Skin contains various populations of stem cells (SCs). Among these are hair follicle
stem cells (HFSCs) in the bulge region. The behavior of HFSCs deserves to be widely studied due
to the benefits to be derived from their identification, isolation, and
amplification. Skin samples of newborn mice (n = 32) and human adults (n = 10) were used, and the bulge region was isolated
and cultured. The isolation and characterization of cells were conducted
through immunocytochemistry and
immunofluorescence, using mainly CD34 and CD200 monoclonal antibodies.
Initially, cells grew slowly from the explant around the bulge region, accruing
cells with different morphology in both mouse and human, latter being mostly
polygonal; the mouse cells reaching confluence faster (5 to 7 days) than the
human (12 to 15 days). It was possible to isolate into subcultures cells
with small size (10 - 13 μm diameter),
round-shape, scant cytoplasm, central
prominent nucleus and with nucleolus, which formed colonies, maintaining their
phenotype in a high proportion (77% - 83% and
91% in mouse and human, respectively), without showing changes in their
morphology during almost 7 months in the mouse cells, and a month and a half in
the human. These results demonstrate that the selection, the isolation, and the
conditioned mediums allowed population increases of bulge cells and indicate
that cultured cells may retain their sternness in that they maintained their
phenotypic characteristics, expressed specific markers for SCs, and showed a
high proliferative capacity for long periods. Hair follicles, in mice and
humans, are important repositories of multipotent stem cells, due to their
tendency to differentiate into keratinocytes. Human HFSCs, obtained by
depilation, preserve their potential for proliferation and prove to be easily
accessible. This suggests that the bulge cells may present an alternative
source of autologous stem cells for tissue engineering and regenerative
medicine.
References
[1]
Weissman, I.L., Anderson, D.J. and Gage, F. (2001) Stem and Progenitor Cells: Origins, Phenotypes, Lineage Commitments, and Transdifferentiations. Annual Review of Cell and Developmental Biology, 17, 387-403.
https://doi.org/10.1146/annurev.cellbio.17.1.387
[2]
Abbas, O. and Mahalingam, M. (2009) Epidermal Stem Cells: Practical Perspectives and Potential Uses. British Journal of Dermatology, 161, 228-236.
https://www.ncbi.nlm.nih.gov/pubmed/19548960
https://doi.org/10.1111/j.1365-2133.2009.09250.x
[3]
Botchkarev, V.A., Botchkareva, N.V., Nakamura, M., Huber, O., Funa, K., Lauster, R., Paus, R. and Gilchrest, B.A. (2001) Noggin Is Required for Induction of the Hair Follicle Growth Phase in Postnatal Skin. The FASEB Journal, 15, 2205-2214.
https://www.ncbi.nlm.nih.gov/pubmed/11641247
https://doi.org/10.1096/fj.01-0207com
[4]
Blanpain, C., Lowry, W.E., Geoghegan, A., Polak, L. and Fuchs, E. (2004) Self-Renewal, Multipotency, and the Existence of Two Cell Populations within an Epithelial Stem Cell Niche. Cell, 118, 635-648. https://www.ncbi.nlm.nih.gov/pubmed/15339667
https://doi.org/10.1016/j.cell.2004.08.012
[5]
Yan, X. and Owens, D.M. (2008) The Skin: A Home to Multiple Classes of Epithelial Progenitor Cells. Stem Cell Reviews, 4, 113-118.
https://www.ncbi.nlm.nih.gov/pubmed/18491239
https://doi.org/10.1007/s12015-008-9022-4
[6]
Schmidt-Ullrich, R. and Paus, R. (2005) Molecular Principles of Hair Follicle Induction and Morphogenesis. BioEssays, 27, 247-261.
https://www.ncbi.nlm.nih.gov/pubmed/15714560
https://doi.org/10.1002/bies.20184
[7]
Yang, C.C. and Cotsarelis, G. (2010) Review of Hair Follicle Dermal Cells. Journal of Dermatological Science, 57, 2-11.
https://www.ncbi.nlm.nih.gov/pubmed/20022473
https://doi.org/10.1016/j.jdermsci.2009.11.005
[8]
Zhang, Y., Xiang, M., Wang, Y., Yan, J., Zeng, Y., Yu, J. and Yang, T. (2006) Bulge Cells of Human Hair Follicles: Segregation, Cultivation and Properties. Colloids and Surfaces B. Biointerfaces, 47, 50-56.
