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3D bioprinting of elastic cartilage tissues that are mechanically and structurally comparable to their native counterparts, while exhibiting favorable cellular behavior, is an unmet challenge. A practical solution for this problem is the multi-material bioprinting of thermoplastic polymers and cell-laden hydrogels using multiple nozzles. However, the processing of thermoplastic polymers requires high temperatures, which can damage hydrogel-encapsulated cells. In this study, the authors developed waterborne polyurethane (WPU)-polycaprolactone (PCL) composites to allow multi-material co-printing with cell-laden gelatin methacryloyl (GelMA) hydrogels. These composites can be extruded at low temperatures (50-60 °C) and high speeds, thereby reducing heat/shear damage to the printed hydrogel-capsulated cells. Furthermore, their hydrophilic nature improved the cell behavior in vitro. More importantly, the bioprinted structures exhibited good stiffness and viscoelasticity compared to native elastic cartilage. In summary, this study demonstrated low-temperature multi-material bioprinting of WPU-PCL-based constructs with good mechanical properties, degradation time-frames, and cell viability, showcasing their potential in elastic cartilage bio-fabrication and regeneration to serve broad biomedical applications in the future., (© 2024 Wiley‐VCH GmbH.)

Objective: To explore the feasibility of three-dimensional (3D) bioprinted adipose-derived stem cells (ADSCs) combined with gelatin methacryloyl (GelMA) to construct tissue engineered cartilage., Methods: Adipose tissue voluntarily donated by liposuction patients was collected to isolate and culture human ADSCs (hADSCs). The third generation cells were mixed with GelMA hydrogel and photoinitiator to make biological ink. The hADSCs-GelMA composite scaffold was prepared by 3D bioprinting technology, and it was observed in general, and observed by scanning electron microscope after cultured for 1 day and chondrogenic induction culture for 14 days. After cultured for 1, 4, and 7 days, the composite scaffolds were taken for live/dead cell staining to observe cell survival rate; and cell counting kit 8 (CCK-8) method was used to detect cell proliferation. The composite scaffold samples cultured in cartilage induction for 14 days were taken as the experimental group, and the composite scaffolds cultured in complete medium for 14 days were used as the control group. Real-time fluorescent quantitative PCR (qRT-PCR) was performed to detect cartilage formation. The relative expression levels of the mRNA of cartilage matrix gene [(aggrecan, ACAN)], chondrogenic regulatory factor (SOX9), cartilage-specific gene [collagen type Ⅱ A1 (COLⅡA1)], and cartilage hypertrophy marker gene [collagen type ⅩA1 (COLⅩA1)] were detected. The 3D bioprinted hADSCs-GelMA composite scaffold (experimental group) and the blank GelMA hydrogel scaffold without cells (control group) cultured for 14 days of chondrogenesis were implanted into the subcutaneous pockets of the back of nude mice respectively, and the materials were taken after 4 weeks, and gross observation, Safranin O staining, Alcian blue staining, and collagen type Ⅱ immunohistochemical staining were performed to observe the cartilage formation in the composite scaffold., Results: Macroscope and scanning electron microscope observations showed that the hADSCs-GelMA composite scaffolds had a stable and regular structure. The cell viability could be maintained at 80%-90% at 1, 4, and 7 days after printing, and the differences between different time points were significant ( P <0.05). The results of CCK-8 experiment showed that the cells in the scaffold showed continuous proliferation after printing. After 14 days of chondrogenic induction and culture on the composite scaffold, the expressions of ACAN, SOX9, and COLⅡA1 were significantly up-regulated ( P <0.05), the expression of COLⅩA1 was significantly down-regulated ( P <0.05). The scaffold was taken out at 4 weeks after implantation. The structure of the scaffold was complete and clear. Histological and immunohistochemical results showed that cartilage matrix and collagen type Ⅱ were deposited, and there was cartilage lacuna formation, which confirmed the formation of cartilage tissue., Conclusion: The 3D bioprinted hADSCs-GelMA composite scaffold has a stable 3D structure and high cell viability, and can be induced differentiation into cartilage tissue, which can be used to construct tissue engineered cartilage in vivo and in vitro .

Finding an ideal scaffold is always an important issue in the field of cartilage tissue engineering. Both decellularized extracellular matrix and silk fibroin have been used as natural biomaterials for tissue regeneration. In this study, a secondary crosslinking method of γ irradiation and ethanol induction was used to prepare decellularized cartilage extracellular matrix and silk fibroin (dECM-SF) hydrogels with biological activity. Furthermore, the dECM-SF hydrogels were cast in custom-designed molds to produce a three-dimensional multi-channeled structure to improve internal connectivity. The adipose-derived stromal cells (ADSC) were seeded on the scaffolds, cultured in vitro for 2 weeks, and implanted in vivo for another 4 and 12 weeks. The double crosslinked dECM-SF hydrogels exhibited an excellent pore structure after lyophilization. The multi-channeled hydrogel scaffold presents higher water absorption ability, surface wettability, and no cytotoxicity. The addition of dECM and a channeled structure could promote chondrogenic differentiation of ADSC and engineered cartilage formation, confirmed by H&E, safranin O staining, type II collagen immunostaining, and qPCR assay. In conclusion, the hydrogel scaffold fabricated by the secondary crosslinking method has good plasticity and can be used as a scaffold for cartilage tissue engineering. The multi-channeled dECM-SF hydrogel scaffolds possess a chondrogenic induction activity that promotes engineered cartilage regeneration of ADSC in vivo. [ABSTRACT FROM AUTHOR]