Histological Analysis and Orientation of Endothelial Cells on 'Stripped' Human Descemet's Membrane and Endothelial Sheets

Abstract

Abstract: : Purpose: Human Descemet's membrane with endothelial tissue can be successfully removed from human cornea. Endothelial counts on such material is favourable for transplantation. Such tissue may be a viable alternative to penetrating keratoplasty for corneal endothelial failure and also used for research into corneal endothelial disease. A better understanding of the histology of this tissue is required particularly for further research and orientation of endothelial cells is essential for successful transplantation. Methods: Descemet's and endothelial sheets were stained with markers including for Keratocytes and counterstained with Haematoxylin and Eosin. Descemet's and Endothelial sheets were stripped and placed into standard culture media. Over a period of 2-4 weeks after formation of a 'roll' the material was placed in 10% formalin and sectioned in wax with H&E staining. The orientation of the endothelial cells whether they were on the 'outside of the roll' or on the 'inside' was confirmed. Trypan blue vital staining was also placed directly onto the 'rolled' sheets and observed for staining. Results: Keratocyte markers suggests that there were few keratocytes in the 'stripped ' material. There was a confluent layer of endothelial cells. Descemet's membrane appears intact throughout. There was a lack of Trypan blue staining of the Descemet's endothelial sheets suggesting that the endothelium was on the 'outside of the roll'and was confirmed histologically. Conclusion: Staining confirms an intact endothelial monolayer and Descemet's membrane. There appears to be a cleavage plane when 'stripping' such tissue. This is in agreement with the known loose attachment of Descemet's membrane to the stroma. The Descemet's and endothelial sheets appear to naturally orientate themselves over time onto the 'outside surface of the roll'. The roll is probably formed by the elastic nature of Descemet's membrane. Orientation could be confirmed per operatively by staining with Trypan blue. 

Keywords: 371 cornea: endothelium • 607 transplantation • 370 cornea: basic science