine (8-oxo-dG) and thus protects DNA from oxidative damage. Overexpression of SIRT3 by adenovirus-mediated transduction reversed the effects of TGF-β1 on ROS production and mitochondrial DNA damage and inhibited TGF-β1-induced myofibroblast differentiation. To determine the antifibrotic role of SIRT3 in vivo, we used the bleomycin-induced mouse model of pulmonary fibrosis. Compared with wild-type controls, Sirt3-knockout mice showed exacerbated fibrosis after intratracheal instillation of bleomycin. Increased lung fibrosis was associated with decreased levels of OGG1 and concomitant accumulation of 8-oxo-dG and increased mitochondrial DNA damage. In contrast, the transgenic mice with whole body Sirt3 overexpression were protected from bleomycin-induced mtDNA damage and development of lung fibrosis. These data demonstrate a critical role of SIRT3 in the control of myofibroblast differentiation and lung fibrosis.

scholarly journals Tetrandrine modulates SQSTM1-mediated selective autophagy and protects pulmonary fibrosis

2021 ◽  
Author(s):  
Yuanyuan Liu ◽  
Wenshan Zhong ◽  
Jinming Zhang ◽  
Weimou Chen ◽  
Ye Lu ◽  
...  
Keyword(s):  
Pulmonary Fibrosis ◽  
Lung Fibrosis ◽  
Smooth Muscle Actin ◽  
Lung Fibroblasts ◽  
Therapeutic Effects ◽  
The Novel ◽  
Induced Expression ◽  
Tgf Β1

Background and Purpose Idiopathic pulmonary fibrosis is a progressive fatal disease characterized by interstitial remodeling, with high lethality and a lack of effective medical therapies. Tetrandrine has been proposed to present anti-fibrotic effects, but the efficacy and mechanisms of tetrandrine against lung fibrosis has not been systematically evaluated. We sought to study the potential therapeutic effects and mechanisms of tetrandrine in lung fibrosis. Experimental Approach The anti-fibrotic effects of tetrandrine were evaluated in bleomycin-induced mouse models and TGF-β1-stimulated murine lung fibroblasts. We performed Chromatin Immunoprecipitation (ChIP), Immunoprecipitation (IP) and mRFP-GFP-MAP1LC3B adenovirus construct to investigate the novel mechanisms of tetrandrine-induced autophagy. Key Results Tetrandrine decreased TGF-β1-induced expression of α-smooth muscle actin, fibronectin, vimentin and type 1 collagen and proliferation in fibroblasts. Tetrandrine restored TGF-β1-induced impaired autophagy, accompanied by the up-regulation and enhanced interaction of SQSTM1 and MAP1LC3-Ⅱ. ChIP studies revealed that NRF2 bound to SQSTM1 promoter in tetrandrine-induced autophagy. Furthermore, TGF-β1-induced phosphorylated mTOR was inhibited by tetrandrine, with reduced activation levels of Rheb. In vivo tetrandrine suppressed the bleomycin-induced expression of fibrotic markers and improved pulmonary function. Conclusion and Implications Our data suggest that tetrandrine might be recognized as a novel autophagy inducer, thus attenuating lung fibrosis. Tetrandrine should be investigated as a novel therapeutic strategy for IPF.

scholarly journals Repetitive intratracheal bleomycin models several features of idiopathic pulmonary fibrosis

2010 ◽  
Vol 299 (4) ◽  
pp. L442-L452 ◽  
Author(s):  
Amber L. Degryse ◽  
Harikrishna Tanjore ◽  
Xiaochuan C. Xu ◽  
Vasiliy V. Polosukhin ◽  
Brittany R. Jones ◽  
...  
Keyword(s):  
Single Dose ◽  
Idiopathic Pulmonary Fibrosis ◽  
Pulmonary Fibrosis ◽  
Lung Fibrosis ◽  
Collagen Content ◽  
Lung Fibroblasts ◽  
Tunel Staining ◽  
Mesenchymal Transition ◽  
Cell Counts ◽  
Reporter Mice

Single-dose intratracheal bleomycin has been instrumental for understanding fibrotic lung remodeling, but fails to recapitulate several features of idiopathic pulmonary fibrosis (IPF). Since IPF is thought to result from recurrent alveolar injury, we aimed to develop a repetitive bleomycin model that results in lung fibrosis with key characteristics of human disease, including alveolar epithelial cell (AEC) hyperplasia. Wild-type and cell fate reporter mice expressing β-galactosidase in cells of lung epithelial lineage were given intratracheal bleomycin after intubation, and lungs were harvested 2 wk after a single or eighth biweekly dose. Lungs were evaluated for fibrosis and collagen content. Bronchoalveolar lavage (BAL) was performed for cell counts. TUNEL staining and immunohistochemistry were performed for pro-surfactant protein C (pro-SP-C), Clara cell 10 (CC-10), β-galactosidase, S100A4, and α-smooth muscle actin. Lungs from repetitive bleomycin mice had marked fibrosis with prominent AEC hyperplasia, similar to usual interstitial pneumonia (UIP). Compared with single dosing, repetitive bleomycin mice had greater fibrosis by scoring, morphometry, and collagen content; increased TUNEL+ AECs; and reduced inflammatory cells in BAL. Sixty-four percent of pro-SP-C+ cells in areas of fibrosis expressed CC-10 in the repetitive model, suggesting expansion of a bronchoalveolar stem cell-like population. In reporter mice, 50% of S100A4+ lung fibroblasts were derived from epithelial mesenchymal transition compared with 33% in the single-dose model. With repetitive bleomycin, fibrotic remodeling persisted 10 wk after the eighth dose. Repetitive intratracheal bleomycin results in marked lung fibrosis with prominent AEC hyperplasia, features reminiscent of UIP.

