Development of novel intelligent tissue culture surfaces to accelerate cell proliferation and recover cell sheet under serum free conditions, using physiologically active peptides

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In order to evolve advanced method of cell sheet engineering, we have developed novel temperature responsive polymers grafted tissue culture polystyrene (TCPS), copolymerizing 2-carboxyisopropylacrylamide monomer as following immobilization of physiologically active peptides with IPAAm monomer (PIPAAm-CIPAAm-TCPS). The novel culture surfaces, PIPAAm-CIPSSm-TCPS, modified with RGD peptides attain culturing cells even under serum free conditions, and cells were successfully recovered as a sheet from the surfaces by lowering temperature. Cell culture onto the novel surfaces is useful for cells to avoid contamination of prion protein and virus during cell culture due to serum free condition. RGD and PHSRN modified surface exhibited more cell adhesive property than only RDG. In contrast, introduction of RDG and Insulin onto the novel culture surface achieve acceleration of proliferation of cultured cells. RGD peptides I inked with biotin were introduced onto avidin proteins modified onto the novel surfaces. Resulting RGD modified surfaces also showed cell adhesion and proliferation properties even under serum free conditions, strongly suggesting various physiologically active peptides, which contain biotin component, are easily immobilized onto the novel surfaces modified with avidin. Determination of appropriate physiologically active peptides for corneal epithelium cells, retinal pigment epithelium cells, alveolar cells and cardiac muscle cells may promote clinical application utilizing these kinds of cell sheets.