Chiko Shimbori

Extracellular matrix microenvironment contributes actively to pulmonary fibrosis.

Purpose of review The purpose of this review is to describe the contribution of an altered, profibrotic extracellular matrix (ECM) microenvironment to pulmonary fibrosis and how it changes cell behaviour and actively drives disease progression. Recent findings Idiopathic pulmonary fibrosis is a chronic and fatal disease of unknown cause. It is characterized by proliferation and accumulation of fibroblasts and myofibroblasts in clusters, termed fibroblastic foci, and extensive ECM deposition. Recent evidence from in-vivo and ex-vivo experimental studies has highlighted that the abnormal ECM in fibrotic lungs alters the behaviour of epithelial and mesenchymal cells. This profibrotic ECM microenvironment is characterized by altered biochemical and biomechanical properties and stores abundant amount of growth factors. By this, the ‘fibrotic ECM’ can drive progressive fibrogenesis in the lungs without any further initiating trigger. These concepts indicate a more complicated dynamic and active role of the fibrotic ECM than previously thought and offer many novel therapeutic targets. Summary The fibrotic ECM microenvironment is an active contributor to the development and progression of pulmonary fibrosis and a promising therapeutic target.

Stretch-induced Activation of Transforming Growth Factor-β1 in Pulmonary Fibrosis.

RATIONALE Recent findings suggesting transforming growth factor (TGF)-β1 activation by mechanical stimuli in vitro raised the question of whether this phenomenon was relevant in vivo in the context of pulmonary fibrosis. OBJECTIVES To explore the effect of mechanical stress on TGF-β1 activation and its signaling pathway in rat and human fibrotic lung tissue using a novel ex vivo model. METHODS Rat lung fibrosis was induced using transient gene expression of active TGF-β1. Lungs were harvested at Day 14 or 21 and submitted to various stimuli in a tissue bath equipped with a force transducer and servo-controlled arm. MEASUREMENTS AND MAIN RESULTS Fibrotic lung strips responded to tensile force by releasing active TGF-β1 from latent stores with subsequent increase in tissue phospho-Smad2/3. In contrast, measurable active TGF-β1 and phospho-Smad2/3 were not induced by mechanical stress in nonfibrotic lungs. Protease inhibition did not affect the release of active TGF-β1. A TGF-β1 receptor inhibitor, Rho-associated protein kinase inhibitor, and αv integrin inhibitor all attenuated mechanical stretch-induced phospho-Smad2/3 in fibrotic lung strips. Furthermore, the induction of phospho-Smad2/3 was enhanced in whole fibrotic rat lungs undergoing ventilation pressure challenge compared with control lungs. Last, tissue slices from human lung with usual interstitial pneumonia submitted to mechanical force showed an increase in TGF-β1 activation and induction of phospho-Smad2/3 in contrast with human nonfibrotic lungs. CONCLUSIONS Mechanical tissue stretch contributes to the development of pulmonary fibrosis via mechanotransduced activation of TGF-β1 in rodent and human pulmonary fibrosis.

Fibrocytes in pulmonary fibrosis: a brief synopsis.

Fibrocytes are bone marrow-derived, circulating mesenchymal progenitor cells that play a role in several fibrotic disorders, including lung fibrosis. They are attracted to injured tissue by various chemokines. It is likely that fibrocytes play a detrimental role in tissue homeostasis and promote fibrosis, although this paradigm needs further confirmation. This would make fibrocytes a possible novel treatment target for fibrotic disorders. Fibrocytes also have some potential as a biomarker for idiopathic pulmonary fibrosis (IPF) and other diseases, but the promising preliminary data from single centre studies still require independent validation. Despite several, as yet, unresolved issues, it has become clear that fibrocytes are more than an incidental finding in lung injury and repair, and may hold great promise for the future of IPF management.

Fibroblast growth factor-1 attenuates TGF-β1-induced lung fibrosis.

