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Meidong Zhu, Li Wen, Gregory Moloney, Gerard Sutton, Jane Treloggen, Constantinos Petsoglou; Laboratory exploration of Ripasudil in descemetorhexis with a human ex vivo model. Invest. Ophthalmol. Vis. Sci. 2019;60(9):2171.
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© ARVO (1962-2015); The Authors (2016-present)
To investigate the effect of Ripasudil on corneal endothelial cell (CEC) survival and migration after descemetorhexis on a human ex vivo model
Eleven human corneoscleral buttons, including 1 individual and 5 pairs, were obtained from New South Wales Tissue Bank. To minimize the effects of donor variation, either the individual tissue was divided into two halves or paired tissue from the same donor was assigned into either Ripasudil or control group.A 2mm diameter descemetorhexis was created by a peeling technique.The tissues were incubated in either 50ml organ culture storage medium containing 10µM Ripasudil or 50µl Dimethyl Sulfoxide (DMSO) for 2 days prior to wound creation then for 14 day after wounding.Tissues were assessed at Day 3, 7 and 14 for morphology and cell migration after wounding. CEC viability and density were assessed at day 3 and 14. Immunohistochemistry with vimentin and Na+/K+/ATPase markers was conducted at the end of study.
Ripasudil prevented CEC death and appeared to maintain better morphology compared to DMSO. However, it did not improve cell proliferation. There were no difference in cell density changes in the periphery between 2 groups at day 14 compared to day 3 (740/mm2 vs 644/mm2, p>0.05). However significant changes were detected in the areas close to the edge of the wound between the two groups (426 vs 1124/mm2, p<0.01) with more cell loss in DMSO group. The viability of CEC decreased significantly in DMSO group after 14 day culture compared to Ripasudil group (viabilities of Ripasudil vs DMSO were 95.7%±4.9 vs 95.0%±2.8 and 81.7%±12.6 vs 60.0%±14.1 in periphery and wound edge, respectively).In Ripasudil group, migration of cells over the wound edge was observed with an altered fibroblastic phenotype similar to that captured with confocal microscopy in patients treated with Ripasudil. A trend was noticed where corneal endothelium from younger donors resulted in better cell migration than older corneas. Immunohistochemistry showed some increased Vimentin filaments and Na+/K+/ATPase positive cells around wound edge and cells migrated across denuded areas.
Ex vivo modelling demonstrated that Ripasudil maintained better endothelial morphology and promoted cell migration. These effects might be via transformation of endothelial cells into a more motile phenotype.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
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