A Novel Random Flexible Nanoscale Topography for Cell Culture

Abstract

Abstract: : Purpose: We have documented that anisotropic nanoscale topography modulates a variety of human corneal epithelial cell (HCEC) behaviors, including orientation, mobility, adhesion, and g–protein signaling. However, random substrate topographies are also of interest because they more closely mimic the topographic features found in the native corneal basement membrane. An ideal random substrate would be comparatively easy and inexpensive to fabricate at a variety of length scales. Methods: Gold nanotubules are fabricated by electroless deposition onto track–etched polycarbonate membranes. This deposition process results in a 10–100nm layer of gold coating the entire membrane. When the topmost gold and 0.5–1 micron of the underlying polymer substrate is removed, the resulting surface consists of gold nanotubules ("gold grass") projecting from a solid polycarbonate surface. A self–assembled monolayer of mercapto–hexadecanoic acid is adsorbed to the gold for improved cell attachment. Cells were plated at a density of 10,000 cells/cm². Assays were performed to determine the effect of topographic cueing provided by nanoscale gold grass surfaces on cell viability, morphology, and proliferation. Results: Gold grass suitable for cell culture was fabricated with outer diameters ranging from 30 to 800nm with an average (center to center) spacing from 400 to 2000nm. Viability of HCECs on gold grass surfaces after 24 hours is comparable to planar gold, electroless gold–coated membranes and tissue culture polystyrene. HCECs extend filopodia that attach to and bend the gold nanotubules, and exhibit time–dependent spreading and proliferation in a manner separate from the three control surfaces. Conclusions: Gold grass provides a viable, unique, and easy–to–fabricate substrate for cell culture. The size and thickness of the nanotubules can be readily adjusted, and the flexible surface is easy to chemically modify. Because a large amount of material can be manufactured at once, these substrates may be particularly suited to the study of topographic cueing on protein expression where large numbers of cells are required. Support: NEI Grant RO1–12253–01, NSF MRSEC Grants DMR–9632527 and CTS–9703207

Keywords: cornea: epithelium • extracellular matrix