Gelatin-tyramine addition and low hydrogel density improves cell attachment, migration, and metabolic activity in vitro and tissue response in vivo in enzymatically crosslinkable dextran-hyaluronic acid hydrogels.

Hydrogels are receiving increasing attention for their use in 3D cell culture, tissue engineering, and bioprinting applications. Each application places specific mechanical and biological demands on these hydrogels. We developed a hydrogel toolbox based on enzymatically crosslinkable polysaccharides via tyramine (TA) moieties, allowing for rapid and tunable crosslinking with well-defined stiffness and high cell viability. Including gelatin modified with TA moieties (Gel-TA) improved the hydrogels' biological properties; 3 T3 fibroblasts and HUVECs attached to and proliferated on the enriched hydrogels at minute Gel-TA concentrations, in contrast to bare or unmodified gelatin-enriched hydrogels. Moreover, we were able to switch HUVECs from a quiescent to a migratory phenotype simply by altering the ligand concentration, demonstrating the potential to easily control cell fate. In encapsulation studies, Gel-TA significantly improved the metabolic activity of 3 T3 fibroblasts in soft hydrogels. Furthermore, we showed rapid migration and network formation in Gel-TA enriched hydrogels in contrast to a non-migratory behavior in non-enriched polysaccharide hydrogels. Finally, low hydrogel density significantly improves tissue response in vivo with large infiltration and low fibrotic reaction. Further development by adding ECM proteins, peptides, and growth factor adhesion sites will lead to a toolbox for hydrogels tailored toward their desired application.

Poly(acrylic acid) as an adhesion promoter for UV-assisted thermoplastic bonding: Application for the in vitro construction of human blood vessels.

In this study, we introduced a novel adhesion bonding method for fabricating thermoplastic microdevices using poly(acrylic acid) (PAA) as a UV-assisted adhesion promoter. The bonding mechanism was based on the covalent cross-links between poly(methyl methacrylate) (PMMA) and PAA via the free radicals in their carbon backbone generated under UV irradiation. The water contact angle and Fourier-transformed infrared (FTIR) analysis were performed to analyze the surface characteristics of the PAA-coated PMMA. PMMAs were bonded under UV treatment for 60 s with the highest bond strength of around 1.18 MPa. The PMMA microdevice was leak-proof for over 200 h. Besides, clog-free PMMA microdevices with various-sizes microchannels were performed to demonstrate such a high applicable bonding method for microdevice fabrication. Moreover, PMMAs were bonded with other thermoplastics with a bond strength of around 0.5 MPa. Notably, collagen was easily coated inside the PMMA microchannels via electrostatic interaction between PAA and collagen which is beneficial for on-device cell culture. As a result, a layered co-culture model of smooth muscle cells (SMCs) and human umbilical vein endothelial cells (HUVECs) was realized inside simple straight microchannels mimicking human blood vessel wall. Therefore, the introduced bonding method could pave the way for fabricating microdevice for cell-related applications.

Chitosan-polydopamine hydrogel complex: a novel green adhesion agent for reversibly bonding thermoplastic microdevice and its application for cell-friendly microfluidic 3D cell culture.

Owing to biocompatible characteristics and supporting cell growth capability, hydrogels have been widely used for scaffold fabrication and surface coating for cell culture. To employ the advantages of hydrogels, in the present study, we introduce a biocompatible chitosan (CS)-polydopamine (pDA) hydrogel complex as a green adhesion agent for the reversible bonding of thermoplastics assisted by UV irradiation. Poly(methyl methacrylate) (PMMA) substrates were bonded due to the covalent bond network formed between the amine groups of either CS or pDA in the hydrogel complex and the aldehyde groups of the oxidized PMMA surface via the Schiff-base reaction during the UV irradiation. Furthermore, the introduced method allowed for reversible bonding, which is highly appropriate for the fabrication of microdevices for cell-related applications. Surface characterizations such as water contact angle measurement, scanning electron microscopy analysis (SEM), atomic force microscopy analysis (AFM), and Fourier-transform infrared microscopy analysis (FTIR) were performed to confirm the successful coating of the hydrogel complex on the PMMA surface. Moreover, the bonding between two PMMAs or PMMA with other thermoplastics was successfully investigated with high bond strengths ranging from 0.4 to 0.7 MPa. The potential for reversible bonding of this method was verified by repeating the bonding/debonding cycle of the bonded PMMAs for three times, which maintained the bond strength at approximately 0.5 MPa. The compatibility of the bonding method in biological applications was examined by culturing mesenchymal stem cells (MSCs) inside a microchannel where multiple uniform-sized MSC spheroids were successfully formed. Then, spheroids were harvested for off-chip experiments enabled by the reversibility of the introduced bonding strategy. The bonding strategy employing a green hydrogel complex as a cell-friendly and eco-friendly adhesion agent could have a high impact on the fabrication of microdevices suitable for advanced organ-on-a-chip studies.

