Dataset Information

Cytokines induce an early steroid resistance in airway smooth muscle cells: novel role of interferon regulatory factor-1.

ABSTRACT: We have previously shown that long-term treatment of airway smooth muscle (ASM) cells with a combination of TNF-alpha and IFN-gamma impaired steroid anti-inflammatory action through the up-regulation of glucocorticoid receptor beta isoform (GRbeta) (Mol Pharmacol 2006;69:588-596). We here found that steroid actions could also be suppressed by short-term exposure of ASM cells to TNF-alpha and IFN-gamma (6 h) as shown by the abrogated glucocorticoid responsive element (GRE)-dependent gene transcription; surprisingly, neither GRalpha nuclear translocation nor GRbeta expression was affected by cytokine mixture. The earlier induction of CD38, a molecule recently involved in asthma, seen with TNF-alpha and IFN-gamma combination but not with cytokine alone, was also completely insensitive to steroid pretreatment. Chromatin-immunoprecipitation (IP) and siRNA strategies revealed not only increased binding of interferon regulatory factor 1 (IRF-1) transcription factor to CD38 promoter, but also its implication in regulating CD38 gene transcription. Interestingly, the capacity of fluticasone to completely inhibit TNF-alpha-induced IRF-1 expression, IRF-1 DNA binding, and transactivation activities was completely lost in cells exposed to TNF-alpha and IFN-gamma in combination. This early steroid dysfunction seen with cytokine combination could be reproduced by enhancing IRF-1 cellular levels using constitutively active IRF-1, which dose-dependently inhibited GRE-dependent gene transcription. Consistently, reducing IRF-1 cellular levels using siRNA approach significantly restored steroid transactivation activities. Collectively, our findings demonstrate for the first time that IRF-1 is a novel alternative GRbeta-independent mechanism mediating steroid dysfunction induced by pro-asthmatic cytokines, in part via the suppression of GRalpha activities.

SUBMITTER: Tliba O

PROVIDER: S-EPMC2274949 | biostudies-literature | 2008 Apr

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Cytokines induce an early steroid resistance in airway smooth muscle cells: novel role of interferon regulatory factor-1.

Tliba Omar O Damera Gautam G Banerjee Audreesh A Gu Su S Baidouri Hasna H Keslacy Stefan S Amrani Yassine Y

American journal of respiratory cell and molecular biology 20071018 4

We have previously shown that long-term treatment of airway smooth muscle (ASM) cells with a combination of TNF-alpha and IFN-gamma impaired steroid anti-inflammatory action through the up-regulation of glucocorticoid receptor beta isoform (GRbeta) (Mol Pharmacol 2006;69:588-596). We here found that steroid actions could also be suppressed by short-term exposure of ASM cells to TNF-alpha and IFN-gamma (6 h) as shown by the abrogated glucocorticoid responsive element (GRE)-dependent gene transcri ...[more]

PMID: 17947510

Similar Datasets

Project description:To elucidate the regulation of glucocorticoid receptor (GR) signaling under pro-asthmatic conditions, cultured human airway smooth muscle (HASM) cells were treated with proinflammatory cytokines or GR ligands alone and in combination, and then examined for induced changes in ligand-dependent and -independent GR activation and downstream signaling events. Ligand stimulation with either cortisone or dexamethsone (DEX) acutely elicited GR translocation to the nucleus and, comparably, ligand-independent stimulation either with the Th2 cytokine, IL-13, or the pleiotropic cytokine combination, IL-1β/TNFα, also acutely evoked GR translocation. The latter response was potentiated by combined exposure of cells to GR ligand and cytokine. Similarly, treatment with either DEX or IL-13 alone induced GR phosphorylation at its serine-211 residue (GR(Ser211)), denoting its activated state, and combined treatment with DEX+IL-13 elicited heightened and sustained GR(Ser211) phosphorylation. Interestingly, the above ligand-independent GR responses to IL-13 alone were not associated with downstream GR binding to its consensus DNA sequence or GR transactivation, whereas both DEX-induced GR:DNA binding and transcriptional activity were significantly heightened in the presence of IL-13, coupled to increased recruitment of the transcriptional co-factor, MED14. The stimulated GR signaling responses to DEX were prevented in IL-13-exposed cells wherein GR(Ser211) phosphorylation was suppressed either by transfection with specific serine phosphorylation-deficient mutant GRs or treatment with inhibitors of the MAPKs, ERK1/2 and JNK. Collectively, these novel data highlight a heretofore-unidentified homeostatic mechanism in HASM cells that involves pro-asthmatic cytokine-driven, MAPK-mediated, non-ligand-dependent GR activation that confers heightened glucocorticoid ligand-stimulated GR signaling. These findings raise the consideration that perturbations in this homeostatic cytokine-driven GR signaling mechanism may be responsible, at least in part, for the insensirtivity to glucocorticoid therapy that is commonly seen in individuals with severe asthma.

