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Transcriptional profiling of putative human epithelial stem cells.

Koçer SS, Djurić PM, Bugallo MF, Simon SR, Matic M - BMC Genomics (2008)

Bottom Line: This study demonstrates that alpha 6+/MHCI- cells have additional characteristics attributed to stem cells.Our study is important for understanding self renewal and differentiation of epidermal stem cells, and for elucidating signaling pathways involved in those processes.The generated data base may serve those working with other human epithelial tissue progenitors.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biochemistry and Cell Biology, State University of New York at Stony Brook, Stony Brook, NY, USA. Salih.Kocer@yale.edu

ABSTRACT

Background: Human interfollicular epidermis is sustained by the proliferation of stem cells and their progeny, transient amplifying cells. Molecular characterization of these two cell populations is essential for better understanding of self renewal, differentiation and mechanisms of skin pathogenesis. The purpose of this study was to obtain gene expression profiles of alpha 6+/MHCI+, transient amplifying cells and alpha 6+/MHCI-, putative stem cells, and to compare them with existing data bases of gene expression profiles of hair follicle stem cells. The expression of Major Histocompatibility Complex (MHC) class I, previously shown to be absent in stem cells in several tissues, and alpha 6 integrin were used to isolate MHCI positive basal cells, and MHCI low/negative basal cells.

Results: Transcriptional profiles of the two cell populations were determined and comparisons made with published data for hair follicle stem cell gene expression profiles. We demonstrate that presumptive interfollicular stem cells, alpha 6+/MHCI- cells, are enriched in messenger RNAs encoding surface receptors, cell adhesion molecules, extracellular matrix proteins, transcripts encoding members of IFN-alpha family proteins and components of IFN signaling, but contain lower levels of transcripts encoding proteins which take part in energy metabolism, cell cycle, ribosome biosynthesis, splicing, protein translation, degradation, DNA replication, repair, and chromosome remodeling. Furthermore, our data indicate that the cell signaling pathways Notch1 and NF-kappaB are downregulated/inhibited in MHC negative basal cells.

Conclusion: This study demonstrates that alpha 6+/MHCI- cells have additional characteristics attributed to stem cells. Moreover, the transcription profile of alpha 6+/MHCI- cells shows similarities to transcription profiles of mouse hair follicle bulge cells known to be enriched for stem cells. Collectively, our data suggests that alpha 6+/MHCI- cells may be enriched for stem cells. This study is the first comprehensive gene expression profile of putative human epithelial stem cells and their progeny that were isolated directly from neonatal foreskin tissue. Our study is important for understanding self renewal and differentiation of epidermal stem cells, and for elucidating signaling pathways involved in those processes. The generated data base may serve those working with other human epithelial tissue progenitors.

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MHCI negative cells express low levels of Ki67. (A) A representative flow cytometric analysis of the expression of proliferation antigen Ki67 in MHCI positive, and MHCI negative cells. Gates were set using isotype control antibodies and single color control antibodies. In this experiment 3.9% of MHCI positive cells express Ki67, while only 0.9% of MHCI negative cells express Ki67. Although the exact values of proliferating population may vary from experiment to experiment the ratio of MHCI positive and MHCI negative proliferating populations stay constant. This result demonstrate quiescent nature of MHCI negative cells. (B) Single positive isotype control for PE. (C) Single positive control for FITC. (D) Secondary control.
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Figure 1: MHCI negative cells express low levels of Ki67. (A) A representative flow cytometric analysis of the expression of proliferation antigen Ki67 in MHCI positive, and MHCI negative cells. Gates were set using isotype control antibodies and single color control antibodies. In this experiment 3.9% of MHCI positive cells express Ki67, while only 0.9% of MHCI negative cells express Ki67. Although the exact values of proliferating population may vary from experiment to experiment the ratio of MHCI positive and MHCI negative proliferating populations stay constant. This result demonstrate quiescent nature of MHCI negative cells. (B) Single positive isotype control for PE. (C) Single positive control for FITC. (D) Secondary control.

Mentions: During tissue homeostasis stem cells are infrequently dividing; thus, and one of the characteristics of stem cells is their quiescence in situ. To determine the proliferative status of MHCI- and MHCI+populations, we analyzed the expression of nuclear proliferation antigen Ki67, which is a marker for actively cycling cells. The data presented reflect keratinocyte proliferation since in normal epidermis, non keratinocytes are found to be non-cycling [27,28]. Although the absolute values of Ki67 may vary depending on the total number of gated cells, the ratio of Ki67 expressing MHCI- and MHCI+ cells is held constant. Flow cytometric analysis showed that MHCI+ cells expressed more than four time higher levels of Ki67 than MHCI- cells (Figure 1). Only 0.9% of MHCI- cells expressed Ki67, while 3.9% of MHCI+cells were in the cell cycle. Given the low percentages of MHCI-cells in the basal layer, it is clear that the bulk of cell production in the epidermis is accomplished through divisions of transient amplifying cells, a finding which is in accordance with the established view of epidermal homeostasis.


