Novel method of forming human embryoid bodies in a polystyrene dish surface-coated with a temperature-responsive methylcellulose hydrogel.

A temperature-responsive hydrogel composed of aqueous methylcellulose (MC) blended with distinct concentrations of PBS was prepared and characterized. The developed MC hydrogel underwent a sol-gel reversible transition upon heating or cooling at approximately 32 degrees C. This temperature-responsive hydrogel was employed to coat the surface of a polystyrene dish and used to cultivate human embryonic stem (hES) cell clumps for the formation of embryoid bodies (EBs) in liquid suspension culture (LSC-MC/PS). The conventional hanging drop culture (HDC) and LSC in the uncoated polystyrene dish (LSC-PS) or in the Corning Ultralow-Attachment plate (LSC-ULAP) were used as controls. The results indicated that LSC-PS failed to generate EBs in an efficient manner, whereas the efficiencies of EB formation observed in LSC-ULAP and LSC-MC/PS were significantly greater than in HDC. The hES cells within the EBs were shown to express molecular markers specific for representative cells from the three embryonic germ layers. These results indicated that the MC-coated dish can be used to produce a large scale of hES cell derivatives through the formation of EBs.