scholarly journals IL-25 contributes to lung fibrosis by directly acting on alveolar epithelial cells and fibroblasts

2019 ◽  
Vol 244 (9) ◽  
pp. 770-780 ◽  
Author(s):  
Xuefeng Xu ◽  
Sa Luo ◽  
Biyun Li ◽  
Huaping Dai ◽  
Jinglan Zhang
Keyword(s):  
Epithelial Cells ◽  
Idiopathic Pulmonary Fibrosis ◽  
Pulmonary Fibrosis ◽  
Lung Fibrosis ◽  
Lung Fibroblast ◽  
Alveolar Epithelial Cells ◽  
Lung Fibroblasts ◽  
Rt Pcr ◽  
Alveolar Epithelial

Interleukin (IL)-25 is shown to potentiate type-2 immunity and contribute to chronic airway inflammation and remodeling in allergic airway diseases. However, the role of IL-25 in idiopathic pulmonary fibrosis (IPF), dominated by nonatopic type-2 immunity, still remains largely unclear. Herein, we detected the expression levels of IL-25 and IL-17BR (IL-25’s receptor) by using lung tissue samples gained from IPF patients and normal subjects. Also, by directly intranasal (IN) instillation of IL-25 to mice, we examined the potential roles and mechanisms of IL-25 in the development of lung fibrosis. Furthermore, we tested whether IL-25 can directly activate human lung fibroblast by in vitro cell culture. Immunohistochemical, Western blot, and real-time reverse transcription-polymerase chain reaction (RT-PCR) showed that the mRNA and protein levels of IL-25 and IL-17BR are significantly higher in IPF patients when compared with normal controls. Intranasal instillation of IL-25 to mice markedly induces the expressions of alveolar IL-5 and IL-13. Furthermore, immunohistochemical analysis showed that the main components of the extracellular matrix including collagen I, collagen III and fibronectin are notably induced by IL-25 instillation in lung parenchyma (especially in alveolar epithelial cells [AECs]). Also, IL-25 potentiates the expression of connective tissue growth factor (CTGF) in AECs and the recruitment of lung fibroblast. By using Cell Counting Kit-8 and EDU incorporation assay, we found that IL-25 markedly enhances the proliferation of lung fibroblast. Finally, IL-25 potentiates fibroblast to produce several fibrogenic genes including collagen I/III, fibronectin, CTGF, α smooth muscle (α-SMA) and tissue inhibitor of metalloproteinase (TIMP)-1 as determined by RT-PCR assay. Collectively, we concluded that IL-25 is increased in IPF lungs and contributes to lung fibrosis by directly mediating AECs/fibroblast activation. Impact statement Our work focused on alveolar epithelial cells (AECs)-derived type-2 cytokine (interleukin [IL]-25) in the pathogenesis of idiopathic pulmonary fibrosis (IPF). We showed that IL-25 and IL-17BR (IL-25’s receptor) is upregulated in lung tissues (especially in AECs and lung fibroblasts) of IPF patients and contributes to lung fibrosis by directly activating lung fibroblasts and modulating epithelial–mesenchymal transition (EMT) of AECs. We suggest that IL-25 may be one of the master switches hidden in the milieu of abnormal epithelial–mesenchymal crosstalk. Treatment targeting IL-25 may be the potential and novel method for IPF patients.

scholarly journals Tetrandrine Modulates Rheb-mTOR Signaling-Mediated Selective Autophagy and Protects Pulmonary Fibrosis

2021 ◽  
Vol 12 ◽  
Author(s):  
Yuanyuan Liu ◽  
Wenshan Zhong ◽  
Jinming Zhang ◽  
Weimou Chen ◽  
Ye lu ◽  
...  
Keyword(s):  
Pulmonary Fibrosis ◽  
Lung Fibrosis ◽  
Smooth Muscle Actin ◽  
Lung Fibroblasts ◽  
Therapeutic Effects ◽  
The Novel ◽  
Induced Expression ◽  
Tgf Β1

Idiopathic pulmonary fibrosis is a progressive fatal disease characterized by interstitial remodeling, with high lethality and a lack of effective medical therapies. Tetrandrine has been proposed to present anti-fibrotic effects, but the efficacy and mechanisms have not been systematically evaluated. We sought to study the potential therapeutic effects and mechanisms of tetrandrine against lung fibrosis. The anti-fibrotic effects of tetrandrine were evaluated in bleomycin-induced mouse models and TGF-β1-stimulated murine lung fibroblasts. We performed Chromatin Immunoprecipitation (ChIP), Immunoprecipitation (IP), and mRFP-GFP-MAP1LC3B adenovirus construct to investigate the novel mechanisms of tetrandrine-induced autophagy. Tetrandrine decreased TGF-β1-induced expression of α-smooth muscle actin, fibronectin, vimentin, and type 1 collagen and proliferation in fibroblasts. Tetrandrine restored TGF-β1-induced impaired autophagy flux, accompanied by enhanced interaction of SQSTM1 and MAP1LC3-Ⅱ. ChIP studies revealed that tetrandrine induced autophagy via increasing binding of NRF2 and SQSTM1 promoter. Furthermore, tetrandrine inhibited TGF-β1-induced phosphorylation of mTOR by reducing activation of Rheb. In vivo tetrandrine suppressed the bleomycin-induced expression of fibrotic markers and improved pulmonary function. Our data suggest that protective effect of tetrandrine against lung fibrosis might be through promoting Rheb-mTOR and NRF2-SQSTM1 mediated autophagy. Tetrandrine may thus be potentially employed as a novel therapeutic medicine against IPF.

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