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Atopic dermatitis-like disease and associated lethal myeloproliferative disorder arise from loss of Notch signaling in the murine skin.

Dumortier A, Durham AD, Di Piazza M, Vauclair S, Koch U, Ferrand G, Ferrero I, Demehri S, Song LL, Farr AG, Leonard WJ, Kopan R, Miele L, Hohl D, Finke D, Radtke F - PLoS ONE (2010)

Bottom Line: The AD-like associated inflammation is accompanied by a myeloproliferative disorder (MPD) characterized by an increase in immature myeloid populations in the bone marrow and spleen.Genetic studies demontrated that G-CSF mediates the MPD as well as changes in the bone marrow microenvironment leading to osteopenia.Our data demonstrate a critical role for Notch in repressing TSLP production in keratinocytes, thereby maintaining integrity of the skin and the hematopoietic system.

View Article: PubMed Central - PubMed

Affiliation: Ecole Polytechnique Fédérale de Lausanne (EPFL SV ISREC), Lausanne, Switzerland.

ABSTRACT

Background: The Notch pathway is essential for proper epidermal differentiation during embryonic skin development. Moreover, skin specific loss of Notch signaling in the embryo results in skin barrier defects accompanied by a B-lymphoproliferative disease. However, much less is known about the consequences of loss of Notch signaling after birth.

Methodology and principal findings: To study the function of Notch signaling in the skin of adult mice, we made use of a series of conditional gene targeted mice that allow inactivation of several components of the Notch signaling pathway specifically in the skin. We demonstrate that skin-specific inactivation of Notch1 and Notch2 simultaneously, or RBP-J, induces the development of a severe form of atopic dermatitis (AD), characterized by acanthosis, spongiosis and hyperkeratosis, as well as a massive dermal infiltration of eosinophils and mast cells. Likewise, patients suffering from AD, but not psoriasis or lichen planus, have a marked reduction of Notch receptor expression in the skin. Loss of Notch in keratinocytes induces the production of thymic stromal lymphopoietin (TSLP), a cytokine deeply implicated in the pathogenesis of AD. The AD-like associated inflammation is accompanied by a myeloproliferative disorder (MPD) characterized by an increase in immature myeloid populations in the bone marrow and spleen. Transplantation studies revealed that the MPD is cell non-autonomous and caused by dramatic microenvironmental alterations. Genetic studies demontrated that G-CSF mediates the MPD as well as changes in the bone marrow microenvironment leading to osteopenia.

Significance: Our data demonstrate a critical role for Notch in repressing TSLP production in keratinocytes, thereby maintaining integrity of the skin and the hematopoietic system.

