Human metapneumovirus (hMPV) is a major cause of lower respiratory tract infections (LRTIs) in infants, elderly and immunocompromised patients. In this study, we show that hMPV can infect in a similar manner epithelial cells representative of different tracts of the airways. hMPV-induced expression of chemokines IL-8 and RANTES in primary small alveolar epithelial cells (SAE) and in a human alveolar type II-like epithelial cell line (A549) was similar, suggesting that A549 cells can be used as a model to study lower airway epithelial cell responses to hMPV infection. A549 secreted a variety of CXC and CC chemokines, cytokines and type I interferons, following hMPV infection. hMPV was also a strong inducer of transcription factors belonging to nuclear factor (NF)-kappaB, interferon regulatory factors (IRFs) and signal transducers and activators of transcription (STATs) families, which are known to orchestrate the expression of inflammatory and immunomodulatory mediators.

Human metapneumovirus (HMPV) is a recently discovered pathogen that causes a significant proportion of respiratory infections in young infants, the elderly and immunocompromised patients. Very little is known regarding the cellular signalling elicited by this virus in airway epithelial cells, the target of HMPV infection. In this study, we investigated the role of the RNA helicases retinoic acid inducible gene-I (RIG-I) and melanoma differentiation-associated gene-5 (MDA-5) as the main pattern recognition receptors (PRRs) involved in viral detection and subsequent expression of proinflammatory and antiviral genes. HMPV infection readily induced RIG-I and MDA-5 gene and protein expression in A549 cells, a type II-like alveolar epithelial cell line. Expression of dominant-negative (DN) RIG-I or downregulation of RIG-I gene expression using small interfering RNA (siRNA) significantly decreased HMPV-induced beta interferon (IFN-b), interleukin (IL)-8 and RANTES gene transcription, by inhibiting viral-induced activation of nuclear factor (NF)-kB and interferon regulatory factor (IRF), leading to enhanced viral replication. On the other hand, MDA-5 did not seem to play a significant role in HMPV-induced cellular responses. Mitochondrial antiviral signalling protein (MAVS), an adaptor protein linking both RIG-I and MDA-5 to downstream activation of IRF-3 and NF-kB, was also necessary for HMPVinduced cellular signalling. Expression of a DN MAVS significantly reduced IFN-b and chemokine gene transcription, by inhibiting NF-kB-and IRF-dependent gene transcription, in response to HMPV infection. Our results show that HMPV activates the RIG-I-MAVS signalling pathway in airway epithelial cells, leading to the expression of important proinflammatory and antiviral molecules involved in the innate immune response to viruses. INTRODUCTIONHuman metapneumovirus (HMPV) is a recently identified respiratory RNA virus belonging to the family Paramyxoviridae. HMPV is responsible for a significant proportion of upper and lower respiratory tract infections in young children, the elderly and immunocompromised patients, causing bronchiolitis, croup, asthma exacerbation and even pneumonia (Boivin et al., 2002;Esper et al., 2003;Williams et al., 2004), second only to respiratory syncytial virus (RSV) (Kahn, 2006;Principi et al., 2006;Williams et al., 2004), also a paramyxovirus. While there is emerging literature on the clinical and epidemiological features of HMPV infection, very little is known regarding the cellular signalling activated by this pathogen in infected cells. We have recently demonstrated that HMPV is a potent stimulus for cytokine, chemokine and type I interferon (IFN) production in cultured human alveolar epithelial cells, and that HMPV-induced chemokine expression is dependent on viral replication (Bao et al., 2007). We have also shown that HMPV infection induces activation of nuclear factor (NF)-kB and interferon regulatory factor (IRF) transcription factors (Bao et al., 2007), which have been shown to play a fundamental ...

Human metapneumovirus (hMPV) is a major cause of lower respiratory tract infections in infants, elderly and immunocompromised patients. Little is known about the response to hMPV infection of airway epithelial cells, which play a pivotal role in initiating and shaping innate and adaptive immune responses. In this study, we analyzed the transcriptional profiles of airway epithelial cells infected with hMPV using high-density oligonucleotide microarrays. Of the 47,400 transcripts and variants represented on the Affimetrix GeneChip Human Genome HG-U133 plus 2 array, 1601 genes were significantly altered following hMPV infection. Altered genes were then assigned to functional categories and mapped to signaling pathways. Many up-regulated genes are involved in the initiation of pro-inflammatory and antiviral immune responses, including chemokines, cytokines, type I interferon and interferon-inducible proteins. Other important functional classes up-regulated by hMPV infection include cellular signaling, gene transcription and apoptosis. Notably, genes associated with antioxidant and membrane transport activity, several metabolic pathways and cell proliferation were down-regulated in response to hMPV infection. Real-time PCR and Western blot assays were used to confirm the expression of genes related to several of these functional groups. The overall result of this study provides novel information on host gene expression upon infection with hMPV and also serves as a foundation for future investigations of genes and pathways involved in the pathogenesis of this important viral infection. Furthermore, it can facilitate a comparative analysis of other paramyxoviral infections to determine the transcriptional changes that are conserved versus the one that are specific to individual pathogens.