Toward a Nerve Regeneration Device: Templating Cell Organization on a Peripheral Nervous System-Compatible Polymer, and a Model for Spatial Control of Cells in Three Dimensions

Abstract

Methods were developed to template cell adhesion, proliferation, and extracellular matrix (ECM) assembly on biomedical polymers in order to generate aligned, native-like ECM that may serve as a tissue regeneration scaffold. The silicon microfabrication techniques of photolithography and chemical vapor deposition were modified to pattern a cell-adhesive, two-component interface consisting of a zirconium oxide (ZrO2) layer terminated with a self-assembled monolayer of phosphonates (SAMP) on commercially available polymer films. Polymers such as polycaprolactone fumarate (PCLF)--a material employed in nerve regeneration scaffolds--proved incompatible with photolithographic reagents; a shadow masking technique was developed to synthesize the patterned ZrO2 / SAMP interface on PCLF. A laser ablated polymer stencil mask was used for shadow masking and proved particularly surface conforming. It was thus useful for patterning curved substrates. ZrO2 / SAMP patterns were characterized by XPS, SEM, EDS, and AFM, and mechanical testing showed that construction of the interface did not change the elastic modulus of the polymer substrate. The ZrO2 / SAMP interface templated alignment of NIH3T3 fibroblast cells plated on patterned polymer substrates and resulted in alignment of fibronectin assembled by these cells.

Laser ablated holes were introduced to polyetheretherketone (PEEK) films to probe an improvement in diffusion when 2D polymer layers were stacked, as one means to translate cell alignment on 2D layers to a 3D structure. Perforated PEEK substrates were functionalized with the patterned ZrO2 / SAMP interface, which caused alignment of cells after 3 days. Compared to non-perforated substrates, perforated PEEK films allowed for some increase in diffusion of a vital dye to cells when layers were stacked; perforated, patterned substrates allowed for maintenance of cell alignment in a stacked construct for 4 days. The model of stacked, patterned polymer films to achieve cell alignment in 3D may inform the construction of an aligned-ECM / polymer composite tissue regeneration scaffold based on ZrO2 / SAMP patterning to template cell-assembly of ECM on designer polymers for nerve regeneration.