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Purpose: In the present study we have examined the effects of human vitreous and hyalocyte-conditioned medium on human umbilical vein endothelial cells (HUVEC). Methods: Human umbilical vein endothelial cells were cultured and treated in quadruplicate with human vitreous and hyalocyte-conditioned medium at different concentrations. Cell numbers were counted on days 1, 3, 5 and 7. Morphological changes and the viability of cells after treatment were also monitored. Results: The results indicate that both human vitreous and hyalocyte-conditioned medium inhibit proliferation and reduce the viability of HUVEC in vitro.These inhibitory effects were dose- and time-dependent. Conclusions: The observations suggest that human hyalo-cytes and vitreous contain anti-angiogenic factors that influence vascular endothelial cell growth. These results, combined with those of previous studies, may yield important information about the functional role of vitreous and hyalocytes in intraocular vascular regression.

Background:Human umbilical endothelial cells were used to model the vascular component of the blood–retinal barriers and to examine the capacity of glial cultures to modulate endothelial cell resistivity in vitro. Methods:Endothelial cell resistivity was monitored with and without cocultured human retinal glia. Immunohistochemistry indicated that both macroglia and microglia were present in one culture, while only macroglia were detectable in the second culture. Results:Both cocultures produced increased resistivity in the target endothelial cells; however a further significant increase in resistivity was noted with the glial coculture containing microglia. The results suggest that the presence of microglia significantly increases the capacity of astrocytes and Müller cells to modulate endothelial cell resistivity.

Background:Current methods for the isolation and culture of adult retinal pigment epithelium (RPE) provide successful primary culture. However, most methods result in contamination with other cell types and low cell yields. The isolation and culture of human foetal RPE presents further problems associated with the limited size of the eye cup and adherence among cells. Reliable methods are necessary for the culture of human RPE and subsequent functional studies. Methods:The present procedure is based on mechanical peeling of the whole RPE layer under the dissecting microscope. Dissected pieces are subsequently explanted to a 35 mm culture dish and are cultured with Dulbecco's modified Eagle's medium containing 10% foetal bovine serum. Peroxidase immunohistochemical methods were used to investigate cell phenotypes. Results:Primary cultures were obtained within 10–14 days with high yields, good viability and purity in subsequent culture. Cultured cells were vimentin and cytokeratin positive and CD31 negative. Conclusions:This mechanical dissection technique is recommended for the isolation of foetal and young adult RPE cells, while the enzyme digestion method is preferred for aged adult tissue.