in terms of GMP requirements, alternative procedures and cultivation systems, like
the spheroid- or microcarrier-based expansion in stirred single-use bioreactors, are
said to be the platforms for future cell therapeutic productions (see Sect. 2.2).
2.2 Dynamic Approach (3D Cultures)
As mentioned in Sect. 2.1, hMSCs are typically expanded under adherent conditions
as a monolayer in 2D culture systems. However, isolation and growth of hMSCs on
rigid tissue culture plastic have been described as promoting spreading of cells rich
in actin-myosin stress fibers [23, 24]. Indeed, the static 2D culture systems represent
an artificial environment which significantly differs from those of the MSC in vivo
niche. Therefore, different efforts have been made over the years to establish
dynamic 3D culture systems working with spheroids (see Sect. 2.2.1) or
microcarriers (see Sect. 2.2.2). In dynamic bioreactor systems (stirred, wavemixed, orbitally shaken, hollow fiber and fixed bed types), the culture medium is
continuously agitated to provide a uniform environment, preventing the formation of
physiochemical gradients and improving mass and heat transfer. Special attention is
currently being paid to SU versions, which significantly improve patient safety
[25]. Even though different studies have recently shown the applicability of SU
systems for MC-based hMSC production processes, challenges still exist.
For this reason, it makes sense to characterize the different bioreactor systems
using appropriate process engineering and cell cultivation technique methods prior
to usage or during process development, simultaneously assisting in the development of a “Digital Twin.” Several studies have been published that provide engineering parameters relating to mixing time, oxygen mass transfer, and power input
for various SU bioreactor types. However, when considering the heterogeneous
distribution of MCs, spheroids and hydrodynamics, and a detailed analysis of the
fluid flow pattern, the MC distribution and the cell growth become worthwhile.
Numerical methods, such as Computational Fluid Dynamics (CFD) and kinetic
growth models, are complementary methods to the experimental investigations
and increase the process knowledge of hMSC production methods. Thus, numerical
models can be used to support process development and scale-up.
2.2.1 Growth in Spheroids
hMSCs are often expanded in stirred SU bioreactors as self-assembling cell aggregates or spheroids that mimic the in situ conditions. Thus, compared to 2D monolayer cultures, 3D structures consisting of multiple cell-to-cell contact points are
obtained. However, due to their heterogeneous nature, spheroids have been more
successfully employed to study complex 3D cell structures and cell differentiation
[26] than for hMSC mass expansion in stirred SU bioreactors, as indicated by the
limited number of publications in this area (see Table 3).
Numerical Methods for the Design and Description of In Vitro Expansion. . .
193
the spheroid- or microcarrier-based expansion in stirred single-use bioreactors, are
said to be the platforms for future cell therapeutic productions (see Sect. 2.2).
2.2 Dynamic Approach (3D Cultures)
As mentioned in Sect. 2.1, hMSCs are typically expanded under adherent conditions
as a monolayer in 2D culture systems. However, isolation and growth of hMSCs on
rigid tissue culture plastic have been described as promoting spreading of cells rich
in actin-myosin stress fibers [23, 24]. Indeed, the static 2D culture systems represent
an artificial environment which significantly differs from those of the MSC in vivo
niche. Therefore, different efforts have been made over the years to establish
dynamic 3D culture systems working with spheroids (see Sect. 2.2.1) or
microcarriers (see Sect. 2.2.2). In dynamic bioreactor systems (stirred, wavemixed, orbitally shaken, hollow fiber and fixed bed types), the culture medium is
continuously agitated to provide a uniform environment, preventing the formation of
physiochemical gradients and improving mass and heat transfer. Special attention is
currently being paid to SU versions, which significantly improve patient safety
[25]. Even though different studies have recently shown the applicability of SU
systems for MC-based hMSC production processes, challenges still exist.
For this reason, it makes sense to characterize the different bioreactor systems
using appropriate process engineering and cell cultivation technique methods prior
to usage or during process development, simultaneously assisting in the development of a “Digital Twin.” Several studies have been published that provide engineering parameters relating to mixing time, oxygen mass transfer, and power input
for various SU bioreactor types. However, when considering the heterogeneous
distribution of MCs, spheroids and hydrodynamics, and a detailed analysis of the
fluid flow pattern, the MC distribution and the cell growth become worthwhile.
Numerical methods, such as Computational Fluid Dynamics (CFD) and kinetic
growth models, are complementary methods to the experimental investigations
and increase the process knowledge of hMSC production methods. Thus, numerical
models can be used to support process development and scale-up.
2.2.1 Growth in Spheroids
hMSCs are often expanded in stirred SU bioreactors as self-assembling cell aggregates or spheroids that mimic the in situ conditions. Thus, compared to 2D monolayer cultures, 3D structures consisting of multiple cell-to-cell contact points are
obtained. However, due to their heterogeneous nature, spheroids have been more
successfully employed to study complex 3D cell structures and cell differentiation
[26] than for hMSC mass expansion in stirred SU bioreactors, as indicated by the
limited number of publications in this area (see Table 3).
Numerical Methods for the Design and Description of In Vitro Expansion. . .
193