Solid multifunctional granular bioink for constructing chondroid basing on stem cell spheroids and chondrocytes

Stem cell spheroids are advanced building blocks to produce chondroid. However, the multi-step operations including spheroids preparation, collection and transfer, the following 3D printing and shaping limit their application in 3D printing. The present study fabricates an 'ALL-IN-ONE' bioink based on granular hydrogel to not only produce adipose derived stem cell (ASC) spheroids, but also realize the further combination of chondrocytes and the subsequent 3D printing. Microgels (6-10 mu m) grafted with beta-cyclodextrin (beta-CD) (MG(beta-CD)) were assembled and crosslinked by in-situ polymerized poly (N-isopropylacrylamide) (PNIPAm) to form bulk granular hydrogel. The host-guest action between beta-CD of microgels and PNIPAm endows the hydrogel with stable, shear-thinning and self-healing properties. After creating caves, ASCs aggregate spontaneously to form numerous spheroids with diameter of 100-200 mu m inside the hydrogel. The thermosensitive porous granular hydrogel exhibits volume change under different temperature, realizing further adsorbing chondrocytes. Then, the granular hydrogel carrying ASC spheroids and chondrocytes is extruded by 3D printer at room temperature to form a tube, which can shrink at cell culture temperature to enhance the resolution. The subsequent ASC spheroids/chondrocytes co-culture forms cartilage-like tissue at 21 d in vitro, which further matures subcutaneously in vivo, indicating the application potential of the fully synthetic granular hydrogel ink toward organoid culture.