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Xian-jin Yi, Yuan Wang, Fu-Shin X. Yu; Corneal Epithelial Tight Junctions and Their Response to Lipopolysaccharide Challenge. Invest. Ophthalmol. Vis. Sci. 2000;41(13):4093-4100.
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purpose. To investigate the expression and cellular distribution of putative
tight junction (TJ) proteins occludin, ZO-1, ZO-2, and claudin-1 in rat
corneal epithelium and alterations of TJs in cultured human corneal
epithelial cells in response to lipopolysaccharide (LPS) challenge.
methods. Immunohistochemistry was used to determine tissue distribution of
occludin, ZO-1, ZO-2, and claudin-1 in the rat cornea. Reverse
transcription–polymerase chain reaction was used to reveal the
expression of mRNAs for claudins in simian virus (SV)40-immortalized
human corneal epithelial (THCE) cells. To assess epithelial
response to LPS challenge, THCE cells were cultured on the upper
chamber of Transwell filters (Costar, Cambridge, MA), transepithelial
electrical resistance (TER) was measured using a voltohmmeter.
Immunocytochemistry and immunoblotting were used to assess alteration
in the levels and localization of TJ-associated proteins occludin,
ZO-1, and ZO-2 in LPS-treated THCE cells.
results. Occludin, ZO-1, and ZO-2 were found at the cell borders of the
superficial layer, whereas claudin-1 was localized mainly in the basal
and wing cell layers of rat corneal epithelium. In addition to
claudin-1, the transcripts for several other isotypes of claudins-2,
-3, -7, -9, -14, and -15 were identified in THCE cells. Treatment of
cultured THCE cells with LPS caused a dose- and time-dependent increase
in monolayer permeability as assessed by TER measurements. The maximal
decrease of TER was observed at approximately 6 to 9 hours after LPS
challenge. The TER was then recovered gradually and returned to
baseline after 24 hours. Examination of specific proteins associated
with TJs by immunoblot analysis and immunomicroscopy revealed changes
in the expression levels and localization of some of these proteins
after their exposure to LPS. Specifically, LPS challenge resulted in a
decrease in the levels of ZO-1 and ZO-2 compared with untreated cells.
Reduction of the ZO-2 level was associated with the disappearance of
ZO-2 staining from cell borders in 6-hour LPS-treated cells.
conclusions. Occludin, ZO-1, and ZO-2, but not claudin-1, are components of corneal
epithelial TJs. LPS induces breakdown of the epithelial barrier through
disruption of TJs, and ZO-1 and ZO-2 are targets for the
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