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The degradation of airway tight junction protein under acidic conditions is probably mediated by transient receptor potential vanilloid 1 receptor.

Xu R, Li Q, Zhou J, Zhou XD, Perelman JM, Kolosov VP - Biosci. Rep. (2013)

Bottom Line: 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 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)].

View Article: PubMed Central - PubMed

Affiliation: *Department of Respiratory Medicine, the Second Affiliated Hospital, Chongqing Medical University, Chongqing 400010, China.

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)].

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MTT assay for cell viability16HBE cells propagated in pH=7.4 culture medium were set as negative controls. 16HBE cells were exposed to different concentrations of hydrochloric acid (pH=7.4, 7.0, 6.0, 5.0, 4.0) and for different exposure durations (4, 8, 12 h). Data were represented as mean ± S.D.; n=6, *P<0.05 versus control.
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Figure 1: MTT assay for cell viability16HBE cells propagated in pH=7.4 culture medium were set as negative controls. 16HBE cells were exposed to different concentrations of hydrochloric acid (pH=7.4, 7.0, 6.0, 5.0, 4.0) and for different exposure durations (4, 8, 12 h). Data were represented as mean ± S.D.; n=6, *P<0.05 versus control.

Mentions: To evaluate the cell viability after exposure to different acidic culture medium, conventional MTT assay was performed to evaluate influence of cell viability at low pH. The result demonstrated a significant reduction of cell viability at pH=5.0 and 4.0 for 12 h. Thus, 8 h was selected as the appropriate exposure time (Figure 1).


The degradation of airway tight junction protein under acidic conditions is probably mediated by transient receptor potential vanilloid 1 receptor.

Xu R, Li Q, Zhou J, Zhou XD, Perelman JM, Kolosov VP - Biosci. Rep. (2013)

MTT assay for cell viability16HBE cells propagated in pH=7.4 culture medium were set as negative controls. 16HBE cells were exposed to different concentrations of hydrochloric acid (pH=7.4, 7.0, 6.0, 5.0, 4.0) and for different exposure durations (4, 8, 12 h). Data were represented as mean ± S.D.; n=6, *P<0.05 versus control.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3814059&req=5

Figure 1: MTT assay for cell viability16HBE cells propagated in pH=7.4 culture medium were set as negative controls. 16HBE cells were exposed to different concentrations of hydrochloric acid (pH=7.4, 7.0, 6.0, 5.0, 4.0) and for different exposure durations (4, 8, 12 h). Data were represented as mean ± S.D.; n=6, *P<0.05 versus control.
Mentions: To evaluate the cell viability after exposure to different acidic culture medium, conventional MTT assay was performed to evaluate influence of cell viability at low pH. The result demonstrated a significant reduction of cell viability at pH=5.0 and 4.0 for 12 h. Thus, 8 h was selected as the appropriate exposure time (Figure 1).

Bottom Line: 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 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)].

View Article: PubMed Central - PubMed

Affiliation: *Department of Respiratory Medicine, the Second Affiliated Hospital, Chongqing Medical University, Chongqing 400010, China.

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)].

Show MeSH
Related in: MedlinePlus