The creation of single and multilayered adult stem cells (ASCs) sheets is presented. The stem cell sheets preserve the cell-cell and cell-extracellular matrices and are developed by utilizing a thermally reversible methylcellulose (MC) coated tissue culture polystyrene (TCPS) dish. This technique is an improvement and a simplification of earlier noninvasive cell retrieval methods based on the use of a temperature-responsive poly(N-isopropylacrylamide) (PIPAAm) coated TCPS dishes. The optimal combination of MC-water-salt was determined to be 12-14% of MC (mol. wt. of 15,000) in water with 0.5× PBS (~150 mOsm). This solution exhibited a gel formation temperature of ~32 °C. The addition (evenly spread) of 1 ml of 3 mg/ml rat tail type-I (pH adjusted to 7.5) over the MC coated surface at 37 °C improves ASC adhesion and proliferation on the methylcellulose system. Upon confluence, a continuous monolayer ASC sheet was formed on the surface of the MC hydrogel system. When the grown cell sheet was removed from the incubator and exposed to room temperature (~30 °C), it spontaneously and gradually detached from the surface of the thermoresponsive hydrogel, creating an ASC sheet.
Keywords: Adult stem cells; Cell sheet engineering; Temperature-responsive polymers; Tissue engineering.