https://www.ncbi.nlm.nih.gov/pubmed/?term=10.1016%2Fj.colsurfb.2005.11.017
https://doi.org/10.1016/j.colsurfb.2005.11.017
[9]
Ohyama, M. (2007) Hair Follicle Bulge: A Fascinating Reservoir of Epithelial Stem Cells. Journal of Dermatological Science, 46, 81-89.
https://www.ncbi.nlm.nih.gov/pubmed/?term=10.1016%2Fj.jdermsci.2006.12.002
https://doi.org/10.1016/j.jdermsci.2006.12.002
[10]
Zouboulis, C.C., Adjaye, J., Akamatsu, H., Moe-Behrens, G. and Niemann, C. (2008) Human Skin Stem Cell and the Ageing Process. Experimental Gerontology, 43, 986-997. https://www.ncbi.nlm.nih.gov/pubmed/18809487
https://doi.org/10.1016/j.exger.2008.09.001
[11]
Forni, M.F., Trombetta-Lima, M. and Sogayar, M.C. (2012) Stem Cells in Embryonic Skin Development. Biological Research, 45, 215-222.
https://doi.org/10.4067/S0716-97602012000300003
https://www.ncbi.nlm.nih.gov/pubmed/23283431
[12]
Ito, M., Liu, Y., Yang, Z., Nguyen, J., Liang, F., Morris, R.J. and Cotsarelis, G. (2005) Stem Cells in the Hair Follicle Bulge Contribute to Wound Repair But Not to Homeostasis of the Epidermis. Nature Medicine, 11, 1351-1354.
https://www.ncbi.nlm.nih.gov/pubmed/16288281
https://doi.org/10.1038/nm1328
[13]
Tiede, S., Kloepper, J., Bodo, E., Tiwari, S., Kruse, C. and Paus, R. (2007) Hair Follicle Stem Cells: Walking the Maze. European Journal of Cell Biology, 86, 355-376.
https://www.ncbi.nlm.nih.gov/pubmed/?term=10.1016%2Fj.ejcb.2007.03.006
https://doi.org/10.1016/j.ejcb.2007.03.006
[14]
Zhou, J., Jia, L., Yang, Y., Peng, S., Cao, Y. and Duan, E. (2004) Enrichment and Characterization of Mouse Putative Epidermal Stem Cells. Cell Biology International, 28, 523-529. https://doi.org/10.1016/j.cellbi.2004.04.007
[15]
Jiang, S., Zhao, L., Purandare, B. and Hantash, B.M. (2010) Differential Expression of Stem Cell Markers in Human Follicular Bulge and Interfollicular Epidermal Compartments. Histochemistry and Cell Biology, 133, 455-465.
https://www.ncbi.nlm.nih.gov/pubmed/15261160
https://doi.org/10.1007/s00418-010-0684-z
[16]
Zurita, M., Aguayo, C., Oya, S. and Vaquero, J. (2007) Implicación de factores neurotróficos en la transdiferenciación neuronal de células madre mesenquimales adultas. Mpfre Medicina, 18, 201-208.
https://dialnet.unirioja.es/servlet/articulo?codigo=2470199
[17]
Amoh, Y., Mii, S., Aki, R., Hamada, Y., Kawahara, K., Hoffman, R.M. and Katsuoka, K. (2012) Multipotent Nestin-Expressing Stem Cells Capable of Forming Neurons Are Located in the Upper, Middle and Lower Part of the Vibrissa Hair Follicle. Cell Cycle, 11, 3513-3517. https://doi.org/10.4161/cc.21803
https://www.ncbi.nlm.nih.gov/pubmed/?term=10.4161%2Fcc.21803
[18]
Tiede, S., Koop, N., Kloepper, J.E., Fässler, R. and Paus, R. (2009) Nonviral in Situ Green Fluorescent Protein Labeling and Culture of Primary, Adult Human Hair Follicle Epithelial Progenitor Cells. Stem Cells, 27, 2793-2803.
https://www.ncbi.nlm.nih.gov/pubmed/?term=10.1002%2Fstem.213
https://doi.org/10.1002/stem.213
[19]
Molina, B.J., Giansante, E. and Finol, H.J. (2018) Identification of Bulge Stem Cells in Mouse and Human Hair Follicles. Microscopy Research, 6, 19-29.
