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Changes in the gene expression pattern of cytokeratins in human respiratory epithelial cells during culture

  • Laryngology
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European Archives of Oto-Rhino-Laryngology and Head & Neck Aims and scope Submit manuscript

Abstract

The replacement of extensive tracheal defects resulting from intensive care medicine, trauma or large resections is still challenged by the re-epithelialization of an autologous or alloplastic trachea replacement. Therefore, this study was performed to investigate the potential of culture-expanded human respiratory epithelial cells (hREC) to regenerate a functional epithelium for tracheal tissue engineering. hREC from seven male nasal turbinates were freshly isolated, expanded on a collagenous matrix and subsequently cultured in high-density multi-layers to allow epithelial differentiation. The composition of epithelial cells in native respiratory epithelial tissue and culture-expanded hREC was analyzed by histological staining with Alcian blue and by immunohistochemical staining of cytokeratin pairs CK1/10 and CK5/14 with the antibodies 34βE12 and CD44v6. Differentiation of culture-expanded hREC was further characterized by gene expression analysis of cytokeratins CK5, CK13, CK14 and CK18 using semi-quantitative real-time RT-PCR technique. Histological and immunohistochemical staining of culture-expanded hREC demonstrated basal cells covering the collagenous matrix. These cells formed a cellular multi-layer, which was composed of a basal layer of undifferentiated basal cells and an upper layer of cells differentiating along the squamous metaplasia and ciliated cell lineage. Lineage development of culture-expanded hREC was further documented by the induction of cytokeratins CK13 and CK18. Our results suggest that culture-expanded hREC have the potential to colonize collagen-coated biomaterials and to regenerate epithelial cell types for tracheal tissue engineering.

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References

  1. Har-El G, Shaha A, Chaudry R, Lucente FE (1993) Resection of tracheal stenosis with end-to-end anastomosis. Ann Otol Rhinol Laryngol 102:670–674

    CAS  PubMed  Google Scholar 

  2. Cull DL, Lally KP, Mair EA, Daidone M, and Parsons DS (1990) Tracheal reconstruction with polytetrafluoroethylene graft in dogs. Ann Thorac Surg 50:899–901

    CAS  PubMed  Google Scholar 

  3. Grillo HC (1970) Surgery of the trachea. Curr Probl Surg:3–59

    Google Scholar 

  4. Suh SW, Kim J, Baek CH, Han J, and Kim H (2001) Replacement of a tracheal defect with autogenous mucosa lined tracheal prosthesis made from polypropylene mesh. Asaio J 47:496–500

    CAS  PubMed  Google Scholar 

  5. Evans MJ, Van Winkle LS, Fanucchi MV, and Plopper CG (2001) Cellular and molecular characteristics of basal cells in airway epithelium. Exp Lung Res 27:401–415

    Article  CAS  PubMed  Google Scholar 

  6. Fahy JV (2001) Remodeling of the airway epithelium in asthma. Am J Respir Crit Care Med 164:S46–51

    CAS  PubMed  Google Scholar 

  7. Otto WR (2002) Lung epithelial stem cells. J Pathol 197:527–535

    Article  CAS  PubMed  Google Scholar 

  8. Hassid S, Salmon I, Brugmans M, Dawance S, Kiss R, Gabius HJ, Danguy A (1997) Histochemical study of the epithelia of nasal polyps by biotinylated lectins and neoglycoprotein. A comparison with the normal human respiratory epithelium. Eur J Morphol 35:79–86

    Article  CAS  PubMed  Google Scholar 

  9. Moll R, Franke WW, Schiller DL, Geiger B, Krepler R (1982) The catalog of human cytokeratins: patterns of expression in normal epithelia, tumors and cultured cells. Cell 31:11–24

    Article  CAS  PubMed  Google Scholar 

  10. Cooper D, Schermer A, Sun TT (1985) Classification of human epithelia and their neoplasms using monoclonal antibodies to keratins: strategies, applications, and limitations. Lab Invest 52:243–256

    CAS  PubMed  Google Scholar 

  11. Purkis PE, Steel JB, Mackenzie IC, Nathrath WB, Leigh IM, Lane EB (1990) Antibody markers of basal cells in complex epithelia. J Cell Sci 97:39–50

    PubMed  Google Scholar 

  12. Stosiek P, Kasper M, Moll R (1992) Changes in cytokeratin expression accompany squamous metaplasia of the human respiratory epithelium. Virchows Arch A Pathol Anat Histopathol 421:133–141

    CAS  PubMed  Google Scholar 

  13. Jetten AM, George MA, Smits HL, Vollberg TM (1989) Keratin 13 expression is linked to squamous differentiation in rabbit tracheal epithelial cells and down-regulated by retinoic acid. Exp Cell Res 182:622–634

    CAS  PubMed  Google Scholar 

  14. Brezillon S, Dupuit F, Hinnrasky J, Marchand V, Kalin N, Tummler B, Puchelle E (1995) Decreased expression of the CFTR protein in remodeled human nasal epithelium from non-cystic fibrosis patients. Lab Invest 72:191–200

