Effects of Environmental Oxygen Concentration on Paracellular Permeability and the Expression and Localization of Zonula Occludens–1 in Human Corneal Epithelial Cells

Abstract

Abstract: : Purpose: Atmospheric oxygen plays a role in maintenance of the structure and function of the cornea as a result of the anatomic location of this tissue. The effect of oxygen concentration on the barrier function of corneal epithelial cells was examined. Methods: Simian virus 40–transformed human corneal epithelial (HCE) cells were cultured for 4 days to allow them to form an epithelial barrier. The cells were then deprived of serum and subsequently incubated for 24 h in the presence of various concentrations (1, 21, or 60%) of oxygen gas. The barrier function of the cells was monitored by determination of transepithelial resistance. HCE cells incubated in the presence of 1, 21, or 60% oxygen were also either fixed and subjected to immunofluorescence staining with antibodies to zonula occludens–1 (ZO–1), a component of tight junctions, or lysed and subjected to immunoblot analysis with antibodies to this protein. Results: Transepithelial resistance decreased during incubation of HCE cells under the hypoxic condition (1% oxygen) but was stable in cells cultured under normoxic (21%) or hyperoxic (60%) conditions. Immunofluorescence analysis revealed that the abundance and localization of ZO–1 at tight junctions decreased in response to exposure of cells to hypoxia. Immunoblot analysis showed that the amount of ZO–1 was reduced in cells after culture with 1% oxygen for 24 or 48 h, and was increased in cells cultured with 60% oxygen for 48 h, compared with that apparent in cells exposed to 21% oxygen. Conclusions: Hypoxia impaired the barrier function of corneal epithelial cells, likely as a result of down–regulation and redistribution of the tight–junction protein ZO–1.

Keywords: cornea: endothelium • hypoxia • cell-cell communication