ABSTRACT
Rhinoviral infection is an important trigger of acute inflammatory exacerbations in patients with underlying airway disease. We have previously established that interleukin-1ß (IL-1ß) is central in the communication between epithelial cells and monocytes during the initiation of inflammation. In this study we explored the roles of IL-1ß and its signaling pathways in the responses of airway cells to rhinovirus-1B (RV-1B) and further determined how responses to RV-1B were modified in a model of bacterial coinfection. Our results revealed that IL-1ß dramatically potentiated RV-1B-induced proinflammatory responses, and while monocytes did not directly amplify responses to RV-1B alone, they played an important role in the responses observed with our coinfection model. MyD88 is the essential signaling adapter for IL-1ß and most Toll-like receptors. To examine the role of MyD88 in more detail, we created stable MyD88 knockdown epithelial cells using short hairpin RNA (shRNA) targeted to MyD88. We determined that IL-1ß/MyD88 plays a role in regulating RV-1B replication and the inflammatory response to viral infection of airway cells. These results identify central roles for IL-1ß and its signaling pathways in the production of CXCL8, a potent neutrophil chemoattractant, in viral infection. Thus, IL-1ß is a viable target for controlling the neutrophilia that is often found in inflammatory airway disease and is exacerbated by viral infection of the airways.
Subject(s)
Interleukin-1beta/physiology , Myeloid Differentiation Factor 88/metabolism , Picornaviridae Infections/metabolism , Rhinovirus/isolation & purification , Signal Transduction , Blotting, Western , Cell Communication , Cell Line , Cytopathogenic Effect, Viral , Enzyme-Linked Immunosorbent Assay , Gene Knockdown Techniques , Humans , Myeloid Differentiation Factor 88/genetics , Picornaviridae Infections/virology , Polymerase Chain Reaction , Rhinovirus/pathogenicityABSTRACT
Human neutrophils express Toll-like receptor 4 (TLR4) at low levels, and the role of this receptor in neutrophil responses to microbial stimuli has been questioned. Genetic manipulation of these cells to enable the study of the role of proteins such as TLR4 in their function is challenging. Here, we show that primary human neutrophils rapidly express novel proteins such as enhanced green fluorescent protein (eGFP) after transduction with lentivirus. Stimulation of transduced neutrophils with lipopolysaccharide (LPS) resulted in increased cell survival, which was inhibited when neutrophils were transduced with a lentivirus encoding a dominant negative (dn) TLR4 protein. LPS-induced survival was also inhibited by lentiviruses encoding dnMyD88 or a truncated TRIF (Toll/interleukin-1R homologous domain-containing adapter protein inducing interferon-beta) molecule, whilst, in contrast, neutrophil survival was enhanced by overexpression of kinase-mutated interleukin-1 receptor-associated kinase 1 (kmIRAK-1), which activated nuclear factor (NF)-kappaB. These studies provide proof of the role of TLR4 in human neutrophil biology, have begun to elucidate TLR-dependent pathways regulating neutrophil survival, and demonstrate that neutrophils can be genetically manipulated to enhance or inhibit survival.
