4D-bioprinting of vascularized bone tissue and evaluation of blood vessel and bone formation in an orthotopic bone defect model

• Applicants: Professor Dr. Günter Finkenzeller (†); Dr. Peter Koltay
• Subject Area: Orthopaedics, Traumatology, Reconstructive Surgery ; Biological Process Engineering
• Term: from 2015 to 2023
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 263422750

In the first funding period, we developed important basics for the bioprinting of vascularized bone tissue. In this context, we identified two suitable hydrogels, which were able to support blood vessel formation starting from bioprinted endothelial cells (HUVECs) and osteogenic differentiation of mesenchymal stem cells (MSCs), respectively. Moreover, we developed a 3D bioprinter which combines two bioprinting technologies, namely Drop on Demand (DoD) and extrusion printing.Using this bioprinter, we printed stabile cubes with edge lengths of 1 cm. In these cubes, the vascular component was realized by DoD printing of high density cell suspensions of HUVECs in fibrin-hydrogels, whereas the bone component was realized by extrusion printing of MSCs in a hydrogel composed of fibrin, gelatin, hyaluronic acid, glycerol and hydroxyapatite (osteo-Hydrogel). We have been able to show in vitro, as well as in vivo in a subcutaneous implantation model, that the bioprinted HUVECs were able to form blood vessels and the bioprinted MSCs were able to form a calcified bone matrix.The elastic modulus of the bioprinted cubes was around 1 kPa, which corresponds to native human soft tissue. However, the rigidity of human native bone is around 1x105 kPA and therefore about 100.000 times higher than the actual E-modulus of our current constructs.Therefore, on major goal of this continued project is to develop a combined printing procedure for printing cell-containing hydrogels and stability-generating thermoplastics and/or calcium phosphate cements (CPC) in order to produce artificial vascularized bone tissue with a physiological relevant rigidity which should resemble human native bone. The second goal is the implementation of the so called 4D-bioprinting (“time” as the fourth dimension) in which the temporal and spatial maturation of the construct should be controlled by the spatially resolved printing of growth factors, differentiation factors and/or additional cell entities. The third major goal of the continued project is the evaluation of the 4D-combination constructs in a physiologically-relevant orthotopic bone defect model in terms of vascularization and bone formation. In this context, we also want to investigate whether the quantity and/or quality of blood vessel formation and bone formation can be steered via modulation of the elastic moduli of the printed constructs.

• DFG Programme: Research Grants