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Optimizing in vitro culture conditions leads to a significantly shorter production time of human dermo-epidermal skin substitutes

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Abstract

Introduction

Autologous dermo-epidermal skin substitutes (DESS) generated in vitro represent a promising therapeutic means to treat full-thickness skin defects in clinical practice. A serious drawback with regard to acute patients is the relatively long production time of 3–4 weeks. With this experimental study we aimed to decrease the production time of DESS without compromising their quality.

Methods

Two in vitro steps of DESS construction were varied: the pre-cultivation time of fibroblasts in hydrogels (1, 3, and 6 days), and the culture time of keratinocytes (3, 6, and 12 days) before transplantation of DESS on nude rats. Additionally, the impact of the air–liquid interface culture during 3 days before transplantation was investigated. 3 weeks after transplantation, the macroscopic appearance was evaluated and histological sections were produced to analyze structure and thickness of epidermis and dermis, the stratification of the epidermis, and the presence of a basal lamina.

Results

Optimal DESS formation was obtained with a fibroblast pre-cultivation time of 6 days. The minimal culture time of keratinocytes on hydrogels was also 6 days. The air–liquid interface culture did not improve graft quality.

Conclusion

By optimizing our in vitro culture conditions, it was possible to very substantially reduce the production time for DESS from 21 to 12 days. However, pre-cultivation of fibroblasts in the dermal equivalent and proliferation of keratinocytes before transplantation remain crucial for an equilibrated maturation of the epidermis and cannot be completely skipped.

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Acknowledgments

This work was financially supported by the EU-FP6 project EuroSTEC (soft tissue engineering for congenital birth defects in children, contract: LSHB-CT-2006-037409), by the EU-FP7 project EuroSkinGraft (FP7/2007-2013: grant agreement No. 279024), by the EU-FP7 (MultiTERM, grant agreement No. 238551), and by the University of Zurich. We are particularly grateful to the Foundation Gaydoul and the sponsors of “DonaTissue” (Thérèse Meier, Robert Zingg) for their generous financial support and interest in our work.

Conflict of interest

None.

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Authors and Affiliations

  1. Tissue Biology Research Unit, University Children’s Hospital Zurich, Zurich, Switzerland

    Luca Pontiggia, Agnieszka Klar, Sophie Böttcher-Haberzeth, Thomas Biedermann & Ernst Reichmann

  2. Department of Pediatric Surgery, University Children’s Hospital Zurich, Steinwiesstrasse 75, 8032, Zurich, Switzerland

    Sophie Böttcher-Haberzeth & Martin Meuli

Authors
  1. Luca Pontiggia
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  2. Agnieszka Klar
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  3. Sophie Böttcher-Haberzeth
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  4. Thomas Biedermann
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  5. Martin Meuli
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  6. Ernst Reichmann
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Corresponding author

Correspondence to Martin Meuli.

Additional information

L. Pontiggia and A. Klar contributed equally to this paper.

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Pontiggia, L., Klar, A., Böttcher-Haberzeth, S. et al. Optimizing in vitro culture conditions leads to a significantly shorter production time of human dermo-epidermal skin substitutes. Pediatr Surg Int 29, 249–256 (2013). https://doi.org/10.1007/s00383-013-3268-x

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  • DOI: https://doi.org/10.1007/s00383-013-3268-x

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