Project description:BackgroundCorticosteroids remain a key therapy for treating children with asthma. Patients with severe asthma are insensitive, resistant, or refractory to corticosteroids and have poorly controlled symptoms that involve airway inflammation, airflow obstruction, and frequent exacerbations. While the pathways that mediate corticosteroid insensitivity in asthma remain poorly defined, recent studies suggest that enhanced Th1 pathways, mediated by TNFα and IFNγ, may play a role. We previously reported that the combined effects of TNFα and IFNγ promote corticosteroid insensitivity in developing human airway smooth muscle (ASM).MethodsTo further understand the effects of TNFα and IFNγ on corticosteroid sensitivity in the context of neonatal and pediatric asthma, we performed RNA sequencing (RNA-seq) on human pediatric ASM treated with fluticasone propionate (FP), TNFα, and/or IFNγ.ResultsWe found that TNFα had a greater effect on gene expression (~ 1000 differentially expressed genes) than IFNγ (~ 500 differentially expressed genes). Pathway and transcription factor analyses revealed enrichment of several pro-inflammatory responses and signaling pathways. Interestingly, treatment with TNFα and IFNγ augmented gene expression with more than 4000 differentially expressed genes. Effects of TNFα and IFNγ enhanced several pro-inflammatory genes and pathways related to ASM and its contributions to asthma pathogenesis, which persisted in the presence of corticosteroids. Co-expression analysis revealed several gene networks related to TNFα- and IFNγ-mediated signaling, pro-inflammatory mediator production, and smooth muscle contractility. Many of the co-expression network hubs were associated with genes that are insensitive to corticosteroids.ConclusionsTogether, these novel studies show the combined effects of TNFα and IFNγ on pediatric ASM and implicate Th1-associated cytokines in promoting ASM inflammation and hypercontractility in severe asthma.

Project description:Rationale: Steroids are the mainstay of asthma therapy. However, it is unclear whether the benefits of steroids in asthma are merely based on anti-inflammatory properties. Steroids may also alter gene expression of airway smooth muscle (ASM). Hypothesis and Aims: We hypothesized that the transcriptomic profile of the ASM layer in endobronchial biopsies of atopic asthma patients changes by oral steroid therapy. First, we examined the change in ASM transcriptomic profile in endobronchial biopsies after 14 days of oral steroid therapy. Second, we investigated the association between changes in ASM transcriptomic profile and airway function. Methods: 12 atopic steroid-free asthma patients were included in this double-blind intervention study. Endobronchial biopsies were taken before and after 14 days of oral prednisolon (n=6) or placebo (n=6). RNA of laser-dissected ASM was sequenced (RNA-Seq) using the GS FLX+ System (454/Roche). Gene networks were identified using Ingenuity Pathway Analysis. RNA-Seq reads were assumed to follow a negative binomial distribution. At the current sample size the estimated false discovery rate was approximately 3%. Results: 15 genes were significantly changed by 14 days of oral prednisolon. 2 of these genes (FAM129A, SYNPO2) were associated with the methacholine PC20 (r=0.637, p=0.035; r=0.662, p=0.027). Pathway analysis revealed 3 gene networks that were associated with cellular functions including cellular growth, proliferation, and development. Conclusion: Oral prednisolon changes the gene expression profile of the ASM layer in asthma. This indicates that steroids also exert effects on the transcriptomic level of ASM in addition to their anti-inflammatory properties, which can promote improved airway function. The current randomized, double-blind, parallel, placebo-controlled intervention study comprised 4 visits. At visit 1, asthma patients were screened according to the in- and exclusion criteria prior to enrollment. Additionally, spirometry and methacholine bronchoprovocation test were performed. At visit 2, FEV1 reversibility was measured and endobronchial biopsies were collected during a bronchoscopy. Asthma patients were then prescribed oral prednisolon at a dose of 0.5 mg/kg per day or placebo for 14 consecutive days. The dosage and dosing scheme was based on international recommendations for the treatment of acute exacerbations [1]. On the 11th day after visit 2, the patients visited the lung function laboratory for spirometry and methacholine bronchoprovocation test. Finally, at visit 4 (15th day after visit 2) FEV1 reversibility was measured and endobronchial biopsies were collected by bronchoscopy. Airway smooth muscle was collected from the biopsies by laser capture microdissection and total RNA isolated. cDNA was prepared using the Ovation RNA-Seq System (NuGEN). RNA-Seq was performed using the GS FLX+ instrument (454/Roche). Sequence reads were mapped against the human genome (hg19; UCSC). Comparison of the numbers of reads per gene between the prednisolon and placebo study group was carried out with the R package DESeq.

Dataset Information

Cytokines induce an early steroid resistance in airway smooth muscle cells: novel role of interferon regulatory factor-1.

Publications

Cytokines induce an early steroid resistance in airway smooth muscle cells: novel role of interferon regulatory factor-1.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets