During wound healing, outer root sheath (ORS) cells of hair follicles can substitute for interfollicular epidermal keratinocytes and thus act as precursor cells for interfollicular epidermal keratinocytes. Owing to improved culture techniques, ORS cells can be induced to develop highly differentiated epidermal equivalents, which are close to the normal human epidermis in terms of histological, ultrastructural, biochemical and immunohistological criteria. Such epidermal equivalents provide a versatile system for various applications in vitro, e.g. the study of epidermal homeostasis, cell interactions, pigmentation as well as toxicity testing and metabolism of xenobiotics. The easy and repeated availability of ORS cells, their successful multiplication in culture irrespective of the age of the hair follicle donor as well as the extended tissue normalization of epidermal equivalents prepared with ORS cells prompted us to test the usefulness of autologous epidermal equivalents for the treatment of recalcitrant chronic wounds. Autologous grafting of such epidermal equivalents in more than 50 recalcitrant leg ulcers of a mainly vascular origin resulted in an initial take rate of around 90%, with subsequent complete closure of the ulcers in about 45% and a significant size reduction in another 40% within 8 weeks. These positive results are probably due to the large compartment of proliferative cells as well as to the well-developed horny layer, which prevents rapid disintegration of the grafts. Practical advantages of this technology are its noninvasiveness and thus repeated availability, the fact that surgical facilities are not necessary and the short immobilization period after grafting, allowing a strategy of sequential application in an outpatient setting as an alternative to surgical autografting.

Keywords:
Outer root sheath cells, Organotypic cultures, Grafts, Leg ulcers
1.
Argyris, T.S. (1976) Kinetics of epidermal production during epidermal regeneration following abrasion in mice. Am J Pathol 83:324–340.
2.
Bell, E., B. Ivarsson, C. Merril (1979) Production of a tissue-like structure by contraction of collagen lattices by human fibroblasts of different proliferative potential in vitro. Proc Natl Acad Sci USA 76:1274–1278.
3.
Berthod, F., O. Damour (1997) In vitro reconstructed skin models for wound coverage in deep burns. Br J Dermatol 136:809–816.
4.
Boyce, S.T., R.G. Ham (1983) Calcium-regulated differentiation of normal human epidermal keratinocytes in chemically defined clonal culture and serum-free serial culture. J Invest Dermatol 81:33s–40s.
5.
Cotsarelis, G., T.T. Sun, R.M. Lavker (1990) Label-retaining cells reside in the bulge area of pilosebaceous unit: Implications for follicular stem cells, hair cycle, and carcinogenesis. Cell 61:1329–1337.
6.
Fuchs, E., J.A. Segre (2000) Stem cells: A new lease on life. Cell 100:143–155.
7.
Fusenig, N.E. (1994) Epithelial-mesenchymal interactions regulate keratinocyte growth and differentiation in vitro; in Leigh, I., B. Lane, F. Watt (eds): The Keratinocyte Handbook. Cambridge, Cambridge University Press, pp 71–94.
8.
Gallico, G.G., N.E. O’ Connor, S. Banks-Schlegel, et al. (1981) Grafting of burns with cultured epithelium prepared from autologous epithelial cells. Lancet 1:75–78.
9.
Gallico, G.G., N.E. O’Connor, C.C. Compton, O. Kehinde, H. Green (1984) Permanent coverage of large burn wounds with autologous cultured human epithelium. N Engl J Med 311:448–451.
10.
Holbrook, K.A., K. Wolff (1993) The structure and development of the skin; in Fitzpatrick T.B. (ed): Dermatology in General Medicine. New York, McGraw-Hill, pp 97–145.
11.
Hunziker, T., A. Limat (1999) Cultured keratinocyte grafts. Curr Probl Dermatol 27:57–64.
12.
Kantor, R.R.S., J.J. Mattes, K.O. Lloyd (1987) Biochemical analysis of two cell surface glycoprotein complexes, very common antigen 1 and very common antigen 2. Relationship to very late activation T cell antigens. J Biol Chem 262:15158–15165.
13.
Limat, A., T. Hunziker, C. Boillat, K. Bayreuther, F. Noser (1989) Post-mitotic human dermal fibroblasts efficiently support the growth of human follicular keratinocytes. J Invest Dermatol 92:758–762.
14.
Limat, A., H. Hunziker, D. Breitkreutz, N.E. Fusenig, L.R. Braathen (1994) Organotypic cocultures as models to study cell-cell and cell-matrix interactions of human hair follicle cells. Skin Pharmacol 7:47–54.
15.
Limat, A., C.E. Klein (1993) Expression of epidermal integrins in human organotypic keratinocyte cultures. Exp Dermatol 2:196–203.