Idiopathic pulmonary fibrosis (IPF) is characterized by progressive fibroblast and myofibroblast proliferation, and extensive deposition of extracellular matrix (ECM). Fibroblast growth factor‐1 (FGF‐1) belongs to the FGF family and has been shown to inhibit fibroblast collagen production and differentiation into myofibroblasts, and revert epithelial–mesenchymal transition by inhibiting TGF‐β1 signalling pathways. However, the precise role of FGF‐1 in pulmonary fibrosis has not yet been elucidated. In this study, we explore the mechanisms underlying the anti‐fibrogenic effect of FGF‐1 in pulmonary fibrosis in vitro and in vivo by prolonged transient overexpression of FGF‐1 (AdFGF‐1) and TGF‐β1 (AdTGF‐β1) using adenoviral vectors. In vivo, FGF‐1 overexpression markedly attenuated TGF‐β1‐induced pulmonary fibrosis in rat lungs when given both concomitantly, or delayed, by enhancing proliferation and hyperplasia of alveolar epithelial cells (AECs). AdFGF‐1 also attenuated the TGF‐β1 signalling pathway and induced FGFR1 expression in AECs. In vitro, AdFGF‐1 prevented the increase in α‐SMA and the decrease in E‐cadherin induced by AdTGF‐β1 in normal human lung fibroblasts, primary human pulmonary AECs, and A549 cells. Concomitantly, AdTGF‐β1‐induced Smad2 phosphorylation was significantly reduced by AdFGF‐1 in both cell types. AdFGF‐1 also attenuated the increase in TGFβR1 protein and mRNA levels in fibroblasts. In AECs, AdFGF‐1 decreased TGFβR1 protein by favouring TGFβR1 degradation through the caveolin‐1/proteasome pathway. Furthermore, FGFR1 expression was increased in AECs, whereas it was decreased in fibroblasts. In serum of IPF patients, FGF‐1 levels were increased compared to controls. Interestingly, FGF‐1 expression was restricted to areas of AEC hyperplasia, but not α‐SMA‐positive areas in IPF lung tissue. Our results demonstrate that FGF‐1 may have preventative and therapeutic effects on TGF‐β1‐driven pulmonary fibrosis via inhibiting myofibroblast differentiation, inducing AEC proliferation, regulating TGF‐β1 signalling by controlling TGFβR1 expression and degradation, and regulating FGFR1 expression. Thus, modulating FGF‐1 signalling represents a potential therapy for the treatment of pulmonary fibrosis. Copyright

Molecular classification of idiopathic pulmonary fibrosis: Personalized medicine, genetics and biomarkers

Idiopathic pulmonary fibrosis (IPF) is a chronic and progressive fibrotic lung disease associated with high morbidity and poor survival. Characterized by substantial disease heterogeneity, the diagnostic considerations, clinical course and treatment response in individual patients can be variable. In the past decade, with the advent of high‐throughput proteomic and genomic technologies, our understanding of the pathogenesis of IPF has greatly improved and has led to the recognition of novel treatment targets and numerous putative biomarkers. Molecular biomarkers with mechanistic plausibility are highly desired in IPF, where they have the potential to accelerate drug development, facilitate early detection in susceptible individuals, improve prognostic accuracy and inform treatment recommendations. Although the search for candidate biomarkers remains in its infancy, attractive targets such as MUC5B and MPP7 have already been validated in large cohorts and have demonstrated their potential to improve clinical predictors beyond that of routine clinical practices. The discovery and implementation of future biomarkers will face many challenges, but with strong collaborative efforts among scientists, clinicians and the industry the ultimate goal of personalized medicine may be realized.

GRP78 and CHOP modulate macrophage apoptosis and the development of bleomycin‐induced pulmonary fibrosis

Endoplasmic reticulum (ER) stress and the unfolded protein response (UPR) have been associated with fibrotic lung disease, although exactly how they modulate this process remains unclear. Here we investigated the role of GRP78, the main UPR regulator, in an experimental model of lung injury and fibrosis. Grp78+/−, Chop−/− and wild type C57BL6/J mice were exposed to bleomycin by oropharyngeal intubation and lungs were examined at days 7 and 21. We demonstrate here that Grp78+/− mice were strongly protected from bleomycin‐induced fibrosis, as shown by immunohistochemical analysis, collagen content and lung function measurements. In the inflammatory phase of this model, a reduced number of lung macrophages associated with an increased number of TUNEL‐positive cells were observed in Grp78+/− mice. Dual immunohistochemical and in situ hybridization experiments showed that the macrophage population from the protected Grp78+/− mice was also strongly positive for cleaved caspase‐3 and Chop mRNA, respectively. In contrast, the administration of bleomycin to Chop−/− mice resulted in increased quasi‐static elastance and extracellular matrix deposition associated with an increased number of parenchymal arginase‐1‐positive macrophages that were negative for cleaved caspase‐3. The data presented indicate that the UPR is activated in fibrotic lung tissue and strongly localized to macrophages. GRP78‐ and CHOP‐mediated macrophage apoptosis was found to protect against bleomycin‐induced fibrosis. Overall, we demonstrate here that the fibrotic response to bleomycin is dependent on GRP78‐mediated events and provides evidence that macrophage polarization and apoptosis may play a role in this process. Copyright

Lysyl Oxidase–Like 1 Protein Deficiency Protects Mice from Adenoviral Transforming Growth Factor-β1–induced Pulmonary Fibrosis

&NA; Idiopathic pulmonary fibrosis (IPF) is a progressive disease characterized by excessive deposition of extracellular matrix (ECM) in the lung parenchyma. The abnormal ECM deposition slowly overtakes normal lung tissue, disturbing gas exchange and leading to respiratory failure and death. ECM cross‐linking and subsequent stiffening is thought to be a major contributor of disease progression and also promotes the activation of transforming growth factor (TGF)‐&bgr;1, one of the main profibrotic growth factors. Lysyl oxidase‐like (LOXL) 1 belongs to the cross‐linking enzyme family and has been shown to be up‐regulated in active fibrotic regions of bleomycin‐treated mice and patients with IPF. We demonstrate in this study that LOXL1‐deficient mice are protected from experimental lung fibrosis induced by overexpression of TGF‐&bgr;1 using adenoviral (Ad) gene transfer (AdTGF‐&bgr;1). The lack of LOXL1 prevented accumulation of insoluble cross‐linked collagen in the lungs, and therefore limited lung stiffness after AdTGF‐&bgr;1. In addition, we applied mechanical stretch to lung slices from LOXL1+/+ and LOXL1−/− mice treated with AdTGF‐&bgr;1. Lung stiffness (Youngs modulus) of LOXL1−/− lung slices was significantly lower compared with LOXL1+/+ lung slices. Moreover, the release of activated TGF‐&bgr;1 after mechanical stretch was significantly lower in LOXL1−/− mice compared with LOXL1+/+ mice after AdTGF‐&bgr;1. These data support the concept that cross‐linking enzyme inhibition represents an interesting therapeutic target for drug development in IPF.

