Abstract
Acidic airway microenvironment is one of the representative pathophysiological features of chronic inflammatory respiratory diseases. Epithelial barrier function is maintained by TJs (tight junctions), which act as the first physical barrier against the inhaled substances and pathogens of airway. As previous studies described, acid stress caused impaired epithelial barriers and led the hyperpermeability of epithelium. However, the specific mechanism is still unclear. We have showed previously the existence of TRPV (transient receptor potential vanilloid) 1 channel in airway epithelium, as well as its activation by acidic stress in 16HBE cells. In this study, we explored the acidic stress on airway barrier function and TJ proteins in vitro with 16HBE cell lines. Airway epithelial barrier function was determined by measuring by TER (trans-epithelial electrical resistance). TJ-related protein [claudin-1, claudin-3, claudin-4, claudin-5, claudin-7 and ZO-1 (zonula occluden 1)] expression was examined by western blotting of insoluble fractions of cell extraction. The localization of TJ proteins were visualized by immunofluorescent staining. Interestingly, stimulation by pH 6.0 for 8 h slightly increased the epithelial resistance in 16HBE cells insignificantly. However, higher concentration of hydrochloric acid (lower than pH 5.0) did reduce the airway epithelial TER of 16HBE cells. The decline of epithelial barrier function induced by acidic stress exhibited a TRPV1-[Ca2+]i-dependent pathway. Of the TJ proteins, claudin-3 and claudin-4 seemed to be sensitive to acidic stress. The degradation of claudin-3 and claudin-4 induced by acidic stress could be attenuated by the specific TRPV1 blocker or intracellular Ca2+ chelator BAPTA/AM [1,2-bis-(o-aminophenoxy)ethane-N,N,N',N'-tetra-acetic acid tetrakis(acetoxymethyl ester)].
Highlights
The airway epithelium of human respiratory mucosa acts as the first physical barrier against the inhaled substances and pathogens [1,2]
Effect of acidification on airway epithelial barrier function (A) 16HBE cells exposed to pH 7.4 culture medium for 8 h were set as negative control
Values are mean +− S.D.; n = 6. *P < 0.05. #P > 0.05. (B) Fold changes of trans-epithelial electrical resistance (TER) values in 16HBE cells after acidic stress. 16HBE cells after exposure to pH 7.4 culture medium for 8 h were set as base line
Summary
The airway epithelium of human respiratory mucosa acts as the first physical barrier against the inhaled substances and pathogens [1,2]. The epithelial barrier function is highly related with the integrity of TJs (tight junctions). As termed ‘barrier function’, epithelial TJs, most apically located of the intercellular junctional complexes, inhibit solute, water and inflammatory factors through the paracellular space [3,4]. TJs separate the apical domain from basolateral and maintain the cell polarity in human airway epithelium [5]. Claudins are considered to be the most important components of the TJs at the interface of the basolateral and apical membranes of polarized epithelial cells. They determine the barrier properties of the cell–cell contact between two neighbouring epithelial cells and regulate the paracellular permeability.
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