Toll-like receptor 7 activation reduces the contractile response of airway smooth muscle

Abstract

Viral respiratory infections are a major cause of asthma exacerbations. The mechanisms by which such infections aggravate airway inflammation and hyperresponsiveness are complex and not fully understood. Toll-like receptor 7 is particularly relevant in the defence against common respiratory viruses, as it recognizes single-stranded viral RNA. The present study was designed to investigate the effect of Toll-like receptor 7 stimulation on airway smooth muscle reactivity. The presence of Toll-like receptor 7 within guinea pig airways was ensured with immunohistochemistry. The effects induced by 3 days of culture of tracheal segments with the Toll-like receptor 7 agonist R837 or the Toll-like receptor 7/8 agonist R848 were evaluated in a myograph organ bath system. The intracellular mechanisms involved were dissected using inhibitors of intracellular mitogen-activated protein kinase (MAPK) activity. Toll-like receptor 7 immunoreactivity was observed across the epithelial cell layer and in the airway smooth muscle cells. Treatment with R837 and R848 reduced the airway contractile responses to 5-hydroxytryptamine (5-HT). This effect was abolished upon treatment with inhibitors of the p38 MAPK pathway and nuclear factor (NF)-κB pathways. According to the present model, activation of Toll-like receptor 7 might prevent development of airway hyperresponsiveness by acting on the airway smooth muscle. The presented data support the idea that individuals with defect Toll-like receptor 7 function might be more prone to respond to virus infections with asthmatic exacerbations. Further, they suggest that inhaled Toll-like receptor 7 ligand might be an effective treatment alternative for asthma.

Introduction

Asthma is a common disorder of the respiratory tract characterized by inflammation, hyperresponsiveness and remodelling (Ward et al., 2002). Airway hyperresponsiveness is defined as an exaggerated airway narrowing in response to bronchoconstrictor agents (Okazawa et al., 1996). It is believed to be connected to ongoing airway inflammation, and there is a strong link to viral infections (Shelhamer et al., 1995, Tauro et al., 2008). The mechanisms behind this relation remain unclear.

Toll-like receptors are a set of intracellular and extracellular receptors which recognize microbial patterns. So far, ten different Toll-like receptors, Toll-like receptors 1–10, have been identified in humans. These receptors are crucial in the defence against pathogens (Medzhitov, 2001). The Toll-like receptors that primarily recognize viral products are Toll-like receptors 3, -7, -8 and -9. Of these Toll-like receptor 7 is of special interest since synthetic Toll-like receptor 7/Toll-like receptor 8 ligands seem to be effective in preventing the development of the asthmatic phenotype in in vivo models of asthma (Quarcoo et al., 2004, Sel et al., 2007). These ligands have also been demonstrated to inhibit the inflammatory reaction in response to allergens when the asthmatic phenotype has been established (Moisan et al., 2006, Sel et al., 2007). Further, Toll-like receptor 7/8 ligands appear to inhibit the increase in airway smooth muscle mass and the development of goblet cell hyperplasia, which are known to be associated with chronic asthma (Camateros et al., 2007).

We have previously developed a murine model of isolated airways and used it to demonstrate that long-term exposure to inflammatory mediators may alter the contractile responses to 5-hydroxytryptamine (5-HT) and other contractile mediators (Adner et al., 2002, Bryborn et al., 2004, Zhang et al., 2004, Zhang et al., 2007). In the airways, 5-HT is released by pulmonary neuroendocrine cells, platelets and, in some species, mast cells (Buckner et al., 1991, Cazzola and Matera, 2000). While the involvement of 5-HT in the development of airway hyperresponsiveness is debated, an anti-asthmatic effect of the 5-HT inhibitor ketanserin has been described in asthmatic patients (Cazzola et al., 1990) and increased levels of free 5-HT have been correlated to asthma severity (Lechin et al., 1996). Furthermore, the plasma concentration of free 5-HT increases during asthma exacerbations (Lechin et al., 1996), all justifying our use of 5-HT for evaluation of airway hyperreactivity. Increasing evidence suggests that deficiencies in the TLR system may be a cause of disease or aggravate an ongoing disease process (Abdollahi-Roodsaz et al., 2008, Zhang et al., 2006). If toll-like receptor 7 activation induces a protective effect against virus infection, a deficient Toll-like receptor 7 response or lack of proper activation in response to viruses could be an underlying cause of exacerbation of airway disease.

The intention of the present study was to use our in vitro model to investigate if Toll-like receptor 7 might have any direct effects on 5-HT-induced smooth muscle contractions.