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(Investigative Ophthalmology and Visual Science. 2008;49:113-119.)
© 2008 by The Association for Research in Vision and Ophthalmology, Inc.
DOI:  10.1167/iovs.07-0353
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Upregulation of Tight-Junctional Proteins in Corneal Epithelial Cells by Corneal Fibroblasts in Collagen Vitrigel Cultures

Ji-Ae Ko,1 Yang Liu,1 Ryoji Yanai,1 Tai-ichiro Chikama,1,2 Toshiaki Takezawa,3 and Teruo Nishida1,2

1From the Departments of Ophthalmology and 2Ocular Pathophysiology, Yamaguchi University Graduate School of Medicine, Ube City, Yamaguchi, Japan; and the 3Transgenic Animal Research Center, National Institute of Agrobiological Sciences, Tsukuba City, Ibaraki, Japan.

PURPOSE. To investigate the effects of corneal fibroblasts on the differentiation of corneal epithelial cells in a coculture system based on a collagen vitrigel membrane.

METHODS. Simian virus 40–transformed human corneal epithelial (HCE) cells and human corneal fibroblasts were cultured on opposite sides of a collagen vitrigel membrane. The distribution of HCE cells and corneal fibroblasts on the collagen membrane was determined by immunofluorescence staining and immunoblot analysis of marker proteins. Expression of the tight-junctional proteins ZO-1, occludin, and claudin and of the adherens-junctional proteins E- and N-cadherin in HCE cells was determined at the mRNA and protein levels by reverse transcription–polymerase chain reaction analysis and immunoblot analysis, respectively.

RESULTS. The abundance of ZO-1, occludin, and claudin mRNA and proteins in HCE cells was markedly increased by coculture with corneal fibroblasts. The expression of E- or N-cadherin did not differ between HCE cells cultured with corneal fibroblasts and those cultured without them. PD98059, a specific inhibitor of signaling by extracellular signal regulated kinase (ERK), prevented the upregulation of tight-junctional proteins in HCE cells by corneal fibroblasts.

CONCLUSIONS. Human corneal fibroblasts regulated the expression of tight-junctional proteins in HCE cells, suggesting that corneal fibroblasts may play an important role in the differentiation of corneal epithelial cells.






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