Study of mechanotransduction in the endothelium of stored human corneas

AbstractPurposeCorneal endothelial cells (ECs) are continuously exposed to two physical stimuli: intraocular pressure (IOP) on their apical surface and contact with the Descemet membrane on their basal surface. We suppose that IOP could act on mechanoreceptors which could induce mechanotransduction in ECs: YAP (Yes Associated Protein) a transcriptional coactivator and Zo‐1 included in apical tight‐junctions. In numerous cell types, mechanical stimuli induce YAP nuclear translocation, which regulates genes expression involved in cell survival.AimTo study the localization of YAP in human ECs and its link with Zo‐1 homeostasis, using our corneal bioreactor (BR) capable of restoring a transcorneal pressure gradient equivalent to the IOP.MethodsHuman corneas were placed in the BR, during 14 days at 21 mmHg with circulation of a storage medium (CorneaMax, Eurobio, France). For control, paired corneas were stored in a flask without IOP in the same medium. Immunolabelling for YAP and Zo‐1 was done on paraformaldehyde of methanol fixed whole corneas by incubating primary and secondary antibodies on the endothelial side only. After flat mount, the entire endothelium was observed by epifluorescence microscope (IX81, Olympus, Japan).ResultsSeveral endothelial areas of corneas stored in the BR had regular hexagonal EC, well‐organized tight‐junctions, and a homogeneous nuclear localization of YAP. Corneas stored without IOP presented some areas without EC and other areas covered by EC with irregular shape, disorganized tight‐junctions and an heterogeneous localization of YAP, with some cells having focal expression in their nucleus and others cytoplasmic expression.ConclusionsThe bioreactor is a promising tool to study the role of mechanotransduction in human EC survival and function. IOP seems able alter expression of YAP and of Zo‐1. IOP‐induced nuclear translocation of YAP in various corneal areas seems linked with integrity of tight‐junctions.