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S. M. Sharma, T. Fuchsluger, R. Dana, U. V. Jurkunas; Comparison of Different Feeder Layers for ex vivo Cultivation of Oral Epithelium for Corneal Surface Reconstruction. Invest. Ophthalmol. Vis. Sci. 2009;50(13):6177.
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© ARVO (1962-2015); The Authors (2016-present)
To investigate the propagation of ex vivo cultivated oral and corneal epithelial cells on amniotic membrane with and without different feeder layers, and to compare the cell sheet characteristics between these different culture conditions.
Oral and limbal epithelial cells were isolated by microdissection followed by digestion with dispase and trypsin/EDTA. The cells were seeded on intact amniotic membrane and co-cultured with (1) mouse 3T3 fibroblast (2) human dermal fibroblast (3) human bone marrow fibroblast feeder layers, and (4) with no feeder layer, for three weeks. The cultivated sheets were characterized by immunofluorescence with epithelial, stem cell, and cell junction markers.
The cultivated sheets from corneal and oral mucosa specimens formed stratified epithelium. There were no discernible morphological differences between cells grown with and without feeder layers for both oral and corneal cells. Immunohistochemistry showed positive staining for cytokeratin-3, the differentiation marker for corneal epithelium, and integrin- ß 1, putative stem cell marker, in all epithelial cells, including the ones grown without a feeder layer. Figure 1 shows immunoflourescent staining of integrin ß-1 (green), with propidium iodide nuclear stain (red) in cultured human oral epithelial cells grown with different feeder cells or without feeder cells.
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