Current Eye Research, Early Online, 1–11, 2014 ! Informa Healthcare USA, Inc. ISSN: 0271-3683 print / 1460-2202 online DOI: 10.3109/02713683.2014.978477 RESEARCH REPORT Culture and Characterization of Oral Mucosal Epithelial Cells on a Fibrin Gel for Ocular Surface Reconstruction Radhika Sheth 1,2 , Michael H. Neale 1,2 , Alex J. Shortt 1,2,3 , Isobel Massie 2 , Amanda J. Vernon 1,2 and Julie T. Daniels 1,2 1 Cells for Sight, Transplantation and Research Programme, University College London Institute of Ophthalmology, London, UK, 2 Ocular Biology and Therapeutics Division, University College London Institute of Ophthalmology, London, UK, and 3 Moorfields Eye Hospital National Health Service Foundation Trust, London, UK ABSTRACT Aim of the study: To develop a clinical grade fibrin gel for the culture of oral mucosal epithelial cells (OMEC) intended for ocular surface reconstruction in the treatment of limbal stem cell deficiency (LSCD). Materials and methods: Transparent fibrin gels composed of fibrinogen and thrombin were developed for the culture of epithelial cells. Oral mucosa was harvested from the buccal region of healthy volunteers and cultured as explants on fibrin gels. Tranexamic acid (TA), a clinically approved anti-fibrinolytic agent was added to prevent the fibrin gel from digesting due to cellular activity. The gels were stained for p63a (as a marker of poorly differentiated epithelial cells), CK19, CK13 and CK3 (expressed by OMEC). Epithelial cell stratification was observed using hematoxylin–eosin staining. Results: Addition of TA prevented gels from dissolving during the culture period. OMEC proliferated on the fibrin gel and attained confluence over a 2-week period (±2 d) and exhibited a typical epithelial, cobblestone morphology. Basal OMEC exhibited positive staining for p63a while the superficial cells exhibited positive staining for CK3. The cells expressed a strong immunoreactivity for CK19 and CK13 suggesting that they retained a normal oral epithelial phenotype. Conclusion: Fibrin gels, maintained in the presence of TA, to control the rate of substrate degradation, provide a more robust yet transparent substrate for the culture and transplantation of cultured OMEC. The fibrin gels are easily standardized, the components commercially available, and produced from clinically approved materials. The resulting stratified OMEC-derived epithelium displays characteristics similar to that of a human cornea, e.g. CK3 expression. The conventional dependence on a murine feeder layer for support of epithelial cells is unnecessary with this technique and hence, provides for an attractive alternative for treatment of LSCD. Keywords: Fibrin gels, limbal stem cell deficiency, ocular surface reconstruction, oral mucosal epithelial cells, tranexamic acid INTRODUCTION The cornea is the clear tissue structure at the front of the eye. Corneal transparency and integrity are fundamental to maintaining vision. The corneal epi- thelium is separated from the bulbar conjunctiva by the limbus at the periphery of the cornea. It represents 10% of the thickness of the cornea and is composed of proliferating and terminally differentiated cells form- ing a non-keratinized, stratified squamous tissue. Limbal epithelial stem cells (LESC) are unipotent stem cells (SC) located in the basal layer of the limbus Correspondence: Radhika Sheth, Cells for Sight, Transplantation and Research Programme, University College London Institute of Ophthalmology, London, UK. Tel: 00 44 207 6086 996. Fax: 00 442 076 086 887. E-mail: radhika.sheth@ucl.ac.uk Received 28 March 2014; revised 1 October 2014; accepted 12 October 2014; published online 6 November 2014 1 Curr Eye Res Downloaded from informahealthcare.com by University College London on 11/13/14 For personal use only.