C/EBPβ Acetylation is Involved in Idiopathic Pulmonary Fibrosis

Background: IPF is a progressive lung disease, characterized by excessive deposition of ECM. C/EBPβ is involved in the development of pulmonary fibrosis. However, the regulation of C/EBPβ in the context of pulmonary fibrosis is not clear. The study is to identify the C/EBPβ acetylation in IPF．Methods: Lung from six IPF and six control samples were selected in this study. We investigated the expression of C/EBPβ in lungs with Immunochemistry. Moreover, the expression of C/EBPβ mRNA via Real Time-PCR and its protein expression via Western Blot were performed. Meanwhile, the levels of collagen-I and α-SMA as markers of pulmonary fibrosis were also determined by Western Blot. Furthermore, we confirmed the relationship between α-SMA and acetylated C/EBPβ by Co-Immunoprecipitation. Results: We found the elevated C/EBPβ mostly locating in fibroblast foci in lungs of IPF. And the expression of C/EBPβ RNA and protein were obviously increased in IPF (P <0.05), in which the proteins of α-SMA and collagen-I were enhanced (P <0.05). Furthermore, the stronger acetylation of C/EBPβ binging to the α-SMA gene was shown in lung fibrosis (P <0.05). Conclusions: The increased expression of C/EBPβ acetylation associated with α-SMA expression is involved in the development of pulmonary fibrosis.

Inhibition of mast cells: a novel mechanism by which nintedanib may elicit anti-fibrotic effects

BackgroundIdiopathic pulmonary fibrosis (IPF) is a chronic and progressive lung disease which presents a grave prognosis for diagnosed patients. Nintedanib (a triple tyrosine kinase inhibitor) and pirfenidone (unclear mechanism of action) are the only approved therapies for IPF, but have limited efficacy. The pathogenic mechanisms of this disease are not fully elucidated; however, a role for mast cells (MCs) has been postulated.ObjectivesThe aim of this work was to investigate a role for MCs in IPF and to understand whether nintedanib or pirfenidone could impact MC function.Methods and resultsMCs were significantly elevated in human IPF lung and negatively correlated with baseline lung function (FVC). Importantly, MCs were positively associated with the number of fibroblast foci, which has been linked to increased mortality. Furthermore, MCs were increased in the region immediately surrounding the fibroblast foci, and co-culture studies confirmed a role for MC–fibroblast crosstalk in fibrosis. Nintedanib but not pirfenidone inhibited recombinant stem cell factor (SCF)–induced MC survival. Further evaluation of nintedanib determined that it also inhibited human fibroblast-mediated MC survival. This was likely via a direct effect on ckit (SCF receptor) since nintedanib blocked SCF-stimulated ckit phosphorylation, as well as downstream effects on MC proliferation and cytokine release. In addition, nintedanib ablated the increase in lung MCs and impacted high tissue density frequency (HDFm) in a rat bleomycin model of lung fibrosis.ConclusionNintedanib inhibits MC survival and activation and thus provides a novel additional mechanism by which this drug may exert anti-fibrotic effects in patients with IPF.

Matrix-transmitted paratensile signaling enables myofibroblast–fibroblast cross talk in fibrosis expansion

While the concept of intercellular mechanical communication has been revealed, the mechanistic insights have been poorly evidenced in the context of myofibroblast–fibroblast interaction during fibrosis expansion. Here we report and systematically investigate the mechanical force-mediated myofibroblast–fibroblast cross talk via the fibrous matrix, which we termed paratensile signaling. Paratensile signaling enables instantaneous and long-range mechanotransduction via collagen fibers (less than 1 s over 70 μm) to activate a single fibroblast, which is intracellularly mediated by DDR2 and integrin signaling pathways in a calcium-dependent manner through the mechanosensitive Piezo1 ion channel. By correlating in vitro fibroblast foci growth models with mathematical modeling, we demonstrate that the single-cell-level spatiotemporal feature of paratensile signaling can be applied to elucidate the tissue-level fibrosis expansion and that blocking paratensile signaling can effectively attenuate the fibroblast to myofibroblast transition at the border of fibrotic and normal tissue. Our comprehensive investigation of paratensile signaling in fibrosis expansion broadens the understanding of cellular dynamics during fibrogenesis and inspires antifibrotic intervention strategies targeting paratensile signaling.

Toll-like receptor 4 activation attenuates profibrotic response in control lung fibroblasts but not in fibroblasts from patients with IPF

Idiopathic pulmonary fibrosis (IPF) is a devastating lung disease with a median survival of 3 yr. IPF deteriorates upon viral or bacterial lung infection although pulmonary infection (pneumonia) in healthy lungs rarely induces fibrosis. Bacterial lipopolysaccharide (LPS) activates Toll-like receptor 4 (TLR4), initiating proinflammatory pathways. As TLR4 has already been linked to hepatic fibrosis and scleroderma, we now investigated the role of TLR4 in IPF fibroblasts. Lung tissue sections from patients with IPF were analyzed for TLR4 expression. Isolated normal human lung fibroblasts (NL-FB) and IPF fibroblasts (IPF-FB) were exposed to LPS and transforming growth factor-β (TGF-β) before expression analysis of receptors, profibrotic mediators, and cytokines. TLR4 is expressed in fibroblast foci of IPF lungs as well as in primary NL-FB and IPF-FB. As a model for a gram-negative pneumonia in the nonfibrotic lung, NL-FB and IPF-FB were coexposed to LPS and TGF-β. Whereas NL-FB produced significantly less connective tissue growth factor upon costimulation compared with TGF-β stimulation alone, IPF-FB showed significantly increased profibrotic markers compared with control fibroblasts after costimulation. Although levels of antifibrotic prostaglandin E2 were elevated after costimulation, they were not responsible for this effect. However, significant downregulation of TGF-β receptor type 1 in control fibroblasts seems to contribute to the reduced profibrotic response in our in vitro model. Normal and IPF fibroblasts thus differ in their profibrotic response upon LPS-induced TLR4 stimulation.