Nano- and Microscale Holes Modulate Cell-Substrate Adhesion, Cytoskeletal Organization, and $-\beta 1$ Integrin Localization in Sv40 Human Corneal Epithelial Cells

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Abstract
Human corneal epithelial cells (HCECs) interface with a basement membrane in vivo that possesses complex nanoscale topographic features. We report that synthetic substrates patterned with nano- and microscale holes differentially modulate the proliferation, shape and adhesion of SV40 human corneal epithelial cells (SV40-HCECs) as a function of feature size: 1) Cell proliferation was inhibited on nanoscale features (features size less than 800 nm in pitch) compared to microscale features or planar substrates in identical culture conditions. 2)Cells on nanoscale holes had a stellate morphology compared to those on microscale features that were more evenly spread. 3) Cells adhered more to nanoscale features than to microscale features when exposed to shear stress in a laminar flow chamber. Transmission electron microscopy showed that cells cultured on the 400 nm pitch patterns had longer and more numerous filopodia and retraction fibers than cells cultured on the 1600 nm pitch patterns. Immunogold labeling of -beta 1 integrins revealed that these receptors were localized at the cell periphery and in the aforementioned cytoskeletal elements. Our findings indicate that surface discontinuities and the activation of mechanochemical cell signaling mechanisms may contribute to the observed responses exhibited by SV40-HCECs cultured on nano- and microscale topography