Introduction: Posttranscriptional gene regulation (PTR) critically controls inflammation by modulating messenger RNA (mRNA) turn over and translation rates. RNA-binding proteins (RBP) coordinate PTR by binding to conserved sequences of targeted mRNAs. The majority of pathogenetic genes expressed in immune and structural cells in chronic lung inflammation are amenable to RBP-coordinated PTR, yet the role of RBPs in this setting remains elusive. Objectives: The overall aim of the study is to characterize the expression pattern and activation state of a panel of oxidative stress-regulated RBPs in patients with asthma, COPD and in relevant control subjects. RBP expression was evaluated by: immunohistochemistry (IHC) with validated antibodies using standard immunoper oxidase techniques on formalin-fixed, paraffin embedded tissues obtained from bronchial mucosa and peripheral lung samples of well-phenotyped, mild to moderate stable COPD patients (n=10), compared with age/gender/smoking history-matched smokers with nor mal lung function (n=12); by Western blot (WB) and real-time PCR in the human airway epithelial cell line BEAS-2B stimulated with H2O2 (100 lM, 24 h, n=3); by real-time PCR in PBMC of COPD patients (n=5)/controls (n=7) characterized as for the IHC study. Study was approved by local Ethics Committees. Statistical significance was assessed by analysis of variance and Kruskal–Wallis tests. Results: We identified by IHC three of the RBPs mainly regulating inflammatory transcripts: tristetraprolin (TTP), Hu antigen R (HuR) and heterogeneous nuclear ribonucleoprotein D (HnRNPD, also ter med AUF-1) in the airways’ samples of COPD patients and controls. Expression of AUF-1, an RBP functionally linked to accelerated decay of inflammatory transcripts, was significantly (P=.015) lower in bronchial epithelium of COPD samples vs. controls, while remaining comparable between groups in bronchiolar epithelium, glandular cells and alveolar macrophages. Decreased AUF-1 expression was also found by WB analysis of BEAS2B cells treated with H2O2; in PBMC, RBP mRNA levels did not differ between groups.

Differential expression of RNA-binding proteins in airway epithelium in chronic lung inflammation

Ricciardi L;Caramori G;
2017-01-01

Abstract

Introduction: Posttranscriptional gene regulation (PTR) critically controls inflammation by modulating messenger RNA (mRNA) turn over and translation rates. RNA-binding proteins (RBP) coordinate PTR by binding to conserved sequences of targeted mRNAs. The majority of pathogenetic genes expressed in immune and structural cells in chronic lung inflammation are amenable to RBP-coordinated PTR, yet the role of RBPs in this setting remains elusive. Objectives: The overall aim of the study is to characterize the expression pattern and activation state of a panel of oxidative stress-regulated RBPs in patients with asthma, COPD and in relevant control subjects. RBP expression was evaluated by: immunohistochemistry (IHC) with validated antibodies using standard immunoper oxidase techniques on formalin-fixed, paraffin embedded tissues obtained from bronchial mucosa and peripheral lung samples of well-phenotyped, mild to moderate stable COPD patients (n=10), compared with age/gender/smoking history-matched smokers with nor mal lung function (n=12); by Western blot (WB) and real-time PCR in the human airway epithelial cell line BEAS-2B stimulated with H2O2 (100 lM, 24 h, n=3); by real-time PCR in PBMC of COPD patients (n=5)/controls (n=7) characterized as for the IHC study. Study was approved by local Ethics Committees. Statistical significance was assessed by analysis of variance and Kruskal–Wallis tests. Results: We identified by IHC three of the RBPs mainly regulating inflammatory transcripts: tristetraprolin (TTP), Hu antigen R (HuR) and heterogeneous nuclear ribonucleoprotein D (HnRNPD, also ter med AUF-1) in the airways’ samples of COPD patients and controls. Expression of AUF-1, an RBP functionally linked to accelerated decay of inflammatory transcripts, was significantly (P=.015) lower in bronchial epithelium of COPD samples vs. controls, while remaining comparable between groups in bronchiolar epithelium, glandular cells and alveolar macrophages. Decreased AUF-1 expression was also found by WB analysis of BEAS2B cells treated with H2O2; in PBMC, RBP mRNA levels did not differ between groups.
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11570/3230729
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