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Laboratory science
Comparison of preservation and transportation protocols for preloaded Descemet membrane endothelial keratoplasty
  1. Vito Romano1,2,
  2. Mohit Parekh3,
  3. Alessandro Ruzza3,
  4. Colin E Willoughby1,4,
  5. Stefano Ferrari3,
  6. Diego Ponzin3,
  7. Stephen B Kaye1,4,
  8. Hannah J Levis4
  1. 1 Department of Ophthalmology, St Paul’s Eye Unit, Royal Liverpool University Hospital, Liverpool, UK
  2. 2 Instituto Universitario Fernandez-Vega, Universidad de Oviedo and Fundacion de Investigacion on Oftalmologica, Oviedo, Spain
  3. 3 International Center for Ocular Physiopathology, Veneto Eye Bank Foundation, Venice, Italy
  4. 4 Department of Eye and Vision Science, University of Liverpool Institute of Ageing and Chronic Disease, Liverpool, UK
  1. Correspondence to Dr Hannah J Levis, Department of Eye and Vision Science, Institute of Ageing and Chronic Disease, University of Liverpool, Liverpool, UK; h.levis{at}liverpool.ac.uk

Abstract

Background/aims Descemet membrane endothelial keratoplasty (DMEK) preparation is technically demanding and is a limiting factor for uptake of this kind of surgery. Supply methods that simplify the procedure for surgeons are key to increasing uptake. This study compares two different shipping protocols for DMEK.

Methods An 8.5 mm DMEK graft was punched, marked and loaded for transportation in two different conditions: (A) endothelium trifolded inwards in organ culture conditions (n=7) and (B) endothelium rolled outwards in hypothermic conditions (n=7). Tissues were shipped from Italy to the UK, then analysed for orientation, endothelial cell density, denuded areas, cell mortality, triple viability staining (Hoechst/ethidium homodimer/calcein AM (HEC)), immunolocalisation of ZO-1 and Na/K-ATPase proteins, visualisation of actin filaments using phalloidin and histological analysis using H&E on paraffin-embedded sections.

Results All tissues clearly showed the mark used for graft orientation. After shipping in condition A, there was an increase in cell mortality of 8.1% and in denuded areas of 22.4%, whereas for condition B there was an increase in cell mortality of 14.2% and in denuded areas of 34.3% after shipping. HEC staining revealed areas of viable cells and apoptotic cells, with large denuded areas found in the periphery for condition B and within folds for condition A.

Conclusions Prestripped preloaded DMEK grafts retained sufficient viable cells for transplantation, with condition A (endothelium-in) offering the advantage of greater flexibility of use due to a longer shelf-life. HEC analysis provides further detailed information as to the status of DMEK grafts and should be used in future similar studies.

  • cornea
  • dystrophy
  • experimental laboratory
  • treatment surgery

This is an Open Access article distributed in accordance with the terms of the Creative Commons Attribution (CC BY 4.0) license, which permits others to distribute, remix, adapt and build upon this work, for commercial use, provided the original work is properly cited. See: http://creativecommons.org/licenses/by/4.0/

https://doi.org/10.1136/bjophthalmol-2017-310906

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    Footnotes

    • Contributors All authors contributed: (1) substantial contributions to the conception or design of the work; or the acquisition, analysis or interpretation of data for the work; (2) drafting the work or revising it critically for important intellectual content; (3) final approval of the version to be published; (4) and agreement to be accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved.

    • Funding The study was supported by a Fight for Sight Small Grant awarded to VR and 2017 Eye Bank Association of America (EBAA) Richard Lindstrom Research Grant to MP.

    • Competing interests None declared.

    • Ethics approval Ethical permission for use of human tissue in research was obtained from the Research Ethics Committee (UK; 16/EM0090).

    • Provenance and peer review Not commissioned; externally peer reviewed.