Stem cell spheroids production for wound healing with a reversible porous hydrogel

Adipose derived stem cell (ASC) spheroids show enhanced cellular viability and paracrine function to promote tissue regeneration and cell therapy effect. However, efficient preparation of numerous ASC spheroids, as well as convenient application on skin wound remains a challenge. The present study explored a new function of frequently-used polymer, Poly (N-isopropylacrylamide) (PNIPAM), developing a simple but efficient method based on PNIPAM to produce numerous ASC spheroids and apply them towards wound healing. PNIPAM-based porous hydrogel that crosslinked with disulfide bonds was fabricated, showing pore size of 300 μm at 37 °C. ASCs seeded into porous hydrogel spontaneously aggregated inside the inner pores to form a large number of spheroids. Higher cell seeding density could produce larger size of spheroids, which showed up-regulated paracrine secretion. After spheroids production, the porous hydrogel became a solution within 25 min after the addition of glutathione at 15 °C. The PNIPAM-based polymer solution carrying the produced ASC spheroids gelled again within 80 s at 33 °C and could tightly attach on skin. The in vivo wound healing evaluation illustrated that ASC spheroids promoted M2 polarization of macrophages, and effectively inhibited the local inflammatory response of wounds, thus accelerating wound healing. Thus, for one thing, this method was expected to realize the industrialized manufacturing of a large number of stem cell spheroids. For another, the spheroids produced by the present method could be easily collected and applied for biomedical engineering.