https://doi.org/10.4236/mr.2018.63003
[20]
Magerl, M., Kauser, S., Paus, R. and Tobin, D.J. (2002) Simple and Rapid Method to Isolate and Culture Follicular Papillae from Human Scalp Hair Follicles. Experimental Dermatology, 11, 381-385. https://www.ncbi.nlm.nih.gov/pubmed/12190949
https://doi.org/10.1034/j.1600-0625.2002.110414.x
[21]
Xu, X., Lyle, S., Liu, Y., Solky, B. and Cotsarelis, G. (2003) Differential Expression of Cyclin D1 in the Human Hair Follicle. The American Journal of Pathology, 163, 969-978. https://www.ncbi.nlm.nih.gov/pubmed/12937137
https://doi.org/10.1016/S0002-9440(10)63456-6
[22]
Sieber-Blum, M. and Hu, Y. (2008) Mouse Epidermal Neural Crest Stem Cell (EPI- NCSC) Cultures. Journal of Visualized Experiments, 9, e772.
https://www.ncbi.nlm.nih.gov/pubmed/19066583
https://doi.org/10.3791/772
[23]
Nowak, J.A. and Fuchs, E. (2009) Isolation and Culture of Epithelial Stem Cells. Methods in Molecular. Biology, 482, 215-232.
https://www.ncbi.nlm.nih.gov/pubmed/19089359
https://doi.org/10.1007/978-1-59745-060-7_14
[24]
Usovetskii, I.A., Burunova, V.V., Kovtun, N.E., Suzdal’tseva, Y.G., Krasil’nikova, Y.B., Korotkii, N.G. and Yarygin, K.N. (2009) Preparation of Mixed Keratinocyte and Melanocyte Cultures from Biopsy Specimens of Pigmented Skin Sites of Patients with Vitiligo Translated. Bulletin of Experimental Biology and Medicine, 148, 103-105. https://doi.org/10.1007/s10517-009-0646-4
https://www.ncbi.nlm.nih.gov/pubmed/?term=10.1007%2Fs10517-009-0646-4
[25]
Savkovic, V., Dieckmann, C., Milkova, L. and Simon, J.C. (2012) Improved Method of Differentiation, Selection and Amplification of Human Melanocytes from the Hair Follicle Cell Pool. Experimental Dermatology, 21, 948-950.
https://www.ncbi.nlm.nih.gov/pubmed/23171457
https://doi.org/10.1111/exd.12038
[26]
Inoue, K. and Yoshimura, K. (2013) Isolation and Characterization of Human Hair Follicle Epithelial Cells. Methods in Molecular. Biology, 946, 411-421.
https://www.ncbi.nlm.nih.gov/pubmed/23179847
https://doi.org/10.1007/978-1-62703-128-8_26
[27]
Wang, H.-T., Chen, B., Tao, K., Tang, C.-W. and Hu, D.-H. (2007) Human Hair Follicle Bulge Cells: Potential Germ Cells for Skin Tissue Engineering. Journal of US-China Medical Science, 4, 1-8.
https://www.ncbi.nlm.nih.gov/pubmed/?term=10.4252%2Fwjsc.v7.i4.711
[28]
Blazejewska, E.A., Schlötzer-Schrehardt, U., Zenkel, M., Bachmann, B., Chankiewitz, E., Jacobi, C. and Kruse, F.E. (2009) Corneal Limbal Microenvironment Can Induce Transdifferentiation of Hair Follicle Stem Cells into Corneal Epithelial-Like Cells. Stem Cells, 27, 642-652. https://www.ncbi.nlm.nih.gov/pubmed/19074417
https://doi.org/10.1634/stemcells.2008-0721
[29]
Yu, H., Fang, D., Kumar, S.M., Li, L., Nguyen, T.K., Acs, G., Herlyn, M. and Xu, X. (2006) Isolation of a Novel Population of Multipotent Adult Stem Cells from Human Hair Follicles. The American Journal Pathology, 168, 1879-1888.
https://www.ncbi.nlm.nih.gov/pubmed/16723703
https://doi.org/10.2353/ajpath.2006.051170
[30]
Drewa, T., Joachimiak, R., Kaznica, A., Sarafian, V. and Sir, J. (2009) Primary Cultures from Rat Vibrissae as a Potential Cell Source for in Vitro Construction of Urinary Bladder Wall Grafts. Transplantation Proceedings, 41, 1932-1935.
https://www.ncbi.nlm.nih.gov/pubmed/19545759
https://doi.org/10.1016/j.transproceed.2009.02.091
[31]
Nobakht, M., Najafzadeh, N., Safari, M., Roshandel, N.R., Delaviz, H., Joghataie, M.T., Bakhtiyari, M., Asalgoo, S. and Safar, F. (2010) Bulge Cells of Rat Hair Follicles: Isolation, Cultivation, Morphological and Biological Features. Yakhteh Medicine Journal, 1, 51-58.
