Dynamic three-dimensional aggregate cultivation for the expansion of mesenchymal stem cells

Abstract: For the use of mesenchymal stem cells (MSCs) in cell based therapies extensive expansion is necessary to obtain sufficient cell number for patient treatments. This expansion phase still remains one of the key challenges since long-term cultivation and excessive passaging in two-dimensional conditions result in a loss of essential stem cell properties. These functional damages and alterations often result in low survival rate of cells, changes of surface marker profiles, and reduced differentiation capacity, which also influences the success rate in clinical applications. The cultivation of MSCs in three-dimensional aggregates positively influences stem cell properties, thus large scale cultivation of MSC aggregates is highly desirable. It is well known that MSC physiologically reside in 3 D microenvironment under hypoxic conditions. “Classical” cell culture protocols do not mimic these requirements resulting in loss of functional properties and genetic instability. Therefore, we developed a scaffold-free aggregate cultivation process for the expansion of human MSCs in a stirred tank reactor under hypoxic conditions [1]. According to computational fluid dynamics calculations the aggregates were exposed to comparatively high average shear stress of 0.2 Pa. However, the cells exposed a viability of 78 – 86% and maintained their surface marker profile and differentiation potential after cultivation. From these initial experiments, we postulate that expansion of MSC in 3D aggregates in stirred tank bioreactors is suitable for large-scale production of MSCs for cell based therapy applications.