Abstract

DO.02.11

Phenotypic characterization of corneal epithelium after transplantation of cultivated limbal epithelium in patients with limbal stem cell deficiency

Pauklin M., Brockmann-Ahmed S., Steuhl K.-P., Meller D.
Zentrum für Augenheilkunde, Universitätsklinikum Essen

Objective: Limbal stem cell deficiency (LSCD) is characterized by impaired visus and ingrowths of an inflamed fibrovascular tissue (pannus) with a conjunctival phenotype into cornea. Good clinical results have been achieved by transplantation of in vitro cultivated limbal epithelium although the reconstructed corneal surface has hardly ever been characterized. Our aim was to characterize the reconstructed cornea of LSCD patients after transplantation of in vitro cultivated limbal epithelium.
Methods: Four patients with total LSCD were treated by transplantation of limbal epithelium cultured on intact AM. Three of these patients had suffered a chemical burn and one a perforating injury. An additional penetrating keratoplasty was performed due to deep stromal scarring at an average interval of 8.5 months after limbal epithelium transplantation. The expression of epithelial lineage markers (keratin 3 (K3), K12, K19, MUC5AC), inflammatory and vascularisation markers (IL-1a, IL-1b, ICAM-1, VCAM-1 and VEGF) was analyzed by Real-Time PCR, Western blotting and immunofluorescence in obtained corneal buttons. The expression was compared to normal corneal, conjunctival and pathological pannus samples.
Results: Three of the analyzed four cases were graded clinically as a success and one as a partially successful procedure due to focal ingrowths of superficial fine blood vessels. Nevertheless, a stable epithelium consisting of 5-7 cell layers was reconstructed in all cases. The expression of lineage markers K3/12 and K19 and the expression of all inflammatory markers except VCAM-1 were similar to healthy cornea in the successful cases. The inflammatory markers were reduced in comparison to pannus tissue. Goblet cells were found only in the peripheral cornea of the partially successful case. This sample showed an overall higher expression of inflammatory markers than healthy cornea.
Conclusions: A normal corneal phenotype and a reduction of inflammatory markers were achieved in most cases by transplantation of limbal epithelium cultured on intact AM.

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