Functional reconstruction of corneal endothelium using nanotopography for tissue-engineering applications

Dysfunction in the corneal endothelium, which controls the hydration and transparency of the cornea, is one of the common reasons for transplantation. A tissue-engineered corneal endothelium is of interest for corneal regeneration and for in vitro testing of ocular drugs. In the native environment, corneal endothelial cells interact with the nanotopography of the underlying Descemet's membrane. This study showed that nanotopography enhanced bovine corneal endothelial cell (BCEC) responses, creating a monolayer which resembled the healthy corneal endothelium. Topographies of different geometries were first tested to identify those that would elicit the most significant responses. A BCEC monolayer was then generated on both micro- and nanoscale pillars and wells. The BCEC monolayer cultured on topographies exhibited polygonal geometries with well-developed tight junction proteins. Scanning electron microscopy revealed that cells on pillars showed a higher density of microvilli, which was similar to native corneal endothelium. BCECs on nanopillars displayed a lower coefficient of variation of area (0.31) that was within the range of healthy corneal endothelium. More importantly, a BCEC monolayer cultured on nanopillars also had an enhanced Na(+)/K(+)-ATPase immunofluorescence expression, mRNA upregulation and a higher Na(+)/K(+)-ATPase activity. These results suggest that nanopillar substrate topography may provide relevant topographical cues, which could significantly enhance the formation and function of corneal endothelium.