Influenza A virus cell tropism and cytokine response in primary tracheal epithelial cell cultures

The primary site of replication for influenza A virus (IAV) in humans is the respiratory epithelia. Unlike transformed cell lines, primary differentiated tracheal epithelial cell (TEC) cultures closely model the in vivo environment with respect to the presence of distinct cell types, cell polarity, and ultrastructural organization. TEC cultures of mouse and hamster origin were used as an in vitro model to investigate the interactions between IAV and its target cell types. Hamster TEC culture experiments focused on the cell tropism and virus receptor distribution for both laboratory-adapted and clinical virus isolates. Virus receptors alpha2,3- and alpha2,6-linked sialic acid were predominantly expressed on non-ciliated cells, correlating with viral antigen expression. Clinical isolates displayed a stricter cell-type specificity for alpha2,6-linked sialic acid expressing cells than laboratory-adapted strains at late timepoints post infection. Hamster TEC and mouse TEC (mTEC) cultures differ in their cell-type distribution of alpha2,3 and alpha2,6 sialic acid, and thus different virus tropism patterns were observed during infection. mTEC cultures abundantly express alpha2,3-linked sialic acid exclusively on ciliated cells, while alpha2,6-linked sialic acid expression is not observed. Viral antigen of the IAV strain rWSN is expressed exclusively in ciliated cells in mTECs, correlating with sialic acid expression patterns.; For mTEC cultures, the investigations focused on the role of the IAV NS1 protein as a regulator of cytokine production and cytokine sensitivity during primary airway infection. A virus containing a mutation in the RNA binding domain of NS1 (rWSN NS1 R38A) resulted in increased inflammatory cytokine production during mTEC infection. In addition, rWSN NS1 R38A displayed decreased replication after cytokine pretreatment of primary cells, suggesting that this protein is a critical regulator of several aspects of cytokine production during influenza infection of primary airway epithelial cells. The studies utilized two different primary cell culture systems and provided insights into viral cell tropism and the innate immune responses of airway epithelial cells initiated by IAV infection.