Epithelial alternate exposure to air and liquid further improves human corneas stored in the active storage machine

Abstract

AbstractPurposeThe standard version of our corneal active storage machine (ASM) maintains corneas viability much better than standard organ culture over 1 and 3 months (Am J Transplant 2019, Transplantation 2020). While it aims at recreating a physiologic environment (IOP and medium renewal), corneas were nevertheless immersed from both sides, probably contributing to slightly higher thickness than in vivo.AimTo assess whether epithelial alternate exposure to air and liquid (AirLifting, AL) could further improve quality of corneas stored into the ASM.MethodsTwenty paired human corneas (initial ECD > 2000 cells/mm2 and <10% difference between right and left) were randomized to be stored for 1 month in the ASM either fully immersed or with AL. All other parameters were similar: IOP (21 mmHg), endothelial medium flow (2.6 μL/min) and nature (CMax, Eurobio). AL was done with CMax (6 cycles air‐liquid/min). Specular microscopy ECD, OCT thickness, transparency were monitored at Day (D)2, D26, D28. Pancorneal viable ECD (Hoechst/Ethidium/Calcein‐AM), histology, immunostaining, western blot for epithelial cells and endothelial cells (ECs) structure/functions proteins were done at D28.ResultsAt D2, ECD was 2412 ± 222 versus 2385 ± 251 cells/mm2 (p = 0.349) respectively with and without AL and 2109 ± 165 versus 2119 ± 199 cells/mm2 (p = 0.621) at D26. The viable ECD at D28 was 2170 ± 321 versus 2185 ± 275cells/mm2 (p = 0.794). All corneas remained thin transparent during storage and at D28 only, corneas were thinner with AL: 667 ± 60μm versus 695 ± 55μm without (p = 0.048). Epithelial quality was similar, but the epithelium was thicker with AL (49 ± 20) than without (23 ± 7μm) (p = 0.001) and had more cell layers. Endothelial cells structure/functions were similar.ConclusionsBy mimicking blinking, AL enhances epithelium structure during storage into the ASM. It may also slightly help controlling corneal thickness. ASM with AL may be useful for penetrating and anterior keratoplasty as well as for research works.