Effective Treatment of Psoriasis with Narrow-Band UVB Phototherapy Is Linked to Suppression of the IFN and Th17 Pathways  Emőke Rácz, Errol P. Prens, Dorota Kurek, Marius Kant, Dick de Ridder, Sabine Mourits, Ewout M. Baerveldt, Zeliha Ozgur, Wilfred F.J. van IJcken, Jon D. Laman, Frank J. Staal, Leslie van der Fits  Journal of Investigative Dermatology  Volume 131, Issue 7, Pages 1547-1558 (July 2011) DOI: 10.1038/jid.2011.53 Copyright © 2011 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 1 Correlation view of expression profiles of the different epidermal RNA samples. The degree of similarity between global gene expression profiles was assessed using the OmniViz package. The red squares indicate positive pairwise correlations (equality in gene expression between clusters) and blue squares indicate negative pairwise correlations. A, B, patient pools; D, sample taken at 50% psoriasis area and severity index (PASI) reduction; E, sample taken after completion of narrow-band ultraviolet-B (NB-UVB) therapy; L, lesional samples; NL, nonlesional samples; T0, sample taken before the first irradiation. Journal of Investigative Dermatology 2011 131, 1547-1558DOI: (10.1038/jid.2011.53) Copyright © 2011 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 2 Successful narrow-band ultraviolet-B (NB-UVB) therapy of psoriasis is associated with suppression of the Th17 pathway. (a) Th17 pathway. Gray indicates gene downregulation. (b) Epidermal S100A9 (S100 calcium binding protein A9) mRNA expression was measured by real-time PCR (RT-PCR). ABL1 (Abelson murine leukemia viral (v-abl) oncogene homolog 1) was used as an internal control gene. Black bars show lesional and gray bars show nonlesional psoriatic samples; psoriasis area and severity index (PASI) 50% is sample taken at 50% PASI score reduction. Error bars indicate SEM, n=4 patients. (c) β-Defensin-2 in psoriasis during NB-UVB therapy, as determined by immunofluorescent staining with a monoclonal anti-β-defensin-2 antibody in lesional skin biopsies taken before, during, and after NB-UVB therapy. Scale bar=200μm. (d–f) A single NB-UVB irradiation counteracts the effects of IL-22 in vitro. Normal skin biopsies were obtained from healthy volunteers. Biopsies were cultured with or without recombinant human IL-22 (50ngml−1) in the culture medium. After 16hours in culture, biopsies were irradiated with a single NB-UVB dose of 600mJcm−2. The cultured biopsies were collected 6hours after UV irradiation. Phosphorylated signal transducer and activator of transcription 3 (STAT3) staining of the collected biopsies was performed using a monoclonal phospho-STAT3 antibody. Phospho-STAT3-positive cells in the epidermis were counted, and represented as mean±SEM (n=4) (e) and a representative example is shown (d). Scale bar for panel d=20μm. (f) Epidermal β-defensin-2 mRNA expression was measured by real-time PCR (RT-PCR). ABL1 was used as an internal control gene. Error bars indicate SEM, n=4 subjects. Journal of Investigative Dermatology 2011 131, 1547-1558DOI: (10.1038/jid.2011.53) Copyright © 2011 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 3 Successful narrow-band ultraviolet-B (NB-UVB) therapy of psoriasis is accompanied by inhibition of IFN signaling pathways and induction of normal epidermal differentiation. (a) IFN signaling pathways. Gray color indicates gene downregulation. (b, d) Epidermal IFN induced with helicase C domain 1/melanoma differentiation associated gene 5 (IFIH1/MDA5; b) and corneodesmosin (CDSN; d) mRNA expression was measured by real-time PCR (RT-PCR). ABL1 (Abelson murine leukemia viral (v-abl) oncogene homolog 1) was used as an internal control gene. Black bars show lesional and gray bars show nonlesional psoriatic samples; psoriasis area and severity index (PASI) 50% is sample taken at 50% PASI score reduction. Error bars indicate SEM, n=5 patients. (c, e) MxA (c) and transglutaminase