A comparative study on different Descemet membrane endothelial keratoplasty graft preparation techniques

Purpose: To compare different Descemet membrane endothelial keratoplasty (DMEK) graft preparation methods. Methods: Stripping from the trabecular meshwork (M1) using epithelial spatula; stripping by scoring the peripheral endothelium (M2) using Sinskey hook; stripping by punch method (M3) using donor trephine; Submerged hydro-separation (M4); and pneumatic dissection method (M5) were evaluated. Preparation time, costs, endothelial cell loss (ECL) postpreparation, cell death and morphology were compared. Hoechst/Ethidium/Calcien AM (HEC) staining and Zonula Occludens-1 (ZO-1) expression were analysed. Statistical analysis was performed using one-way anova and; Tukey as post hoc test. Results: A total of 35 corneas (seven per group) were used. Endothelial cell loss (ECL) represented as Mean (SD), in M1, M2, M3, M4 and M5 was 2.7 (5.0), 3.0 (7.4), 1.2 (7.4), 3.3 (7.3) and 4.1 (7.1)%, respectively not showing any difference between the groups (p = 0.96). A significantly higher cell death (p < 0.05) was observed in M4 and M5 compared with M1, M2 and M3. Graft preparation time was significantly shorter in M4 and M5 and longest in M3 (p < 0.05). M3 was the most expensive preparation technique. Minimum pleomorphic cells were observed in M1, M2 and M3, whereas moderate pleomorphism was seen in M4 and M5. Hoechst, Ethidium homodimer and Calcein AM (HEC) staining showed high Ethidium positivity (dead cells) in M4 and M5 with minimum positivity in M1, M2 and M3. Zonula Occludens-1 (ZO-1) was expressed in all the conditions except the denuded areas. Conclusion: Graft preparation using Sinskey hook (M2) and donor punch (M3) are reliable methods in terms of efficiency and quality with acceptable range of ECL. The preparation time and associated costs could be a limitation for M3.