Transcriptional profiling of putative human epithelial stem cells.

Koçer SS, Djurić PM, Bugallo MF, Simon SR, Matic M - BMC Genomics (2008)

MHCI negative cells express low levels of Ki67. (A) A representative flow cytometric analysis of the expression of proliferation antigen Ki67 in MHCI positive, and MHCI negative cells. Gates were set using isotype control antibodies and single color control antibodies. In this experiment 3.9% of MHCI positive cells express Ki67, while only 0.9% of MHCI negative cells express Ki67. Although the exact values of proliferating population may vary from experiment to experiment the ratio of MHCI positive and MHCI negative proliferating populations stay constant. This result demonstrate quiescent nature of MHCI negative cells. (B) Single positive isotype control for PE. (C) Single positive control for FITC. (D) Secondary control.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2536675&req=5

Figure 1: MHCI negative cells express low levels of Ki67. (A) A representative flow cytometric analysis of the expression of proliferation antigen Ki67 in MHCI positive, and MHCI negative cells. Gates were set using isotype control antibodies and single color control antibodies. In this experiment 3.9% of MHCI positive cells express Ki67, while only 0.9% of MHCI negative cells express Ki67. Although the exact values of proliferating population may vary from experiment to experiment the ratio of MHCI positive and MHCI negative proliferating populations stay constant. This result demonstrate quiescent nature of MHCI negative cells. (B) Single positive isotype control for PE. (C) Single positive control for FITC. (D) Secondary control.
Mentions: During tissue homeostasis stem cells are infrequently dividing; thus, and one of the characteristics of stem cells is their quiescence in situ. To determine the proliferative status of MHCI- and MHCI+populations, we analyzed the expression of nuclear proliferation antigen Ki67, which is a marker for actively cycling cells. The data presented reflect keratinocyte proliferation since in normal epidermis, non keratinocytes are found to be non-cycling [27,28]. Although the absolute values of Ki67 may vary depending on the total number of gated cells, the ratio of Ki67 expressing MHCI- and MHCI+ cells is held constant. Flow cytometric analysis showed that MHCI+ cells expressed more than four time higher levels of Ki67 than MHCI- cells (Figure 1). Only 0.9% of MHCI- cells expressed Ki67, while 3.9% of MHCI+cells were in the cell cycle. Given the low percentages of MHCI-cells in the basal layer, it is clear that the bulk of cell production in the epidermis is accomplished through divisions of transient amplifying cells, a finding which is in accordance with the established view of epidermal homeostasis.

Bottom Line: This study demonstrates that alpha 6+/MHCI- cells have additional characteristics attributed to stem cells.Our study is important for understanding self renewal and differentiation of epidermal stem cells, and for elucidating signaling pathways involved in those processes.The generated data base may serve those working with other human epithelial tissue progenitors.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biochemistry and Cell Biology, State University of New York at Stony Brook, Stony Brook, NY, USA. Salih.Kocer@yale.edu

ABSTRACT

Background: Human interfollicular epidermis is sustained by the proliferation of stem cells and their progeny, transient amplifying cells. Molecular characterization of these two cell populations is essential for better understanding of self renewal, differentiation and mechanisms of skin pathogenesis. The purpose of this study was to obtain gene expression profiles of alpha 6+/MHCI+, transient amplifying cells and alpha 6+/MHCI-, putative stem cells, and to compare them with existing data bases of gene expression profiles of hair follicle stem cells. The expression of Major Histocompatibility Complex (MHC) class I, previously shown to be absent in stem cells in several tissues, and alpha 6 integrin were used to isolate MHCI positive basal cells, and MHCI low/negative basal cells.

Results: Transcriptional profiles of the two cell populations were determined and comparisons made with published data for hair follicle stem cell gene expression profiles. We demonstrate that presumptive interfollicular stem cells, alpha 6+/MHCI- cells, are enriched in messenger RNAs encoding surface receptors, cell adhesion molecules, extracellular matrix proteins, transcripts encoding members of IFN-alpha family proteins and components of IFN signaling, but contain lower levels of transcripts encoding proteins which take part in energy metabolism, cell cycle, ribosome biosynthesis, splicing, protein translation, degradation, DNA replication, repair, and chromosome remodeling. Furthermore, our data indicate that the cell signaling pathways Notch1 and NF-kappaB are downregulated/inhibited in MHC negative basal cells.

Conclusion: This study demonstrates that alpha 6+/MHCI- cells have additional characteristics attributed to stem cells. Moreover, the transcription profile of alpha 6+/MHCI- cells shows similarities to transcription profiles of mouse hair follicle bulge cells known to be enriched for stem cells. Collectively, our data suggests that alpha 6+/MHCI- cells may be enriched for stem cells. This study is the first comprehensive gene expression profile of putative human epithelial stem cells and their progeny that were isolated directly from neonatal foreskin tissue. Our study is important for understanding self renewal and differentiation of epidermal stem cells, and for elucidating signaling pathways involved in those processes. The generated data base may serve those working with other human epithelial tissue progenitors.

Show MeSH
Related in: MedlinePlus