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

Rhinovirus (RV) infections account for approximately two thirds of all virus-induced asthma exacerbations and often result in an impaired response to b2 agonist therapy. Using an in vitro model of RV infection, we investigated the mechanisms underlying RV-induced b2 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 b2 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 b2 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, PGF2a and PGI2 had the ability to cause b2 adrenoceptor desensitization on ASMCs. RV-induced conditioned medium from HBECs depleted of PGE2 did not prevent ASMC b2 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 b2 adrenoceptor desensitization was mediated through ASMC derived COX-2 induced prostaglandins. Since ASMC prostaglandin production is unlikely to be caused by RVinduced 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 b2 adrenoceptor desensitization on ASMC as did the RNA extracted from RV-induced conditioned medium. Viral RNA but not epithelial RNA caused b2 adrenoceptor desensitization confirming that viral RNA and not endogenous human RNA was responsible. It was deduced that the mechanism by which b2 adrenoceptor desensitization occurs was by pattern recognition receptor activation of COX-2 induced prostaglandins. Citation: Van Ly D, Faiz A, Jenkins C, Crossett B, Black JL, et al. (2013) Characterising the Mechanism of Airway Smooth Muscle b2 Adrenoceptor Desensitization by Rhinovirus Infected Bronchial Epithelial Cells. PLoS ONE 8(2): e56058. doi:10.1371/journal.pone. 0056058 Editor: Vladimir V. Kalinichenko, Cincinnati Children’s Hospital Medical Center, United States of America Received October 11, 2012; Accepted January 4, 2013; Published February 15, 2013 Copyright: 2013 Van Ly et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: JKB is supported by a National Health and Medical Research Council Career Development Fellowship #1032695. JLB is supported by a National Health and Medical Research Council Senior Principal Research Fellowship #571098. BGGO is supported by a National Health and Medical Research Council Career Development Fellowship APP1026880. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: dvan7422@gmail.com