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Lrig1 expression defines a distinct multipotent stem cell population in mammalian epidermis.

Jensen KB, Collins CA, Nascimento E, Tan DW, Frye M, Itami S, Watt FM - Cell Stem Cell (2009)

Bottom Line: Lrig1 is a marker of human interfollicular epidermal stem cells and helps maintain stem cell quiescence.Lrig1 is a Myc target gene; loss of Lrig1 increases the proliferative capacity of stem cells in culture and results in epidermal hyperproliferation in vivo.Lrig1-expressing cells can give rise to all of the adult epidermal lineages in skin reconstitution assays.

View Article: PubMed Central - PubMed

Affiliation: Laboratory for Epidermal Stem Cell Biology, Wellcome Trust Centre for Stem Cell Research, University of Cambridge, Cambridge, UK.

ABSTRACT
Lrig1 is a marker of human interfollicular epidermal stem cells and helps maintain stem cell quiescence. We show that, in mouse epidermis, Lrig1 defines the hair follicle junctional zone adjacent to the sebaceous glands and infundibulum. Lrig1 is a Myc target gene; loss of Lrig1 increases the proliferative capacity of stem cells in culture and results in epidermal hyperproliferation in vivo. Lrig1-expressing cells can give rise to all of the adult epidermal lineages in skin reconstitution assays. However, during homeostasis and on retinoic acid stimulation, they are bipotent, contributing to the sebaceous gland and interfollicular epidermis. beta-catenin activation increases the size of the junctional zone compartment, and loss of Lrig1 causes a selective increase in beta-catenin-induced ectopic hair follicle formation in the interfollicular epidermis. Our results suggest that Lrig1-positive cells constitute a previously unidentified reservoir of adult mouse interfollicular epidermal stem cells.

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Epidermal Reconstitution by Lrig1-Expressing Cells(A–C) Keratinocytes isolated from the back skin of wild-type adult telogen (7-week-old) mice were sorted into α6 integrin-high (I) and -low (II) populations (A). Populations I (B) and II (C) were gated into three further populations on the basis of Sca1 and CD34 expression, yielding a total of six discrete populations of keratinocytes (1–3 in [B]; 4–6 in [C]).(D) RNA from each of the six populations or total live cells (low forward/side scatter; all) were analyzed using Q-PCR for the genes indicated (mean ± SEM, n = 5).(E–P) Epidermal cells from telogen back skin of GFP-expressing mice were fractionated based on Sca1, α6 integrin, and CD34 expression as in (A)–(C) or Lrig1 (as in Figure 2E) for epidermal reconstitution experiments by mixing GFP-positive and GFP-negative epidermal cells. Four cell populations were compared: α6 high (ItgA6H), CD34 positive, Sca1 negative (population 1; enriched for bulge cells; [E], [I], and [M]); α6 high, CD34 negative, Sca1 negative (population 2; enriched for Lrig1-positive cells; [F], [J], and [N]); α6 high, CD34 negative, SCA1 positive (population 3; [G], [K], and [O]); or Lrig1 positive ([H], [L], [P]). (E–H) Dermal view of grafts; (I-P) Sections of grafts labeled with GFP antibody (brown) and hematoxylin counterstain (blue). Interfollicular epidermis (IFE), hair follicles (HF), and sebaceous glands (SG) are indicated. Scale bars, 100 μm (I–L) and 25 μm (M–P).
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fig3: Epidermal Reconstitution by Lrig1-Expressing Cells(A–C) Keratinocytes isolated from the back skin of wild-type adult telogen (7-week-old) mice were sorted into α6 integrin-high (I) and -low (II) populations (A). Populations I (B) and II (C) were gated into three further populations on the basis of Sca1 and CD34 expression, yielding a total of six discrete populations of keratinocytes (1–3 in [B]; 4–6 in [C]).(D) RNA from each of the six populations or total live cells (low forward/side scatter; all) were analyzed using Q-PCR for the genes indicated (mean ± SEM, n = 5).(E–P) Epidermal cells from telogen back skin of GFP-expressing mice were fractionated based on Sca1, α6 integrin, and CD34 expression as in (A)–(C) or Lrig1 (as in Figure 2E) for epidermal reconstitution experiments by mixing GFP-positive and GFP-negative epidermal cells. Four cell populations were compared: α6 high (ItgA6H), CD34 positive, Sca1 negative (population 1; enriched for bulge cells; [E], [I], and [M]); α6 high, CD34 negative, Sca1 negative (population 2; enriched for Lrig1-positive cells; [F], [J], and [N]); α6 high, CD34 negative, SCA1 positive (population 3; [G], [K], and [O]); or Lrig1 positive ([H], [L], [P]). (E–H) Dermal view of grafts; (I-P) Sections of grafts labeled with GFP antibody (brown) and hematoxylin counterstain (blue). Interfollicular epidermis (IFE), hair follicles (HF), and sebaceous glands (SG) are indicated. Scale bars, 100 μm (I–L) and 25 μm (M–P).

Mentions: To allow isolation of junctional zone cells from trypsinized and Lrig1- epidermis, we developed a second sorting strategy. Undifferentiated (low forward and side scatter) epidermal keratinocytes were sorted into six distinct populations based on CD34, α6 integrin, and Sca1 expression (Jensen et al., 2008; Figures 3A–3C). Epidermal cells were divided into α6 integrin-high (I in Figure 3A) or -low (II in Figure 3A) populations (Blanpain et al., 2004; Jensen et al., 2008; Silva-Vargas et al., 2005). These two populations were each fractionated into three further groups—CD34 positive, Sca1 negative (1 in Figure 3B; 4 in Figure 3C); CD34 negative, Sca1 negative (2 in Figure 3B; 5 in Figure 3C); or CD34 negative, Sca1 positive (3 in Figure 3B; 6 in Figure 3C)—and RNA was isolated from all six populations. The color coding of the different populations in Figures 3B and 3C matches the colors in Figure 3D. Cells in population 2 were, as expected, enriched for Lrig1 expression (Figures 3A–3D and Figures S2A–S2F) and had the same RNA expression profile as cells sorted with Lrig1 antibodies (Figures S2D–S2F; cf. Figure 2G). We conclude that sorting CD34-negative, Sca1-negative, α6 integrin-high cells represents an alternative strategy for enriching Lrig1-positive cells.


Lrig1 expression defines a distinct multipotent stem cell population in mammalian epidermis.

Jensen KB, Collins CA, Nascimento E, Tan DW, Frye M, Itami S, Watt FM - Cell Stem Cell (2009)

Epidermal Reconstitution by Lrig1-Expressing Cells(A–C) Keratinocytes isolated from the back skin of wild-type adult telogen (7-week-old) mice were sorted into α6 integrin-high (I) and -low (II) populations (A). Populations I (B) and II (C) were gated into three further populations on the basis of Sca1 and CD34 expression, yielding a total of six discrete populations of keratinocytes (1–3 in [B]; 4–6 in [C]).(D) RNA from each of the six populations or total live cells (low forward/side scatter; all) were analyzed using Q-PCR for the genes indicated (mean ± SEM, n = 5).(E–P) Epidermal cells from telogen back skin of GFP-expressing mice were fractionated based on Sca1, α6 integrin, and CD34 expression as in (A)–(C) or Lrig1 (as in Figure 2E) for epidermal reconstitution experiments by mixing GFP-positive and GFP-negative epidermal cells. Four cell populations were compared: α6 high (ItgA6H), CD34 positive, Sca1 negative (population 1; enriched for bulge cells; [E], [I], and [M]); α6 high, CD34 negative, Sca1 negative (population 2; enriched for Lrig1-positive cells; [F], [J], and [N]); α6 high, CD34 negative, SCA1 positive (population 3; [G], [K], and [O]); or Lrig1 positive ([H], [L], [P]). (E–H) Dermal view of grafts; (I-P) Sections of grafts labeled with GFP antibody (brown) and hematoxylin counterstain (blue). Interfollicular epidermis (IFE), hair follicles (HF), and sebaceous glands (SG) are indicated. Scale bars, 100 μm (I–L) and 25 μm (M–P).
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Related In: Results  -  Collection