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Related in: MedlinePlus

Loss of Notch signaling in post-natal epidermis leads to a severe form of atopic dermatitis and lethality.(A) Representative photograph of Control (Ctrl) and N1N2K5 mice 38 days post first injection of tamoxifen showing loss of hair, thick, dry, and scaly skin. (B) Survival curve of control (Ctrl, n = 20) and N1N2K5 (n = 24) mice after tamoxifen injection. The survival curve is the combined result of 3 individual experiments. (C) Southern blot analysis of genomic DNA from scraped epidermis from control (Ctrl, n = 2) and N1N2K5 mice (n = 3) showing the floxed and the recombined (Recomb) alleles of Notch1 and Notch2 respectively. The recombination efficiency is >70% for both genes. Three individual experiments were performed. (D) Representative HE staining on control (Ctrl) and N1N2K5 dorsal skin sections showing a thickened epidermal layer (e) a massively infiltrated dermis (d) with large epidermoid cysts (c) from degenerated hair follicles (hf) and absence of subcutis (sc) above the muscles (m). Asterisks indicate enlarged regions of the skin showing acanthosis, hyperkeratosis and spongiosis of the epidermis as well as eosinophil infiltrates (arrows) around dilated blood vessels (bv) in the dermis (n = 8, 4 individual experiments were performed). (E) Goldner's Trichrome (GT) readily shows the spongiosis and hyperkeratosis (n = 7, 4 individual experiments were performed). (F) Toluidine blue (TB) staining on control (Ctrl) and N1N2K5 skin sections showing massive infiltration of mast cells (dark blue) (n = 7, 4 individual experiments were performed). (G) Quantitative RT-PCR on dermis-derived RNA for the T helper specific cytokine IL-4 from Ctrl and N1N2K5 mice. The experiment was performed in triplicates (n = 3 per sample group, two individual experiments were performed). (H) A 16-fold increase in serum IgE levels is observed in N1N2K5 compared to Ctrl mice. The experiment was performed in triplicates (n = 3 per sample group, three individual experiments). (* p<0.01; ** p<0.001). [Scale bars: 50 µm].
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pone-0009258-g001: Loss of Notch signaling in post-natal epidermis leads to a severe form of atopic dermatitis and lethality.(A) Representative photograph of Control (Ctrl) and N1N2K5 mice 38 days post first injection of tamoxifen showing loss of hair, thick, dry, and scaly skin. (B) Survival curve of control (Ctrl, n = 20) and N1N2K5 (n = 24) mice after tamoxifen injection. The survival curve is the combined result of 3 individual experiments. (C) Southern blot analysis of genomic DNA from scraped epidermis from control (Ctrl, n = 2) and N1N2K5 mice (n = 3) showing the floxed and the recombined (Recomb) alleles of Notch1 and Notch2 respectively. The recombination efficiency is >70% for both genes. Three individual experiments were performed. (D) Representative HE staining on control (Ctrl) and N1N2K5 dorsal skin sections showing a thickened epidermal layer (e) a massively infiltrated dermis (d) with large epidermoid cysts (c) from degenerated hair follicles (hf) and absence of subcutis (sc) above the muscles (m). Asterisks indicate enlarged regions of the skin showing acanthosis, hyperkeratosis and spongiosis of the epidermis as well as eosinophil infiltrates (arrows) around dilated blood vessels (bv) in the dermis (n = 8, 4 individual experiments were performed). (E) Goldner's Trichrome (GT) readily shows the spongiosis and hyperkeratosis (n = 7, 4 individual experiments were performed). (F) Toluidine blue (TB) staining on control (Ctrl) and N1N2K5 skin sections showing massive infiltration of mast cells (dark blue) (n = 7, 4 individual experiments were performed). (G) Quantitative RT-PCR on dermis-derived RNA for the T helper specific cytokine IL-4 from Ctrl and N1N2K5 mice. The experiment was performed in triplicates (n = 3 per sample group, two individual experiments were performed). (H) A 16-fold increase in serum IgE levels is observed in N1N2K5 compared to Ctrl mice. The experiment was performed in triplicates (n = 3 per sample group, three individual experiments). (* p<0.01; ** p<0.001). [Scale bars: 50 µm].

Mentions: The function of Notch signaling during skin homeostasis was characterized using mice bearing homozygously floxed alleles for Notch1, for Notch2, for both Notch1 and Notch2 [17], and for RBP-J [18]. These mice were crossed to transgenic mice expressing a tamoxifen inducible Cre-recombinase (CreERT) under the control of the Keratin5 promoter [19] (hereafter: N1K5, N2K5, N1N2K5, RBP-JK5). Eight day-old mutant mice and corresponding littermate controls, lacking the CreERT transgene, were injected with tamoxifen for 5 consecutive days and analyzed 30 to 40 days post injection (Figure 1). Gene specific deletion efficiency was assessed by Southern blot analysis of genomic DNA isolated from the epidermis of the different gene targeted mice at the time point of analysis. The recombination efficiency in N1K5 and N2K5 was >80% (data not shown), and 70% in N1N2K5 mice for both the Notch1 and the Notch2 genes (Figure 1C). Postnatal skin specific inactivation of Notch1 resulted in loss of skin appendages, deregulation of several epidermal differentiation markers and hyperproliferation (Figure S1) as previously reported [5]. In contrast, postnatal skin-specific loss of Notch2 did not lead to any apparent phenotype (Figure S1), which is similar to data describing embryonic inactivation of Notch2 [7]. This suggests that either Notch2 has no function in murine skin, or that its loss is fully compensated by redundant Notch1 signaling. This question was addressed by analyzing mice with simultaneous inactivation of both Notch1 and Notch2 in the epidermis. Moreover, the phenotype of these mice was compared to mice in which RBP-J, the downstream mediator for all Notch receptors, was inactivated.


Atopic dermatitis-like disease and associated lethal myeloproliferative disorder arise from loss of Notch signaling in the murine skin.

Dumortier A, Durham AD, Di Piazza M, Vauclair S, Koch U, Ferrand G, Ferrero I, Demehri S, Song LL, Farr AG, Leonard WJ, Kopan R, Miele L, Hohl D, Finke D, Radtke F - PLoS ONE (2010)

Loss of Notch signaling in post-natal epidermis leads to a severe form of atopic dermatitis and lethality.(A) Representative photograph of Control (Ctrl) and N1N2K5 mice 38 days post first injection of tamoxifen showing loss of hair, thick, dry, and scaly skin. (B) Survival curve of control (Ctrl, n = 20) and N1N2K5 (n = 24) mice after tamoxifen injection. The survival curve is the combined result of 3 individual experiments. (C) Southern blot analysis of genomic DNA from scraped epidermis from control (Ctrl, n = 2) and N1N2K5 mice (n = 3) showing the floxed and the recombined (Recomb) alleles of Notch1 and Notch2 respectively. The recombination efficiency is >70% for both genes. Three individual experiments were performed. (D) Representative HE staining on control (Ctrl) and N1N2K5 dorsal skin sections showing a thickened epidermal layer (e) a massively infiltrated dermis (d) with large epidermoid cysts (c) from degenerated hair follicles (hf) and absence of subcutis (sc) above the muscles (m). Asterisks indicate enlarged regions of the skin showing acanthosis, hyperkeratosis and spongiosis of the epidermis as well as eosinophil infiltrates (arrows) around dilated blood vessels (bv) in the dermis (n = 8, 4 individual experiments were performed). (E) Goldner's Trichrome (GT) readily shows the spongiosis and hyperkeratosis (n = 7, 4 individual experiments were performed). (F) Toluidine blue (TB) staining on control (Ctrl) and N1N2K5 skin sections showing massive infiltration of mast cells (dark blue) (n = 7, 4 individual experiments were performed). (G) Quantitative RT-PCR on dermis-derived RNA for the T helper specific cytokine IL-4 from Ctrl and N1N2K5 mice. The experiment was performed in triplicates (n = 3 per sample group, two individual experiments were performed). (H) A 16-fold increase in serum IgE levels is observed in N1N2K5 compared to Ctrl mice. The experiment was performed in triplicates (n = 3 per sample group, three individual experiments). (* p<0.01; ** p<0.001). [Scale bars: 50 µm].
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2823782&req=5

