Expansion of Human Neural Precursor Cells in Large‐Scale Bioreactors for the Treatment of Neurodegenerative Disorders

Abstract

The transplantation of in vitro expanded human neural precursor cells (hNPCs) represents a potential new treatment alternative for individuals suffering from incurable neurodegenerative disorders such as Parkinson's disease (PD) and Huntington's disease (HD). However, in order for cell restorative therapy to have widespread therapeutic significance, it will be necessary to generate unlimited quantities of clinical grade hNPCs in a standardized method. We report here that we have developed a serum‐free medium and scale‐up protocols that allow for the generation of clinical quantities of human telencephalon‐derived hNPCs in 500‐mL computer‐controlled suspension bioreactors. The average hNPC aggregate diameter in the bioreactors was maintained below a target value of 500 μm by controlling the liquid shear field. The human cells, which were inoculated at 105 cells/mL, exhibited a doubling time of 84 h, underwent a 36‐fold expansion over the course of 18 days, and maintained an average viability of over 90%. The bioreactor‐derived hNPCs retained their nestin expression following expansion and were able to differentiate into glial and neuronal phenotypes under defined conditions. It has also been demonstrated that these hNPCs differentiated to a GABAergic phenotype that has recently been shown to be able to restore functional behavior in rat models of HD and neuropathic pain (Mukhida, K. et al. Stem Cells 2007; DOI 10.1634/stemcells.2007–0326). This study demonstrates that clinical quantities of hNPCs can be successfully and reproducibly generated under standardized conditions in computer‐controlled suspension bioreactors.

Antitumor Effects of CD40 Ligand-Expressing Endothelial Progenitor Cells Derived From Human Induced Pluripotent Stem Cells in a Metastatic Breast Cancer Model

“…For clinical applications, more robust and costeffective cell expansion technologies will be required. In this aspect, expansion methods such as automated bioreactors and rotary culture machines hold promise for generating iPS cell derivatives in bulk [28][29][30][31].…”

Bioprocess Development for the Expansion of Embryonic Stem Cells

“…In smaller vessels, surface aeration as the main mode of oxygen supply is typically sufficient and commonly used due to its simplicity. However, as culture volumes increase and cell densities increase, the surface area to volume ratios decrease and surface aeration may no longer be sufficient to ensure oxygen transport to the cells (Gilbertson et al, 2006;Baghbaderani et al, 2008). Options to increase oxygen transport include increasing agitation rate of the impeller, sparging, or medium additives such as Perfluorocarbons (PFCs) which increase oxygen solubility within the liquid medium.…”

“…There have been thorough reviews elsewhere which have described several approaches to experimentally determine k L a (Garcia-Ochoa & Gomez., 2009). One example that we have used, described by Baghbaderani et al (2008), is as follows. Briefly, the assumption is made that at some point in time the reactor reaches steady state during which any oxygen entering the medium would be immediately consumed by the cells (that is, C O2 equals zero in the medium).…”

“…This method, however, obviously does not take into account any dimensional effects of the system. As such, Baghbaderani et al (2008) have recommended a correlation put forth by Aunins et al (1989) for a 500mL Corning spinner which takes the following form:…”

Small-Scale Bioreactors for the Culture of Embryonic Stem Cells

“…High-throughput cell culture experiments at a more economical scale could then be performed for culture optimisation (Kostov et al 2001) and allow for factorial experimental design. While substantial research has been completed in the area of scaled-up production (Baghbaderani et al 2008;Fernandes-Platzgummer et al 2011;Garcia-Ochoa and Gomez 2009;Gilbertson et al 2006;Park et al 2010;Youn et al 2005;Yu et al 2009) to increase cell cultures to clinically meaningful numbers such as the estimated 1 billion cells per patient required for stem cell therapy (Docherty et al 2007), there has been little development in scaling down mammalian cell bioreactor systems. Many scaled-down bioreactors in the literature have been developed to culture Escherichia coli (bacteria) or Saccharomyces cerevisiae (yeast).…”