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fig3: Epidermal Reconstitution by Lrig1-Expressing Cells(A–C) Keratinocytes isolated from the back skin of wild-type adult telogen (7-week-old) mice were sorted into α6 integrin-high (I) and -low (II) populations (A). Populations I (B) and II (C) were gated into three further populations on the basis of Sca1 and CD34 expression, yielding a total of six discrete populations of keratinocytes (1–3 in [B]; 4–6 in [C]).(D) RNA from each of the six populations or total live cells (low forward/side scatter; all) were analyzed using Q-PCR for the genes indicated (mean ± SEM, n = 5).(E–P) Epidermal cells from telogen back skin of GFP-expressing mice were fractionated based on Sca1, α6 integrin, and CD34 expression as in (A)–(C) or Lrig1 (as in Figure 2E) for epidermal reconstitution experiments by mixing GFP-positive and GFP-negative epidermal cells. Four cell populations were compared: α6 high (ItgA6H), CD34 positive, Sca1 negative (population 1; enriched for bulge cells; [E], [I], and [M]); α6 high, CD34 negative, Sca1 negative (population 2; enriched for Lrig1-positive cells; [F], [J], and [N]); α6 high, CD34 negative, SCA1 positive (population 3; [G], [K], and [O]); or Lrig1 positive ([H], [L], [P]). (E–H) Dermal view of grafts; (I-P) Sections of grafts labeled with GFP antibody (brown) and hematoxylin counterstain (blue). Interfollicular epidermis (IFE), hair follicles (HF), and sebaceous glands (SG) are indicated. Scale bars, 100 μm (I–L) and 25 μm (M–P).
Mentions: To allow isolation of junctional zone cells from trypsinized and Lrig1- epidermis, we developed a second sorting strategy. Undifferentiated (low forward and side scatter) epidermal keratinocytes were sorted into six distinct populations based on CD34, α6 integrin, and Sca1 expression (Jensen et al., 2008; Figures 3A–3C). Epidermal cells were divided into α6 integrin-high (I in Figure 3A) or -low (II in Figure 3A) populations (Blanpain et al., 2004; Jensen et al., 2008; Silva-Vargas et al., 2005). These two populations were each fractionated into three further groups—CD34 positive, Sca1 negative (1 in Figure 3B; 4 in Figure 3C); CD34 negative, Sca1 negative (2 in Figure 3B; 5 in Figure 3C); or CD34 negative, Sca1 positive (3 in Figure 3B; 6 in Figure 3C)—and RNA was isolated from all six populations. The color coding of the different populations in Figures 3B and 3C matches the colors in Figure 3D. Cells in population 2 were, as expected, enriched for Lrig1 expression (Figures 3A–3D and Figures S2A–S2F) and had the same RNA expression profile as cells sorted with Lrig1 antibodies (Figures S2D–S2F; cf. Figure 2G). We conclude that sorting CD34-negative, Sca1-negative, α6 integrin-high cells represents an alternative strategy for enriching Lrig1-positive cells.

Bottom Line: Lrig1 is a marker of human interfollicular epidermal stem cells and helps maintain stem cell quiescence.Lrig1 is a Myc target gene; loss of Lrig1 increases the proliferative capacity of stem cells in culture and results in epidermal hyperproliferation in vivo.Lrig1-expressing cells can give rise to all of the adult epidermal lineages in skin reconstitution assays.

View Article: PubMed Central - PubMed

Affiliation: Laboratory for Epidermal Stem Cell Biology, Wellcome Trust Centre for Stem Cell Research, University of Cambridge, Cambridge, UK.

ABSTRACT
Lrig1 is a marker of human interfollicular epidermal stem cells and helps maintain stem cell quiescence. We show that, in mouse epidermis, Lrig1 defines the hair follicle junctional zone adjacent to the sebaceous glands and infundibulum. Lrig1 is a Myc target gene; loss of Lrig1 increases the proliferative capacity of stem cells in culture and results in epidermal hyperproliferation in vivo. Lrig1-expressing cells can give rise to all of the adult epidermal lineages in skin reconstitution assays. However, during homeostasis and on retinoic acid stimulation, they are bipotent, contributing to the sebaceous gland and interfollicular epidermis. beta-catenin activation increases the size of the junctional zone compartment, and loss of Lrig1 causes a selective increase in beta-catenin-induced ectopic hair follicle formation in the interfollicular epidermis. Our results suggest that Lrig1-positive cells constitute a previously unidentified reservoir of adult mouse interfollicular epidermal stem cells.

Show MeSH
Related in: MedlinePlus