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The expression pattern of keratin 24 in tissue-engineered dermo-epidermal human skin substitutes in an in vivo model

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Abstract

Aims and objectives

The use of autologous tissue-engineered skin substitutes is a promising approach to cover large skin defects in patients. Preclinical investigation is pivotal to test and improve the quality of these bio-engineered substitutes. In the skin, the epidermis, formed mainly by keratinocytes, provides the first physical barrier protecting from the environment. Proper keratinocyte differentiation and, thus, formation of a stratified epidermis is essential for this function. Keratins, the main structural support of keratinocytes, play a vital role regarding differentiation of keratinocytes. Here, we examined the expression pattern of a recently described keratinocyte differentiation marker, namely Keratin 24, in our skin substitutes.

Materials and Methods

Human epidermal keratinocytes, melanocytes, dermal fibroblasts, palmar fibroblasts or sweat gland cells were used to prepare skin substitutes. Fibroblast-containing collagen hydrogels were prepared, and keratinocytes or sweat gland cells and melanocytes were seeded onto the hydrogels. The generated tissue-engineered dermo-epidermal skin analogs were transplanted onto full-thickness skin wounds created on the back of immuno-incompetent rats. The skin substitutes were excised at different time points and histologically examined with regard to Keratin 24 expression.

Results

We observed the expression of Keratin 24 in keratinocytes of the upper stratum spinosum of the epidermis. In particular, we observed an intensified expression of Keratin 24 13 weeks after transplantation compared to 4 weeks after transplantation. Importantly, we noticed a markedly higher presence of Keratin 24 in more spinous layers if we used palmar fibroblasts or sweat gland cells in our skin substitutes compared non-palmar fibroblasts or epidermal keratinocytes.

Conclusion

Our observations prove that the keratinocyte differentiation marker Keratin 24 is expressed in our dermo-epidermal skin substitutes in a normal pattern. This highlights that our bio-engineered skin analogs mature and reach homeostasis in an in vivo assay. These findings harbor favorable implications regarding future clinical application.

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References

  1. Berman B, Viera MH, Amini S, Huo R, Jones IS (2008) Prevention and management of hypertrophic scars and keloids after burns in children. J Craniofac Surg 19(4):989–1006

    Article  PubMed  Google Scholar 

  2. Schiestl C, Stiefel D, Meuli M (2010) Giant naevus, giant excision, eleg(i)ant closure? Reconstructive surgery with integra artificial skin to treat giant congenital melanocytic naevi in children. J Plast Reconstr Aesthet Surg 63(4):610–615

    Article  CAS  PubMed  Google Scholar 

  3. Biedermann T, Boettcher-Haberzeth S, Reichmann E (2013) Tissue engineering of skin for wound coverage. Eur J Pediatr Surg 23(5):375–382

    Article  PubMed  Google Scholar 

  4. Meuli M, Raghunath M (1997) Burns (Part 2). Tops and flops using cultured epithelial autografts in children. Pediatr Surg Int 12(7):471–477

    CAS  PubMed  Google Scholar 

  5. Braziulis E, Diezi M, Biedermann T, Pontiggia L, Schmucki M, Hartmann-Fritsch F, Luginbühl J, Schiestl C, Meuli M, Reichmann E (2012) Modified plastic compression of collagen hydrogels provides an ideal matrix for clinically applicable skin substitutes. Tissue Eng Part C Methods 18(6):464–474

    Article  CAS  PubMed  Google Scholar 

  6. Klar AS, Güven S, Biedermann T, Luginbühl J, Böttcher-Haberzeth S, Meuli-Simmen C, Meuli M, Martin I, Scherberich A, Reichmann E (2014) Tissue-engineered dermo-epidermal skin grafts prevascularized with adipose-derived cells. Biomaterials 35(19):5065–5078

    Article  CAS  PubMed  Google Scholar 

  7. Boyce ST, Lloyd CM, Kleiner MC, Swope VB, Abdel-Malek Z, Supp DM (2017) Restoration of cutaneous pigmentation by transplantation to mice of isogeneic human melanocytes in dermal-epidermal engineered skin substitutes. Pigment Cell Melanoma Res. doi:10.1111/pcmr.12609

    PubMed  Google Scholar 

  8. Madison KC (2003) Barrier function of the skin: “la raison d’être” of the epidermis. J Invest Dermatol 121(2):231–241

    Article  CAS  PubMed  Google Scholar 

  9. Wu N, Gidrol X (2014) The wind rose of human keratinocyte cell fate. Cell Mol Life Sci 71(24):4697–4702

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Henry J, Toulza E, Hsu CY, Pellerin L, Balica S, Mazereeuw-Hautier J, Paul C, Serre G, Jonca N, Simon M (2012) Update on the epidermal differentiation complex. Front Biosci (Landmark Ed) 17:1517–1532

    Article  CAS  Google Scholar 

  11. Candi E, Schmidt R, Melino G (2005 Apr) The cornified envelope: a model of cell death in the skin. Nat Rev Mol Cell Biol 6(4):328–340

    Article  CAS  PubMed  Google Scholar 

  12. Alonso L, Fuchs E (2003) Stem cells of the skin epithelium. Proc Natl Acad Sci USA 100(Suppl 1):11830–11835

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. McLean WH, Lane EB (1995) Intermediate filaments in disease. Curr Opin Cell Biol 7(1):118–125

