Ekatherina Vassina, Martin Leverkus, Shida Yousefi, Lasse R

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Increased Expression and a Potential Anti-Inflammatory Role of TRAIL in Atopic Dermatitis Ekatherina Vassina, Martin Leverkus, Shida Yousefi, Lasse R. Braathen, Hans-Uwe Simon, Dagmar Simon Journal of Investigative Dermatology Volume 125, Issue 4, Pages 746-752 (October 2005) DOI: 10.1111/j.0022-202X.2005.23878.x Copyright © 2005 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 1 Expression of the tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) in different subsets of blood cells. Cells were isolated from atopic dermatitis patients and normal donors. Monocytes, CD8+ cells, and CD4+ cells were analyzed within PBMC and identified using anti-CD14, anti-CD8, and anti-CD4 monoclonal antibody (mAb), respectively. TRAIL expression on neutrophils and eosinophils was analyzed using purified cell populations. TRAIL expression was measured by flow cytometry using mouse anti-TRAIL mAb (red). Control IgG1 mAb staining is shown in gray. Three representative and independent experiments for each cell type. Statistical analysis of these experiments is provided in Table I. Journal of Investigative Dermatology 2005 125, 746-752DOI: (10.1111/j.0022-202X.2005.23878.x) Copyright © 2005 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 2 Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) expression in atopic dermatitis (AD). (A) Representative double immunofluorescence stainings of skin biopsy specimens from AD patients using CD4+, CD8+, and TRAIL mAb. White bars, 10 μM. (B) Bars indicate mean numbers of total and TRAIL-positive CD4+ and CD8+ cells of 10 patients in the dermal infiltrate of lesional skin before and after treatment with topical tacrolimus. Journal of Investigative Dermatology 2005 125, 746-752DOI: (10.1111/j.0022-202X.2005.23878.x) Copyright © 2005 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 3 Representative immunofluorescence stainings of interleukin-1 receptor antagonist (IL-1Ra) in the skin. Staining with an anti-IL-1RA Ab reveals strong positive cells throughout the epidermis of lesional AD skin. Nuclei were counterstained with propidium iodide. After treatment, few keratinocytes in the suprabasal layers stained weakly positive. Weak staining was also observed in normal skin. The white dashed line indicates the border between the epidermis and the dermis. The original magnification is indicated in each panel. Journal of Investigative Dermatology 2005 125, 746-752DOI: (10.1111/j.0022-202X.2005.23878.x) Copyright © 2005 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 4 Interleukin-1 receptor antagonist (IL-1Ra) expression following tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) stimulation of keratinocytes. (A) Following preincubation with either diluent alone or 40 μM zVAD-fmk for 1 h, HaCaT cells were treated with recombinant TRAIL (500 ng per mL) for the indicated times. Multiprobe RPA (RiboQuant) were performed using 5 μg of total RNA. Steady-state levels for IL-1Ra were determined by normalizing against the expression of the L32 housekeeping gene. (B) TRAIL-induced IL-1Ra secretion in HaCaT cells. Cells were preincubated with either diluent alone (control) in the presence or absence of 10 μg per mL of TRAIL-R2-Fc or control IgG for 1 h, and subsequently treated with 500 ng per mL recombinant TRAIL for 24 h. Cell-free supernatants were collected and assayed for IL-1Ra by ELISA. (C) TRAIL-induced IL-1Ra secretion in primary keratinocytes. Cells were preincubated with either diluent alone (control) in the presence or absence of 10 μg per mL of TRAIL-R2-Fc or control IgG for 1 h, and subsequently treated with the indicated concentrations of recombinant TRAIL for 24 h. Cell-free supernatants were collected and assayed for IL-1Ra by ELISA. Two independent experiments are shown. Journal of Investigative Dermatology 2005 125, 746-752DOI: (10.1111/j.0022-202X.2005.23878.x) Copyright © 2005 The Society for Investigative Dermatology, Inc Terms and Conditions