Immobilization of Cell-Adhesive Peptides to Temperature-Responsive Surfaces Facilitates Both Serum-Free Cell Adhesion and Noninvasive Cell Harvest

We have developed temperature-responsive cell culture surfaces to harvest intact cell sheets for tissue- engineering applications. Both cost and safety issues (e.g., prions, bovine spongiform encephalopathy) are compelling reasons to avoid use of animal-derived materials, including serum, in such culture. In the present study, synthetic cell-adhesive peptides are immobilized onto temperature- responsive polymer-grafted surfaces, and cell adhesion and detachment under serum-free conditions were examined. The temperature-responsive polymer poly(N-isopropylacrylamide) (PIPAAm) was functionalized by copolymerization with a reactive comonomer having both a carboxyl group and an isopropylacrylamide group. These copolymers were covalently grafted onto tissue culture- grade polystyrene dishes. Synthetic cell-adhesive peptides were then immobilized onto these surfaces via carboxyl groups. Bovine aortic endothelial cells both adhered and spread on these surfaces even under serum-free conditions at 37°C, similar to those in 10% serum-supplemented culture. Spread cells promptly detached from the surfaces on lowering culture temperatures below the lower critical solution temperature of the polymer, 32°C. These surfaces would be useful for serumfree culture for tissue-engineering applications.