    Article  CAS  PubMed  Google Scholar 

  14. Coulombe PA (2017) The molecular revolution in cutaneous biology: keratin genes and their associated disease: diversity, opportunities, and challenges. J Invest Dermatol 137(5):e67-e71

    Article  PubMed  PubMed Central  Google Scholar 

  15. Irvine AD, McLean WH (1999) Human keratin diseases: the increasing spectrum of disease and subtlety of the phenotype–genotype correlation. Br J Dermatol 140(5):815–828

    Article  CAS  PubMed  Google Scholar 

  16. Moll R, Divo M, Langbein L (2008) The human keratins: biology and pathology. Histochem Cell Biol 129(6):705–733

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Sprecher E, Itin P, Whittock NV, McGrath JA, Meyer R, DiGiovanna JJ, Bale SJ, Uitto J, Richard G (2002) Refined mapping of Naegeli–Franceschetti–Jadassohn syndrome to a 6 cM interval on chromosome 17q11.2-q21 and investigation of candidate genes. J Invest Dermatol 119(3):692–698

    Article  CAS  PubMed  Google Scholar 

  18. Min M, Chen XB, Wang P, Landeck L, Chen JQ, Li W, Cai SQ, Zheng M, Man XY (2017) Role of keratin 24 in human epidermal keratinocytes. PLoS One 12(3):e0174626

    Article  PubMed  PubMed Central  Google Scholar 

  19. Böttcher-Haberzeth S, Biedermann T, Pontiggia L, Braziulis E, Schiestl C, Hendriks B, Eichhoff OM, Widmer DS, Meuli-Simmen C, Meuli M, Reichmann E (2013) Human eccrine sweat gland cells turn into melanin-uptaking keratinocytes in stratifying dermo-epidermal skin substitutes. J Invest Dermatol 133(2):316–324

    Article  PubMed  Google Scholar 

  20. Biedermann T, Pontiggia L, Böttcher-Haberzeth S, Tharakan S, Braziulis E, Schiestl C, Meuli M, Reichmann E (2010) Human eccrine sweat gland cells can reconstitute a stratified epidermis. J Invest Dermatol 130(8):1996–2009

    Article  CAS  PubMed  Google Scholar 

  21. Biedermann T, Böttcher-Haberzeth S, Klar AS, Widmer DS, Pontiggia L, Weber AD, Weber DM, Schiestl C, Meuli M, Reichmann E (2015) The influence of stromal cells on the pigmentation of tissue-engineered dermo-epidermal skin grafts. Tissue Eng Part A 21(5–6):960–969

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Klar AS, Biedermann T, Michalak K, Michalczyk T, Meuli-Simmen C, Scherberich A, Meuli M, Reichmann E (2017) Human adipose mesenchymal cells inhibit melanocyte differentiation and the pigmentation of human skin via increased expression of TGF-β1. J Invest Dermatol

  23. Knapp AC, Franke WW, Heid H, Hatzfeld M, Jorcano JL, Moll R (1986) Cytokeratin No. 9, an epidermal type I keratin characteristic of a special program of keratinocyte differentiation displaying body site specificity. J Cell Biol 103(2):657–667

    Article  CAS  PubMed  Google Scholar 

  24. Compton CC, Nadire KB, Regauer S, Simon M, Warland G, O’Connor NE, Gallico GG, Landry DB (1998) Cultured human sole-derived keratinocyte grafts re-express site-specific differentiation after transplantation. Differentiation 64(1):45–53

    Article  CAS  PubMed  Google Scholar 

  25. Yamaguchi Y, Itami S, Tarutani M, Hosokawa K, Miura H, Yoshikawa K (1999) Regulation of keratin 9 in nonpalmoplantar keratinocytes by palmoplantar fibroblasts through epithelial-mesenchymal interactions. J Invest Dermatol 112(4):483–488

    Article  CAS  PubMed  Google Scholar 

  26. Pontiggia L, Biedermann T, Böttcher-Haberzeth S, Oliveira C, Braziulis E, Klar AS, Meuli-Simmen C, Meuli M, Reichmann E (2014) De novo epidermal regeneration using human eccrine sweat gland cells: higher competence of secretory over absorptive cells. J Invest Dermatol 134(6):1735–1742

    Article  CAS  PubMed  Google Scholar 

  27. Watt FM J (1983) Involucrin and other markers of keratinocyte terminal differentiation. J Invest Dermatol 81(1 Suppl):100 s–3 s

    Article  CAS  Google Scholar 

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Acknowledgements

This work was financially supported by 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), and the Clinical Research Priority Programs (CRPP) of the Faculty of Medicine of the University of Zurich. We are particularly grateful to the Foundation Gaydoul and the sponsors of “DonaTissue” (Thérèse Meier, Robert Zingg, the Vontobel Foundation, and the Werner Spross Foundation) for their generous financial support and interest in our work.

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Correspondence to Agnes S. Klar.

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Conflict of interest

ER and MM are co-founding members and shareholders of “Cutiss AG”, a company to fund the further development of the tissue-engineered skin substitutes. All other authors declare that they have no conflict of interest.

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Klar, A.S., Michalak, K., Böttcher-Haberzeth, S. et al. The expression pattern of keratin 24 in tissue-engineered dermo-epidermal human skin substitutes in an in vivo model. Pediatr Surg Int 34, 237–244 (2018). https://doi.org/10.1007/s00383-017-4198-9

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