Human mesenchymal stem cells attenuate early damage in a ventilated pig model of acute lung injury

Acute lung injury/acute respiratory distress syndrome (ALI/ARDS) is a major cause of global morbidity and mortality. Mesenchymal stem cells (MSC) have shown promise in treating inflammatory lung conditions. We hypothesised that human MSC (hMSC) can improve ALI/ARDS through their anti-inflammatory actions. We subjected pigs (n=6) to intravenous oleic acid (OA) injury, ventilation and hMSC infusion, while the controls (n=5) had intravenous OA, ventilation and an infusion vehicle control. hMSC were infused 1h after the administration of OA. The animals were monitored for additional 4h. Nuclear translocation of nuclear factor-light chain enhancer of activated B cells (NF-κB), a transcription factor that mediates several inflammatory pathways was reduced in hMSC treated pigs compared to controls (p=0.04). There was no significant difference in lung injury, assessed by histological scoring in hMSC treated pigs versus controls (p=0.063). There was no difference in neutrophil counts between hMSC-treated pigs and controls. Within 4h, there was no difference in the levels of IL-10 and IL-8 pre- and post-treatment with hMSC. In addition, there was no difference in hemodynamics, lung mechanics or arterial blood gases between hMSC treated animals and controls. Subsequent studies are required to determine if the observed decrease in inflammatory transcription factors will translate into improvement in inflammation and in physiological parameters over the long term.

Macitentan reduces progression of TGF-β1-induced pulmonary fibrosis and pulmonary hypertension

Idiopathic pulmonary fibrosis (IPF) is a progressive disease with an unknown cause. Two drugs, nintedanib and pirfenidone, have been shown to slow, but not stop, disease progression. Pulmonary hypertension (PH) is a frequent complication in IPF patients and is associated with poor prognosis. Macitentan is a dual endothelin receptor antagonist that is approved for pulmonary arterial hypertension treatment. We hypothesised that using macitentan to treat animals with pulmonary fibrosis induced by adenoviral vector encoding biologically active transforming growth factor-β1 (AdTGF-β1) would improve the PH caused by chronic lung disease and would limit the progression of fibrosis. Rats (Sprague Dawley) which received AdTGF-β1 were treated by daily gavage of macitentan (100 mg·kg−1·day−1), pirfenidone (0.5% food admix) or a combination from day 14 to day 28. Pulmonary artery pressure (PAP) was measured before the rats were killed, and fibrosis was subsequently evaluated by morphometric measurements and hydroxyproline analysis. AdTGF-β1 induced pulmonary fibrosis associated with significant PH. Macitentan reduced the increase in PAP and both macitentan and pirfenidone stopped fibrosis progression from day 14 to day 28. Macitentan protected endothelial cells from myofibroblast differentiation and apoptosis whereas pirfenidone only protected against fibroblast-to-myofibroblast differentiation. Both drugs induced apoptosis of differentiated myofibroblasts in vitro and in vivo. Our results demonstrate that dual endothelin receptor antagonism was effective in both PH and lung fibrosis whereas pirfenidone only affected fibrosis. Macitentan reduces the progression of TGF-β1-induced pulmonary fibrosis and pulmonary hypertension, while pirfenidone only slows fibrosis progression http://ow.ly/4tt230kHQOn

Matrix abnormalities in pulmonary fibrosis

Idiopathic pulmonary fibrosis (IPF) is a devastating, progressive disease, marked by excessive scarring, which leads to increased tissue stiffness, loss in lung function and ultimately death. IPF is characterised by progressive fibroblast and myofibroblast proliferation, and extensive deposition of extracellular matrix (ECM). Myofibroblasts play a key role in ECM deposition. Transforming growth factor (TGF)-β1 is a major growth factor involved in myofibroblast differentiation, and the creation of a profibrotic microenvironment. There is a strong link between increased ECM stiffness and profibrotic changes in cell phenotype and differentiation. The activation of TGF-β1 in response to mechanical stress from a stiff ECM explains some of the influence of the tissue microenvironment on cell phenotype and function. Understanding the close relationship between cells and their surrounding microenvironment will ultimately facilitate better management strategies for IPF. Cellular changes cause matrix aberrations that further drive cellular changes; this sustains and progresses fibrosis http://ow.ly/TpHq30kgEUX