[32]
Hejazian, L.B., Esmaeilzade, B., Moghanni, G.F., Moradi, F., Hejazian, M.B., Aslani, A., Bakhtiari, M., Soleimani, M. and Nobakht, M. (2012) The Role of Biodegradable Engineered Nanofiber Scaffolds Seeded with Hair Follicle Stem Cells for Tissue Engineering. Iran Biomedical Journal, 16, 193-201.
https://www.ncbi.nlm.nih.gov/pubmed/23183618
[33]
Ramot, Y., Bíró, T., Tiede, S., Tóth, B.I., Langan, E.A., Sugawara, K., Foitzik, K., Ingber, A., Goffin, V., Langbein, L. and Paus, R. (2010) Prolactina Novel Neuroendocrine Regulator of Human Keratin Expression in Situ. FASEB Journal, 24, 1768-1779.
https://www.ncbi.nlm.nih.gov/pubmed/?term=10.1096%2Ffj.09-146415
https://doi.org/10.1096/fj.09-146415
[34]
Amoh, Y., Li, L., Katsuoka, K., Penman, S. and Hoffman, R.M. (2005) Multipotent Nestin-Positive, Keratin-Negative Hair-Follicle Bulge Stem Cells Can Form Neurons. PNAS, 102, 5530-5534. https://doi.org/10.1073/pnas.0501263102
https://www.ncbi.nlm.nih.gov/pubmed/?term=10.1073%2Fpnas.0501263102
[35]
Liu, F., Uchugonova, A., Kimura, H., Zhang, C., Zhao, M., Zhang, L., Koenig, K., Duong, J., Aki, R., Saito, N., Mii, S., Amoh, Y., Katsuoka, K. and Hoffman, R.M. (2011) The Bulge Area Is the Major Hair Follicle Source of Nestin-Expressing Pluripotent Stem Cells Which Can Repair the Spinal Cord Compared to the Dermal Papilla. Cell Cycle, 10, 830-839. https://doi.org/10.4161/cc.10.5.14969
https://www.ncbi.nlm.nih.gov/pubmed/?term=10.4161%2Fcc.10.5.14969
[36]
Gartner, S., Liu, Y. and Natesan, S. (2012) De Novo Generation of Cells within Human Nurse Macrophages and Consequences Following HIV-1 Infection. PLoS ONE, 7, e40139. https://www.ncbi.nlm.nih.gov/pubmed/22911696
https://doi.org/10.1371/journal.pone.0040139
[37]
Castellana, D., Paus, R. and Perez-Moreno, M. (2014) Macrophages Contribute to the Cyclic Activation of Adult Hair Follicle Stem Cells. PLoS Biology, 23, e1002002.
https://www.ncbi.nlm.nih.gov/pubmed/?term=10.1371%2Fjournal.pbio.1002002
https://doi.org/10.1371/journal.pbio.1002002
[38]
Magerl, M., Kauser, S., Paus, R. and Tobin, D.J. (2002) Simple and Rapid Method to Isolate and Culture Follicular Papillae from Human Scalp Hair Follicles. Experimental Dermatology, 11, 381-385.
https://www.ncbi.nlm.nih.gov/pubmed/?term=10.1034%2Fj.1600-0625.2002.110414.x
https://doi.org/10.1034/j.1600-0625.2002.110414.x
[39]
Wu, M., Sun, Q., Guo, X. and Liu, H. (2012) hMSCs Possess the Potential to Differentiate into DP Cells in Vivo and in Vitro. Cell Biology International Reports, 19, e00019. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3475446
https://doi.org/10.1042/CBR20120003
[40]
Barrandon, Y. and Green, H. (1985) Cell Size as a Determinant of the Clone Forming Ability of Human Keratinocytes. PNAS, 82, 5390-5394.
https://www.ncbi.nlm.nih.gov/pubmed/2410922
https://doi.org/10.1073/pnas.82.16.5390
[41]
Inoue, K., Aoi, N., Sato, T., Yamauchi, Y., Suga, H., Eto, H., Kato, H., Araki, J. and Yoshimura, K. (2009) Differential Expression of Stem-Cell-Associated Markers in Human Hair Follicle Epithelial Cells. Laboratory Investigation, 89, 844-856.
https://www.ncbi.nlm.nih.gov/pubmed/?term=10.1038%2Flabinvest.2009.48
https://doi.org/10.1038/labinvest.2009.48
[42]
Oh, J.H., Moheb, P., Farkas, D.L. and Tajbakhsh, J. (2011) Towards Expansion of Human Hair Follicle Stem Cells in Vitro. Cell Proliferation, 44, 244-253.
