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IRC-SET 2018 pp 249-261 | Cite as

Biofabrication of Organotypic Full-Thickness Skin Constructs

  • Abby Chelsea LeeEmail author
  • Yihua Loo
  • Andrew Wan
Conference paper

Abstract

Organotypic skin constructs have gained significant research and commercial interest in the face of EU bans on animal-tested cosmetic products. In this project, a simplified, full-thickness organotypic skin construct was prepared using fibroblasts encapsulated in a synthetic peptide hydrogel matrix, over which keratinocytes were allowed to proliferate, differentiate and stratify. The self-assembling peptide was synthesized using solid phase chemistry. The peptide was then used to prepare hydrogels of varying concentrations and the formulation was optimized based on gelation kinetics and mechanical strength. The long-term biocompatibility of this matrix with dermal fibroblasts was also evaluated. Finally, the in vitro skin constructs were characterized using histology and electron microscopy. Potential primers to evaluate gene expression of epithelial biomarkers were also identified. In conclusion, the peptide hydrogel is an appropriate matrix for culturing organotypic skin constructs due to its stability and low cytotoxicity. Building on this model, more elaborate systems can be cultured with the addition of more cell types. These biological constructs can potentially be used to screen therapeutic candidates, as well as to evaluate the effects of compounds on skin tissue viability, permeability and cellular gene expression.

Keywords

Organotypic skin construct Ultra-small peptides  Peptide hydrogel 

Notes

Acknowledgements

This work was supported by the Youth Research Program (YRP) at the Institute of Bioengineering and Nanotechnology (Biomedical Research Council, Agency for Science, Technology and Research, Singapore).

References

  1. 1.
    Shamir, E., & Ewald, A. (2014). Three-dimensional organotypic culture: Experimental models of mammalian biology and disease. Nature Reviews Molecular Cell Biology, 15(10), 647–664.  https://doi.org/10.1038/nrm3873.CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Weigelt, B., Lo, A. T., Park, C. C., Gray, J. W., & Bissell, M. J. (2010). HER2 signaling pathway activation and response of breast cancer cells to HER2-targeting agents is dependent strongly on the 3D microenvironment. Breast Cancer Research and Treatment, 122, 35–43.  https://doi.org/10.1007/s10549-009-0502-2.CrossRefPubMedGoogle Scholar
  3. 3.
    Kenny, P. A., Lee, G. Y., Myers, C. A., Neve, R. M., Semeiks, J. R., Spellman, P. T., et al. (2007). The morphologies of breast cancer cell lines in three-dimensional assays correlate with their profiles of gene expression. Molecular Oncology, 1, 84–96.  https://doi.org/10.1016/j.molonc.2007.02.004.CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    EUR-Lex—32009R1223—EN—EUR-Lex. (2017). Eur-lex.europa.eu. Retrieved December 28, 2017, from http://eur-lex.europa.eu/legal-content/EN/TXT/?uri=CELEX:32009R1223.
  5. 5.
    Panchagnula, R., Stemmer, K., & Ritschel, W. A. (1997). Animal models for transdermal drug delivery. Retrieved from https://www.ncbi.nlm.nih.gov/pubmed/9379782.
  6. 6.
    Andersen, T., Auk-Emblem, P., & Dornish, M. (2015). 3D cell culture in alginate hydrogels. Microarrays, 4(2), 133–161.  https://doi.org/10.3390/microarrays4020133.CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Lee, K., & Mooney, D. (2001). Hydrogels for tissue engineering. Chemical Reviews, 101(7), 1869–1880.  https://doi.org/10.1021/cr000108x.CrossRefPubMedGoogle Scholar
  8. 8.
    Rosso, F., Giordano, A., Barbarisi, M., & Barbarisi, A. (2004). From cell-ECM interactions to tissue engineering. Journal of Cellular Physiology, 199(2), 174–180.  https://doi.org/10.1002/jcp.10471.CrossRefPubMedGoogle Scholar
  9. 9.
    Timpson, P., Mcghee, E., Erami, Z., Nobis, M., Quinn, J., Edward, M., et al. (2011). Organotypic collagen I assay: A malleable platform to assess cell behaviour in a 3-dimensional context. Journal of Visualized Experiments, (56). http://dx.doi.org/10.3791/3089.
  10. 10.
    Hauser, C., Deng, R., Mishra, A., Loo, Y., Khoe, U., Zhuang, F., et al. (2011). Natural tri- to hexapeptides self-assemble in water to amyloid β-type fiber aggregates by unexpected α-helical intermediate structures. Proceedings of the National Academy of Sciences, 108(4), 1361–1366.  https://doi.org/10.1073/pnas.1014796108.CrossRefGoogle Scholar
  11. 11.
    Loo, Y., Lakshmanan, A., Ni, M., Toh, L., Wang, S., & Hauser, C. (2015). Peptide bioink: Self-assembling nanofibrous scaffolds for three-dimensional organotypic cultures. Nano Letters, 15(10), 6919–6925.  https://doi.org/10.1021/acs.nanolett.5b02859.CrossRefPubMedGoogle Scholar
  12. 12.
    Kirin, S., Noor, F., Metzler-Nolte, N., & Mier, W. (2017). Manual solid–phase peptide synthesis of metallocene–peptide bioconjugates.Google Scholar
  13. 13.
    Sriram, G., Bigliardi, P., & Bigliardi-Qi, M. (2015). Fibroblast heterogeneity and its implications for engineering organotypic skin models in vitro. European Journal of Cell Biology, 94(11), 483–512.  https://doi.org/10.1016/j.ejcb.2015.08.00.CrossRefPubMedGoogle Scholar
  14. 14.
    Eves, P., Haycock, J., Layton, C., Wagner, M., Kemp, H., Szabo, M., et al. (2003). Anti-inflammatory and anti-invasive effects of α-melanocyte-stimulating hormone in human melanoma cells. British Journal of Cancer, 89(10), 2004–2015.  https://doi.org/10.1038/sj.bjc.6601349.CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Nature Singapore Pte Ltd. 2019

Authors and Affiliations

  1. 1.Raffles InstitutionSingaporeSingapore
  2. 2.Institute of Bioengineering and NanotechnologySingaporeSingapore

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