Developmental Changes in Patterns of Distribution of Fibronectin and Tenascin-C in the Chicken Cornea: Evidence for Distinct and Independent Functions during Corneal Development and Morphogenesis

Abstract

The cornea forms the tough and transparent anterior part of the eye and by accurate shaping forms the major refractive element for vision. Its largest component is the stroma, a dense collagenous connective tissue positioned between the epithelium and the endothelium. In chicken embryos, the stroma initially develops as the primary stroma secreted by the epithelium, which is then invaded by migratory neural crest cells. These cells secrete an organised multi-lamellar collagenous extracellular matrix (ECM), becoming keratocytes. Within individual lamellae, collagen fibrils are parallel and orientated approximately orthogonally in adjacent lamellae. In addition to collagens and associated small proteoglycans, the ECM contains the multifunctional adhesive glycoproteins fibronectin and tenascin-C. We show in embryonic chicken corneas that fibronectin is present but is essentially unstructured in the primary stroma before cell migration and develops as strands linking migrating cells as they enter, maintaining their relative positions as they populate the stroma. Fibronectin also becomes prominent in the epithelial basement membrane, from which fibronectin strings penetrate into the stromal lamellar ECM at right angles. These are present throughout embryonic development but are absent in adults. Stromal cells associate with the strings. Since the epithelial basement membrane is the anterior stromal boundary, strings may be used by stromal cells to determine their relative anterior–posterior positions. Tenascin-C is organised differently, initially as an amorphous layer above the endothelium and subsequently extending anteriorly and organising into a 3D mesh when the stromal cells arrive, enclosing them. It continues to shift anteriorly in development, disappearing posteriorly, and finally becoming prominent in Bowman’s layer beneath the epithelium. The similarity of tenascin-C and collagen organisation suggests that it may link cells to collagen, allowing cells to control and organise the developing ECM architecture. Fibronectin and tenascin-C have complementary roles in cell migration, with the former being adhesive and the latter being antiadhesive and able to displace cells from their adhesion to fibronectin. Thus, in addition to the potential for associations between cells and the ECM, the two could be involved in controlling migration and adhesion and subsequent keratocyte differentiation. Despite the similarities in structure and binding capabilities of the two glycoproteins and the fact that they occupy similar regions of the developing stroma, there is little colocalisation, demonstrating their distinctive roles.