Effect of mechanical stretching and substrate stiffness on the morphology, cytoskeleton and nuclear shape of corneal endothelial cells

Abstract

Due to the limited capacity of corneal endothelial cells (CECs) division, corneal endothelial diseases have become a great challenge. The cornea is subjected to various mechanical stimuli in vivo, which may have a positive or negative influence. Thus, it is significant to gain an insight into the mechanism of mechanobiology of CECs for seeking more possible treatment. The purpose of this study was to determine the impacts of mechanical stretch and substrate stiffness on the morphology and fundamental cell behavior of CECs. Rabbit corneal endothelial cells (RCECs) were subjected to a 5% mechanical stretch or cultured on substrates of different stiffness. The impacts of mechanical stimulus on cell area, aspect ratio, circularity, cell density, nuclear shape, cytoskeleton, and cell viability were investigated. The expressions of the corneal endothelium-related markers ZO-1 and Na+/K+-ATPase were also evaluated by confocal immunofluorescence microscopy in the stiffness group. Our results suggested that mechanical stretch promoted the rearrangement of the cytoskeleton while decreasing the cell circularity, nuclear area, and cell density as well as cell viability. RCECs cultured on 10 ​kPa substrates, which was close to the physiological stiffness of rabbit Descemet's membrane (DM), showed better cell morphology, more stable actin cytoskeleton assembly, and more robust expression of the functional marker compared with other softer or stiffer substrates. In summary, mechanical stretch and substrate stiffness have profound influences on the morphology and function of CECs, which may have implications for the understanding and possible treatment of corneal endothelial diseases.