The use of electron beam lithographic graft-polymerization on thermoresponsive polymers for regulating the directionality of cell attachment and detachment

A simple process for nano-patterned cell culture substrates by direct graft-polymerization has been developed using an electron beam (EB) lithography system requiring no photo-masks or EB-sensitive resists. The compound N-isopropylacrylamide (IPAAm) was locally polymerized and grafted directly by EB lithographic exposure onto hydrophilic polyacrylamide (PAAm)-grafted glass surfaces. The size of the surface grafted polymers was controlled by varying the area of EB dose, and a minimal stripe pattern with a 200 nm line-width could be fabricated onto the surface. On the stripe-patterned surfaces, above the lower critical solution temperature (LCST), the cells initially adhered and spread with an orientation along the pattern direction. The magnitude of the spreading angle and elongation of adhered cells depended on the pattern intervals of the grafted PIPAAm. When culture temperature was lower than the LCST, cultured cells detached from the surfaces with strong shrinkage along the pattern direction, and sometimes folded and became parallel with the stripe pattern. This patterned cell recovery technique may be useful for the construction of muscle cell sheets with efficient shrinkage/relaxation in a specific direction and spheroidal 3D cell structures, with application to tissue engineering and microfluidic cellular devices.