Cell-Cell and Cell-Matrix Interactions at the Presumptive Stem Cell Niche of the Chick Corneal Limbus.

Abstract

(1) Background: Owing to its ready availability and ease of acquisition, developing chick corneal tissue has long been used for research purposes. Here, we seek to ascertain the three-dimensional microanatomy and spatiotemporal interrelationships of the cells (epithelial and stromal), extracellular matrix, and vasculature at the corneo-scleral limbus as the site of the corneal stem cell niche of the chicken eye. (2) Methods: The limbus of developing (i.e., embryonic days (E) 16 and 18, just prior to hatch) and mature chicken eyes was imaged using scanning electron microscopy (SEM), transmission electron microscopy (TEM), and the volume electron microscopy technique, serial-block face SEM (SBF-SEM), the latter technique allowing us to generate three-dimensional reconstructions from data sets of up to 1000 serial images; (3) Results: Data revealed that miniature limbal undulations of the embryonic basement membrane, akin to Palisades of Vogt (PoV), matured into distinct invaginations of epithelial cells that extended proximally into a vascularized limbal stroma. Basal limbal epithelial cells, moreover, occasionally exhibited a high nuclear:cytoplasmic ratio, which is a characteristic feature of stem cells. SBF-SEM identified direct cell-cell associations between corneal epithelial and stromal cells at the base of structures akin to limbal crypts (LCs), with cord-like projections of extracellular matrix extending from the basal epithelial lamina into the subjacent stroma, where they made direct contact with stomal cells in the immature limbus. (4) Conclusion: Similarities with human tissue suggest that the corneal limbus of the mature chicken eye is likely the site of a corneal stem cell niche. The ability to study embryonic corneas pre-hatch, where we see characteristic niche-like features emerge, thus provides an opportunity to chart the development of the limbal stem cell niche of the cornea.