https://www.ncbi.nlm.nih.gov/pubmed/21535265
https://doi.org/10.1111/j.1365-2184.2011.00754.x
[43]
Wang, X., Shi, Y., Zhou, Q., Liu, X., Xu, S. and Lei, T. (2012) Detailed Histological Structure of Human Hair Follicle Bulge Region at Different Ages: A Visible Niche for Nesting Adult Stem Cells. Journal of Huazhong University of Science and Technology Medical Science, 32, 648-656.
https://doi.org/10.1007/s11596-012-1012-8
https://www.ncbi.nlm.nih.gov/pubmed/?term=10.1007%2Fs11596-012-1012-8
[44]
Freshney, I.R. (2005) Culture of Animal Cells a Manual of Basic Technique. 5nd Edition, John Wiley & Sons, Inc., Hoboken, 642.
https://doi.org/10.1002/9780471747598
[45]
Bruder, S.P., Jaiswal, N. and Haynesworth, S.E. (1997) Growth Kinetics, Self-Renewal, and the Osteogenic Potential of Purified Human Mesenchymal Stem Cells during Extensive Subcultivation and Following Cryopreservation. Journal of Cellular Biochemistry, 64, 278-294. https://www.ncbi.nlm.nih.gov/pubmed/9027588
https://doi.org/10.1002/(SICI)1097-4644(199702)64:2<278::AID-JCB11>3.0.CO;2-F
[46]
Nath, M., Offers, M., Hummel, M. and Seissler, J. (2011) Isolation and in Vitro Expansion of Lgr6-Positive Multipotent Hair Follicle Stem Cells. Cell Tissue Research, 344, 435-444. https://doi.org/10.1007/s00441-011-1165-y
https://www.ncbi.nlm.nih.gov/pubmed/?term=10.1007%2Fs00441-011-1165-y
[47]
Bajpai, V.K., Mistriotis, P. and Andreadis, S.T. (2012) Clonal Multipotency and Effect of Long-Term in Vitro Expansion on Differentiation Potential of Human Hair Follicle Derived Mesenchymal Stem Cells. Stem Cell Research, 8, 74-84.
https://www.ncbi.nlm.nih.gov/pubmed/?term=10.1016%2Fj.scr.2011.07.003
https://doi.org/10.1016/j.scr.2011.07.003
[48]
Driskell, R.R., Juneja, V.R., Connelly, J.T., Kretzschmar, K., Tan, D.W. and Watt, F.M. (2012) Clonal Growth of Dermal Papilla Cells in Hydrogels Reveals Intrinsic Differences between Sox2-Positive and -Negative Cells in Vitro and in Vivo. Journal Investigative Dermatology, 132, 1084-1093.
https://www.ncbi.nlm.nih.gov/pubmed/?term=10.1038%2Fjid.2011.428
https://doi.org/10.1038/jid.2011.428
[49]
Beyer, N. and Da Silva, L. (2006) Mesenchymal Stem Cells: Isolation in Vitro Expansion and Characterization. In: Wobus, A.M. and Boheler, K.R., Eds., Stem Cells, Vol. 174, Handbook of Experimental Pharmacology, Springer-Verland, New York, 249-282. https://www.ncbi.nlm.nih.gov/pubmed/16370331
https://doi.org/10.1007/3-540-31265-X_11
[50]
Vaissiere, C., Chevallay, B., Herbage, C. and Damour, G. (2000) Comparative Analysis of Different Collagen-Based Biomaterials as Scaffolds for Long-Term Culture of Human Fibroblasts. Medical & Biological Engineering Computing, 38, 205-210.
https://www.ncbi.nlm.nih.gov/pubmed/?term=10.1007%2Fbf02344778
https://doi.org/10.1007/BF02344778
[51]
Liu, Y., Lyle, S., Yang, Z. and Cotsarelis, G. (2003) Keratin 15 Promoter Targets Putative Epithelial Stem Cells in the Hair Follicle Bulge. Journal Investigative Dermatology, 121, 963-968.
https://www.ncbi.nlm.nih.gov/pubmed/?term=10.1046%2Fj.1523-1747.2003.12600.x
https://doi.org/10.1046/j.1523-1747.2003.12600.x
[52]
Ohyama, M., Terunuma, A., Tock, C.L., Radonovich, M.F., PiseMasison, C.A. and Hopping, S.B. (2006) Characterization and Isolation of Stem Cell-Enriched Human Hair Follicle Bulge Cells. The Journal Clinical Investigation, 116, 249-260.
https://www.ncbi.nlm.nih.gov/pubmed/16395407
https://doi.org/10.1172/JCI26043
