Effective cell sheet preparation using thermoresponsive polymer brushes with various graft densities and chain lengths

Various cell sheets have been used as effective and useful cellular tissues in tissue engineering and regenerative therapy. Poly(N-isopropylacrylamide) (PNIPAAm)-modified surfaces have been investigated for effective cell sheet preparation. In this study, the effective PNIPAAm graft density and chain length of PNIPAAm brushes for various cell types were investigated. The PNIPAAm brush-grafted glass was prepared via silanization and subsequent atom transfer radical polymerization (ATRP). The density of the PNIPAAm brushes was modulated by changing the ATRP initiator and co-adsorber composition, while the PNIPAAm brush length was modulated by changing the monomer concentration in the ATRP. The hydrophilicity of the PNIPAAm brushes increased with increasing PNIPAAm brush length because long PNIPAAm brushes tended to hydrate. Fibronectin adsorption increased with decreasing PNIPAAm brush concentration because the exposed hydrophobic co-adsorber in the dilute PNIPAAm brush enhanced the adsorption of fibronectin. The cell-sheet fabrication ability was investigated using six types of PNIPAAm brushes. An endothelial cell sheet was fabricated using a dense, short PNIPAAm brush. NIH/3T3 sheets can be fabricated using three types of PNIPAAm brushes: dense-long PNIPAAm brushes, moderately dense-short PNIPAAm brushes, and dilute-long PNIPAAm brushes. MDCK cell sheets could not be prepared using the PNIPAAm brushes. A549 cell sheets were prepared using a dense-short PNIPAAm brush and moderately dense-short PNIPAAm brushes. These results indicate that the optimal PNIPAAm brush conditions for cell sheet preparation vary depending on cell type. Thus, modulation of PNIPAAm brush density and length is an effective approach for preparing target cell sheets.