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Dedifferentiation derived cells exhibit phenotypic and functional characteristics of epidermal stem cells.

Zhang C, Fu X, Chen P, Bao X, Li F, Sun X, Lei Y, Cai S, Sun T, Sheng Z - J. Cell. Mol. Med. (2010)

Bottom Line: Furthermore, the percentages of CK19 and beta1-integrin+ cells detected by flow cytometric analysis were increased after grafting (P < 0.01) and CK10+ cells in grafted sheets decreased (P < 0.01).Then we isolated these cells on the basis of rapid adhesion to type IV collagen and found that there were 4.56% adhering cells (dedifferentiation-derived cells) in the grafting group within 10 min.These results suggested that the characteristics of dedifferentiation-derived cells cultured in vitro were similar to epidermal stem cells.

View Article: PubMed Central - PubMed

Affiliation: Wound Healing and Cell Biology Laboratory, Burns Institute, The First Affiliated Hospital, General Hospital of PLA, Trauma Center of Postgraduate Medical College, Beijing, PR China.

ABSTRACT
Differentiated epidermal cells can dedifferentiate into stem cells or stem cell-like cells in vivo. In this study, we report the isolation and characterization of dedifferentiation-derived cells. Epidermal sheets eliminated of basal stem cells were transplanted onto the skin wounds in 47 nude athymic (BALB/c-nu/nu) mice. After 5 days, cells negative for CK10 but positive for CK19 and beta1-integrin emerged at the wound-neighbouring side of the epidermal sheets. Furthermore, the percentages of CK19 and beta1-integrin+ cells detected by flow cytometric analysis were increased after grafting (P < 0.01) and CK10+ cells in grafted sheets decreased (P < 0.01). Then we isolated these cells on the basis of rapid adhesion to type IV collagen and found that there were 4.56% adhering cells (dedifferentiation-derived cells) in the grafting group within 10 min. The in vitro phenotypic assays showed that the expressions of CK19, beta1-integrin, Oct4 and Nanog in dedifferentiation-derived cells were remarkably higher than those in the control group (differentiated epidermal cells) (P < 0.01). In addition, the results of the functional investigation of dedifferentiation-derived cells demonstrated: (1) the numbers of colonies consisting of 5-10 cells and greater than 10 cells were increased 5.9-fold and 6.7-fold, respectively, as compared with that in the control (P < 0.01); (2) more cells were in S phase and G2/M phase of the cell cycle (proliferation index values were 21.02% in control group, 45.08% in group of dedifferentiation); (3) the total days of culture (28 days versus 130 days), the passage number of cells (3 passages versus 20 passages) and assumptive total cell output (1 x 10(5) cells versus 1 x 10(12) cells) were all significantly increased and (4) dedifferentiation-derived cells, as well as epidermal stem cells, were capable of regenerating a skin equivalent, but differentiated epidermal cells could not. These results suggested that the characteristics of dedifferentiation-derived cells cultured in vitro were similar to epidermal stem cells. This study may also offer a new approach to yield epidermal stem cells for wound repair and regeneration.

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Detection of X- and Y-chromosome specific sequences by PCR. M, Marker; lane 1, control (differentiated epidermal cells); lane 2, positive control (epidermal stem cells); lane 3, grafting group (dedifferentiation-derived cells); lane 4, mouse skin tissue; lane 5, no DNA (negative control).
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fig03: Detection of X- and Y-chromosome specific sequences by PCR. M, Marker; lane 1, control (differentiated epidermal cells); lane 2, positive control (epidermal stem cells); lane 3, grafting group (dedifferentiation-derived cells); lane 4, mouse skin tissue; lane 5, no DNA (negative control).

Mentions: To discriminate dedifferentiated-derived cells of human origin from mouse stem cells of the host, polymerase chain reaction was performed on DNA extracted from above initial adherent cells and from mouse skin tissue. Y-chromosome-specific sequences could be detected in the adhering cells isolated from ultrathin epidermal sheet (grafted and ungrafted) and full-thickness epidermal sheets. However, in mouse skin tissue, there were no Y-chromosome-specific sequences (Fig. 3). These results demonstrate that cells isolated from xenografts were originally from human male donors and are not mouse stem cells coming from the host that have attached to the graft.


