Fabrication of 3D-Printed Interpenetrating Hydrogel Scaffolds for Promoting Chondrogenic Differentiation.

The limited self-healing ability of cartilage necessitates the application of alternative tissue engineering strategies for repairing the damaged tissue and restoring its normal function. Compared to conventional tissue engineering strategies, three-dimensional (3D) printing offers a greater potential for developing tissue-engineered scaffolds. Herein, we prepared a novel photocrosslinked printable cartilage ink comprising of polyethylene glycol diacrylate (PEGDA), gelatin methacryloyl (GelMA), and chondroitin sulfate methacrylate (CSMA). The PEGDA-GelMA-CSMA scaffolds possessed favorable compressive elastic modulus and degradation rate. In vitro experiments showed good adhesion, proliferation, and F-actin and chondrogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) on the scaffolds. When the CSMA concentration was increased, the compressive elastic modulus, GAG production, and expression of F-actin and cartilage-specific genes (COL2, ACAN, SOX9, PRG4) were significantly improved while the osteogenic marker genes of COL1 and ALP were decreased. The findings of the study indicate that the 3D-printed PEGDA-GelMA-CSMA scaffolds possessed not only adequate mechanical strength but also maintained a suitable 3D microenvironment for differentiation, proliferation, and extracellular matrix production of BMSCs, which suggested this customizable 3D-printed PEGDA-GelMA-CSMA scaffold may have great potential for cartilage repair and regeneration in vivo.

Lyophilized Scaffolds Fabricated from 3D-Printed Photocurable Natural Hydrogel for Cartilage Regeneration.

Repair of cartilage defects is highly challenging in clinical treatment. Tissue engineering provides a promising approach for cartilage regeneration and repair. As a core component of tissue engineering, scaffolds have a crucial influence on cartilage regeneration, especially in immunocompetent large animal and human. Native polymers, such as gelatin and hyaluronic acid, have known as ideal biomimetic scaffold sources for cartilage regeneration. However, how to precisely control their structure, degradation rate, and mechanical properties suitable for cartilage regeneration remains a great challenge. To address these issues, a series of strategies were introduced in the current study to optimize the scaffold fabrication. First, gelatin and hyaluronic acid were prepared into a hydrogel and 3D printing was adopted to ensure precise control in both the outer 3D shape and internal pore structure. Second, methacrylic anhydride and a photoinitiator were introduced into the hydrogel system to make the material photocurable during 3D printing. Finally, lyophilization was used to further enhance mechanical properties and prolong degradation time. According to the current results, by integrating photocuring 3D printing and lyophilization techniques, gelatin and hyaluronic acid were successfully fabricated into human ear- and nose-shaped scaffolds, and both scaffolds achieved shape similarity levels over 90% compared with the original digital models. The scaffolds with 50% infill density achieved proper internal pore structure suitable for cell distribution, adhesion, and proliferation. Besides, lyophilization further enhanced mechanical strength of the 3D-printed hydrogel and slowed its degradation rate matching to cartilage regeneration. Most importantly, the scaffolds combined with chondrocytes successfully regenerated mature cartilage with typical lacunae structure and cartilage-specific extracellular matrixes both in vitro and in the autologous goat model. The current study established novel scaffold-fabricated strategies for native polymers and provided a novel natural 3D scaffold with satisfactory outer shape, pore structure, mechanical strength, degradation rate, and weak immunogenicity for cartilage regeneration.

Development of a xeno-free substrate for human embryonic stem cell growth.

Traditionally, human embryonic stem cells (hESCs) are cultured on inactivated live feeder cells. For clinical application using hESCs, there is a requirement to minimize the risk of contamination with animal components. Extracellular matrix (ECM) derived from feeder cells is the most natural way to provide xeno-free substrates for hESC growth. In this study, we optimized the step-by-step procedure for ECM processing to develop a xeno-free ECM that supports the growth of undifferentiated hESCs. In addition, this newly developed xeno-free substrate can be stored at 4°C and is ready to use upon request, which serves as an easier way to amplify hESCs for clinical applications.