Patterned poly(chlorotrifluoroethylene) guides primary nerve cell adhesion and neurite outgrowth

Abstract

Central nervous system (CNS) neurons, unlike those of the peripheral nervous system, do not spontaneously regenerate following injury. Recently it has been shown that in the developing CNS, a combination of cell-adhesive and cell-repulsive cues guide growing axons to their targets. We hypothesized that by mimicking these guidance signals, we could guide nerve cell adhesion and neurite outgrowth in vitro. Our objective was to direct primary nerve cell adhesion and neurite outgrowth on poly(chlorotrifluoroethylene) (PCTFE) surfaces by incorporating alternating patterns of cell-adhesive (peptide) and nonadhesive (polyethylene glycol; PEG) regions. PCTFE was surface-modified with lithium PEG-alkoxide, demonstrating the first report of metal-halogen exchange with an alkoxide and PCTFE. Titanium and then gold were sputtered onto PEG-modified films, using a shadow-masking technique that creates alternating patterns on the micrometer scale. PCTFE-Au regions then were modified with one of two cysteine-terminated laminin-derived peptides, C-GYIGSR or C-SIKVAV. Hippocampal neuron cell-surface interactions on homogeneously modified surfaces showed that neuron adhesion was decreased significantly on PEG-modified surfaces and was increased significantly on peptide-modified surfaces. Cell adhesion was greatest on CGYIGSR surfaces while neurite length was greatest on CSIKVAV surfaces and PLL/laminin positive controls, indicating the promise of peptides for enhanced cellular interactions. On patterned surfaces, hippocampal neurons adhered and extended neurites preferentially on peptide regions. By incorporating PEG and peptide molecules on the surface, we were able to simultaneously mimic cell-repulsive and cell-adhesive cues, respectively, and maintain the biopatterning of primary CNS neurons for over 1 week in culture.

Length-scale mediated adhesion and directed growth of neural cells by surface-patterned poly(ethylene glycol) hydrogels

“…Many investigators have since used this method to either test neuronal preferences for various substrates or to confine neurons to specific areas of a substrate. Common methods for patterning neuronal growth have been based on UV photolithography [15, 16], microcontact printing [17], masked sputtering [18], microfluidic patterning [19], ink-jet printing [20], and fiber aligning [21], among others. Few methods, however, have thus far exploited a spatial resolution comparable to the characteristic size of a typical axon.…”

Biomaterial surface patterning of self-assembled monolayers for controlling neuronal cell behaviour

“…There have been many techniques used to modify the surface of a biomaterial and they mainly fall under two major categories, chemical and topographical (Stevens and George 2005; Molnar, Hirsch-Kuchma et al 2006). Surface chemistry and topography (surface structure) of biomaterial substrates have a great influence on regulating cell behavior, such as adhesion, migration, orientation, guidance, differentiation, proliferation, gene expression and protein synthesis (Kleinfeld, Kahler et al 1988; Dulcey, Georger et al 1991; Ravenscroft, Bateman et al 1998; Stenger, Hickman et al 1998; Jenney and Anderson 1999; Saneinejad and Shoichet 2000; Craighead, James et al 2001; Andersson, Olsson et al 2003). This is due to the fact that during natural development the same types of extracellular signals are regulating many cellular functions.…”

High-density electrode array for imaging in vitro electrophysiological activity

“…The first one consists of patterning the cell networks on MEAs by using adhesion promoters/inhibitors or guiding microstructures (Heiduschka et al, 2001;Martinoia et al, 1999;Saneinejad and Shoichet, 2000;Yeung et al, 2001). The second methodology relies on confinement of individual neurons over each electrode, the so-called neuro-cages, built in a way allowing the formation of networks (Maher et al, 1999).…”