Ex vivo expansion of human labial mucosal epithelium for corneal surface reconstruction.

Purpose: To achieve ex vivo expansion of human labial mucosal epithelium using the explant method and simplified formulation of the culture media. Methods: Labial mucosa was obtained from six patients during conjunctival fornix reconstruction after signing informed contest. The substantia propria was severed, and the remaining tissue was into 1 × 1 mm explants. Explants were cultured in EpiLife (0.06 mM Ca) and DMEM/F12 (1:1) (1.05 mM Ca) media supplemented with 5% fetal bovine serum, hydrocortisone 5 µg/mL, human recombinant epidermal growth factor (hrEGF) 10 ng/ml, insulin 5 µg/ml, and standard antifungal‐antibiotic. Besides routine monitoring, primary cells were double stained with anti‐p75/p63, anti‐integrin b1/vimentin, and anti‐ZO‐1/CK10 antibodies. Results: Explant outgrowths had cobblestone epithelial phenotype in all samples in both media; however, cell growths were delayed in the low‐Ca medium. In the high‐Ca medium, all the cultures had more unfastened cells and debris. Integrin b1, CK10, and p75 had no expression in all the primary cultures. p63 expression was registered in 39.2% and 34.7% (median, n = 3, ns) cells for EpiLife and DMEM/F12 media respectively. Expression of vimentin was found as very low in both media. ZO‐1 expression was calculated as 5.18 and 17.5 mkm (median, n = 3, ns) length per cells for EpiLife and media respectively. Conclusions: It was confirmed that human labial mucosal epithelium could be cultured from a small biopsy without feeder cells in the culture media contained only three specific growth factors. Better cell morphology and lowered differentiation status were obtained in the 0.06Ca medium. Thus it was considered that the cell source, culture technique, and media formulations are suitable for autologous transplantation of cultured labial mucosal epithelium in patients with limbal stem cell deficiency. [ABSTRACT FROM AUTHOR]