Alginate exopolysaccharide acts as a ligand for P.aeruginosa invasion of lung epithelial cells through its interaction with p63
Abstract number: P1749
Background: The interplay of P. aeruginosa with the airway epithelial cells is a crucial step in the pathogenesis of the respiratory infections caused by this microorganism. Both, bacterial and host factors modulate this interaction. In this work, we investigated the role of the alginate exopolysaccharide, a key virulence factor associated to severe respiratory infections, and the surfactant protein A (SP-A), a broad spectrum opsonin with membrane permeabilisation properties in this interaction.
Methods: Bacterial invasion capacity of human bronchoepithelial cells (16HBEo-) and human type II pneumocytes (A549) monolayers was determined by standard invasion assays in presence of SP-A with the type strain PAO1, an isogenic alginate hyperproducing mucA mutant (PAOMA), an isogenic alginate deficient algD mutant and 2 clinical isolates (PAA2 and PAC20). The role of p63 in P. aeruginosa internalization by respiratory epithelial cells was studied using standard invasion assays in presence of a p63 blocking antibody or using p63 silenced cells by siRNA. Alginate binding assays were performed measuring purified P. aeruginosa FITC-labelled alginate union to A549 monolayers by fluorescence spectroscopy.
Results: Opsonisation of P. aeruginosa with SP-A did not affect the invasion of the airway epithelial cells by the microorganism. However, SP-A inhibited P. aeruginosa internalization by airway epithelial cells through direct interaction of the protein with the cells, suggesting that the microorganism attached to the cells through a specific receptor for SP-A such as p63. Specific anti p63 antibodies blocked the invasion of epithelial cells by alginate-producing P. aeruginosa strains. Furthermore, the invasive capacity of these strains was reduced in p63 silenced cells. Finally, the binding of purified alginate to p63 silenced cells or cells pretreated with p63 blocking antibodies was significantly reduced.
Conclusions: Altogether these results indicate that P. aeruginosa and SP-A share a common receptor, p63, on the surface of respiratory epithelial cells. Alginate exopolysaccharide appears to be involved in the interaction of the bacterium with this receptor and could play an important role during the process of infection by promoting bacterial internalization.
|Session name:||Abstracts of 21st ECCMID / 27th ICC|
|Location:||Milan, Italy, 7 - 10 May 2011|
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