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Characterising the mechanism of airway smooth muscle ?2 adrenoceptor desensitization by rhinovirus infected bronchial epithelial cells.

ABSTRACT: Rhinovirus (RV) infections account for approximately two thirds of all virus-induced asthma exacerbations and often result in an impaired response to ?2 agonist therapy. Using an in vitro model of RV infection, we investigated the mechanisms underlying RV-induced ?2 adrenoceptor desensitization in primary human airway smooth muscle cells (ASMC). RV infection of primary human bronchial epithelial cells (HBEC) for 24 hours produced conditioned medium that caused ?2 adrenoceptor desensitization on ASMCs without an effect on ASMCs viability. Less than 3 kDa size fractionation together with trypsin digestion of RV-induced conditioned medium did not prevent ?2 adrenoceptor desensitization, suggesting it could potentially be mediated by a small peptide or lipid. RV infection of BECs, ASMCs and fibroblasts produced prostaglandins, of which PGE2, PGF2? and PGI2 had the ability to cause ?2 adrenoceptor desensitization on ASMCs. RV-induced conditioned medium from HBECs depleted of PGE2 did not prevent ASMC ?2 adrenoceptor desensitization; however this medium induced PGE2 from ASMCs, suggesting that autocrine prostaglandin production may be responsible. Using inhibitors of cyclooxygenase and prostaglandin receptor antagonists, we found that ?2 adrenoceptor desensitization was mediated through ASMC derived COX-2 induced prostaglandins. Since ASMC prostaglandin production is unlikely to be caused by RV-induced epithelial derived proteins or lipids we next investigated activation of toll-like receptors (TLR) by viral RNA. The combination of TLR agonists poly I:C and imiquimod induced PGE2 and ?2 adrenoceptor desensitization on ASMC as did the RNA extracted from RV-induced conditioned medium. Viral RNA but not epithelial RNA caused ?2 adrenoceptor desensitization confirming that viral RNA and not endogenous human RNA was responsible. It was deduced that the mechanism by which ?2 adrenoceptor desensitization occurs was by pattern recognition receptor activation of COX-2 induced prostaglandins.

SUBMITTER: Van Ly D

PROVIDER: S-EPMC3574065 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Characterising the mechanism of airway smooth muscle β2 adrenoceptor desensitization by rhinovirus infected bronchial epithelial cells.

Van Ly David D Faiz Alen A Jenkins Christine C Crossett Ben B Black Judith L JL McParland Brent B Burgess Janette K JK Oliver Brian G G BG

PloS one 20130215 2

Rhinovirus (RV) infections account for approximately two thirds of all virus-induced asthma exacerbations and often result in an impaired response to β2 agonist therapy. Using an in vitro model of RV infection, we investigated the mechanisms underlying RV-induced β2 adrenoceptor desensitization in primary human airway smooth muscle cells (ASMC). RV infection of primary human bronchial epithelial cells (HBEC) for 24 hours produced conditioned medium that caused β2 adrenoceptor desensitization on ...[more]

PMID: 23457497

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Project description:BACKGROUND AND PURPOSE: ??-Adrenoceptor agonists are important therapeutic agents in the treatment of asthma and chronic obstructive pulmonary disease. The regular use of these drugs has been associated with proasthmatic-like changes that limit their efficacy and increase the risk of severe adverse reactions. We investigated whether the peroxisome-proliferator-activated receptor (PPAR)? agonist rosiglitazone modulated salbutamol-induced ??-adrenoceptor desensitization in vivo and in vitro. EXPERIMENTAL APPROACH: An in vivo model of homologous ??-adrenoceptor desensitization, established in guinea-pigs by administering salbutamol continuously, was used to study the ability of rosiglitazone to prevent ??-adrenoceptor tolerance. In vitro experiments on human bronchial smooth muscle cells were performed to increase the clinical relevance of the study. KEY RESULTS: In tracheal smooth muscle tissues from desensitized animals, we observed a decrease in the protective effect of salbutamol on carbachol-induced contraction, a hyperresponsiveness to cholinergic stimuli, a modest underexpression of ??-adrenoceptor gene and a marked decrease in ?-adrenoceptor number, relative to control values. Treatment with rosiglitazone preserved salbutamol relaxant activity, mitigated carbachol hyperresponsiveness and partially restored ??-adrenoceptor binding sites in tracheal tissues from homologously desensitized animals. The highly selective PPAR? agonist, GW1929, reproduced the effect of rosiglitazone, in vivo. In vitro ??-adrenoceptor desensitization decreased salbutamol-mediated cAMP production, without affecting forskolin responses and ??-adrenoceptor expression. Rosiglitazone and 15-deoxy-?¹²(,)¹?-prostaglandin J? restored salbutamol sensitivity in homologously desensitized cells. CONCLUSIONS AND IMPLICATIONS: These data suggest a potential pharmacodynamic interaction between PPAR? agonists and salbutamol on airway smooth muscle responsiveness, supporting the therapeutic potential of this combination in chronic airway disease.