Nanohybrid biodegradable scaffolds for TGF-ß3 release for the chondrogenic differentiation of human mesenchymal stem cells.

An ideal scaffold for bone tissue engineering should have chondroinductive, biodegradable, and biocompatible properties, as well as the ability to absorb and slowly release the biological molecules. In order to develop such a system to support bone tissue regeneration, in the present study, we developed a three-dimensional poly(L-lactic-co-glycolic acid) (PLGA)/Polycaprolactone (PCL) nanohybrid scaffold embedded with PLGA macroparticles (MPs) conjugated with TGF-ß3 for the growth and chondrogenic differentiation of human mesenchymal stem cells (hMSCs). First, a microfluidic device was used to fabricate porous PLGA MPs with the sizes ranging from 10 to 50 µm. Next, the PLGA MPs were loaded with TGF-ß3, mixed with PCL solution, and then electrospun to obtain PLGA-TGF-ß3 MPs/PCL nanohybrid scaffold. Our results demonstrated that PLGA MPs fabricated using a microfluidic-based approach exhibited enhanced conjugation of TGF-ß3 with over 80% loading efficiency and sustained release of TGF-ß3. Furthermore, the results of glycosaminoglycan (GAG) content measurement and Safranin O staining revealed that the PLGA-TGF-ß3 MPs and PLGA-TGF-ß3 MPs/PCL nanohybrid scaffold can promote the proliferation and chondrogenic differentiation of hMSCs in vitro. Therefore, the PLGA-TGF-ß3 MPs/PCL nanohybrid scaffold could pave the way for cartilage regeneration and have wide applications in regenerative medicine.

Microfluidic Approach to Generate a Tadpole-Egg-Shaped Alginate Fiber and Its Application in Tissue Engineering.

Herein, we introduce a facile microfluidic technique to produce a hybrid alginate fiber with a tadpole-egg shape. A triple-flow polydimethylsiloxane microfluidic device was constructed to allow the formation of oil droplets inside the alginate stream and was instantaneously gelated with the coaxially adjacent CaCl2. The fiber entrapping the uniform oil droplets was dehydrated, leading to the formation of a distinct tadpole-egg-shaped structure. A series of diverse fiber architectures was fabricated in a controlled manner based on the flow rates of the relevant flows. The tadpole-egg-shaped alginate fibers were employed as building blocks to create a three-dimensional microwell template for cell cultures. First, the tadpole-egg-shaped alginate fibers containing the oil droplets were half-dipped into a melted agarose solution. After the solidification of the agarose gel, the alginate fibers were degraded by an ethylenediaminetetraacetic acid (EDTA) solution to generate the hemispherical microwells. Mesenchymal stem cells (MSCs) were cultured in the microwells to generate spheroids, which were induced into chondrocytes using transforming growth factor-ß3. The formed MSC spheroids exhibited a relatively high ratio of cell viability with more than 95% live cells after 14 days of culture. The success of the chondrogenic differentiation was proven based on staining (Safranin O) and the glycosaminoglycan levels. The latter was significantly higher in spheroids that were induced to form chondrocytes compared to those that were not induced after 21 days of differentiation. Second, we investigated the potential of the tadpole-egg-shaped alginate fibers as microcarriers for applications in drug delivery and implantable technologies. It was revealed that the degradation of the Ca-alginate wall of the hybrid fibers to release the oil droplets required an EDTA solution with a concentration of 500 mM for a 15 min period. This result can be used to further develop the tadpole-egg-shaped alginate fibers as uniform microcarriers with multiple compartments.