Bioreactor expansion of human neural precursor cells in serum‐free media retains neurogenic potential

Abstract

Human neural precursor cells (hNPCs), harvested from somatic tissue and grown in vitro, may serve as a source of cells for cell replacement strategies aimed at treating neurodegenerative disorders such as Parkinson's disease (PD), Huntington's disease (HD), and intractable spinal cord pain. A crucial element in a robust clinical production method for hNPCs is a serum-free growth medium that can support the rapid expansion of cells while retaining their multipotency. Here, we report the development of a cell growth medium (PPRF-h2) for the expansion of hNPCs, achieving an overall cell-fold expansion of 10(13) over a period of 140 days in stationary culture which is significantly greater than other literature results. More importantly, hNPC expansion could be scaled-up from stationary culture to suspension bioreactors using this medium. Serial subculturing of the cells in suspension bioreactors resulted in an overall cell-fold expansion of 7.8 x 10(13) after 140 days. These expanded cells maintained their multipotency including the capacity to generate large numbers of neurons (about 60%). In view of our previous studies regarding successful transplantation of the bioreactor-expanded hNPCs in animal models of neurological disorders, these results have demonstrated that PPRF-h2 (containing dehydroepiandrosterone, basic fibroblast growth factor and human leukemia inhibitory factor) can successfully facilitate the production of large quantities of hNPCs with potential to be used in the treatment of neurodegenerative disorders.

Secretome of Undifferentiated Neural Progenitor Cells Induces Histological and Motor Improvements in a Rat Model of Parkinson's Disease

“…hNPCs cell culture was performed as previously described by our group , and according to the protocols and strict ethical guidelines previously established and approved by the Conjoint Health Research Ethics Board (CHREB, University of Calgary, AB, ID: E‐18786) . Regarding conditioned media (CM) collection, at passage 5 (P5), hNPCs were enzymatically dissociated and plated into new tissue culture flasks at a density of 5,000 cells/cm 2 for experiments, and 12,000 cells/cm 2 for proteomic procedures.…”

Modulation of the Mesenchymal Stem Cell Secretome Using Computer-Controlled Bioreactors: Impact on Neuronal Cell Proliferation, Survival and Differentiation

“…Ethical consent was approved by the Conjoint Health Research Ethics Board (CHREB), University of Calgary (ID: E-18786). So, pre-isolated and cryopreserved hNPCs were thawed and placed into a Nunc T-25 flask containing 5 mL of a serum-free medium PPRF-h2 (described in detail in52). After two days, cells were harvested and mechanically dissociated into a single cell suspension, and subcultured into fresh PPRF-h2.…”

Multiplex mapping of chromatin accessibility and DNA methylation within targeted single molecules identifies epigenetic heterogeneity in neural stem cells and glioblastoma

“…Cell culture NSC and GBM spheroid cultures were derived and maintained as previously described (Deleyrolle and Reynolds 2009;Baghbaderani et al 2010;Deleyrolle et al 2011).…”