Search

Bioprinting is a promising tool to fabricate organized cartilage. This study aimed to investigate the printability of gelatin-methacryloyl/gellan gum (gelMA/gellan) hydrogels with and without methacrylated hyaluronic acid (HAMA), and to explore (zone-specific) chondrogenesis of chondrocytes, articular cartilage progenitor cells (ACPCs), and multipotent mesenchymal stromal cells (MSCs) embedded in these bio-inks. The incorporating of HAMA in gelMA/gellan bio-ink increased filament stability, as measured using a filament collapse assay, but did not influence (zone-specific) chondrogenesis of any of the cell types. Highest chondrogenic potential was observed for MSCs, followed by ACPCs, which displayed relatively high proteoglycan IV mRNA levels. Therefore, two-zone constructs were printed with gelMA/gellan/HAMA containing ACPCs in the superficial region and MSCs in the middle/deep region. Chondrogenic differentiation was confirmed, however, printing influence cellular differentiation. ACPC- and MSC-laden gelMA/gellan/HAMA hydrogels are of interest for the fabrication of cartilage constructs. Nevertheless, this study underscores the need for careful evaluation of the effects of printing on cellular differentiation.

Clinical implementation of endochondral bone regeneration (EBR) would benefit from the engineering of devitalized cartilaginous constructs of allogeneic origins. Nevertheless, development of effective devitalization strategies that preserves extracellular matrix (ECM) is still challenging. The aim of this study is to investigate EBR induced by devitalized, soft callus-mimetic spheroids. To challenge the translatability of this approach, the constructs are generated using an allogeneic cell source. Neo-bone formation is evaluated in an immunocompetent rat model, subcutaneously and in a critical size femur defect. Living spheroids are used as controls. Also, the effect of spheroid maturation towards hypertrophy is evaluated. The devitalization procedure successfully induces cell death without affecting ECM composition or bioactivity. In vivo, a larger amount of neo-bone formation is observed for the devitalized chondrogenic group both ectopically and orthotopically. In the femur defect, accelerated bone regeneration is observed in the devitalized chondrogenic group, where defect bridging is observed 4 weeks post-implantation. The authors' results show, for the first time, a dramatic increase in the rate of bone formation induced by devitalized soft callus-mimetics. These findings pave the way for the development of a new generation of allogeneic, "off-the-shelf" products for EBR, which are suitable for the treatment of every patient.

Hydrogels based on triblock copolymers of polyethylene glycol and partially methacrylated poly[N-(2-hydroxypropyl) methacrylamide mono/dilactate] make up an attractive class of biomaterials because of their biodegradability, cytocompatibility, and tunable thermoresponsive and mechanical properties. If these properties are fine-tuned, the hydrogels can be three-dimensionally bioprinted, to generate, for instance, constructs for cartilage repair. This study investigated whether hydrogels based on the polymer mentioned above with a 10% degree of methacrylation (M10P10) support cartilage formation by chondrocytes and whether the incorporation of methacrylated chondroitin sulfate (CSMA) or methacrylated hyaluronic acid (HAMA) can improve the mechanical properties, long-term stability, and printability. Chondrocyte-laden M10P10 hydrogels were cultured for 42 days to evaluate chondrogenesis. M10P10 hydrogels with or without polysaccharides were evaluated for their mechanical properties (before and after UV photo-cross-linking), degradation kinetics, and printability. Extensive cartilage matrix production occurred in M10P10 hydrogels, highlighting their potential for cartilage repair strategies. The incorporation of polysaccharides increased the storage modulus of polymer mixtures and decreased the degradation kinetics in cross-linked hydrogels. Addition of HAMA to M10P10 hydrogels improved printability and resulted in three-dimensional constructs with excellent cell viability. Hence, this novel combination of M10P10 with HAMA forms an interesting class of hydrogels for cartilage bioprinting.

Gelatine methacryloyl (GelMA) hydrogels are widely used in studies aimed at cartilage regeneration. However, the endotoxin content of commercially available GelMAs and gelatines used in these studies is often overlooked, even though endotoxins may influence several cellular functions. Moreover, regulations for clinical use of biomaterials dictate a stringent endotoxin limit. We determined the endotoxin level of five different GelMAs and evaluated the effect on the chondrogenic differentiation of equine mesenchymal stromal cells (MSCs). Cartilage-like matrix production was evaluated by biochemical assays and immunohistochemistry. Furthermore, equine peripheral blood mononuclear cells (PBMCs) were cultured on the hydrogels for 24 h, followed by the assessment of tumour necrosis factor (TNF)-&alpha, and C&ndash, C motif chemokine ligand (CCL)2 as inflammatory markers. The GelMAs were found to have widely varying endotoxin content (two with &gt, 1000 EU/mL and three with &lt, 10 EU/mL), however, this was not a critical factor determining in vitro cartilage-like matrix production of embedded MSCs. PBMCs did produce significantly higher TNF-&alpha, and CCL2 in response to the GelMA with the highest endotoxin level compared to the other GelMAs. Although limited effects on chondrogenic differentiation were found in this study, caution with the use of commercial hydrogels is warranted in the translation from in vitro to in vivo studies because of regulatory constraints and potential inflammatory effects of the content of these hydrogels.

Hydrogels can provide a suitable environment for tissue formation by embedded cells, which makes them suitable for applications in regenerative medicine. However, hydrogels possess only limited mechanical strength, and must therefore be reinforced for applications in load-bearing conditions. In most approaches the reinforcing component and the hydrogel network have poor interactions and the synergetic effect of both materials on the mechanical properties is not effective. Therefore, in the present study, a thermoplastic polymer blend of poly(hydroxymethylglycolide-co-ε-caprolactone)/ poly(ε-caprolactone) (pHMGCL/PCL) was functionalized with methacrylate groups (pMHMGCL/PCL) and covalently grafted to gelatin methacrylamide (gelMA) hydrogel through photopolymerization. The grafting resulted in an at least fivefold increase in interface-binding strength between the hydrogel and the thermoplastic polymer material. GelMA constructs were reinforced with three-dimensionally printed pHMGCL/PCL and pMHMGCL/PCL scaffolds and tested in a model for a focal articular cartilage defect. In this model, covalent bonds at the interface of the two materials resulted in constructs with an improved resistance to repeated axial and rotational forces. Moreover, chondrocytes embedded within the constructs were able to form cartilage-specific matrix both in vitro and in vivo. Thus, by grafting the interface of different materials, stronger hybrid cartilage constructs can be engineered. © 2014 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.