Gene Profiling of Narrowband UVB–Induced Skin Injury Defines Cellular and Molecular Innate Immune Responses Milène Kennedy Crispin, Judilyn Fuentes-Duculan,

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Gene Profiling of Narrowband UVB–Induced Skin Injury Defines Cellular and Molecular Innate Immune Responses Milène Kennedy Crispin, Judilyn Fuentes-Duculan, Nicholas Gulati, Leanne M. Johnson-Huang, Tim Lentini, Mary Sullivan-Whalen, Patricia Gilleaudeau, Inna Cueto, Mayte Suárez-Fariñas, Michelle A. Lowes, James G. Krueger Journal of Investigative Dermatology Volume 133, Issue 3, Pages 692-701 (March 2013) DOI: 10.1038/jid.2012.359 Copyright © 2013 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 1 Differentially expressed antimicrobial peptides and chemokines were validated by immunohistochemistry or quantitative real-time reverse-transcriptase–PCR (qRT-PCR). (a) Representative immunohistochemistry for (i) S100A7, (ii) S100A12, (iii) S100A8/9, (iv) HBD-2, and (v) elafin showed increased S100A7, S100A12, HBD-2, and elafin staining in the epidermis and increased S100A8/9 and HBD-2 in the dermis of irradiated samples. Arrow indicates (i) focal epidermal expression of S100A7 and (ii) S100A12+ dermal cell. (b) qRT-PCR using messenger RNA from paired nonirradiated and irradiated skin (n=9) showed increased expression of (i) CCL2, (ii) CXCL1, (iii) IL-8, and (iv) IL-10 in irradiated skin. All data were normalized to human acidic ribosomal protein housekeeping gene. Error bars indicate SEM. I, irradiated skin; NI, nonirradiated skin. Bars=100μm. **P<0.01, ***P<0.001. Journal of Investigative Dermatology 2013 133, 692-701DOI: (10.1038/jid.2012.359) Copyright © 2013 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 2 Irradiated (I) skin contained increased CD11c+ dendritic cells and CD163+ macrophages. Representative immunohistochemistry and cell counts showed (a, c) increased CD11c+ dendritic cells and (b, c) increased CD163+ macrophages in the dermis of irradiated skin compared with paired nonirradiated (NI) skin. (d) There was no coexpression between CD11c+ myeloid dendritic cells (red) and CD163+ macrophages (green) in irradiated skin. Bar=100μm. **P<0.01. Journal of Investigative Dermatology 2013 133, 692-701DOI: (10.1038/jid.2012.359) Copyright © 2013 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 3 CD11c+ BDCA1− and CD11c+ BDCA3+ dendritic cells (DCs) were increased in NB-UVB-irradiated (I) skin. Representative immunohistochemistry, cell counts, and IF for DC surface markers CD11c, BDCA1, and BDCA3 in nonirradiated (NI) and irradiated (I) skin (n=8–10) showed that (a) BDCA1+ cells decreased 2-fold in I skin, with (b) decreased colocalization of CD11c and BDCA1 in I skin compared with NI skin. (c) BDCA3+ cells increased 4.5-fold. (d) Most BDCA-3+ cells colocalized with CD11c in I skin (yellow cells), but there were many CD11c+ cells that did not colocalize with BDCA-3 (red cells). (e) Inflammatory DCs (CD11c+ BDCA1− BDCA3−) were elevated in I versus NI skin, as calculated by subtracting BDCA1+ and BDCA3+ cells from CD11c+ cells. Bar=100μm. *P<0.05, **P<0.01. Journal of Investigative Dermatology 2013 133, 692-701DOI: (10.1038/jid.2012.359) Copyright © 2013 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 4 CD11c+ cells were immature and expressed inflammatory markers tumor necrosis factor (TNF) and TNF–related apoptosis-inducing ligand (TRAIL). (a) Representative IF showed occasional colocalization of CD11c+ cells with maturation marker DC-LAMP in irradiated skin, and (b) many BDCA-1+ cells expressed DC-LAMP after irradiation. UVB-irradiated skin had increased colocalization of CD11c and (c, d) TNF and (e, f) TRAIL, compared with nonirradiated skin. Bar=100μm. Journal of Investigative Dermatology 2013 133, 692-701DOI: (10.1038/jid.2012.359) Copyright © 2013 The Society for Investigative Dermatology, Inc Terms and Conditions