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Cryopreservation of dermal fibroblasts and keratinocytes in hydroxyethyl starch–based cryoprotectants

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posted on 10.02.2017 by Yahaira Naaldijk, Adiv A. Johnson, Annett Friedrich-Stockigt, Alexandra Stolzing
Background: Preservation of human skin fibroblasts and keratinocytes is essential for the creation of skin tissue banks. For successful cryopreservation of cells, selection of an appropriate cryoprotectant agent (CPA) is imperative. The aim of this study was to identify CPAs that minimize toxic effects and allow for the preservation of human fibroblasts and keratinocytes in suspension and in monolayers. Results: We cryopreserved human fibroblasts and keratinocytes with different CPAs and compared them to fresh, unfrozen cells. Cells were frozen in the presence and absence of hydroxyethyl starch (HES) or dimethyl sulfoxide (DMSO), the latter of which is a commonly used CPA known to exert toxic effects on cells. Cell numbers were counted immediately post-thaw as well as three days after thawing. Cellular structures were analyzed and counted by labeling nuclei, mitochondria, and actin filaments. We found that successful cryopreservation of suspended or adherent keratinocytes can be accomplished with a 10% HES or a 5% HES, 5% DMSO solution. Cell viability of fibroblasts cryopreserved in suspension was maintained with 10% HES or 5% HES, 5% DMSO solutions. Adherent, cryopreserved fibroblasts were successfully maintained with a 5% HES, 5% DMSO solution. Conclusion: We conclude that skin tissue cells can be effectively cryopreserved by substituting all or a portion of DMSO with HES. Given that DMSO is the most commonly used CPA and is believed to be more toxic than HES, these findings are of clinical significance for tissue-based replacement therapies. Therapies that require the use of keratinocyte and fibroblast cells, such as those aimed at treating skin wounds or skin burns, may be optimized by substituting a portion or all of DMSO with HES during cryopreservation protocols.

Funding

This work was supported by the project “Automated Tissue Engineering on Demand” from the Stiftungsprojekt of the Fraunhofer Society.

History

School

  • Mechanical, Electrical and Manufacturing Engineering

Published in

BMC Biotechnology

Citation

NAALDIJK, Y. ... et al, 2016. Cryopreservation of dermal fibroblasts and keratinocytes in hydroxyethyl starch–based cryoprotectants. BMC Biotechnology, 16 (85), 11pp.

Publisher

© The Author(s). Published by BioMed Central

Version

VoR (Version of Record)

Publisher statement

This work is made available according to the conditions of the Creative Commons Attribution 4.0 International (CC BY 4.0) licence. Full details of this licence are available at: http://creativecommons.org/licenses/ by/4.0/

Publication date

2016

Notes

This is an Open Access Article. It is published by BioMed Central under the Creative Commons Attribution 4.0 International Licence (CC BY). Full details of this licence are available at: http://creativecommons.org/licenses/by/4.0/

ISSN

1472-6750

Language

en

Licence

Exports