gms | German Medical Science

Objectives: 3D printing technologies offer a tremendous potential to produce patient-specific implants and treat critical-sized bone defects, which vary in size, shape, and clinical requirements. Although progress has been made for 3D printing of biomaterial-based bone constructs mostly lacking biological active material. For larger-sized bone implants however, an early biologization and vascularization is essential. In this context, bioprinting technologies enable the integration of vital cells or active growth factors into 3D printed constructs. In this study, we established bioinks for digital light based (DLP) bioprinting of bone constructs using methacrylated gelatine (GelMa) hydrogels as basis for the bioink. The bioink was modified using graphene oxide or calcium phosphate nanomaterials to modulate the mechanical and biofunctional properties of the constructs. The 3D model of the porous bone implant was designed computationally. Before bioprinting, human mesenchymal stem cells (hMSC) were integrated in the bioinks. After printing, the impact of bioinks on cell distribution, viability, cell proliferation and differentiation, as well as mechanical properties of constructs was compared over a period of 14 days.

Methods: The model was designed providing a cylindrical shape (d = 7 mm, h = 3.64 mm) and interconnected pores of different sizes. Different bioinks were formulated using GelMa, a photoinitiator and photoabsorber. To enhance osteoinductive properties, calcium phosphate (CaP) nano particles (< 150 nm) were blended to the bioink. In addition, the GelMa bioink was modified using graphene oxide (GO). For the bioprinting process the bioinks were loaded with 3 x 10^6 hMSC/ml bioink. Tissue constructs were printed by DLP using the Lumen X followed by evaluation of the constructs by DNA-quantification, confocal and scanning electron microscopy, real-time polymerase chain reaction (PCR), cryosections followed by alizarin red and sirius red staining and compressive strength testing.

Results and conclusion: In comparison to commercial bioinks, the cell viability was higher in the established GelMa bioink and all nanomaterial containing variations along with an even distribution of cells. Morphological data and DNA quantification indicated the highest cell vitality, respectively, proliferation over time in GelMa basic ink, as well as the best printing resolution of the constructs in comparison to the 3D model. CaP-modified GelMa bioink indicated the best differentiation of hMSC in terms of osteogenic gene expression and calcium deposition. GO increased the stiffness of the printed constructs in control samples printed without cells to analyze the impacts of the nanomaterials on the bioinks per se. However, in initial studies we also observed differences in the stiffness in samples with cells over time. Overall, nanomaterials showed diverse beneficial effects in functionalizing DLP printed bone constructs containing living osteogenic cells.