Stirred culture of peripheral and cord blood hematopoietic cells offers advantages over traditional static systems for clinically relevant applications

Abstract

The ability to culture hematopoietic cells in readily characterizable and scalable stirred systems, combined with the capability to utilize serum‐free medium, will aid the development of clinically attractive bioreactor systems for transplantation therapies. We thus examined the proliferation and differentiation characteristics of peripheral blood (PB) mononuclear cells (MNC), cord blood (CB) MNC, and PB CD34+ cells in spinner flasks and (control) T‐flask cultures in both serum‐containing and serum‐free media. Hematopoietic cultures initiated from all sources examined (PB MNC, CB MNC, and PB CD34+ cells) grew well in spinner vessels with either serum‐containing or serum‐free medium. Culture proliferation in spinner flasks was dependent on both agitator design and RPM as well as on the establishment of critical inoculum densities (ID) in both serum‐containing (2 × 105 MNC/mL) and serum‐free (3 × 105 MNC/mL) media. Spinner flask culture of PB MNC in serum‐containing medium provided superior expansion of total cells and colony‐forming cells (CFC) at high ID (1.2 × 106 cells/mL) as compared to T‐flask controls. Serum‐free spinner culture was comparable, if not superior, to that observed in serum‐containing medium. This is the first report of stirred culture of PB or CB MNC, as well as the first report of stirred CD34+ cell culture. Additionally, this is the first account of serum‐free stirred culture of hematopoietic cells from any source. © 1998 John Wiley & Sons, Inc. Biotechnol Bioeng 59: 534–543, 1998.

Damage mechanisms of suspended animal cells in agitated bioreactors with and without bubble entrainment

“…While the subject matter has now largely reached maturity in the context of large-scale CHO cultures, the emergence of stem-cell bioengineering will likely require revisiting the issue, since both microcarrier (Abranches et al, 2007) and free-suspension (Collins et al, 1998;Kirouac and Zandstra, 2008) bioreactors are now contemplated. Whether adult or embryonic-stem cells, these cells are considerably more sensitive to process parameters, such as mixing, additives, or oxidative damage.…”

Geometric Control of Cardiomyogenic Induction in Human Pluripotent Stem Cells

“…The ability to generate large numbers of HPS cell-derived differentiated cells is critical to developing these strategies. Stirred suspension cultures are a popular approach to scale-up static dish-based cell cultures because they improve culture homogeneity, permit online monitoring and control of pH and dissolved oxygen concentration, and continuous medium perfusion can be easily enabled (Collins, Miller et al 1998;Madlambayan, Rogers et al 2001;Cabrita, Ferreira et al 2003;Kwon, Kim et al 2003;Zandstra, Bauwens et al 2003;Dang, Gerecht-Nir et al 2004;Bauwens, Yin et al 2005). …”

Agitation increases expansion of cord blood hematopoietic cells and promotes their differentiation into myeloid lineage

“…The resultant cells were suspended in the cell culture medium at density of 10 6 cells/ml in T-150 flasks (Falcon, Corning, Tewksbury, MA, USA) and incubated for 24 h at 37°C in a humidified incubator with 5 % CO 2 , before being entered into either suspension or static culture systems. This preincubation was done to accustom the cells to the culture conditions before facing high damages resulted from fluid mechanics (Collins et al 1998a;Liu et al 2006). …”

“…To date, proliferation and differentiation potential of mononuclear cells (MNCs) derived from different sources in diverse suspension cultures have been investigated and comparisons between static and dynamic cultures have been made (Collins et al 1998a;Kwon et al 2003;Shayan et al 2012;Yang et al 2008). However, the accurate influence of shear stress on UCB-MNCs expansion and their functional characteristics still remains undetermined.…”