In silico and in vitro analysis of cation-activated potassium channels in human corneal endothelial cells

The corneal endothelium is the inner cell monolayer involved in the maintenance of corneal transparence by the generation of homeostatic dehydration. The glycosaminoglycans of the corneal stroma develop a continuous swelling pressure that should be counteracted by the corneal endothelial cells through active transport mechanisms to move the water to the anterior chamber. Protein transporters for sodium (Na⁺), potassium (K⁺), chloride (Cl^-) and bicarbonate (HCO₃^-) are involved in this endothelial "pump function", however despite its physiological importance, the efflux mechanism is not completely understood. There is experimental evidence describing transendothelial diffusion of water in the absence of osmotic gradients. Therefore, it is important to get a deeper understanding of alternative models that drive the fluid transport across the endothelium such as the electrochemical gradients. Three transcriptomic datasets of the corneal endothelium were used in this study to analyze the expression of genes that encode proteins that participate in the transport and the reestablishment of the membrane potential across the semipermeable endothelium. Subsequently, the expression of the identified channels was validated in vitro both at mRNA and protein levels. The results of this study provide the first evidence of the expression of KCNN2, KCNN3 and KCNT2 genes in the corneal endothelium. Differences among the level of expression of KCNN2, KCNT2 and KCNN4 genes were found in a differentially expressed gene analysis of the dataset. Taken together these results underscore the potential importance of the ionic channels in the pathophysiology of corneal diseases. Moreover, we elucidate novel mechanisms that might be involved in the pivotal dehydrating function of the endothelium and in others physiologic functions of these cells using in silico pathways analysis.