Article

Bioprocessing of cryopreservation for large-scale banking of human pluripotent stem cells.

Department of Chemical and Biomedical Engineering, Florida State University , Tallahassee, Florida.
BioResearch open access 10/2012; 1(5):205-14. DOI: 10.1089/biores.2012.0224
Source: PubMed

ABSTRACT Human pluripotent stem cell (hPSC)-derived cell therapy requires production of therapeutic cells in large quantity, which starts from thawing the cryopreserved cells from a working cell bank or a master cell bank. An optimal cryopreservation and thaw process determines the efficiency of hPSC expansion and plays a significant role in the subsequent lineage-specific differentiation. However, cryopreservation in hPSC bioprocessing has been a challenge due to the unique growth requirements of hPSC, the sensitivity to cryoinjury, and the unscalable cryopreservation procedures commonly used in the laboratory. Tremendous progress has been made to identify the regulatory pathways regulating hPSC responses during cryopreservation and the development of small molecule interventions that effectively improves the efficiency of cryopreservation. The adaption of these methods in current good manufacturing practices (cGMP)-compliant cryopreservation processes not only improves cell survival, but also their therapeutic potency. This review summarizes the advances in these areas and discusses the technical requirements in the development of cGMP-compliant hPSC cryopreservation process.

Download full-text

Full-text

Available from: Yan Li, Jun 20, 2015
0 Followers
 · 
175 Views
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The effects of cryopreservation on mesenchymal stem cell (MSC) phenotype are not well documented; however this process is of increasing importance for regenerative therapies. This study examined the effect of cryopreservation (10% dimethyl-sulfoxide) on the morphology, viability, gene-expression and relative proportion of MSC surface-markers on cells derived from rat adipose, bone marrow and dental pulp. Cryopreservation significantly reduced the number of viable cells in bone marrow and dental pulp cell populations but had no observable effect on adipose cells. Flow cytometry analysis demonstrated significant increases in the relative expression of MSC surface-markers, CD90 and CD29/CD90 following cryopreservation. sqRT-PCR analysis of MSC gene-expression demonstrated increases in pluripotent markers for adipose and dental pulp, together with significant tissue-specific increases in CD44, CD73 and CD105 following cryopreservation. Cells isolated from different tissue sources did not respond equally to cryopreservation with adipose tissue representing a more robust source of MSCs.
    Cryobiology 08/2014; 69(2). DOI:10.1016/j.cryobiol.2014.08.003 · 1.64 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Neural progenitor cells (NPCs) are usually derived from pluripotent stem cells (PSCs) through the formation of embryoid bodies (EBs), the 3-D aggregate-like structure mimicking embryonic development. Cryo-banking of EBs is a critical step for sample storage, process monitoring, and preservation of intermediate cell populations during the lengthy differentiation procedure of PSCs. However, the impact of microenvironment (including 3-D cell organization and biochemical factors) of EBs on neural lineage commitment post-cryopreservation has not been well understood. In this study, intact EBs (I-E) and dissociated EBs (D-E) were compared for the recovery and neural differentiation after cryopreservation. I-E group showed the enhanced viability and recovery upon thaw compared to D-E group due to the preservation of extracellular matrix, cell-cell contacts, and F-actin organization. Moreover, both I-E and D-E groups showed the increased neuronal differentiation and D-E group also showed the enhanced astrocyte differentiation after thaw, probably due to the modulation of cellular redox state indicated by the expression of reactive oxygen species. In addition, mesenchymal stem cell secretome, known to bear a broad spectrum of protective factors, enhanced EB recovery. Taken together, EB microenvironment plays a critical role in the recovery and neural differentiation post-cryopreservation.
    Tissue Engineering Part C Methods 09/2014; 21(4). DOI:10.1089/ten.TEC.2014.0276 · 4.64 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Because of their multi/pluripotency and immunosuppressive properties mesenchymal stem/stromal cells (MSCs) are important tools for treating immune disorders and for tissue repair. The increasing use of MSCs has led to production processes that need to be in accordance with Good Manufacturing Practice (GMP). In cellular therapy, safety remains one of the main concerns and refers to donor validation, choice of starting material, processes, and the controls used, not only at the batch release level but also during the development of processes. The culture processes should be reproducible, robust, and efficient. Moreover, they should be adapted to closed systems that are easy to use. Implementing controls during the manufacturing of clinical-grade MSCs is essential. The controls should ensure microbiological safety but also avoid potential side effects linked to genomic instability driving transformation and senescence or decrease of cell functions (immunoregulation, differentiation potential). In this rapidly evolving field, a new approach to controls is needed.
    Human gene therapy 10/2010; 22(1):19-26. DOI:10.1089/hum.2010.197 · 3.62 Impact Factor