Cryogenic 3D printing of heterogeneous scaffolds with gradient mechanical strengths and spatial delivery of osteogenic peptide/TGF-ß1 for osteochondral tissue regeneration.

Due to the increasing aging population and the high probability of sport injury among young people nowadays, it is of great demand to repair/regenerate diseased/defected osteochondral tissue. Given that osteochondral tissue mainly consists of a subchondral layer and a cartilage layer which are structurally heterogeneous and mechanically distinct, developing a biomimetic bi-phasic scaffold with excellent bonding strength to regenerate osteochondral tissue is highly desirable. Three-dimensional (3D) printing is advantageous in producing scaffolds with customized shape, designed structure/composition gradients and hence can be used to produce heterogeneous scaffolds for osteochondral tissue regeneration. In this study, bi-layered osteochondral scaffolds were developed through cryogenic 3D printing, in which osteogenic peptide/ß-tricalcium phosphate/poly(lactic-co-glycolic acid) water-in-oil composite emulsions were printed into hierarchically porous subchondral layer while poly(D,L-lactic acid-co-trimethylene carbonate) water-in-oil emulsions were printed into thermal-responsive cartilage frame on top of the subchondral layer. The cartilage frame was further filled/dispensed with transforming growth factor-ß1 loaded collagen I hydrogel to form the cartilage module. Although the continuously constructed osteochondral scaffolds had distinct microscopic morphologies and varied mechanical properties at the subchondral zone and cartilage zone at 37 °C, respectively, the two layers were closely bonded together, showing excellent shear strength and peeling strength. Rat bone marrow derived mesenchymal stem cells (rBMSCs) exhibited high viability and proliferation at both subchondral- and cartilage layer. Moreover, gradient rBMSC osteogenic/chondrogenic differentiation was obtained in the osteochondral scaffolds. This proof-of-concept study provides a facile way to produce integrated osteochondral scaffolds for concurrently directing rBMSC osteogenic/chondrogenic differentiation at different regions.

Cryogenic 3D printing of porous scaffolds for in situ delivery of 2D black phosphorus nanosheets, doxorubicin hydrochloride and osteogenic peptide for treating tumor resection-induced bone defects.

Tumor resection is widely used to prevent tumor growth. However, the defected tissue at the original tumor site also causes tissue or organ dysfunction which lowers the patient's life quality. Therefore, regenerating the tissue and preventing tumor recurrence are highly important. Herein, according to the concept of 'first kill and then regenerate', a versatile scaffold-based tissue engineering strategy based on cryogenic 3D printing of water-in-oil polyester emulsion inks, containing multiple functional agents, was developed, in order to realize the elimination of tumor cells with recurrence suppression and improved tissue regeneration sequentially. To illustrate our strategy, water/poly(lactic-co-glycolic acid)/dichloromethane emulsions containing ß-tricalcium phosphate (ß-TCP), 2D black phosphorus (BP) nanosheets, low-dose doxorubicin hydrochloride (DOX) and high-dose osteogenic peptide were cryogenically 3D printed into hierarchically porous and mechanically strong nanocomposite scaffolds, with multiple functions to treat bone tumor, resection-induced tissue defects. Prompt tumor ablation and long-term suppression of tumor recurrence could be achieved due to the synergistic effects of photothermotherapy and chemotherapy, and improved bone regeneration was obtained eventually due to the presence of bony environment and sustained peptide release. Notably, BP nanosheets in scaffolds significantly reduced the long-term toxicity phenomenon of released DOX during in vivo bone regeneration. Our study also provides insights for the design of multi-functional tissue engineering scaffolds for treating other tumor resection-induced tissue defects.