K (e) protein expression (red staining) during NB-UVB therapy, as determined by immunostaining in lesional psoriatic skin samples taken before, during, and after NB-UVB therapy. Representative examples of five patients are shown. Scale bar=100μm. Journal of Investigative Dermatology 2011 131, 1547-1558DOI: (10.1038/jid.2011.53) Copyright © 2011 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 4 Improvement of the imiquimod-induced psoriasiform dermatitis in mice by narrow-band ultraviolet-B (NB-UVB) is accompanied by a reduction of epidermal phospho-signal transducer and activator of transcription 3 (STAT3) staining and by induction of normal differentiation. BALB/c mice were treated daily with IMQ cream or control cream on the shaved back skin, and irradiated or sham-irradiated every other day with NB-UVB, starting on the first day of imiquimod treatment. (a–c) Mice were killed on day 6. Imiquimod-induced inflammation was studied on sections made from the back skin of the mice. Immunohistochemical staining of myeloid dendritic cells (CD11c, a), granulocytes (GR1, b), and endothelial cells (MECA-20, c) is shown. Scale bars=100μm. (d) Erythema, scaling, and thickness of the back skin were scored daily on a scale from 0 to 4. The cumulative score (erythema plus scaling plus thickness) is shown. Symbols indicate mean score±SEM of three mice per group. (e) The number of phospho-STAT3-positive cells were counted in three photos made of different sections by two independent researchers, and are shown relative to the total number of epidermal cells per section. Error bars indicate the SEM. (f) Immunofluorescent staining for phosphorylated STAT3 of the back skin of the mice. Scale bar=20μm. (g) Keratin 1/10 immunofluorescent staining of the back skin is shown. Scale bar=100μm. Journal of Investigative Dermatology 2011 131, 1547-1558DOI: (10.1038/jid.2011.53) Copyright © 2011 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 5 Short-term epidermal effects of narrow-band ultraviolet-B (NB-UVB). (a) Epidermal mRNA expression of cryptochrome 1 (CRY1) before and 6hours after the first irradiation with NB-UVB (UVB dose was 70% of the minimal erythema dose (MED)), as measured by real-time PCR (RT-PCR). ABL1 (Abelson murine leukemia viral (v-abl) oncogene homolog 1) was used as an internal control gene. Black bars: before irradiation, gray bars: 6hours after irradiation. Error bars indicate SEM, n=6 patients. (b, c) Immunohistochemical staining for cyclobutane pyrimidine dimers (CPDs) in lesional psoriatic skin (b) before and (c) 15minutes after irradiation with 70% MED. Dotted line indicates dermal/epidermal junction. Scale bars=200μm. Journal of Investigative Dermatology 2011 131, 1547-1558DOI: (10.1038/jid.2011.53) Copyright © 2011 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 6 Comparison of epidermal gene modulation in nonlesional skin of narrow-band ultraviolet-B (NB-UVB)- and ustekinumab-treated patients with psoriasis. Nonlesional epidermal samples were collected from patients with psoriasis before treatment and at 50% psoriasis area and severity index (PASI) reduction after treatment with NB-UVB or with ustekinumab, a monoclonal anti-IL12/23p40 antibody. The expression of (a) S100 calcium binding protein A7 (S100A7), (b) β-defensin-2, (c) IFN induced with helicase C domain 1 (IFIH1), and (d) GATA3 mRNA was quantified using real-time PCR (RT-PCR), with Abelson murine leukemia viral (v-abl) oncogene homolog 1 (ABL1) as an internal control gene (in ustekinumab-treated patients, n=8) or with microarray (NB-UVB-treated patients, n=2, pools of 4 patients each). Expression values at 50% PASI reduction are shown as percentages of the expression values before treatment in both sets of data. Journal of Investigative Dermatology 2011 131, 1547-1558DOI: (10.1038/jid.2011.53) Copyright © 2011 The Society for Investigative Dermatology, Inc Terms and Conditions