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. 2011 May 3;108(18):7431-6.
doi: 10.1073/pnas.1012720108. Epub 2011 Apr 18.

Identifying the cellular origin of squamous skin tumors

Gaëlle Lapouge  1 , Khalil Kass Youssef, Benoit Vokaer, Younes Achouri, Cindy Michaux, Panagiota A Sotiropoulou, Cédric Blanpain
Affiliations

Identifying the cellular origin of squamous skin tumors

Gaëlle Lapouge et al. Proc Natl Acad Sci U S A. .

Abstract

Squamous cell carcinoma (SCC) is the second most frequent skin cancer. The cellular origin of SCC remains controversial. Here, we used mouse genetics to determine the epidermal cell lineages at the origin of SCC. Using mice conditionally expressing a constitutively active KRas mutant (G12D) and an inducible CRE recombinase in different epidermal lineages, we activated Ras signaling in different cellular compartments of the skin epidermis and determined from which epidermal compartments Ras activation induces squamous tumor formation. Expression of mutant KRas in hair follicle bulge stem cells (SCs) and their immediate progeny (hair germ and outer root sheath), but not in their transient amplifying matrix cells, led to benign squamous skin tumor (papilloma). Expression of KRas(G12D) in interfollicular epidermis also led to papilloma formation, demonstrating that squamous tumor initiation is not restricted to the hair follicle lineages. Whereas no malignant tumor was observed after KRas(G12D) expression alone, expression of KRas(G12D) combined with the loss of p53 induced invasive SCC. Our studies demonstrate that different epidermal lineages including bulge SC are competent to initiate papilloma formation and that multiple genetic hits in the context of oncogenic KRas are required for the development of invasive SCC.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.
Fig. 1.
Targeting KRasG12D expression in bulge SCs and their progeny induces papilloma formation. (A) Bulge SCs and their HF progeny targeted by K19 promoter are highlighted in dark blue. (B) YFP immunostaining in the absence of TAM administration. (C–G) Lineage tracings of K19CREER/RYFP mice are shown in the bulge (C), HG (D), and ORS (E and F) of the back skin and in the bulge of the lips (G) 1 wk after 10 mg TAM administration. (H–M) Papilloma formation in K19CREER/KRasLSL-G12D mice. (H and K) Macroscopic pictures of tumors seen in the K19CREER/KRasLSL-G12D lips (H), face, and back skin (K) 4 mo after TAM administration. Expression of K5 (I and L) and of K1 (J and M) in papilloma from lips and back skin arising from K19CREER/KRasLSL-G12D mice. Bu, bulge; IRS, inner root sheath; Mx, matrix. (Scale bars, 50 μm.)
Fig. 2.
Fig. 2.
Transit amplifying HF matrix cells do not give rise to papilloma upon KRasG12D expression. (A) Matrix cells and their progeny targeted by Shh promoter are highlighted in blue. (B) YFP immunostaining in the absence of TAM administration. (C–G) Lineage tracing in ShhCREER/RYFP mice showed the presence of YFP cells in the matrix cells of the lips (C), HF (D), matrix (Mx), and hair shaft (HS) (E and F); inner root sheath (IRS) of the back skin (G) shows that ShhCREER marked TA matrix cells and their HF progeny 1 wk after 2.5 mg TAM administration. (D–H) Comparison between wild-type and ShhCREER/KRasLSL-G12D back skin 4 mo after 2.5 mg TAM administration. Hematoxylin-eosin staining (H) and expression of K5 (I) and K6 (J), (H) Oil-red-O staining in back skin, show no epidermal defect or tumor formation in mice expressing KRasG12D in matrix cells. Bu, bulge; Mx, matrix. (Scale bars, 50 μm.)
Fig. 3.
Fig. 3.
KRasG12D expression in the IFE induces papilloma formation. (A) Interfollicular suprabasal cells and rare basal cells targeted by involucrin promoter are highlighted in dark pink. (B) YFP immunostaining in the absence of TAM administration. (C–E) Lineage tracing of InvCREER/RosaYFP back skin mice 1 wk after 5 mg TAM administration. Costaining of YFP and β4, K5, and K1 shows that InvCREER mostly targets the suprabasal differentiated cells of the IFE, as well as some basal cells (D, arrow). (F–K) Papilloma formation in InvCREER/KRasLSL-G12D mice. (F and I) Macroscopic picture of tumors in the lips (F) and back skin (I) of InvCREER/KRasLSL-G12D mice 4 mo after TAM administration. Expression of K5 (G and J) and K1 (H and K) in papilloma from the back skin and lips of InvCREER/KRasLSL-G12D mice. (L–O) Follicular marker expression in papilloma arising from InvCREER/KRasLSL-G12D mice. Expression of K17 (L and M), a follicular marker, and CD34 (N and O), a bulge SC marker in wild-type epidermis (L and N) and papilloma (M and O) from InvCREER/KRasLSL-G12D mice, indicating that tumor differentiation does not reflect their cellular origin. (Scale bars, 50 μm.)
Fig. 4.
Fig. 4.
Combined KRasG12D expression and p53 deletion in the skin epidermis induces SCC. (A) IFE and HF basal cells targeted by K14 promoter at clonal induction are highlighted in dark green. (B) Macroscopic picture of ulcerative back skin lesions from K14CREER/KRasLSL-G12D/p53fl/fl mouse 1 mo after TAM administration. (C and D) Immunostaining of K5 (C) and K1 (D) in K14CREER/KRasLSL-G12D/p53fl/fl back skin tumors show the absence of K1 expression in these tumors. (E–I) Comparison between wild-type back skin and SCC from K14CREER/KRasLSL-G12D/p53fl/fl mice. Hematoxylin-eosin staining (E), expression of K5 (F), K1 (G), laminin5, a basal lamina marker (H), and vimentin, a mesenchymal marker (I), showing that these tumors present all histological and biochemical characteristics of invasive SCC. (Scale bars, 50 μm.)
Fig. 5.
Fig. 5.
KRasG12D expression and p53 deletion in bulge SCs and their HF progeny induce SCC. (A) Macroscopic picture of tumors arising in K19CREER/KRasLSL-G12D/p53fl/fl back skin grafted onto nude mouse. Immunostaining of K5 (B) and K1 (C) in K19CREER/KRasLSL-G12D/p53fl/fl carcinoma. (D–H) Comparison between wild-type back skin, papilloma, and carcinoma arising from grafted K19CREER/KRasLSL-G12D/p53fl/fl back skin. Hematoxylin-eosin staining (D) and expression of K5 (E), K1 (F), laminin5 (G), and vimentin (H) in wild-type back skin and in papilloma and SCC from K19CREER/KRasLSL-G12D/p53fl/fl mice. (Scale bars, 50 μm.)
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