Transcriptome profiling reveals the complexity of pirfenidone effects in idiopathic pulmonary fibrosis.

Despite the beneficial effects of pirfenidone in treating idiopathic pulmonary fibrosis (IPF), it remains unclear if lung fibroblasts (FB) are the main therapeutic target.To resolve this question, we employed a comparative transcriptomic approach and analysed lung homogenates (LH) and FB derived from IPF patients treated with or without pirfenidone.In FB, pirfenidone therapy predominantly affected growth and cell division pathways, indicating a major cellular metabolic shift. In LH samples, pirfenidone treatment was mostly associated with inflammation-related processes. In FB and LH, regulated genes were over-represented in the Gene Ontology node "extracellular matrix". We identified lower expression of cell migration-inducing and hyaluronan-binding protein (CEMIP) in both LH and FB from pirfenidone-treated IPF patients. Plasma levels of CEMIP were elevated in IPF patients compared to healthy controls and decreased after 7 months of pirfenidone treatment. CEMIP expression in FB was downregulated in a glioma-associated oncogene homologue-dependent manner and CEMIP silencing in IPF FB reduced collagen production and attenuated cell proliferation and migration.Cumulatively, our approach indicates that pirfenidone exerts beneficial effects via its action on multiple pathways in both FB and other pulmonary cells, through its ability to control extracellular matrix architecture and inflammatory reactions.

The drug diazaborine blocks ribosome biogenesis by inhibiting the AAA-ATPase Drg1.

The drug diazaborine is the only known inhibitor of ribosome biogenesis and specifically blocks large subunit formation in eukaryotic cells. However, the target of this drug and the mechanism of inhibition were unknown. Here we identify the AAA-ATPase Drg1 as a target of diazaborine. Inhibitor binding into the second AAA domain of Drg1 requires ATP loading and results in inhibition of ATP hydrolysis in this site. As a consequence the physiological activity of Drg1, i.e. the release of Rlp24 from pre-60S particles, is blocked, and further progression of cytoplasmic preribosome maturation is prevented. Our results identify the first target of an inhibitor of ribosome biogenesis and provide the mechanism of inhibition of a key step in large ribosomal subunit formation.

LRP1 promotes synthetic phenotype of pulmonary artery smooth muscle cells in pulmonary hypertension.

Pulmonary hypertension (PH) is characterized by a thickening of the distal pulmonary arteries caused by medial hypertrophy, intimal proliferation and vascular fibrosis. Low density lipoprotein receptor-related protein 1 (LRP1) maintains vascular homeostasis by mediating endocytosis of numerous ligands and by initiating and regulating signaling pathways. Here, we demonstrate the increased levels of LRP1 protein in the lungs of idiopathic pulmonary arterial hypertension (IPAH) patients, hypoxia-exposed mice, and monocrotaline-treated rats. Platelet-derived growth factor (PDGF)-BB upregulated LRP1 expression in pulmonary artery smooth muscle cells (PASMC). This effect was reversed by the PDGF-BB neutralizing antibody or the PDGF receptor antagonist. Depletion of LRP1 decreased proliferation of donor and IPAH PASMC in a ß1-integrin-dependent manner. Furthermore, LRP1 silencing attenuated the expression of fibronectin and collagen I and increased the levels of α-smooth muscle actin and myocardin in donor, but not in IPAH, PASMC. In addition, smooth muscle cell (SMC)-specific LRP1 knockout augmented α-SMA expression in pulmonary vessels and reduced SMC proliferation in 3D ex vivo murine lung tissue cultures. In conclusion, our results indicate that LRP1 promotes the dedifferentiation of PASMC from a contractile to a synthetic phenotype thus suggesting its contribution to vascular remodeling in PH.

Fra2 Overexpression in Mice Leads to Non-allergic Asthma Development in an IL-13 Dependent Manner.

Background: Asthma is a complex chronic inflammatory disease characterised by airway inflammation, remodelling and hyperresponsiveness (AHR). Members of the AP-1 transcription factor family play important roles in the activation of the immune system and the control of cellular responses; however, their role in the development of asthma has not been well studied. We aimed to investigate the role of the lesser known AP-1 family member, Fra2 in experimental asthma. Methods: Phenotypic characterisation and gene expression profiling was performed on Fra2 (TG) overexpressing and wild-type mice. The efficacy of therapeutic interventions in regulating the Fra2 phenotype was determined. Results: Transcriptional profiling of TG mice revealed a high abundance of regulated genes associated with airway remodelling, inflammation and mucus production. A concomitant increase in peribronchial collagen deposition, smooth muscle thickening and mucus production was observed. TG mice possessed increased inflammatory infiltration in the lung, predominantly consisting of eosinophils and T-cells and elevated expression of Th2 cytokines and eotaxin. Furthermore, TG mice possessed severe AHR in response to increasing doses of methacholine. Glucocorticoid treatment led to a partial improvement of the asthma phenotype, whereas blockade of IL-13 via neutralising antibodies ameliorated AHR and mucus production, but had no effect on collagen deposition. Conclusion: We here describe a novel model for non-allergic asthma that does not require the application of exogenous allergens, which mimics several key features of the disease, such as airway inflammation, remodelling and hyperresponsiveness. Fra2 may represent a key molecule coordinating multiple aspects of asthma pathogenesis.

Biomarkers for Pulmonary Vascular Remodeling in Systemic Sclerosis: A Pathophysiological Approach.

Pulmonary arterial hypertension (PAH) is a severe complication of systemic sclerosis (SSc) associated with high morbidity and mortality. There are several biomarkers of SSc-PAH, reflecting endothelial physiology, inflammation, immune activation, extracellular matrix, metabolic changes, or cardiac involvement. Biomarkers associated with diagnosis, disease severity and progression have been identified, however, very few have been tested in a prospective setting. Some antinuclear antibodies such as nucleosome antibodies (NUC), anti-centromere antibodies (CENP-A/B) and anti-U3-ribonucleoprotein (anti-U3-RNP) are associated with PAH while anti-U1-ribonucleoprotein (anti-U1-RNP) is associated with a reduced PAH risk. Anti-endothelin receptor and angiotensin-1 receptor antibodies might be good markers of SSc-PAH and progression of pulmonary vasculopathy. Regarding the markers reflecting immune activation and inflammation, there are many inconsistent results. CXCL-4 was associated with SSc progression including PAH and lung fibrosis. Growth differentiation factor (GDF)-15 was associated with PAH and mortality but is not specific for SSc. Among the metabolites, kynurenine was identified as diagnostic marker for PAH, however, its pathologic role in the disease is unclear. Endostatin, an angiostatic factor, was associated with heart failure and poor prognosis. Established heart related markers, such as N-terminal fragment of A-type natriuretic peptide/brain natriuretic peptide (NT-proANP, NT-proBNP) or troponin I/T are elevated in SSc-PAH but are not specific for the right ventricle and may be increased to the same extent in left heart disease. Taken together, there is no universal specific biomarker for SSc-PAH, however, there is a pattern of markers that is strongly associated with a risk of vascular complications in SSc patients. Further comprehensive, multicenter and prospective studies are warranted to develop reliable algorithms for detection and prognosis of SSc-PAH.