PI3K p110 gamma overexpression in idiopathic pulmonary fibrosis lung tissue and fibroblast cells: in vitro effects of its inhibition

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Comparative proteome analysis of lung tissue from patients with idiopathic pulmonary fibrosis (IPF), non-specific interstitial pneumonia (NSIP) and organ donors

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Targeting Histone Deacetylases in Idiopathic Pulmonary Fibrosis: A Future Therapeutic Option

Idiopathic pulmonary fibrosis (IPF) is a progressive and fatal lung disease with limited therapeutic options, and there is a huge unmet need for new therapies. A growing body of evidence suggests that the histone deacetylase (HDAC) family of transcriptional corepressors has emerged as crucial mediators of IPF pathogenesis. HDACs deacetylate histones and result in chromatin condensation and epigenetic repression of gene transcription. HDACs also catalyse the deacetylation of many non-histone proteins, including transcription factors, thus also leading to changes in the transcriptome and cellular signalling. Increased HDAC expression is associated with cell proliferation, cell growth and anti-apoptosis and is, thus, a salient feature of many cancers. In IPF, induction and abnormal upregulation of Class I and Class II HDAC enzymes in myofibroblast foci, as well as aberrant bronchiolar epithelium, is an eminent observation, whereas type-II alveolar epithelial cells (AECII) of IPF lungs indicate a significant depletion of many HDACs. We thus suggest that the significant imbalance of HDAC activity in IPF lungs, with a “cancer-like” increase in fibroblastic and bronchial cells versus a lack in AECII, promotes and perpetuates fibrosis. This review focuses on the mechanisms by which Class I and Class II HDACs mediate fibrogenesis and on the mechanisms by which various HDAC inhibitors reverse the deregulated epigenetic responses in IPF, supporting HDAC inhibition as promising IPF therapy

Exhalative Breath Markers Do Not Offer for Diagnosis of Interstitial Lung Diseases: Data from the European IPF Registry (eurIPFreg) and Biobank

Background: New biomarkers are urgently needed to facilitate diagnosis in Interstitial Lung Diseases (ILD), thus reducing the need for invasive procedures, and to enable tailoring and monitoring of medical treatment. Methods: In this study we investigated if patients with idiopathic pulmonary fibrosis (IPF; n = 21), non-IPF ILDs (n = 57) and other lung diseases (chronic obstructive pulmonary disease (COPD) n = 24, lung cancer (LC) n = 16) as well as healthy subjects (n = 20) show relevant differences in exhaled NO (FeNO; Niox MINO), or in eicosanoid (PGE2, 8-isoprostane; enzyme-linked immunosorbent assay (ELISA)) levels as measured in exhaled breath condensates (EBC) and bronchoalveolar lavage fluids (BALF). Results: There was no significant difference in FeNO values between IPF, non-IPF ILDs and healthy subjects, although some individual patients showed highly elevated FeNO. On the basis of the FeNO signal, it was neither possible to differentiate between the kind of disease nor to detect exacerbations. In addition, there was no correlation between FeNO values and lung function. The investigation of the eicosanoids in EBCs was challenging (PGE2) or unreliable (8-isoprostane), but worked out well in BALF. A significant increase of free 8-isoprostane was observed in BALF, but not in EBCs, of patients with IPF, hypersensitivity pneumonitis (HP) and sarcoidosis, possibly indicating severity of oxidative stress. Conclusions: FeNO-measurements are not of diagnostic benefit in different ILDs including IPF. The same holds true for PGE2 and 8-isoprostane in EBC by ELISA

Dynamic expression of HOPX in alveolar epithelial cells reflects injury and repair during the progression of pulmonary fibrosis

Abstract Mechanisms of injury and repair in alveolar epithelial cells (AECs) are critically involved in the progression of various lung diseases including idiopathic pulmonary fibrosis (IPF). Homeobox only protein x (HOPX) contributes to the formation of distal lung during development. In adult lung, alveolar epithelial type (AT) I cells express HOPX and lineage-labeled Hopx+ cells give rise to both ATI and ATII cells after pneumonectomy. However, the cell function of HOPX-expressing cells in adult fibrotic lung diseases has not been investigated. In this study, we have established a flow cytometry-based method to evaluate HOPX-expressing cells in the lung. HOPX expression in cultured ATII cells increased over culture time, which was accompanied by a decrease of proSP-C, an ATII marker. Moreover, HOPX expression was increased in AECs from bleomycin-instilled mouse lungs in vivo. Small interfering RNA-based knockdown of Hopx resulted in suppressing ATII-ATI trans-differentiation and activating cellular proliferation in vitro. In IPF lungs, HOPX expression was decreased in whole lungs and significantly correlated to a decline in lung function and progression of IPF. In conclusion, HOPX is upregulated during early alveolar injury and repair process in the lung. Decreased HOPX expression might contribute to failed regenerative processes in end-stage IPF lungs

