Human trabecular endothelium, corneal endothelium, keratocytes, and scleral fibroblasts in primary cell culture. A comparative study of growth characteristics, morphology, and phagocytic activity by light and scanning electron microscopy

Abstract

To obtain baseline data for our experimental investigations in vitro we established a pure cellline culture model of human trabecular endothelium, corneal endothelium, keratocytes, and scleral fibroblasts, using an identical environment for each cell type. We studied the growth characteristics, morphology and phagocytic activity of live cultures by phase-contrast microscopy and time-lapse cinephotomicrography; fixed preparations were examined by light and scanning electron microscopy. From these observations, we were able to characterize the distinctive features of the cells. By correlated ultrastructural and tissue culture studies, we showed that human cadaver eyes cold-stored (4°C) up to 5 days can be used as a tissue source of trabecular endothelium, keratocytes and scleral fibroblasts. Corneal endothelium, however, rarely survived after 4 days of postmortem storage. Our observations revealed that, in primary confluent culture, the trabecular cells grew in a monolayer. Individual cells had flattened, elongated cell bodies, a centrally located, oval nucleus, and a generally smooth cell surface with occasional microvillous projections; in places, adjacent cells and their processes adhered closely one to another. Corneal endothelium at confluency presented a flattened hexagonal profile of cells arranged in a honeycomb pattern, interdigitating cell borders, a centrally located nucleus, prominent perinuclear microvillous projections, and numerous surface invaginations. The keratocytes and scleral fibroblasts had a thin and elongated, spindle-shaped profile with a remarkably smooth cell surface and an oval to oblong nucleus surrounded laterally by scanty cytoplasm; the cell surfaces and processes frequently overlapped but rarely formed firm attachments. In contrast to the monolayer growth of trabecular and corneal endothelium, these cells characteristically grew in superimposed layers. In primary culture and with latex spheres and carmine particles used as markers, the trabecularendothelial cells showed a greater phagocytic capability than did corneal endothelium, keratocytes and scleral fibroblasts. Our preliminary studies indicate that cytochalasin B markedly inhibits the phagocytic activity of the trabecular cells.