Matrix Metalloprotease 9 Mediates Neutrophil Migration into the Airways in Response to Influenza Virus-Induced Toll-Like Receptor Signaling

Abstract

The early inflammatory response to influenza virus infection contributes to severe lung disease and continues to pose a serious threat to human health. The mechanisms by which neutrophils gain entry to the respiratory tract and their role during pathogenesis remain unclear. Here, we report that neutrophils significantly contributed to morbidity in a pathological mouse model of influenza virus infection. Using extensive immunohistochemistry, bone marrow transfers, and depletion studies, we identified neutrophils as the predominant pulmonary cellular source of the gelatinase matrix metalloprotease (MMP) 9, which is capable of digesting the extracellular matrix. Furthermore, infection of MMP9-deficient mice showed that MMP9 was functionally required for neutrophil migration and control of viral replication in the respiratory tract. Although MMP9 release was toll-like receptor (TLR) signaling-dependent, MyD88-mediated signals in non-hematopoietic cells, rather than neutrophil TLRs themselves, were important for neutrophil migration. These results were extended using multiplex analyses of inflammatory mediators to show that neutrophil chemotactic factor, CCL3, and TNFα were reduced in the Myd88^−/− airways. Furthermore, TNFα induced MMP9 secretion by neutrophils and blocking TNFα in vivo reduced neutrophil recruitment after infection. Innate recognition of influenza virus therefore provides the mechanisms to induce recruitment of neutrophils through chemokines and to enable their motility within the tissue via MMP9-mediated cleavage of the basement membrane. Our results demonstrate a previously unknown contribution of MMP9 to influenza virus pathogenesis by mediating excessive neutrophil migration into the respiratory tract in response to viral replication that could be exploited for therapeutic purposes. Influenza-associated morbidity and mortality due to yearly epidemics and sporadic, devastating pandemics are a significant health and economic burden. Severe complications arising from highly virulent viruses are associated with rapid, massive inflammatory cell infiltration. Although neutrophils are the predominant cell population recruited to the lung in response to pandemic influenza viruses, the mechanisms by which they gain entry to the respiratory tract remain unclear. In this study, we show a previously unknown contribution of MMP9 to influenza pathogenesis by mediating excessive neutrophil migration into the lung, which not only controls viral replication, but also contributes to morbidity. The in vivo relevance of MMP9-derived enzymatic activity in neutrophils is controversial and understudied, but our data provide new evidence that innate recognition of influenza virus attracts neutrophils that secrete MMP9, which enables them to traverse the basement membrane of the lung by digesting the extracellular matrix. The dichotomy of MMP9 function in immunity versus pathology provides real challenges for targeting MMP9 for therapeutic purposes. Nevertheless, finding the balance to modulate neutrophil numbers following influenza virus infection will allow for innate immunity to be boosted whilst preventing pathology associated with pandemic strains.