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Value of human amniotic epithelial cells in tissue engineering for cornea

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Abstract

Human amniotic epithelial cells (hAECs) are potentially one of the key players in tissue engineering due to their easy availability. The aim of the present study was to develop an optimal isolation and transportation technique, as well as to determine the immunophenotype and epithelial gene expression of hAECs. Amnion was mechanically peeled off from the chorion and digested with trypsin-ethylenediaminetetraacetic acid. The isolated hAECs were cultured in medium containing 10 ng/mL epidermal growth factor until P4. The epithelial gene expression, cell surface antigen and protein expression of hAECs were analyzed by quantitative polymerase chain reaction, flow cytometry and immunocytochemistry. hAECs were also cultured in adipogenic, osteogenic and neurogenic induction media. The best cell yield of hAECs was seen in the digestion of 15 pieces of amnion (2 × 2 cm) and isolated 30 min after digestion with trypsin. F12:Dulbecco’s modified eagle medium was the best medium for short term storage at 4 °C. hAECs expressed CD9, CD44, CD73 and CD90, and negligibly expressed CD31, CD34, CD45 and CD117. After serial passage, CK3, CK19 and involucrin gene expressions were upregulated, while p63, CK1 and CK14 gene expressions were downregulated. Sustained gene expressions of integrin β1 and CK18 were observed. At initial culture, these cells might have stem-like properties. However, they differentiated after serial passage. Nonetheless, hAECs have epithelial stem cell characteristics and have the potential to differentiate into corneal epithelial cells. Further investigations are still needed to elucidate the mechanism of differentiation involved and to optimize the culture condition for long term in vitro culture.

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References

  1. Parolini O, Alviano F, Bagnara GP et al. Concise review: isolation and characterization of cells from human term placenta: outcome of the first international workshop on placenta derived stem cells. Stem Cells 2008; 26: 300–11.

    Article  PubMed  Google Scholar 

  2. Simat SF, Chua KH, Abdul Rahman H, Tan AE, Tan GC. The stemness gene expression of cultured human amniotic epithelial cells in serial passages. Med J Malaysia 2008; 63 (A): 53–4.

    PubMed  Google Scholar 

  3. Tan GC, Simat SF, Abdul Rahman H, Tan AE, Chua KH. Quantitative RT-PCR approach to evaluate the neurogenic and gliagenic gene expression of cultured human amniotic epithelial cells. Med J Malaysia 2008; 63 (A): 51–2.

    PubMed  Google Scholar 

  4. Miki T, Lehmann T, Cai H, Stolz DB, Strom SC. Stem cell characteristics of amniotic epithelial cells. Stem Cells 2005; 23: 1549–59.

    Article  CAS  PubMed  Google Scholar 

  5. Portmann-Lanz CB, Schoeberlein A, Huber A et al. Placental mesenchymal stem cells as potential autologous graft for pre- and perinatal neuroregeneration. Am J Obstet Gynecol 2006; 194: 664–73.

    Article  CAS  PubMed  Google Scholar 

  6. Hou Y, Huang Q, Liu T, Guo L. Human amniotic epithelial cells can be induced to differentiate into functional insulin-producing cells. Acta Biochim Biophys Sin (Shanghai) 2008; 40: 830–9.

    CAS  Google Scholar 

  7. Ito A, Takizawa Y, Masashige S et al. Proliferation and stratification of keratinocyte on cultured amniotic epithelial cells for tissue engineering. J Biosci Bioeng 2003; 95: 589–93.

    CAS  PubMed  Google Scholar 

  8. Ravishanker R, Bath AS, Roy R. “Amnion bank”- the use of long term glycerol preserved amniotic membranes in the management of superficial and superficial partial thickness burns. Burns 2003; 29: 369–74.

    CAS  PubMed  Google Scholar 

  9. Koizumi N, Inatomi T, Suzuki T, Sotozono C, Kinoshita S. Cultivated corneal epithelial stem cell transplantation in ocular surface disorders. Ophthalmology 2001; 108: 1569–74.

    Article  CAS  PubMed  Google Scholar 

  10. Akle CA, Adinolfi M, Welsh KI, Leibowitz S, McColl I. Immunogenicity of human amniotic epithelial cells after transplantation into volunteers. Lancet 1981; 2: 1003–5.

    Article  CAS  PubMed  Google Scholar 

  11. Boyce ST, Ham RG. Calcium-regulated differentiation of normal human epidermal keratinocytes in chemically defined clonal culture and serum-free serial culture. J Invest Dermatol 1983; 81: 33s-40s.

    Article  CAS  PubMed  Google Scholar 

  12. Jones PH, Watt FM. Separation of human epidermal stem cells from transit amplifying cells on the basis of differences in integrin function and expression. Cell 1993; 73: 713–24.

    Article  CAS  PubMed  Google Scholar 

  13. Michel M, Török N, Godbout MJ et al. Keratin 19 as a biochemical marker of skin stem cells in vivo and in vitro: keratin 19 expressing cells are differentially localized in function of anatomic sites, and their number varies with donor age and culture stage. J Cell Sci 1996; 109: 1017–28.

    CAS  PubMed  Google Scholar 

  14. Sun TT, Eichner R, Nelson WG et al. Keratin classes: molecular markers for different types of epithelial differentiation. J Invest Dermatol 1983; 81: 109s-115s.