Project description:BackgroundThe effectiveness of bronchial thermoplasty (BT) has been reported in patients with severe asthma. This study compared the effects of BT and cryoballoon ablation (CBA) therapy on the airway smooth muscle (ASM).MethodsEight healthy male beagle dogs were included in this experiment. In the preliminary experiment, one dog received BT treatment for both lower lobe bronchus, another dog received CBA treatment for 7 s on the upper and lower lobe of right bronchus, and 30 s on the left upper and lower lobe. The treatments were performed twice at an interval of 1 month. In subsequent experiments, the right lower lobe bronchus was treated with BT, and the left lower lobe bronchus was treated with CBA. The effects of treatment were observed after 1 (n = 3) month and 6 months (n = 3). Hematoxylin-eosin staining, Masson trichrome staining, and immunohistochemical staining were used to compare the effects of BT and CBA therapy on the ASM thickness, collagen fibers synthesis, and M3 receptor expression after treatment. One-way analysis of variance with Dunnett post hoc test was used to analyze the differences among groups.ResultsIn the preliminary experiment, the ASM ablation effect of 30-s CBA was equivalent to that of 7-s CBA (ASM thickness: 30.52 ± 7.75 μm vs. 17.57 ± 15.20 μm, P = 0.128), but the bronchial mucociliary epithelium did not recover, and large numbers of inflammatory cells had infiltrated the mucosal epithelium at 1-month post-CBA with 30-s freezing. Therefore, we chose 7 s as the CBA treatment time in our follow-up experiments. Compared with the control group (35.81 ± 11.02 μm), BT group and CBA group (13.41 ± 4.40 μm and 4.81 ± 4.44 μm, respectively) had significantly decreased ASM thickness after 1 month (P < 0.001). Furthermore, the ASM thickness was significantly lower in the 1-month post-CBA group than in the 1-month post-BT group (P = 0.015). There was no significant difference in ASM thickness between the BT and CBA groups after six months (9.92 ± 4.42 μm vs. 7.41 ± 7.20 μm, P = 0.540). Compared with the control group (0.161 ± 0.013), the average optical density of the ASM M3 receptor was significantly decreased in 6-month post-BT, 1-month post-CBA, and 6-month post-CBA groups (0.070 ± 0.022, 0.072 ± 0.012, 0.074 ± 0.008, respectively; all P < 0.001). There was no significant difference in the average optical density of ASM M3 receptor between the BT and CBA therapy groups after six months (P = 0.613).ConclusionsCBA therapy effectively ablates the ASM, and its ablation effect is equivalent to that of BT with a shorter onset time. A neural mechanism is involved in both BT and CBA therapy.

Project description:Asthma exacerbations are commonly associated with rhinovirus (RV) infection. Interleukin-33 (IL-33) plays an important role during exacerbation by enhancing Type 2 inflammation. Recently we showed that RV infects bronchial smooth muscle cells (BSMCs) triggering production of interferons and IL-33. Here we compared levels of RV-induced IL-33 in BSMCs from healthy and asthmatic subjects, and explored the involvement of pattern-recognition receptors (PRRs) and downstream signalling pathways in IL-33 expression. BSMCs from healthy and severe and non-severe asthmatic patients were infected with RV1B or stimulated with the PRR agonists poly(I:C) (Toll-like receptor 3 (TLR3)), imiquimod (TLR7) and poly(I:C)/LyoVec (retinoic acid-inducible gene 1 (RIG-I)/melanoma differentiation-associated protein 5 (MDA5)). Knockdown of TLR3, RIG-I and MDA5 was performed, and inhibitors targeting TBK1, nuclear factor-κB (NF-κB) and transforming growth factor (TGF)-β-activated kinase 1 (TAK1) were used. Gene and protein expression were assessed. RV triggered IL-33 gene and protein expression in BSMCs. BSMCs from patients with non-severe asthma showed higher baseline and RV-induced IL-33 gene expression compared to cells from patients with severe asthma and healthy controls. Furthermore, RV-induced IL-33 expression in BSMCs from healthy and asthmatic individuals was attenuated by knockdown of TLR3. Inhibition of TAK1, but not NF-κB or TBK1, limited RV-induced IL-33. The cytokine secretion profile showed higher production of IL-33 in BSMCs from patients with non-severe asthma compared to healthy controls upon RV infection. In addition, BSMCs from patients with non-severe asthma had higher levels of RV-induced IL-8, TNF-α, IL-1β, IL-17A, IL-5 and IL-13. RV infection caused higher levels of IL-33 and increased pro-inflammatory and Type 2 cytokine release in BSMCs from patients with non-severe asthma. RV-induced IL-33 expression was mainly regulated by TLR3 and downstream via TAK1. These signalling molecules represent potential therapeutic targets for treating asthma exacerbations.

Dataset Information

Characterising the mechanism of airway smooth muscle ?2 adrenoceptor desensitization by rhinovirus infected bronchial epithelial cells.

Publications

Characterising the mechanism of airway smooth muscle β2 adrenoceptor desensitization by rhinovirus infected bronchial epithelial cells.

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