pone-0009258-g001: Loss of Notch signaling in post-natal epidermis leads to a severe form of atopic dermatitis and lethality.(A) Representative photograph of Control (Ctrl) and N1N2K5 mice 38 days post first injection of tamoxifen showing loss of hair, thick, dry, and scaly skin. (B) Survival curve of control (Ctrl, n = 20) and N1N2K5 (n = 24) mice after tamoxifen injection. The survival curve is the combined result of 3 individual experiments. (C) Southern blot analysis of genomic DNA from scraped epidermis from control (Ctrl, n = 2) and N1N2K5 mice (n = 3) showing the floxed and the recombined (Recomb) alleles of Notch1 and Notch2 respectively. The recombination efficiency is >70% for both genes. Three individual experiments were performed. (D) Representative HE staining on control (Ctrl) and N1N2K5 dorsal skin sections showing a thickened epidermal layer (e) a massively infiltrated dermis (d) with large epidermoid cysts (c) from degenerated hair follicles (hf) and absence of subcutis (sc) above the muscles (m). Asterisks indicate enlarged regions of the skin showing acanthosis, hyperkeratosis and spongiosis of the epidermis as well as eosinophil infiltrates (arrows) around dilated blood vessels (bv) in the dermis (n = 8, 4 individual experiments were performed). (E) Goldner's Trichrome (GT) readily shows the spongiosis and hyperkeratosis (n = 7, 4 individual experiments were performed). (F) Toluidine blue (TB) staining on control (Ctrl) and N1N2K5 skin sections showing massive infiltration of mast cells (dark blue) (n = 7, 4 individual experiments were performed). (G) Quantitative RT-PCR on dermis-derived RNA for the T helper specific cytokine IL-4 from Ctrl and N1N2K5 mice. The experiment was performed in triplicates (n = 3 per sample group, two individual experiments were performed). (H) A 16-fold increase in serum IgE levels is observed in N1N2K5 compared to Ctrl mice. The experiment was performed in triplicates (n = 3 per sample group, three individual experiments). (* p<0.01; ** p<0.001). [Scale bars: 50 µm].
Mentions: The function of Notch signaling during skin homeostasis was characterized using mice bearing homozygously floxed alleles for Notch1, for Notch2, for both Notch1 and Notch2 [17], and for RBP-J [18]. These mice were crossed to transgenic mice expressing a tamoxifen inducible Cre-recombinase (CreERT) under the control of the Keratin5 promoter [19] (hereafter: N1K5, N2K5, N1N2K5, RBP-JK5). Eight day-old mutant mice and corresponding littermate controls, lacking the CreERT transgene, were injected with tamoxifen for 5 consecutive days and analyzed 30 to 40 days post injection (Figure 1). Gene specific deletion efficiency was assessed by Southern blot analysis of genomic DNA isolated from the epidermis of the different gene targeted mice at the time point of analysis. The recombination efficiency in N1K5 and N2K5 was >80% (data not shown), and 70% in N1N2K5 mice for both the Notch1 and the Notch2 genes (Figure 1C). Postnatal skin specific inactivation of Notch1 resulted in loss of skin appendages, deregulation of several epidermal differentiation markers and hyperproliferation (Figure S1) as previously reported [5]. In contrast, postnatal skin-specific loss of Notch2 did not lead to any apparent phenotype (Figure S1), which is similar to data describing embryonic inactivation of Notch2 [7]. This suggests that either Notch2 has no function in murine skin, or that its loss is fully compensated by redundant Notch1 signaling. This question was addressed by analyzing mice with simultaneous inactivation of both Notch1 and Notch2 in the epidermis. Moreover, the phenotype of these mice was compared to mice in which RBP-J, the downstream mediator for all Notch receptors, was inactivated.

Bottom Line: The AD-like associated inflammation is accompanied by a myeloproliferative disorder (MPD) characterized by an increase in immature myeloid populations in the bone marrow and spleen.Genetic studies demontrated that G-CSF mediates the MPD as well as changes in the bone marrow microenvironment leading to osteopenia.Our data demonstrate a critical role for Notch in repressing TSLP production in keratinocytes, thereby maintaining integrity of the skin and the hematopoietic system.

View Article: PubMed Central - PubMed

Affiliation: Ecole Polytechnique Fédérale de Lausanne (EPFL SV ISREC), Lausanne, Switzerland.

ABSTRACT

Background: The Notch pathway is essential for proper epidermal differentiation during embryonic skin development. Moreover, skin specific loss of Notch signaling in the embryo results in skin barrier defects accompanied by a B-lymphoproliferative disease. However, much less is known about the consequences of loss of Notch signaling after birth.

Methodology and principal findings: To study the function of Notch signaling in the skin of adult mice, we made use of a series of conditional gene targeted mice that allow inactivation of several components of the Notch signaling pathway specifically in the skin. We demonstrate that skin-specific inactivation of Notch1 and Notch2 simultaneously, or RBP-J, induces the development of a severe form of atopic dermatitis (AD), characterized by acanthosis, spongiosis and hyperkeratosis, as well as a massive dermal infiltration of eosinophils and mast cells. Likewise, patients suffering from AD, but not psoriasis or lichen planus, have a marked reduction of Notch receptor expression in the skin. Loss of Notch in keratinocytes induces the production of thymic stromal lymphopoietin (TSLP), a cytokine deeply implicated in the pathogenesis of AD. The AD-like associated inflammation is accompanied by a myeloproliferative disorder (MPD) characterized by an increase in immature myeloid populations in the bone marrow and spleen. Transplantation studies revealed that the MPD is cell non-autonomous and caused by dramatic microenvironmental alterations. Genetic studies demontrated that G-CSF mediates the MPD as well as changes in the bone marrow microenvironment leading to osteopenia.

Significance: Our data demonstrate a critical role for Notch in repressing TSLP production in keratinocytes, thereby maintaining integrity of the skin and the hematopoietic system.

Show MeSH
Related in: MedlinePlus