Search

The production of stem cells for clinical applications requires a suitable mass expansion and harvest process, which is implemented in a microcarrier-based bioreactor. Important parameters to consider include the choice of microcarriers for cell expansion, the growth medium and the scale-up strategy. We investigated six different microcarriers to determine whether they can support the growth and harvest of primary human mesenchymal stromal/stem cells (hMSCs) derived from bone marrow, and the immortalized cell line hMSC-TERT. A serum-containing medium (SCM) and a new chemically defined medium (CDM) were compared under dynamic culture conditions. We also investigated bead-to-bead transfer from spinner flasks to a stirred tank reactor as a scale-up strategy. We observed cell type-dependent differences in growth rate and attachment behavior on each microcarrier. Missing serum components in the CDM led to slower cell attachment and growth, whereas microcarriers suitable for the SCM were also suitable for CDM. Glass-coated microcarriers supported hMSC-TERT growth and bead-to-bead transfer in SCM, whereas plasma-treated plastic surfaces promoted cell growth in CDM. We demonstrated that stem cell cultures can be scaled up by bead-to-bead transfer, avoiding the need for pre-cultures in tissue flasks. The replacement of pre-cultures with a monitored bioprocess could therefore facilitate the development of conditions suitable for large-scale stem cell production. [ABSTRACT FROM AUTHOR]

Large-scale human mesenchymal stem cell expansion calls for a bioreaction system, that provides a sufficient growth surface. An alternative to static cultivations systems like cell factories are disposable stirred tank reactors. Here, microcarriers provide the required growth surface, but these make it difficult to achieve a complete homogenization in the bioreactor, while avoiding shear stress. To gain insight into this process, we investigated the impact of different power inputs (0.02–2.6 W m −3 ) on the mixing time (t m ). Whereas t m was inversely proportional to agitation in a one-phase-system, aeration resulted in a constant mixing time at 30–70 rpm. A high microcarrier concentration (30 g L −1 ) and low stirrer speed (30 rpm) in the liquid-solid system caused a 50-fold increase in t m and the formation of a discrete non-mixed upper zone. The effect of the microcarrier concentration on t m became negligible at higher stirrer speeds. In the three-phase system, microcarrier settling was prevented by aeration and a minimal specific power input of 0.6 W m −3 was sufficient for complete homogenization. We confirmed that a low power input during stem cell expansion leads to inhomogeneity, which has not been investigated in the three-phase system up to date. [ABSTRACT FROM AUTHOR]

Large-scale human mesenchymal stem cell expansion calls for a bioreaction system, that provides a sufficient growth surface. An alternative to static cultivations systems like cell factories are disposable stirred tank reactors. Here, microcarriers provide the required growth surface, but these make it difficult to achieve a complete homogenization in the bioreactor, while avoiding shear stress. To gain insight into this process, we investigated the impact of different power inputs (0.02–2.6 W m −3 ) on the mixing time (t m ). Whereas t m was inversely proportional to agitation in a one-phase-system, aeration resulted in a constant mixing time at 30–70 rpm. A high microcarrier concentration (30 g L −1 ) and low stirrer speed (30 rpm) in the liquid-solid system caused a 50-fold increase in t m and the formation of a discrete non-mixed upper zone. The effect of the microcarrier concentration on t m became negligible at higher stirrer speeds. In the three-phase system, microcarrier settling was prevented by aeration and a minimal specific power input of 0.6 W m −3 was sufficient for complete homogenization. We confirmed that a low power input during stem cell expansion leads to inhomogeneity, which has not been investigated in the three-phase system up to date. [ABSTRACT FROM AUTHOR]