    CAS  PubMed  Google Scholar 

  15. Boers JE, Ambergen AW, Thunnissen FB (1998) Number and proliferation of basal and parabasal cells in normal human airway epithelium. Am J Respir Crit Care Med 157:2000–2006

    CAS  PubMed  Google Scholar 

  16. Chomczynski P, Mackey K (1995) Short technical reports. Modification of the TRI reagent procedure for isolation of RNA from polysaccharide- and proteoglycan-rich sources. Biotechniques 19:942–945

    CAS  PubMed  Google Scholar 

  17. Gubler U, Hoffman BJ (1983) A simple and very efficient method for generating cDNA libraries. Gene 25:263–269

    Article  CAS  PubMed  Google Scholar 

  18. Winer J, Jung CK, Shackel I, Williams PM (1999) Development and validation of real-time quantitative reverse transcriptase-polymerase chain reaction for monitoring gene expression in cardiac myocytes in vitro. Anal Biochem 270:41–49

    Article  CAS  PubMed  Google Scholar 

  19. Olze H, Kaschke O, Muller WD (1997) Improving the design of an alloplastic epithelialized tracheal prosthesis. HNO 45:453–459

    Article  CAS  PubMed  Google Scholar 

  20. Grillo HC (2002) Tracheal replacement: a critical review. Ann Thorac Surg 73:1995–2004

    Article  PubMed  Google Scholar 

  21. Kasper M, Gunthert U, Dall P, Kayser K, Schuh D, Haroske G, Muller M (1995) Distinct expression patterns of CD44 isoforms during human lung development and in pulmonary fibrosis. Am J Respir Cell Mol Biol 13:648–656

    CAS  PubMed  Google Scholar 

  22. Wimmel A, Kogan E, Ramaswamy A, Schuermann M (2001) Variant expression of CD44 in preneoplastic lesions of the lung. Cancer 92:1231–1236

    Article  CAS  PubMed  Google Scholar 

  23. Fuchs E (1988) Keratins as biochemical markers of epithelial differentiation. Trends Genet 4:277–281

    Google Scholar 

  24. Kaartinen L, Nettesheim P, Adler KB, and Randell SH (1993) Rat tracheal epithelial cell differentiation in vitro. In Vitro Cell Dev Biol Anim 29A:481–492

    CAS  PubMed  Google Scholar 

  25. Moll R (1993) Cytokeratins as markers of differentiation. Expression profiles in epithelia and epithelial tumors. Veroff Pathol 142:1–197

    CAS  PubMed  Google Scholar 

  26. van Muijen GN, Ruiter DJ, Franke WW, Achtstatter T, Haasnoot WH, Ponec M, Warnaar SO (1986) Cell type heterogeneity of cytokeratin expression in complex epithelia and carcinomas as demonstrated by monoclonal antibodies specific for cytokeratins nos. 4 and 13. Exp Cell Res 162:97–113

    PubMed  Google Scholar 

  27. Chausovsky G, Luchansky M, Figer A, Shapira J, Gottfried M, Novis B, Bogelman G, Zemer R, Zimlichman S, Klein A (1999) Expression of cytokeratin 20 in the blood of patients with disseminated carcinoma of the pancreas, colon, stomach, and lung. Cancer 86:2398–2405

    Article  CAS  PubMed  Google Scholar 

  28. Quinlan RA, Schiller DL, Hatzfeld M, Achtstatter T, Moll R, Jorcano JL, Magin TM, Franke WW (1985) Patterns of expression and organization of cytokeratin intermediate filaments. Ann N Y Acad Sci 455:282–306

    CAS  PubMed  Google Scholar 

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Acknowledgements

This study was supported by the DFG (Ro 2207/1–1 and SI 569). The authors are very grateful to Samuel Vetterlein and Johanna Golla for their excellent assistance.

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Authors and Affiliations

  1. Tissue Engineering Laboratory, University Medical Center Charité, Tucholskystr. 2, 10117, Berlin, Germany

    Michaela Endres, Iris Leinhase, Marek Wentges, Manja Unger, Jochen Ringe & Michael Sittinger

  2. TransTissue Technologies GmbH, Berlin, Germany

    Michaela Endres, Christian Kaps & Jochen Ringe

  3. Department of Otorhinolaryngology, University Medical Center Charité, Berlin, Germany

    Heidi Olze

  4. Department of Otorhinolaryngology and Head and Neck Surgery, Medical University of Luebeck, Luebeck, Germany

    Nicole Rotter

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  1. Michaela Endres
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Correspondence to Michaela Endres.

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Endres, M., Leinhase, I., Kaps, C. et al. Changes in the gene expression pattern of cytokeratins in human respiratory epithelial cells during culture. Eur Arch Otorhinolaryngol 262, 390–396 (2005). https://doi.org/10.1007/s00405-004-0842-9

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  • DOI: https://doi.org/10.1007/s00405-004-0842-9

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