Cytosolic pathogen recognition receptors (RIG-I & MDA-5) and interferon responses to rhinovirus infection in asthma.
Lead Research Organisation:
Imperial College London
Department Name: National Heart and Lung Institute
Abstract
Asthma is an increasing problem; its prevalence has increased over the last decades. Acute attacks (exacerbations) are common, can be fatal and occur despite the best currently available treatments. The majority of asthma exacerbations are related to viral infections and of these approximately 60% are caused by rhinoviruses. It has been shown in previous studies that cells from the lining of the airways in asthmatics respond differently to rhinovirus infection when compared to non asthmatic airway cells. These cells (Bronchial Epithelial Cells or BECs) are more susceptible to infection and this correlated with asthmatics having more prolonged/severe respiratory symptoms. The reasons why remains incompletely understood. The aim of this study is to investigate mechanisms of anti-viral defenses to identify any defect in asthmatic BECs and to see if this is responsible for the increased susceptibility to infection. This may suggest new approaches to prevention and/or treatment of asthma exacerbations. This will involve taking BECs from the airways of asthmatics and non asthmatics by bronchoscopy and then examining how they respond to infection with rhinovirus and compare any differences in response. Any differences will be looked at in greater detail. This work aims to identify new targets for treatments of asthma exacerbations which are currently a major problem for people with asthma.
Technical Summary
This study will investigate the role of RIG-I & MDA-5 in rhinovirus induced interferon production in primary bronchial epithelial cells (BECs) and bronchoalveolar lavage (BAL) cells from atopic asthmatics and normal controls. The aim is to determine whether any difference in expression/activity is responsible for the deficient interferon responses seen in asthma.
Asthma prevalence has risen and the major morbidity, mortality and health care costs are related to exacerbations. The majority of asthma exacerbations are associated with rhinovirus infections and asthmatics have increased susceptibility to rhinovirus infection, with deficient production of the type I interferon (IFN- ) and newly identified Type III (IL-28/29, ?) interferons. Many IFN-? responses are known to be mediated by two groups of pattern recognition receptors, the toll like receptors (TLRs) and the cytosolic receptors: retinoic acid inducible gene-I (RIG-I) and melanoma differentiation antigen-5 (MDA-5), though their importance in rhinovirus infections is unknown. Much less is known about regulation of the IFN- s although TLRs 3, 4, 7/8 and 9 have been reported to be involved, the roles of RIG-I and MDA-5 are unknown.
This study will focus on RIG-I and MDA-5 and will investigate primary BECs and BAL cells from asthmatic and normal subjects. The hypothesis is that RIG-I & MDA-5 are required for induction of type I & III IFNs by rhinovirus in normal healthy cells, and that there is a defect in this pathway in asthmatic cells which accounts for the deficient type I & III IFN responses seen in asthma.
12 atopic asthmatic subjects (mild/moderate severity and permitted to take regular asthma medication) and 12 healthy volunteers (all non smokers) will undergo clinical characterisation and bronchoscopy to obtain primary (BECs) and BAL cells on which to perform laboratory studies.
BECs will be expanded by culture and studied at passage 3-4, BAL cells will be studied fresh. The expression of RIG-I, MDA-5, their adaptor MAVS (IPS-1) and type I & type III IFNs will be determined. Cells will be unstimulated, infected with rhinovirus or UV-inactivated virus and comparison will be made between asthmatics and normals. mRNA expression will be quantified at baseline and 8hrs post infection by qPCR and proteins at baseline and 24-72hrs by ELISA/western blot. Functional relevance will be assessed by blocking expression in normal cells by siRNA and augmenting expression in asthmatic cells by over-expression using CMV driven promoters.
This study may identify new approaches to prevention and/or treatment of asthma exacerbations.
Asthma prevalence has risen and the major morbidity, mortality and health care costs are related to exacerbations. The majority of asthma exacerbations are associated with rhinovirus infections and asthmatics have increased susceptibility to rhinovirus infection, with deficient production of the type I interferon (IFN- ) and newly identified Type III (IL-28/29, ?) interferons. Many IFN-? responses are known to be mediated by two groups of pattern recognition receptors, the toll like receptors (TLRs) and the cytosolic receptors: retinoic acid inducible gene-I (RIG-I) and melanoma differentiation antigen-5 (MDA-5), though their importance in rhinovirus infections is unknown. Much less is known about regulation of the IFN- s although TLRs 3, 4, 7/8 and 9 have been reported to be involved, the roles of RIG-I and MDA-5 are unknown.
This study will focus on RIG-I and MDA-5 and will investigate primary BECs and BAL cells from asthmatic and normal subjects. The hypothesis is that RIG-I & MDA-5 are required for induction of type I & III IFNs by rhinovirus in normal healthy cells, and that there is a defect in this pathway in asthmatic cells which accounts for the deficient type I & III IFN responses seen in asthma.
12 atopic asthmatic subjects (mild/moderate severity and permitted to take regular asthma medication) and 12 healthy volunteers (all non smokers) will undergo clinical characterisation and bronchoscopy to obtain primary (BECs) and BAL cells on which to perform laboratory studies.
BECs will be expanded by culture and studied at passage 3-4, BAL cells will be studied fresh. The expression of RIG-I, MDA-5, their adaptor MAVS (IPS-1) and type I & type III IFNs will be determined. Cells will be unstimulated, infected with rhinovirus or UV-inactivated virus and comparison will be made between asthmatics and normals. mRNA expression will be quantified at baseline and 8hrs post infection by qPCR and proteins at baseline and 24-72hrs by ELISA/western blot. Functional relevance will be assessed by blocking expression in normal cells by siRNA and augmenting expression in asthmatic cells by over-expression using CMV driven promoters.
This study may identify new approaches to prevention and/or treatment of asthma exacerbations.
