A hydrogel model incorporating 3D-plotted hydroxyapatite for osteochondral tissue engineering

The concept of biphasic or multi-layered compound scaffolds has been explored within numerous studies in the context of cartilage and osteochondral regeneration. To date, no system has been identified that stands out in terms of superior chondrogenesis, osteogenesis or the formation of a zone of calcified cartilage (ZCC). Herein we present a 3D plotted scaffold, comprising an alginate and hydroxyapatite paste, cast within a photocrosslinkable hydrogel made of gelatin methacrylamide (GelMA), or GelMA with hyaluronic acid methacrylate (HAMA). We hypothesized that this combination of 3D plotting and hydrogel crosslinking would form a high fidelity, cell supporting structure that would allow localization of hydroxyapatite to the deepest regions of the structure whilst taking advantage of hydrogel photocrosslinking. We assessed this preliminary design in terms of chondrogenesis in culture with human articular chondrocytes, and verified whether the inclusion of hydroxyapatite in the form presented had any influence on the formation of the ZCC. Whilst the inclusion of HAMA resulted in a better chondrogenic outcome, the effect of HAP was limited. We overall demonstrated that formation of such compound structures is possible, providing a foundation for future work. The development of cohesive biphasic systems is highly relevant for current and future cartilage tissue engineering

A Novel Plasma-Based Bioink Stimulates Cell Proliferation and Differentiation in Bioprinted, Mineralized Constructs

Extrusion-based bioprinting, also known as 3D bioplotting, is a powerful tool for the fabrication of tissue equivalents with spatially defined cell distribution. Even though considerable progress has been made in recent years, there is still a lack of bioinks which enable a tissue-like cell response and are plottable at the same time with good shape fidelity. Herein, we report on the development of a bioink which includes fresh frozen plasma from full human blood and thus a donor/patient-specific protein mixture. By blending of the plasma with 3 w/v% alginate and 9 w/v% methylcellulose, a pasty bioink (plasma-alg-mc) was achieved, which could be plotted with high accuracy and furthermore allowed bioplotted mesenchymal stromal cells (MSC) and primary osteoprogenitor cells to spread within the bioink. In a second step, the novel plasma-based bioink was combined with a plottable self-setting calcium phosphate cement (CPC) to fabricate bone-like tissue constructs. The CPC/plasma-alg-mc biphasic constructs revealed open porosity over the entire time of cell culture (35 d), which is crucial for bone tissue engineered grafts. The biphasic structures could be plotted in volumetric and clinically relevant dimensions and complex shapes could be also generated, as demonstrated for a scaphoid bone model. The plasma bioink potentiated that bioplotted MSC were not harmed by the setting process of the CPC. Latest after 7 days, MSC migrated from the hydrogel to the CPC surface, where they proliferated to 20-fold of the initial cell number covering the entire plotted constructs with a dense cell layer. For bioplotted and osteogenically stimulated osteoprogenitor cells, a significantly increased alkaline phosphatase activity was observed in CPC/plasma-alg-mc constructs in comparison to plasma-free controls. In conclusion, the novel plasma-alg-mc bioink is a promising new ink for several forms of bioprinted tissue equivalents and especially gainful for the combination with CPC for enhanced, biofabricated bone-like constructs.</p

Endosteal and Perivascular Subniches in a 3D Bone Marrow Model for Multiple Myeloma

The bone marrow microenvironment is the preferred location of multiple myeloma, supporting tumor growth and development. It is composed of a collection of interacting subniches, including the endosteal and perivascular niche. Current in vitro models mimic either of these subniches. By developing a model combining both niches, this study aims to further enhance the ability to culture primary myeloma cells in vitro. Also, the dependency of myeloma cells on each niche was studied. A 3D bone marrow model containing two subniches was created using 3D bioprinting technology. We used a bioprintable pasty calcium phosphate cement (CPC) scaffold with seeded osteogenic multipotent mesenchymal stromal cells (O-MSCs) to model the endosteal niche, and Matrigel containing both endothelial progenitor cells (EPCs) and MSCs to model the perivascular niche. Within the model containing one or both of the niches, primary CD138+ myeloma cells were cultured and analyzed for both survival and proliferation. The 3D bone marrow model with combined subniches significantly increasing the proliferation of CD138+ myeloma cells compared to both environments separately. The developed model showed an essential role of the perivascular niche over the endosteal niche in supporting myeloma cells. The developed model can be used to study the expansion of primary myeloma cells and their interactions with varying bone marrow subniches

Endosteal and Perivascular Subniches in a 3D Bone Marrow Model for Multiple Myeloma

The bone marrow microenvironment is the preferred location of multiple myeloma, supporting tumor growth and development. It is composed of a collection of interacting subniches, including the endosteal and perivascular niche. Current in vitro models mimic either of these subniches. By developing a model combining both niches, this study aims to further enhance the ability to culture primary myeloma cells in vitro. Also, the dependency of myeloma cells on each niche was studied. A 3D bone marrow model containing two subniches was created using 3D bioprinting technology. We used a bioprintable pasty calcium phosphate cement (CPC) scaffold with seeded osteogenic multipotent mesenchymal stromal cells (O-MSCs) to model the endosteal niche, and Matrigel containing both endothelial progenitor cells (EPCs) and MSCs to model the perivascular niche. Within the model containing one or both of the niches, primary CD138+ myeloma cells were cultured and analyzed for both survival and proliferation. The 3D bone marrow model with combined subniches significantly increasing the proliferation of CD138+ myeloma cells compared to both environments separately. The developed model showed an essential role of the perivascular niche over the endosteal niche in supporting myeloma cells. The developed model can be used to study the expansion of primary myeloma cells and their interactions with varying bone marrow subniches