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Human adipose tissue-derived stromal/stem cells (hASC) represent a valuable source of cells for clinical applications, especially in the field of regenerative medicine. Therefore, it comes as no surprise that interest in hASCs has increased greatly over the last decade. However, in order to use hASCs successfully in clinical applications, in vitro expansion is required. Single-use bioreactors in combination with microcarriers (MC) have been shown to be suitable systems for this task (1-3). However, hASCs are prone to higher shear sensitivity than conventional cell lines (e.g. CHO, BHK) that are normally expanded in these systems. Hence, the goal of this study was to investigate the influence of different shear stress levels on the growth of hASCs in small scale single-use spinner flasks. For this purpose, Computational Fluid Dynamics simulations based on a Euler-Euler and Euler-Lagrange approach were performed to predict the hydrodynamic stresses (0.06 – 0.87 Pa), the residence times (0.4 – 7.3 s) and the circulation times (1.6 - 16.6 s) of the MCs in various high shear zones. The numerical findings were combined with experimental data from cultivation studies (0.29 – 1.1∙106 hASC/mL) in order to develop a segregated mathematical growth model for the prediction of MC-associated hASC growth in small scale single-use spinner flasks.

Cell-based therapies have become increasingly important in the field of regenerative medicine, as global revenues of approximately 1 billion US$ indicate. A special focus in the clinical field is placed on human mesenchymal stem cells (hMSCs), especially on those isolated from the stromal vascular fraction of the adipose tissue. Due to their good clinical efficacy and tolerability, they can be used for allogeneic cell therapies (e.g. Crohn’s disease, graft versus host disease, acute myocardial infarct). However, the required number of therapeutically active human adipose tissue-derived stromal/stem cells (hASCs) for allogeneic applications is in the range of trillion cells per year. Thus, alternatives to the 2-dimensional planar cultivation systems, typically applied to propagate hASCs are urgently required. Different studies have recently demonstrated the applicability of stirred single-use bioreactors as a promising alternative to the planar cultivation systems. In these dynamically mixed culture systems, the required growth surface for the adherent growing cells is provided by microcarriers (MCs), which are suspended in the bioreactor. Although adherent mammalian cells are well investigated in combination with MCs for the production of vaccines, working with hASCs is more complex due to their higher shear sensitivity. This raises the question of how the induced fluid shear stress affects the cell expansion, the cell quality and the formation of MC-cell agglomerates during the cultivation in dynamic bioreactors. For this purpose, MC-based cell expansions were performed in single-use spinner flasks at different impeller speeds below (25, 43 rpm) and above (90 and 120 rpm) the specific suspension criteria NS1u (49 rpm) and NS1 (63 rpm) for polystyrene-based MCs. Cell numbers, metabolites and the formation of MC-cell agglomerates were measured during the cell expansion process. Furthermore, the expression of standard surface markers (CD14, CD20, CD34, CD45, CD73, CD9