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Magnetic separation and characterization of keratinocyte stem cells from human gingiva
Author(s) -
Calenic B.,
Ishkitiev N.,
Yaegaki K.,
Imai T.,
Kumazawa Y.,
Nasu M.,
Hirata T.
Publication year - 2010
Publication title -
journal of periodontal research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.31
H-Index - 83
eISSN - 1600-0765
pISSN - 0022-3484
DOI - 10.1111/j.1600-0765.2010.01284.x
Subject(s) - involucrin , cytokeratin , stem cell , biology , keratinocyte , stem cell marker , microbiology and biotechnology , side population , cellular differentiation , transferrin receptor , pathology , cell , immunology , cell culture , medicine , cancer stem cell , immunohistochemistry , biochemistry , genetics , gene
Calenic B, Ishkitiev N, Yaegaki K, Imai T, Kumazawa Y, Nasu M, Hirata T. Magnetic separation and characterization of keratinocyte stem cells from human gingiva. J Periodont Res 2010; 45: 703–708. © 2010 John Wiley & Sons A/SBackground and Objective: Although keratinocyte stem cells play a key role in tissue homeostasis, wound healing and neoplasia, they remain difficult to identify and characterize. The purpose of this study was to isolate and characterize an oral keratinocyte stem‐cell population. Material and Methods: Oral human keratinocytes obtained from keratinized oral mucosa were magnetically separated using α 6 β 4 integrin and a proliferation‐related marker, CD71. The isolated cell fractions were analyzed for cell size, cell cycle stage (using flow cytometry) and colony‐forming ability. The expression of stem cell markers p63 and cytokeratin 19 and of differentiation markers cytokeratin 10 and involucrin was checked using immunocytochemical analysis. Results: The stem cell CD71 neg fraction had the smallest cell size compared with CD71 pos and fractions [780.7 ± 141.5 (pixels), 1422.9 ± 264.6 (pixels) and 3844.4 ± 220.1 (pixels) respectively, p < 0.01; analysis of variance (ANOVA)]. Also, the CD71 neg subpopulation consistently had the highest colony‐forming ability among the three cell fractions (126.2 ± 21.7 vs. 32.8 ± 4.5 vs. 12.4 ± 2.1 compared with CD71 pos and subpopulations, respectively, p < 0.01; ANOVA). Moreover, the CD71 neg fraction contained more quiescent cells and fewer actively cycling cells than the CD71 pos cell fraction. The candidate stem cells were positive for cytokeratin 19 and p63 keratinocyte stem cell markers, while differentiation markers such as cytokeratin 10 or involucrin were absent. Conclusion: The human gingival CD71 neg cell fraction, separated by a magnetic system, demonstrated several characteristics of gingival keratinocyte stem cells. It is also suggested that a magnetic system may be an important tool in acquiring oral keratinocyte stem cells for research.