Vitronectin is present in epithelial cells of the intact lens and promotes epithelial mesenchymal transition in lens epithelial explants.

PURPOSE: Extracellular matrix (ECM) accumulates during the development of posterior capsule opacification (PCO). Vitronectin, an ECM component that is generally prominent in wound healing, has been detected in PCO specimens. Here we set out to investigate the distribution of vitronectin in the lens and determine how it, and other ECM components, influence the lens epithelial phenotype. METHODS: Rat lens epithelial explants were cultured on vitronectin, fibronectin, and laminin substrata. Explants were monitored for cell migration and the appearance of markers for epithelial mesenchymal transition (EMT), using phase contrast microscopy and immunohistochemistry, respectively. Explants were also monitored for evidence of Smad signaling. Vitronectin expression was analyzed in embryonic and postnatal rodent lens development by immunohistochemistry, western blotting, and in situ hybridization. RESULTS: Vitronectin, like fibronectin and laminin, provided a good substratum for cellular attachment and migration. However, in the case of vitronectin and fibronectin, this was accompanied by a major phenotypic change. On either vitronectin or fibronectin, but not laminin, most of the cells became elongated, spindle-shaped and were strongly reactive for filamentous alpha-smooth muscle actin. In these respects this transition was typical of the well known TGFbeta-induced EMT. In explants cultured on vitronectin and fibronectin, but not laminin, cell nuclei showed prominent reactivity for Smad 2/3. Vitronectin was also shown to be expressed during embryonic and postnatal development. Initially mRNA and protein were detected in all lens cells, however as development progressed, expression became restricted to cells of the epithelium and transition zone. CONCLUSIONS: The results clearly show that lens cell engagement with a vitronectin or a fibronectin, but not laminin, substratum has a potent EMT promoting effect and that Smad 2/3 signaling is involved. Thus when considering strategies to slow or prevent PCO, these results highlight the need to take into account ECM molecules such as vitronectin that have the capacity to promote EMT.

Nance-Horan syndrome protein, NHS, associates with epithelial cell junctions.

Nance-Horan syndrome, characterized by congenital cataracts, craniofacial, dental abnormalities and mental disturbances, is an X-linked disorder with significant phenotypic heterogeneity. Affected individuals have mutations in the NHS (Nance-Horan syndrome) gene typically resulting in premature truncation of the protein. This report underlines the complexity of the regulation of the NHS gene that transcribes several isoforms. We demonstrate the differential expression of the two NHS isoforms, NHS-A and NHS-1A, and differences in the subcellular localization of the proteins encoded by these isoforms. This may in part explain the pleiotropic features of the syndrome. We show that the endogenous and exogenous NHS-A isoform localizes to the cell membrane of mammalian cells in a cell-type-dependent manner and that it co-localizes with the tight junction (TJ) protein ZO-1 in the apical aspect of cell membrane in epithelial cells. We also show that the NHS-1A isoform is a cytoplasmic protein. In the developing mammalian lens, we found continuous expression of NHS that became restricted to the lens epithelium in pre- and postnatal lens. Consistent with the in vitro findings, the NHS-A isoform associates with the apical cell membrane in the lens epithelium. This study suggests that disturbances in intercellular contacts underlie cataractogenesis in the Nance-Horan syndrome. NHS is the first gene localized at TJs that has been implicated in congenital cataracts.

A role for Wnt/beta-catenin signaling in lens epithelial differentiation.

The differentiation of epithelial cells and fiber cells from the anterior and posterior compartments of the lens vesicle, respectively, give the mammalian lens its distinctive polarity. While much progress has been made in understanding the molecular basis of fiber differentiation, little is known about factors that govern the differentiation of the epithelium. Members of the Wnt growth factor family appear to be key regulators of epithelial differentiation in various organ systems. Wnts are ligands for Frizzled receptors and can activate several signaling pathways, of which the best understood is the Wnt/beta-catenin pathway. The presence of LDL-related protein coreceptors (LRPs) 5 or 6 has been shown to be a requirement for Wnt signaling through the beta-catenin pathway. To access the role of this signaling pathway in the lens, we analyzed mice with a null mutation of lrp6. These mice had small eyes and aberrant lenses, characterized by an incompletely formed anterior epithelium resulting in extrusion of the lens fibers into the overlying corneal stroma. We also showed that multiple Wnts, including 5a, 5b, 7a, 7b, 8a, 8b, and Frizzled receptors 1, 2, 3, 4, and 6, were detected in the lens. Expression of these molecules was generally present throughout the lens epithelium and extended into the transitional zone, where early fiber elongation occurs. In addition to both LRP5 and LRP6, we also showed the expression of other molecules involved in Wnt signaling and its regulation, including Dishevelleds, Dickkopfs, and secreted Frizzled-related proteins. Taken together, these results indicate a role for Wnt signaling in regulating the differentiation and behavior of lens cells.