Flagellin potentiates the Ca2+-dependent electrolytes secretion from airway submucosal gland in a TLR4-independent, TLR5-dependent and cGK-dependent manner

[Background] Airway surface fluids are mainly secreted from submucosal glands and play important roles in the airway defense via the upregulation of mucociliary transport. Although Toll-like receptors (TLRs) recognize distinct pathogen-associated molecular patterns, it is not well known whether TLRs directly upregulates the airway secretions. Recently, we have reported that TLR4 works as one of important secretagogue in the airways (AJRCMB 2011). In the present study, we focused on the TLR5 signaling as other candidate for a potentiator of airway secretion. [Methods] Freshly isolated swine tracheal submucosal gland cells were prepared and measured their secretory activities as ionic currents by applying a patch-clamp technique. The expression of TLR5 was estimated by both the immunofluorescent staining and RT-PCR. The intracellular mechanisms were investigated by applying some inhibitors of NOS or cGMP-dependent protein kinase (cGK). [Results] Flagellin, a TLR5 ligand, showed significant potentiating effects on ACh-evoked ionic currents. These effects were abolished by the pretreatment with anti-TLR5 antibody, but not by specific TLR4 antagonist. Two different inhibitors of each NOS and cGK abolished these effects. We also revealed that the expression of TLR5 on the tracheal submucosal gland cells. [Conclusion] Our study showed that flagellin caused the potentiating effects on airway secretions in a TLR5-dependent, TLR4-independent manner and that NO/cGMP/cGK pathway was involved in these intracellular mechanisms. These finding suggest that TLR5 also works as a potentiator of airway secretion in the airway mucosal immune systems.