The effect of Me2SO overexposure during cryopreservation on HOS TE85 and hMSC viability, growth and quality

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Morris, Timothy J. ; Picken, Andrew ; Sharp, Duncan M.C. ; Slater, Nigel K.H. ; Hewitt, Christopher J. ; Coopman, Karen (2016)
  • Publisher: Elsevier BV
  • Journal: Cryobiology, volume 73, issue 3, pages 367-375 (issn: 0011-2240)
  • Related identifiers: doi: 10.1016/j.cryobiol.2016.09.004, doi: 10.17863/CAM.7994
  • Subject: toxicity | Agricultural and Biological Sciences(all) | bioprocessing | Biochemistry, Genetics and Molecular Biology(all) | human mesenchymal stem cells | dimethylsulfoxide | HOS TE85 | Medicine(all) | cryopreservation

With the cell therapy industry continuing to grow, the ability to preserve clinical grade cells, including mesenchymal stem cells (MSCs), whilst retaining cell viability and function remains critical for the generation of off-the-shelf therapies. Cryopreservation of MSCs, using slow freezing, is an established process at lab scale. However, the cytotoxicity of cryoprotectants, like Me$_{2}$SO, raises questions about the impact of prolonged cell exposure to cryoprotectant at temperatures >0 °C during processing of large cell batches for allogenic therapies prior to rapid cooling in a controlled rate freezer or in the clinic prior to administration. Here we show that exposure of human bone marrow derived MSCs to Me$_{2}$SO for ≥1 h before freezing, or after thawing, degrades membrane integrity, short-term cell attachment efficiency and alters cell immunophenotype. After 2 h's exposure to Me$_{2}$SO at 37 °C post-thaw, membrane integrity dropped to ∼70% and only ∼50% of cells retained the ability to adhere to tissue culture plastic. Furthermore, only 70% of the recovered MSCs retained an immunophenotype consistent with the ISCT minimal criteria after exposure. We also saw a similar loss of membrane integrity and attachment efficiency after exposing osteoblast (HOS TE85) cells to Me$_{2}$SO before, and after, cryopreservation. Overall, these results show that freezing medium exposure is a critical determinant of product quality as process scale increases. Defining and reporting cell sensitivity to freezing medium exposure, both before and after cryopreservation, enables a fair judgement of how scalable a particular cryopreservation process can be, and consequently whether the therapy has commercial feasibility. The authors would like to acknowledge the Engineering and Physical Sciences Research Council (EPSRC; UK, EP/F500491/1) and Bioprocessing Research Industry Club (BBSRC/BRIC; UK, BB/I017602/1) for their support and funding.
  • References (44)
    44 references, page 1 of 5

    [1] M. Aye, C. Di Giorgio, M. De Mo, A. Botta, J. Perrin, C. Courbiere, Assessment of the genotoxicity of three cryoprotectants used for human oocyte vitrification: dimethyl sulfoxide, ethylene glycol and propylene glycol, Food Chem. Toxicol. 48 (2010) 1905e1912.

    [2] Biocision, Coolcell lx. Available at http://biocision.com/products/CoolCell-LXFreezing-Container/. Accessed: 2014-08-13.

    [3] A.J. Caplan, D. Correa, The MSC: an injury drugstore, Cell Stem Cell 9 (2011) 11e15.

    [4] J. Carmen, S.R. Burger, M. McCaman, J.A. Rowley, Developing assays to address identity, potency, purity and safety: cell characterization in cell therapy process development, Regen. Med. 7 (2012) 85e100.

    [5] A.K. Chan, T.R. Heathman, K. Coopman, C.J. Hewitt, Multiparameter flow cytometry for the characterisation of extracellular markers on human mesenchymal stem cells, Biotechnol. Lett. 36 (2014) 731e741.

    [6] M.A. Cox, J. Kastrup, M. Hrubisko, Historical perspectives and the future of adverse reactions associated with haemopoietic stem cells cryopreserved with dimethyl sulfoxide, Cell Tissue Bank. 13 (2012) 203e215.

    [7] F. Deng, H. Lei, Y. Hu, L. He, H. Fu, R. Feng, P. Feng, W. Huang, X. Wang, J. Chang, Combination of retinoic acid, dimethyl sulfoxide and 5-azacytidine promotes cardiac differentiation of human fetal liver-derived mesenchymal stem cells, Cell Tissue Bank. 17 (2016) 147e159.

    [8] M. Dominici, K. Le Blanc, I. Mueller, I. Slaper-Cortenbach, F. Marini, D. Krause, R. Deans, A. Keating, D. Prockop, E. Horwitz, Minimal criteria for defining multipotent mesenchymal stromal cells. The International Society for Cellular Therapy position statement, Cytotherapy 8 (2006) 315e317.

    [9] G.M. Fahy, Cryoprotectant toxicity neutralization, Cryobiology 60 (2010) S45eS53.

    [10] M. François, I.B. Copland, S. Yuan, R. Romieu-Mourez, E.K. Waller, Cryopreserved mesenchymal stromal cells display impaired immunosuppressive properties as a result of heat-shock response and impaired interferon-g licensing, Cytotherapy 12 (2012) 147e152.

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