Epithelial Endoplasmic Reticulum Stress and Apoptosis in Sporadic Idiopathic Pulmonary Fibrosis

Rationale: The molecular pathomechanisms underlying idiopathic pulmonary fibrosis (IPF) are elusive, but chronic epithelial injury has recently been suggested as key event

Tubastatin ameliorates pulmonary fibrosis by targeting the TGFβ-PI3K-Akt pathway

<div><p>Background</p><p>Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive and fatal disease. Histone deacetylase 6 (HDAC6) alters function and fate of various proteins via deacetylation of lysine residues, and is implicated in TGF-β1-induced EMT (epithelial-mesenchymal transition). However, the role of HDAC6 in pulmonary fibrosis is unknown.</p><p>Methods</p><p>HDAC6 expression in IPF and control lungs was assessed by quantitative real-time PCR (qRT-PCR) and immunoblots. Lung fibroblasts were treated with TGF-β1 ± HDAC6 inhibitors (Tubacin, Tubastatin, ACY1215, or MC1568), and fibrotic markers such as type I collagen were assessed using qRT-PCR and immunoblots. Mice were treated with bleomycin (oropharyngeal aspiration; single dose) ± Tubastatin (intraperitoneally injection; daily for 21 days), and lung collagen expression was gauged using immunoblots and trichrome staining. In a separate experiment, HDAC6 wild-type (WT) and knockout (KO) mice were administered bleomycin, and lungs were evaluated in the same manner.</p><p>Results</p><p>HDAC6 expression was deregulated in IPF lungs. Among the HDAC6 inhibitors tested, only Tubastatin significantly repressed TGF-β1-induced expression of type-1 collagen in lung fibroblasts, and this finding was coupled with decreased Akt phosphorylation and increased Akt-PHLPP (PH domain and Leucine rich repeat Protein Phosphatase) association. Tubastatin repressed TGF-β1-induced S6K phosphorylation, HIF-1α expression, and VEGF expression. Tubastatin also repressed TGF-β1-induced inhibition of LC3B-II (a marker of autophagosome formation). In bleomycin-treated mouse lungs, HDAC6 expression was increased, and Tubastatin repressed type-1 collagen expression. However, in HDAC6 KO mice, bleomycin-induced type-1 collagen expression was not repressed compared to WT mice. Knockdown of HDAC6, as well as HDAC10, another potential Tubastatin target, did not inhibit TGF-β1-induced collagen expression in lung fibroblasts.</p><p>Conclusions</p><p>HDAC6 expression is altered during lung fibrogenesis. Tubastatin represses TGF-β1-induced collagen expression, by diminishing Akt phosphorylation and regulating downstream targets such as HIF-1α-VEGF axis and autophagy. Tubastatin-treated WT mice are protected against bleomycin-induced fibrosis, but HDAC6 KO mice are not. Our data suggest that Tubastatin ameliorates pulmonary fibrosis, by targeting the TGFβ-PI3K-Akt pathway, likely via an HDAC6-independent mechanism.</p></div

Tubastatin decreases TGF-β1-induced expression of type-1 collagen.

<p>(A-E) Subconfluent NHLFs (A, C, D, E) or IPF fibroblasts (B) were pretreated with an HDAC6 inhibitor (Tubacin, Tubastatin, ACY1215, or MC1568) for 6 hours and then co-treated with TGF-β1 and the HDAC6 inhibitor. (A, B) Protein expression levels were assessed using immunoblots at 48 hours. D: DMSO, Tc: Tubacin, Ts: Tubastatin, A: ACY1215, M: MC1568; Col-1: type-1 collagen, α-SMA: α-smooth muscle actin (C) Gene expression levels for collagen-1 (col1a1) were assessed using qRT-PCR at 24 hours. * p<0.05 compared to the control; # p<0.05 compared to the TGF-β1. (D) Correlation between the degree of α-tubulin acetylation and collagen-1 protein expression in experiments involving Tubastatin treatment. There was a modest inverse correlation between the degree of α-tubulin acetylation and collagen-1 expression (r² = 0.7602 [p<0.0001] for pooled results of six independent experiments.). (E) An MTT assay showing that Tubastatin does not affect NHLF viability. Values for cell viability were normalized to the control group (= DMSO treatment).</p