16.
Limat, A., D. Mauri, T. Hunziker (1996) Successful treatment of chronic leg ulcers with epidermal equivalents generated from cultured autologous outer root sheath cells. J Invest Dermatol 107:128–135.
17.
Limat, A., F.K. Noser (1986) Serial cultivation of single keratinocytes from the outer root sheath of human scalp hair follicles. J Invest Dermatol 87:485–488.
18.
Limat, A., D. Salomon, P. Carraux, J.H. Saurat, T. Hunziker (1999) Human melanocytes grown in epidermal equivalents transfer their melanin to follicular outer root sheath keratinocytes. Arch Dermatol Res 291:325–332.
19.
Maas-Szabowski, N., H.J. Stark, N.E. Fusenig (2000) Keratinocyte growth regulation in defined organotypic cultures through IL-1-induced keratinocyte growth factor expression in resting fibroblasts. J Invest Dermatol 114:1075–1084.
20.
Moll, I. (1995) Proliferative potential of different keratinocytes of plucked human hair follicles. J Invest Dermatol 105:14–21.
21.
Myers, S., H. Navsaria, R. Sanders, C. Green, I. Leigh (1995) Transplantation of keratinocytes in the treatment of wounds. Am J Surg 170:75–83.
22.
Pruniéras, M., M. Régnier, D. Woodley (1983) Methods for cultivation of keratinocytes with an air-liquid interface. J Invest Dermatol 81:28s–33s.
23.
Rheinwald, J.G., H. Green (1975) Serial cultivation of strains of human epidermal keratinocytes: The formation of keratinizing colonies from single cells. Cell 6:331–344.
24.
Rochat, A., K. Kobayashi, Y. Barrandon (1994) Location of stem cells of human hair follicles by clonal analysis. Cell 76:1063–1073.
25.
Schön, M.P., A. Limat, B. Hartmann, C.E. Klein (1995) Characterization of an 80-kD membrane glycoprotein (gp80) of human keratinocytes: A marker for commitment to terminal differentiation in vivo and in vitro. J Invest Dermatol 105:418–425.
26.
Smola, H., G. Thiekötter, N.E. Fusenig (1993) Mutual induction of growth factor gene expression by epidermal-dermal interaction. J Cell Biol 122:417–429.
27.
Taylor, G., M.S. Lehrer, P.J. Jensen, T.T. Sun, R.M. Lavker (2000) Involvement of follicular stem cells in forming not only the follicle but also the epidermis. Cell 102:451–461.
28.
Werner, S. (1998) Keratinocyte growth factor: A unique player in epithelial repair processes. Cytokine Growth Factor Rev 9:153–165.
29.
Watt, F.M., B.L. Hogan (2000) Out of Eden: Stem cells and their niches. Science 287:1427–1430.
30.
Watt, F.M., K.L. Reichelt, K. Elgjo (1989) Pentapeptide inhibitor of epidermal mitosis: Production and responsiveness in cultures of normal, transformed and neoplastic human keratinocytes. Carcinogenesis 10:2249–2253.
31.
Wiszniewski, L., A. Limat, J.H. Saurat, P. Meda, D. Salomon (2000) Differential expression of connexins during stratification of human keratinocytes. J Invest Dermatol 115:278–285.
32.
Yang, J.S., R.M. Lavker, T.T. Sun (1993) Upper human hair follicle contains a subpopulation of keratinocytes with superior in vitro proliferative potential. J Invest Dermatol 101:652–659.
© 2002 S. Karger AG, Basel
2002
Copyright / Drug Dosage / Disclaimer
Copyright: All rights reserved. No part of this publication may be translated into other languages, reproduced or utilized in any form or by any means, electronic or mechanical, including photocopying, recording, microcopying, or by any information storage and retrieval system, without permission in writing from the publisher.
Drug Dosage: The authors and the publisher have exerted every effort to ensure that drug selection and dosage set forth in this text are in accord with current recommendations and practice at the time of publication. However, in view of ongoing research, changes in government regulations, and the constant flow of information relating to drug therapy and drug reactions, the reader is urged to check the package insert for each drug for any changes in indications and dosage and for added warnings and precautions. This is particularly important when the recommended agent is a new and/or infrequently employed drug.
Disclaimer: The statements, opinions and data contained in this publication are solely those of the individual authors and contributors and not of the publishers and the editor(s). The appearance of advertisements or/and product references in the publication is not a warranty, endorsement, or approval of the products or services advertised or of their effectiveness, quality or safety. The publisher and the editor(s) disclaim responsibility for any injury to persons or property resulting from any ideas, methods, instructions or products referred to in the content or advertisements.
You do not currently have access to this content.