Cytocompatibility and bioactivity testing in vitro

该子项目的目的是开发一种标准化的基于细胞的测定方法，以测试新植入材料的适用性。对于测试程序，我们将使用间充质干细胞来评估材料成分的生物相容性。此外，我们将开发策略，以改善细胞接种和附着，以及静态和动态培养技术与细胞基质结构的生物力学刺激，以模拟生理培养条件。两组将使用相同的协议分离和培养间充质干细胞。为了获得足够的细胞数量用于检测，将建立并优化动态扩增技术，如Wave技术（混合系统上的Cellbag生物反应器）。除了基于微载体的技术之外，我们还打算使用无颗粒微聚集体培养技术。此外，我们还将研究低氧（5%O2）对骨髓间充质干细胞增殖、分化和衰老的影响，并探讨细胞在不同材料上的接种和贴壁的优化策略。除了细胞对表面特性的附着外，我们还将研究细胞在基质中的向内生长。然后将细胞接种的构建体在专门的生物反应器系统中在动态条件下培养，并分化成特定的组织类型，如肌腱或骨，用于植入物的功能测试。在本项目的这一部分，我们将集中于TP1，TP2和TP5的蛋白质功能化基质与间充质干细胞接种，我们将测试蛋白质对细胞分化的影响。同时，我们将在静态长期培养物中测试不同灭菌方法的影响和效率。

Aim of this subproject is the development of a standardized cell-based assay method in or-der to test the suitability of new implant materials For the test procedure we will use mesenchymal stem cells in order to evaluate the biocompatibility of the material components. Furthermore, we will develop strategies to improve cell seeding and attachement, as well as static and dynamic culture techniques together with biomechanical stimulation of the cell-matrix-constructs in order to mimic physiological culture conditions.Both groups will isolate and characterise mesenchymal stem cells using identical protocols. In order to achieve sufficient cell numbers for testing, dynamic expansion techniques such as the Wave technology (cellbag bioreactors on mixing systems) will be established and optimized. Be-sides of microcarrier-based technologies we intend to use also particle-free microaggregate culture techniques. Besides we will study the influence of hypoxia (5% O2) on proliferation, differentiation and senescence of mesenchymal stem cells.Furthermore we will work on strategies to optimize cell seeding and attachment on different materials. Beside of cellular attachment to surface properties we will study the cellular ingrowth in-side the matrices. The cell-seeded constructs will then be cultured under dynamic conditions in specialized bioreactor systems and differentiated into the particular tissue types, such as tendon or bone, for functional testing of the implant. In this part of the project we will concentrate on protein-functionalized matrices from TP1, TP2, and TP5 seeded with mesenchymal stem cells and we will test the effects of proteins on cell differentiation. In parallel, we will test impact and efficiency of different sterilization methods in static long-term cultures.