Regulation of the Expression of Secretory Component by Human Airway Epithelial Cells in vitro

Abstract

Secretory component (SC) is a member of the immunoglobulin superfamily and is secreted uniquely by epithelial cells. Its primary role is the transcytosis of secretory immunoglobulin A (SIgA) from the basolateral to the apical side of epithelial cells. Membrane bound SC(polymeric immunoglobulin receptor; pIgR) forms a complex with dimeric IgA at the basolateral side of the cell, the complex is transcytosed and SIgA is released following proteolytic cleavage at the luminal side of the cell. When in complex with IgA, SC confers resistance and stability to the antibody. In addition plgR can transcytose by itself and it is proteolytically cleaved and released at the apical side of the cell as free SC. The role of free SC in mucosal protection remains elusive with both anti-inflammatory and anti-infective properties proposed. Previous studies investigating the role of SC in mucosal immunity have shown that SC is upregulated in airway epithelia by the inflammatory cytokines interferon gamma (IFN) and interleukin-4 (IL-4). Both IFNe and IL-4 are found in inflamed airways which suggest that SC could play an important anti-inflammatory role. Bacterial stimuli such as lipopolysaccharide (LPS) and lipoteichoic acid (LTA) have been shown to cause inflammation in vitro and in vivo and therefore the work presented in this thesis aimed to establish whether SC production by human airway epithelial cells in vitro was up regulated following stimulation with LPS or LTA. An immortalized adherent epithelial cell line (CALU-3) that secretes SC constitutively was used as an in vitro model to investigate whether SC expression was up regulated following stimulation with either LPS from E. coli, B. cepacia, P. aeruginosa, or LTA from S. aureus. Analytical techniques were developed to enable the analysis and quantification of SC released from CALU-3 cells. It was found that SC expression was upregulated following stimulation with LPS or LTA. Furthermore it was observed constitutively released SC was detected only after 48 hours, and did not accumulate in culture supernatants. Foetal calf serum and/ or protease inhibitor cocktail was reintroduced into the cell culture model and it was found that following this, SC was detected at the earlier time point of 24 hours and the concentration of SC detected in unstimulated culture supernatants increased as time increased. Additionally medium extracted from CALU-3 cell cultures was shown to degrade SC isolated from colostrum. Previous studies have shown that SC is degraded in the chronically inflamed lungs of CF patients and that this degraded SC no longer binds interleukin-8 (IL-8). The findings presented in this thesis could suggest that as SC is readily degraded by serine proteases invitro, the degradation of SC observed in inflamed airways in vivo could be a result of proteolytic degradation of SC and potentially this could impact SCs immunological function.