Regionally-selective cell colonization of micropatterned surfaces prepared by plasma polymerization of acrylic acid and 1,7-octadiene.

Micropatterned surfaces have been used as a tool for controlling the extent and strength of cell adhesion, the direction of cell growth and the spatial distribution of cells. In this study, chemically micropatterned surfaces were prepared by successive plasma polymerization of acrylic acid (AA) and 1,7-octadiene (OD) through a mask. Rat vascular smooth muscle cells (VSMC), bovine endothelial cells (EC), porcine mesenchymal stem cells (MSC) or human skeletal muscle cells (HSKMC) were seeded on these surfaces in densities from 9,320 cells/cm(2) to 31,060 cells/cm(2). All cell types adhered and grew preferentially on the strip-like AA domains. Between day 1 and 7 after seeding, the percentage of cells on AA domains ranged from 84.5 to 63.3 % for VSMC, 85.3 to 73.5 % for EC, 98.0 to 90.0 % for MSC, and 93.6 to 55.0 % for HSKMC. The enzyme-linked immunosorbent assay (ELISA) revealed that the concentration of alpha-actin per mg of protein was significantly higher in VSMC on AA. Similarly, immunofluorescence staining of von Willebrand factor showed more apparent Weibel-Palade bodies in EC on AA domains. MSC growing on AA had better developed beta-actin cytoskeleton, although they were less stained for hyaluronan receptor (CD44). In accordance with this, MSC on AA contained a higher concentration of beta-actin, although the concentration of CD44 was lower. HSKMC growing on AA had a better developed alpha-actin cytoskeleton. These results based on four cell types suggest that plasma polymerization is a suitable method for producing spatially defined patterned surfaces for controlled cell adhesion, proliferation and maturation.

Plasma polymer coated surfaces for serum-free culture of limbal epithelium for ocular surface disease.

The potential use of plasma polymer coatings as substrates for serum-free expansion of limbal epithelial cells was investigated. Preliminary studies using a human corneal epithelial cell line showed that acrylic acid-coated surfaces performed better than allyl amine and allyl alcohol coated surfaces in terms of cell metabolic activity and confluence as assessed using the MTT assay. Subsequently, the proliferation and maturity of primary human limbal epithelial cells in co-culture with growth arrested 3T3 fibroblasts on a range of acrylic acid plasma coated surfaces, octadiene plasma coated surfaces and tissue culture plastic was investigated using MTT and cytokeratin 3 immunostaining. The cells performed better in the presence of serum on all surfaces. However, the acrylic acid coated surfaces successfully sustained a serum-free fibroblast/epithelial cell co-culture. The metabolic activity of the epithelial cells was superior on the acrylic acid coated surfaces than on tissue culture plastic in serum-free conditions and their levels of differentiation were not significantly higher than in the presence of serum. These results suggest that these surfaces can be used successfully for the serum-free expansion of human limbal epithelial cells.