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Quantitative evaluation of the corneal endothelium in the mouse after grafting

¹ Department of Ophthalmology, University of Aberdeen, Scotland, UK
² Department of Ophthalmology, Charles University, Prague, Czech Republic
³ Institute of Molecular Genetics, Academy of Science, Prague, Czech Republic

Correspondence to:
Correspondence to:
Professor J V Forrester
Department of Ophthalmology, Medical School, University of Aberdeen, Foresterhill, Aberdeen AB25 2ZD, UK; j.forrester{at}abdn.ac.uk

Background/aim: Corneal graft survival depends critically on the quality of the endothelium. In this study the authors aimed to evaluate corneal endothelium in mice at different times after transplantation and to correlate endothelial integrity with corneal graft survival.

Methods: Syngeneic and allogeneic corneal grafts at various times (days 0–60) after engraftment were examined in flat mount preparation by confocal microscopy, by evaluating the hexagonal pattern of the endothelial monolayer using actin staining of the cell cortex. Corneas from untreated mice and from mice, who were grafted after removal of draining lymph nodes served as controls.

Results: In control corneas, more than 90% of the posterior surface was covered by endothelium. Syngeneic grafts were always covered by 54–99% of endothelium. In contrast, the posterior surface of corneal allografts showed great variation in the degree of endothelial cell coverage (0–98%). In addition, clinical opacity grading measure was not a reliable predictor of endothelial coverage.

Conclusion: In corneal allografts there is progressive loss of endothelium over time, unlike with syngeneic grafts. However, in the early stages of allograft rejection, the grade of graft opacity does not accurately reflect the degree of endothelial cell coverage. Although corneal opacity grade is considered the main determinant of graft rejection, the data suggest that both the grade of corneal opacity plus a sufficient post-graft time duration (>8 weeks in the mouse) are required for the diagnosis of irreversible graft rejection.

Abbreviations: eGFP, enhanced green fluorescence protein; LNR, lymph node removal; PBS, phosphate buffered saline; PFA, paraformaldehyde; tPA, tissue plasminogen activator

Keywords: corneal endothelial cells; corneal transplantation; actin; confocal microscopy