The lack of extracellular laminin ß2 chain deposition correlates to the loss of conjunctival epithelial keratin K4 localization in culture

Abstract

Purpose. To examine the effects of external modulation of epithelial-mesenchymal interaction on conjunctival epithelial cell differentiation characteristics. Methods. Keratin K4 and laminin ß2 chain protein localization was examined in an organotypic model which facilitates the comparison of differentiation characteristics of conjunctival epithelium interacting with conjunctival basement membrane or corneal basement membrane. In addition, keratin K4 and laminin ß2 chain localization was examined in primary cultures of conjunctival epithelial cells and fibroblasts. The synthesis and secretion of laminin ß2 chain by conjunctival fibroblasts in culture was determined by western blot analysis and immunoprecipitation. The ability of conjunctival epithelium to respond to exogenous laminin ß2 chain was assayed by culturing epithelial cells on a laminin matrix isolated from human placenta. Results. In culture, conjunctival fibroblasts synthesize and secrete laminin ß2 chain but do not deposit this chain into an extracellular matrix substrate or basement membrane-like structure. The lack of extracellular deposition of this chain correlates to the gradual loss of keratin K4 protein in conjunctival epithelial cell culture. Conjunctival epithelium remains responsive to laminin ß2 chain in vitro because keratin K4 localization can be rescued in these cells by culture on a substrate of exogenous placental laminin. Conclusions. In vitro, alterations in native conjunctival epithelial-mesenchymal interactions results in aberrant basement membrane laminin isoform composition. This, in turn, leads to the loss of adult epithelial cell phenotype characteristics, suggesting that at least some aspects of conjunctival epithelial cell differentiation are regulated by the extracellular matrix.