    Article  CAS  PubMed  Google Scholar 

  15. Schlötzer-Schrehardt U, Kruse FE. Identification and characterization of limbal stem cells. Exp Eye Res 2005; 81: 247–64.

    PubMed  Google Scholar 

  16. Debus E, Weber K, Osborn M. Monoclonal cytokeratin antibodies that distinguish simple from stratified squamous epithelia: characterization on human tissues. EMBOJ 1982; 1: 1641–7.

    CAS  Google Scholar 

  17. Koster MI, Kim S, Mills AA, DeMayo FJ, Roop DR. p63 is the molecular switch for initiation of an epithelial stratification program. Genes Dev 2004; 18: 126–31.

    Article  CAS  PubMed  Google Scholar 

  18. Watt FM. Involucrin and other markers of keratinocyte terminal differentiation. J Invest Dermatol 1983; 81: 100s-103s.

    Article  CAS  PubMed  Google Scholar 

  19. Miller WM, Blanch HW, Wilke CR. A kinetic analysis of hybridoma growth and metabolism in batch and continuous suspension culture: effect of nutrient concentration, dilution rate, and pH. Biotechnol Bioeng 1988; 32: 947–65.

    Article  CAS  PubMed  Google Scholar 

  20. Mazlyzam AL, Aminuddin BS, Fuzina NH et al. Reconstruction of living bilayer human skin equivalent utilizing human fibrin as a scaffold. Burns 2007; 33: 355–63.

    Article  CAS  PubMed  Google Scholar 

  21. Noruddin NA, Saim AB, Chua KH, Idrus R. Human nasal turbinates as a viable source of respiratory epithelial cells using co-culture system versus dispase-dissociation technique. Laryngoscope 2007; 117: 2139–45.

    Article  PubMed  Google Scholar 

  22. Bilic G, Zeisberger SM, Mallik AS, Zimmermann R, Zisch AH. Comparative characterization of cultured human term amnion epithelial and mesenchymal stromal cells for application in cell therapy. Cell Transplant 2008; 17: 955–68.

    Article  PubMed  Google Scholar 

  23. Terada S, Matsuura K, Enosawa S et al. Inducing Proliferation of human amniotic epithelial (HAE) cells for cell therapy. Cell Transplant 2000; 9: 701–4.

    CAS  PubMed  Google Scholar 

  24. Oertel M, Menthena A, Chen YQ, Shafritz DA. Comparison of hepatic properties and transplantation of Thy-1+ and Thy-1− cells isolated from embryonic day 14 rat fetal liver. Hepatology 2007; 46: 1236–45.

    Article  CAS  PubMed  Google Scholar 

  25. Nakamura Y, Muguruma Y, Yahata Y et al. Expression of CD90 on keratinocyte stem/progenitor cells. BrJ Dermatol 2006; 154: 1062–70.

    Article  CAS  Google Scholar 

  26. Li H, Niederkorn JY, Neelam S et al. Immuno-suppressive factors secreted by human amniotic epithelial cells. Invest Ophthalmol Vis Sci 2005; 46: 900–7.

    Article  PubMed  Google Scholar 

  27. Bailo M, Soncini M, Vertua E et al. Engraftment potential of human amnion and chorion cells derived from term placenta. Transplantation 2004; 78: 1439–48.

    Article  PubMed  Google Scholar 

  28. Waseem A, Alexander CM, Steel JB, Lane EB. Embryonic simple epithelial keratins 8 and 18: chromosomal location emphasizes difference from other keratin pairs. New Biol 1990; 2: 464–78.

    CAS  PubMed  Google Scholar 

  29. Pelligrini G, Dellambra E, Golisano O et al. p63 identifies keratinocyte stem cells. Proct Natl Acad Sci USA 2001; 98: 3156–61.

    Article  Google Scholar 

  30. Chen Z, de Paiva CS, Luo L, Kretzer FL, Pflugfelder SC, Li DQ. Characterization of putative stem cell phenotype in human limbal epithelia. Stem Cells 2004; 22: 355–66.

    Article  PubMed  Google Scholar 

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Authors and Affiliations

  1. Department of Pathology, Universiti Kebangsaan Malaysia, Jalan Yaacob Latif, Bandar Tun Razak, 56000, Cheras, Kuala Lumpur, Malaysia

    Simat Siti Fatimah, Sook Luan Ng, Abdul Rahman Hayati & Geok Chin Tan

  2. Department of Physiology, Universiti Kebangsaan Malaysia, Kuala Lumpur, Malaysia

    Kien Hui Chua

  3. Department of Obstetrics and Gynaecology, Universiti Kebangsaan Malaysia, Kuala Lumpur, Malaysia

    Ay Eeng Tan

Authors
  1. Simat Siti Fatimah
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  2. Sook Luan Ng
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  3. Kien Hui Chua
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  4. Abdul Rahman Hayati
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  5. Ay Eeng Tan
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  6. Geok Chin Tan
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Correspondence to Geok Chin Tan.

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Fatimah, S.S., Ng, S.L., Chua, K.H. et al. Value of human amniotic epithelial cells in tissue engineering for cornea. Hum Cell 23, 141–151 (2010). https://doi.org/10.1111/j.1749-0774.2010.00096.x

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  • DOI: https://doi.org/10.1111/j.1749-0774.2010.00096.x

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