Dedifferentiation derived cells exhibit phenotypic and functional characteristics of epidermal stem cells.

Zhang C, Fu X, Chen P, Bao X, Li F, Sun X, Lei Y, Cai S, Sun T, Sheng Z - J. Cell. Mol. Med. (2010)

Detection of X- and Y-chromosome specific sequences by PCR. M, Marker; lane 1, control (differentiated epidermal cells); lane 2, positive control (epidermal stem cells); lane 3, grafting group (dedifferentiation-derived cells); lane 4, mouse skin tissue; lane 5, no DNA (negative control).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3822750&req=5

fig03: Detection of X- and Y-chromosome specific sequences by PCR. M, Marker; lane 1, control (differentiated epidermal cells); lane 2, positive control (epidermal stem cells); lane 3, grafting group (dedifferentiation-derived cells); lane 4, mouse skin tissue; lane 5, no DNA (negative control).
Mentions: To discriminate dedifferentiated-derived cells of human origin from mouse stem cells of the host, polymerase chain reaction was performed on DNA extracted from above initial adherent cells and from mouse skin tissue. Y-chromosome-specific sequences could be detected in the adhering cells isolated from ultrathin epidermal sheet (grafted and ungrafted) and full-thickness epidermal sheets. However, in mouse skin tissue, there were no Y-chromosome-specific sequences (Fig. 3). These results demonstrate that cells isolated from xenografts were originally from human male donors and are not mouse stem cells coming from the host that have attached to the graft.

Bottom Line: Furthermore, the percentages of CK19 and beta1-integrin+ cells detected by flow cytometric analysis were increased after grafting (P < 0.01) and CK10+ cells in grafted sheets decreased (P < 0.01).Then we isolated these cells on the basis of rapid adhesion to type IV collagen and found that there were 4.56% adhering cells (dedifferentiation-derived cells) in the grafting group within 10 min.These results suggested that the characteristics of dedifferentiation-derived cells cultured in vitro were similar to epidermal stem cells.

View Article: PubMed Central - PubMed

Affiliation: Wound Healing and Cell Biology Laboratory, Burns Institute, The First Affiliated Hospital, General Hospital of PLA, Trauma Center of Postgraduate Medical College, Beijing, PR China.

ABSTRACT
Differentiated epidermal cells can dedifferentiate into stem cells or stem cell-like cells in vivo. In this study, we report the isolation and characterization of dedifferentiation-derived cells. Epidermal sheets eliminated of basal stem cells were transplanted onto the skin wounds in 47 nude athymic (BALB/c-nu/nu) mice. After 5 days, cells negative for CK10 but positive for CK19 and beta1-integrin emerged at the wound-neighbouring side of the epidermal sheets. Furthermore, the percentages of CK19 and beta1-integrin+ cells detected by flow cytometric analysis were increased after grafting (P < 0.01) and CK10+ cells in grafted sheets decreased (P < 0.01). Then we isolated these cells on the basis of rapid adhesion to type IV collagen and found that there were 4.56% adhering cells (dedifferentiation-derived cells) in the grafting group within 10 min. The in vitro phenotypic assays showed that the expressions of CK19, beta1-integrin, Oct4 and Nanog in dedifferentiation-derived cells were remarkably higher than those in the control group (differentiated epidermal cells) (P < 0.01). In addition, the results of the functional investigation of dedifferentiation-derived cells demonstrated: (1) the numbers of colonies consisting of 5-10 cells and greater than 10 cells were increased 5.9-fold and 6.7-fold, respectively, as compared with that in the control (P < 0.01); (2) more cells were in S phase and G2/M phase of the cell cycle (proliferation index values were 21.02% in control group, 45.08% in group of dedifferentiation); (3) the total days of culture (28 days versus 130 days), the passage number of cells (3 passages versus 20 passages) and assumptive total cell output (1 x 10(5) cells versus 1 x 10(12) cells) were all significantly increased and (4) dedifferentiation-derived cells, as well as epidermal stem cells, were capable of regenerating a skin equivalent, but differentiated epidermal cells could not. These results suggested that the characteristics of dedifferentiation-derived cells cultured in vitro were similar to epidermal stem cells. This study may also offer a new approach to yield epidermal stem cells for wound repair and regeneration.

Show MeSH
Related in: MedlinePlus