Droplet-based vitrification of adherent human induced pluripotent stem cells on alginate microcarrier influenced by adhesion time and matrix elasticity.

Autor: Meiser I; Fraunhofer Institute for Biomedical Engineering IBMT, 66280, Sulzbach, Saar, Germany. Electronic address: ina.meiser@ibmt.fraunhofer.de., Majer J; Fraunhofer Institute for Biomedical Engineering IBMT, 66280, Sulzbach, Saar, Germany., Katsen-Globa A; Fraunhofer Institute for Biomedical Engineering IBMT, 66280, Sulzbach, Saar, Germany., Schulz A; Fraunhofer Institute for Biomedical Engineering IBMT, 66280, Sulzbach, Saar, Germany., Schmidt K; Fraunhofer Institute for Biomedical Engineering IBMT, 66280, Sulzbach, Saar, Germany., Stracke F; Fraunhofer Institute for Biomedical Engineering IBMT, 66280, Sulzbach, Saar, Germany., Koutsouraki E; Censo Biotechnologies Ltd, Roslin Midlothian, EH25 9RG, United Kingdom., Witt G; Fraunhofer Institute for Translational Medicine and Pharmacology ITMP, ScreeningPort, 22525, Hamburg, Germany., Keminer O; Fraunhofer Institute for Translational Medicine and Pharmacology ITMP, ScreeningPort, 22525, Hamburg, Germany., Pless O; Fraunhofer Institute for Translational Medicine and Pharmacology ITMP, ScreeningPort, 22525, Hamburg, Germany., Gardner J; Censo Biotechnologies Ltd, Roslin Midlothian, EH25 9RG, United Kingdom., Claussen C; Fraunhofer Institute for Translational Medicine and Pharmacology ITMP, ScreeningPort, 22525, Hamburg, Germany., Gribbon P; Fraunhofer Institute for Translational Medicine and Pharmacology ITMP, ScreeningPort, 22525, Hamburg, Germany., Neubauer JC; Fraunhofer Institute for Biomedical Engineering IBMT, 66280, Sulzbach, Saar, Germany; Fraunhofer Project Centre for Stem Cell Process Engineering, 97081, Würzburg, Germany., Zimmermann H; Fraunhofer Institute for Biomedical Engineering IBMT, 66280, Sulzbach, Saar, Germany; Censo Biotechnologies Ltd, Roslin Midlothian, EH25 9RG, United Kingdom; Faculty of Marine Science, Universidad Católica Del Norte, 1781421, Coquimbo, Chile; Chair for Molecular and Cellular Biotechnology / Nanotechnology, Saarland University, 66123, Saarbrücken, Germany.
Jazyk: angličtina
Zdroj: Cryobiology [Cryobiology] 2021 Dec; Vol. 103, pp. 57-69. Date of Electronic Publication: 2021 Sep 25.
DOI: 10.1016/j.cryobiol.2021.09.010
Abstrakt: The gold standard in cryopreservation is still conventional slow freezing of single cells or small aggregates in suspension, although major cell loss and limitation to non-specialised cell types in stem cell technology are known drawbacks. The requirement for rapidly available therapeutic and diagnostic cell types is increasing constantly. In the case of human induced pluripotent stem cells (hiPSCs) or their derivates, more sophisticated cryopreservation protocols are needed to address this demand. These should allow a preservation in their physiological, adherent state, an efficient re-cultivation and upscaling upon thawing towards high-throughput applications in cell therapies or disease modelling in drug discovery. Here, we present a novel vitrification-based method for adherent hiPSCs, designed for automated handling by microfluidic approaches and with ready-to-use potential e.g. in suspension-based bioreactors after thawing. Modifiable alginate microcarriers serve as a growth surface for adherent hiPSCs that were cultured in a suspension-based bioreactor and subsequently cryopreserved via droplet-based vitrification in comparison to conventional slow freezing. Soft (0.35%) versus stiff (0.65%) alginate microcarriers in concert with adhesion time variation have been examined. Findings revealed specific optimal conditions leading to an adhesion time and growth surface (matrix) elasticity dependent hypothesis on cryo-induced damaging regimes for adherent cell types. Deviations from the found optimum parameters give rise to membrane ruptures assessed via SEM and major cell loss after adherent vitrification. Applying the optimal conditions, droplet-based vitrification was superior to conventional slow freezing. A decreased microcarrier stiffness was found to outperform stiffer material regarding cell recovery, whereas the stemness characteristics of rewarmed hiPSCs were preserved.
(Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)
Databáze: MEDLINE
Externí odkaz: