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Interferon-γ links ultraviolet radiation to melanomagenesis in mice.

Zaidi MR, Davis S, Noonan FP, Graff-Cherry C, Hawley TS, Walker RL, Feigenbaum L, Fuchs E, Lyakh L, Young HA, Hornyak TJ, Arnheiter H, Trinchieri G, Meltzer PS, De Fabo EC, Merlino G - Nature (2011)

Bottom Line: We use expression profiling to show that activated neonatal skin melanocytes isolated following a melanomagenic UVB dose bear a distinct, persistent interferon response signature, including genes associated with immunoevasion.UVB-induced melanocyte activation, characterized by aberrant growth and migration, was abolished by antibody-mediated systemic blockade of interferon-γ (IFN-γ), but not type-I interferons.IFN-γ was produced by macrophages recruited to neonatal skin by UVB-induced ligands to the chemokine receptor Ccr2.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Cancer Biology and Genetics, National Cancer Institute, Bethesda, Maryland 20892, USA.

ABSTRACT
Cutaneous malignant melanoma is a highly aggressive and frequently chemoresistant cancer, the incidence of which continues to rise. Epidemiological studies show that the major aetiological melanoma risk factor is ultraviolet (UV) solar radiation, with the highest risk associated with intermittent burning doses, especially during childhood. We have experimentally validated these epidemiological findings using the hepatocyte growth factor/scatter factor transgenic mouse model, which develops lesions in stages highly reminiscent of human melanoma with respect to biological, genetic and aetiological criteria, but only when irradiated as neonatal pups with UVB, not UVA. However, the mechanisms underlying UVB-initiated, neonatal-specific melanomagenesis remain largely unknown. Here we introduce a mouse model permitting fluorescence-aided melanocyte imaging and isolation following in vivo UV irradiation. We use expression profiling to show that activated neonatal skin melanocytes isolated following a melanomagenic UVB dose bear a distinct, persistent interferon response signature, including genes associated with immunoevasion. UVB-induced melanocyte activation, characterized by aberrant growth and migration, was abolished by antibody-mediated systemic blockade of interferon-γ (IFN-γ), but not type-I interferons. IFN-γ was produced by macrophages recruited to neonatal skin by UVB-induced ligands to the chemokine receptor Ccr2. Admixed recruited skin macrophages enhanced transplanted melanoma growth by inhibiting apoptosis; notably, IFN-γ blockade abolished macrophage-enhanced melanoma growth and survival. IFN-γ-producing macrophages were also identified in 70% of human melanomas examined. Our data reveal an unanticipated role for IFN-γ in promoting melanocytic cell survival/immunoevasion, identifying a novel candidate therapeutic target for a subset of melanoma patients.

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UVB-induced melanocyte activation is mediated by interferon-γa, Unsupervised clustering of cDNA microarray analysis of gene expression in FACS-sorted melanocytes from 1 day (P2) or 6 days (P7) following UVB or UVA irradiation, and respective unirradiated controls. The expanded heatmap (right) shows the delayed induced gene subset, which includes multiple genes known to be induced by IFN-γ. Primary mouse keratinocytes (PrKC) were included as controls. All groups included biological triplicates. b, qRT-PCR validation of expression of 4 genes (n=3 samples each) from IFN signature (error bars = s.e.m.). c, Antibody-mediated blockade of interferons by treating pups with intraperitoneal injections of anti-IFN-αR1, anti-IFN-γ, or both in combination, 1 h prior to and 3 days after UVB irradiation at P1. The dorsal skins were harvested (n=3 each group) and analyzed for melanocyte activation. Representative images are shown. E, epidermis; D, dermis. Scale bars = 40 μm.
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Figure 2: UVB-induced melanocyte activation is mediated by interferon-γa, Unsupervised clustering of cDNA microarray analysis of gene expression in FACS-sorted melanocytes from 1 day (P2) or 6 days (P7) following UVB or UVA irradiation, and respective unirradiated controls. The expanded heatmap (right) shows the delayed induced gene subset, which includes multiple genes known to be induced by IFN-γ. Primary mouse keratinocytes (PrKC) were included as controls. All groups included biological triplicates. b, qRT-PCR validation of expression of 4 genes (n=3 samples each) from IFN signature (error bars = s.e.m.). c, Antibody-mediated blockade of interferons by treating pups with intraperitoneal injections of anti-IFN-αR1, anti-IFN-γ, or both in combination, 1 h prior to and 3 days after UVB irradiation at P1. The dorsal skins were harvested (n=3 each group) and analyzed for melanocyte activation. Representative images are shown. E, epidermis; D, dermis. Scale bars = 40 μm.

Mentions: We performed an expression microarray study on melanocytes isolated from dorsal skin of iDct-GFP pups irradiated at P1 with UVB or UVA (Fig. 1e). Doxycycline-induced GFP-labeled melanocytes were isolated via fluorescence-activated cell sorting (FACS) at 1-day and 6-days post-irradiation (ages P2 and P7, respectively); arrays from 1-day post-UV would reflect the acute UV stress response of in vivo melanocytes, while the 6-day post-UV time point should uncover responses persisting after the acute stress response subsides. FACS isolation consistently yielded >95% melanocyte enrichment (Supplementary Fig. 5). Gene expression profiling produced robust data with good reproducibility among biological triplicates (Fig. 2a), and confirmed the absence of detectable levels of contaminating skin cell types, including keratinocytes, fibroblasts and adipocytes (Supplementary Fig. 6).


