Gtn-HPA as a new biomaterial in dental reconstructive surgery - the effect of the injectable hydrogel on the vitality, on the cytokine expression, on the differentiation and on the vascularization of endothelial progenitor cells and osteoblasts in 2D and

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The development of new bone substitute materials (BSM) is of high scientific and clinical interest in dental surgery as they represent an attractive alternative to autogenous bone grafts. Early vascularization of the BSM is an important precondition for bone regeneration. Optimizing the BSM with respect to this process is widely seen as one of the key areas for possible future improvement to optimize healing. Regarding vascularization and osteoblast differentiation, preliminary studies have proven the high relevance of endothelial progenitor cells (EPC) and osteoblasts (OB). In addition, studies have shown that mechanical properties of BSM influence the function, proliferation and differentiation of these cells. In this context, the stiffness of the matrix seems to be on of the key components for bone regeneration.Hydrogels are already widely used as scaffolds for tissue engineering, because of their high permeability for oxygen, nutrients and other water-soluble metabolites through their water-content matrix. This matrix is an excellent environment for cell growth and tissue regeneration. However, a major drawback of existing injectable hydrogel systems is their missing control of the gelation rate which leads to suboptimal vascularization in hard hydrogels or missing mechanical stability in soft hydrogels. Recently, the injectable hydrogel gelatin-hydroxyphenylpropionic acid (Gtn-HPA) has been introduced by our cooperation partner from Harvard-MIT in the context of neural defects. During crosslinking, the concentrations of the enzyme (horseradish peroxidase, HRP) and the oxidant (hydrogen peroxide, H2O2) can independently be controlled to modify the gelation rate of the hydrogel. Thus, the gelation rate of this hydrogel can be fine-tuned for the first-time. With the vision to establish Gtn-HPA as a new BSM in reconstructive dental surgery, it is the aim of this study to evaluate Gtn-HPA as a matrix for the coculture of EPC and OB in 2D and 3D culture. The cytocompatibility of Gtn-HPA will be assessed with a viability assay. Furthermore, optimal stiffness of Gtn-HPA for maximal vascularization and osteogenesis will be examined. Therefore, an analysis of proliferation, differentiation and cytokine expression of EPC and OB in coculture will be performed. In addition, an examination of cell morphology and vascularization will be done by confocal-laser-scanning-microscopy (CLSM) and CD34 immunohistochemistry. By these analyses, a hydrogel system with tunable mechanical properties will be examined regarding the interaction of EPC and OB in 2D and 3D culture, to describe the effects of dimensionality and stiffness of BSM for bone regeneration for the first time and for prospective clinical use.