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T.–Y. Chung, F.H. Casanova, T. Sakimoto, J.A. Javier, E. Albe, J.H. Chang, D.T. Azar; Expression Pattern of EphrinB Ligand and EphB Receptor and Cell–Cell Interaction in Corneal Epithelial and Keratocyte Cell Lines . Invest. Ophthalmol. Vis. Sci. 2005;46(13):2602.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: EphrinB ligands and EphB receptors are involved in blood vessel differentiation and nerve pathfinding. Our purpose was to investigate the expression pattern of ephrinB and EphB in mouse corneal epithelial and keratocyte cell lines. Methods: Excised corneas were immortalized and subcloned to generate wild type mouse corneal epithelial and keratocyte cell lines, which were characterized using immunohistochemistry. Indirect immunofluorescent assays with antibodies against ephrinB ligands (ephrinB1, B2 or B3) and EphB receptors (EphB1, B2, B3, B4 or B6) were performed on the corneal epithelial and keratocyte cell lines as well as mouse neuroblastoma (CCL–131) and endothelioma (CRL–2299) cell lines, which served as controls. Further immunolocalization of ephrinB2 and EphB4 was performed in vivo and in epithelial and keratocyte cell lines seeded at low (1×104 cells/ml) and high (5×104 cells/ml) densities. Results: Positive staining for ephrinB2, B3 and EphB1, B2, B3 and B4 was observed in corneal epithelial cells (with EphB4 showing highest levels of expression and ephrinB3, EphB1 and EphB2 showing moderate expression levels). Positive staining for ephrinB1, B2 and B3 and EphB1, B4 and B6 was observed in keratocytes (with ephrinB1 and EphB4 showing highest level of expression and ephrinB2 and EphB1 showing moderate expression levels). Neuroblastoma cell line showed intense immunolocalization of ephrinB1, B2 and EphB1 and B4. Endothelioma cell line showed intense immunolocalization of ephrinB1, EphB1 and EphB4. EphB4 and ephrinB2 showed diffuse membrane immunolocalization in keratocytes seeded at low density, while EphB4 showed focal areas of intense immunolocalization in keratocytes seeded at high density. Corneal epithelial cells seeded at low and high densities showed diffuse membrane immunolocalization of ephrinB2 and EphB4. Mouse corneal epithelium showed expression of both ephrinB2 and EphB4 (with EphB4 showing higher expression than ephrinB2). Conclusions: Certain members of the ephrinB and EphB families are exclusively expressed in corneal epithelial cells (EphB2 and EphB3) and keratocytes (ephrinB1 and EphB6). Despite similarities in the expression of other ephrinB and EphB family members in corneal epithelial cells and keratocytes (ephrinB2, B3 and EphB1 and B4), they respond differently to changes in cell seeding densities. The differences in the response to cell–cell interactions may be important in the regulation of angiogenesis in the cornea.
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