Interferon-γ links ultraviolet radiation to melanomagenesis in mice.

Zaidi MR, Davis S, Noonan FP, Graff-Cherry C, Hawley TS, Walker RL, Feigenbaum L, Fuchs E, Lyakh L, Young HA, Hornyak TJ, Arnheiter H, Trinchieri G, Meltzer PS, De Fabo EC, Merlino G - Nature (2011)

UVB-induced melanocyte activation is mediated by interferon-γa, Unsupervised clustering of cDNA microarray analysis of gene expression in FACS-sorted melanocytes from 1 day (P2) or 6 days (P7) following UVB or UVA irradiation, and respective unirradiated controls. The expanded heatmap (right) shows the delayed induced gene subset, which includes multiple genes known to be induced by IFN-γ. Primary mouse keratinocytes (PrKC) were included as controls. All groups included biological triplicates. b, qRT-PCR validation of expression of 4 genes (n=3 samples each) from IFN signature (error bars = s.e.m.). c, Antibody-mediated blockade of interferons by treating pups with intraperitoneal injections of anti-IFN-αR1, anti-IFN-γ, or both in combination, 1 h prior to and 3 days after UVB irradiation at P1. The dorsal skins were harvested (n=3 each group) and analyzed for melanocyte activation. Representative images are shown. E, epidermis; D, dermis. Scale bars = 40 μm.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC3140101&req=5

Figure 2: UVB-induced melanocyte activation is mediated by interferon-γa, Unsupervised clustering of cDNA microarray analysis of gene expression in FACS-sorted melanocytes from 1 day (P2) or 6 days (P7) following UVB or UVA irradiation, and respective unirradiated controls. The expanded heatmap (right) shows the delayed induced gene subset, which includes multiple genes known to be induced by IFN-γ. Primary mouse keratinocytes (PrKC) were included as controls. All groups included biological triplicates. b, qRT-PCR validation of expression of 4 genes (n=3 samples each) from IFN signature (error bars = s.e.m.). c, Antibody-mediated blockade of interferons by treating pups with intraperitoneal injections of anti-IFN-αR1, anti-IFN-γ, or both in combination, 1 h prior to and 3 days after UVB irradiation at P1. The dorsal skins were harvested (n=3 each group) and analyzed for melanocyte activation. Representative images are shown. E, epidermis; D, dermis. Scale bars = 40 μm.
Mentions: We performed an expression microarray study on melanocytes isolated from dorsal skin of iDct-GFP pups irradiated at P1 with UVB or UVA (Fig. 1e). Doxycycline-induced GFP-labeled melanocytes were isolated via fluorescence-activated cell sorting (FACS) at 1-day and 6-days post-irradiation (ages P2 and P7, respectively); arrays from 1-day post-UV would reflect the acute UV stress response of in vivo melanocytes, while the 6-day post-UV time point should uncover responses persisting after the acute stress response subsides. FACS isolation consistently yielded >95% melanocyte enrichment (Supplementary Fig. 5). Gene expression profiling produced robust data with good reproducibility among biological triplicates (Fig. 2a), and confirmed the absence of detectable levels of contaminating skin cell types, including keratinocytes, fibroblasts and adipocytes (Supplementary Fig. 6).

Bottom Line: We use expression profiling to show that activated neonatal skin melanocytes isolated following a melanomagenic UVB dose bear a distinct, persistent interferon response signature, including genes associated with immunoevasion.UVB-induced melanocyte activation, characterized by aberrant growth and migration, was abolished by antibody-mediated systemic blockade of interferon-γ (IFN-γ), but not type-I interferons.IFN-γ was produced by macrophages recruited to neonatal skin by UVB-induced ligands to the chemokine receptor Ccr2.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Cancer Biology and Genetics, National Cancer Institute, Bethesda, Maryland 20892, USA.

ABSTRACT
Cutaneous malignant melanoma is a highly aggressive and frequently chemoresistant cancer, the incidence of which continues to rise. Epidemiological studies show that the major aetiological melanoma risk factor is ultraviolet (UV) solar radiation, with the highest risk associated with intermittent burning doses, especially during childhood. We have experimentally validated these epidemiological findings using the hepatocyte growth factor/scatter factor transgenic mouse model, which develops lesions in stages highly reminiscent of human melanoma with respect to biological, genetic and aetiological criteria, but only when irradiated as neonatal pups with UVB, not UVA. However, the mechanisms underlying UVB-initiated, neonatal-specific melanomagenesis remain largely unknown. Here we introduce a mouse model permitting fluorescence-aided melanocyte imaging and isolation following in vivo UV irradiation. We use expression profiling to show that activated neonatal skin melanocytes isolated following a melanomagenic UVB dose bear a distinct, persistent interferon response signature, including genes associated with immunoevasion. UVB-induced melanocyte activation, characterized by aberrant growth and migration, was abolished by antibody-mediated systemic blockade of interferon-γ (IFN-γ), but not type-I interferons. IFN-γ was produced by macrophages recruited to neonatal skin by UVB-induced ligands to the chemokine receptor Ccr2. Admixed recruited skin macrophages enhanced transplanted melanoma growth by inhibiting apoptosis; notably, IFN-γ blockade abolished macrophage-enhanced melanoma growth and survival. IFN-γ-producing macrophages were also identified in 70% of human melanomas examined. Our data reveal an unanticipated role for IFN-γ in promoting melanocytic cell survival/immunoevasion, identifying a novel candidate therapeutic target for a subset of melanoma patients.

Show MeSH
Related in: MedlinePlus