Hydrodynamic effects on BHK cells grown as suspended natural aggregates

Abstract

Baby hamster kidney (BHK) cell aggregates grown in stirred vessels with different working volumes and impeller sizes were characterized. Using batch cultures, the range of agitation rates studied (25-100 rpm) led to aggregates with maximum sizes of 150 mum. Necrotic centers were not observed and cell specific productivity was independent of aggregate size. High cell viability was found for both single and adherent cells without an increase in cell death when agitation rate was increased. The increase in agitation rate affected aggregates by reducing their size and increasing their concentration and cell concentration in aggregates, while increasing the fraction of free cells in suspension. The experimental relationship between aggregate size and power dissipation rate per unit of mass was close to -1/4, suggesting a correlation with a critical turbulence microscale; this was independent of vessel scale and impeller geometry over the range investigated. Viscous stresses in the viscous dissipation subrange (below Kolmogoroff eddies) appear to be responsible for aggregate breakage. Under intense agitation BHK cells grown in the absence of microcarriers existed as aggregates without cell damage, whereas cells grown on the surface of microcarriers were largely reduced. This is a clear advantage for scaleup purposes if aggregates are used as a natural immobilization system in stirred vessels. (c) 1995 John Wiley & Sons, Inc.

Large-Scale Expansion of Mammary Epithelial Stem Cell Aggregates in Suspension Bioreactors

“…Mammospheres were observed in suspension cultures 1 day after inoculation and continued to grow in size until they reached 40-80 µm in diameter, as seen in Figure 2. Similar observations have been found in a number of other aggregate suspension cultures, including neural stem cells, embryonic stem cells, and BHK cells, where aggregates rapidly formed and continued to grow in size throughout the growth phase (21,32). Though mammospheres were also observed in stationary cultures, they were found to be much smaller and resembled cell clusters rather than spherical aggregates.…”

Shear‐Controlled Single‐Step Mouse Embryonic Stem Cell Expansion and Embryoid Body–Based Differentiation

“…To control aggregate size, adequate shear must be exerted to remove the outer cells of an aggregate (6.5 dyn/cm 2 ) [11], while excessive shear stress (15-30 dyn/cm 2 ) may result in cell damage [12,13]. In a similar system, the shear stress exerted at 60 rpm was reported to be 6.25 dyn per cm 2 [8], which may not be enough to effectively dislodge the outer cells of the aggregates, resulting in large aggregate sizes that may lead to nutrient depletion, causing variable and reduced cell growth.…”

“…The control of aggregate size is important in the regulation of cell agglomeration [1] because the centers of very large aggregates (>300 μm) may experience cell death due to limitations in nutrients and oxygen delivery [10]. It has been reported that the minimum shear needed to remove cells cultured on surfaces is 6.5 dyn per cm 2 [11], whereas shear stress levels in the range of 15-30 dyn per cm 2 are shown to cause damage to attached cells [12,13]. Therefore, a culture system that can control aggregate size effectively must exert a shear stress that falls within the aforementioned ranges.…”

Inoculation and growth conditions for high-cell-density expansion of mammalian neural stem cells in suspension bioreactors

“…The pH decreases during experiments using mammalian cells as a result of a buildup of toxic metabolites such as lactate (Xie and Wang, 1996). Increased agitation also causes damage to cells; however, when cells are grown as aggregates, high cell densities can be obtained at high agitation rates (Moreira et al, 1995). Most important, the aggregate microenvironment provides shear protection for the mammalian cells.…”