Diana Rodijk-Olthuis

Psoriasis is associated with increased beta-defensin genomic copy number

Psoriasis is a common inflammatory skin disease with a strong genetic component. We analyzed the genomic copy number polymorphism of the β-defensin region on human chromosome 8 in 179 Dutch individuals with psoriasis and 272 controls and in 319 German individuals with psoriasis and 305 controls. Comparisons in both cohorts showed a significant association between higher genomic copy number for β-defensin genes and risk of psoriasis.

β-Defensin-2 Protein Is a Serum Biomarker for Disease Activity in Psoriasis and Reaches Biologically Relevant Concentrations in Lesional Skin

Background Previous studies have extensively documented antimicrobial and chemotactic activities of beta-defensins. Human beta-defensin-2 (hBD-2) is strongly expressed in lesional psoriatic epidermis, and recently we have shown that high beta-defensin genomic copy number is associated with psoriasis susceptibility. It is not known, however, if biologically and pathophysiologically relevant concentrations of hBD-2 protein are present in vivo, which could support an antimicrobial and proinflammatory role of beta-defensins in lesional psoriatic epidermis. Methodology/Principal Findings We found that systemic levels of hBD-2 showed a weak but significant correlation with beta defensin copy number in healthy controls but not in psoriasis patients with active disease. In psoriasis patients but not in atopic dermatitis patients, we found high systemic hBD-2 levels that strongly correlated with disease activity as assessed by the PASI score. Our findings suggest that systemic levels in psoriasis are largely determined by secretion from involved skin and not by genomic copy number. Modelling of the in vivo epidermal hBD-2 concentration based on the secretion rate in a reconstructed skin model for psoriatic epidermis provides evidence that epidermal hBD-2 levels in vivo are probably well above the concentrations required for in vitro antimicrobial and chemokine-like effects. Conclusions/Significance Serum hBD-2 appears to be a useful surrogate marker for disease activity in psoriasis. The discrepancy between hBD-2 levels in psoriasis and atopic dermatitis could explain the well known differences in infection rate between these two diseases.

Psoriasis Risk Genes of the Late Cornified Envelope-3 Group Are Distinctly Expressed Compared with Genes of Other LCE Groups

Deletion of the late cornified envelope (LCE) genes LCE3B and LCE3C has recently been identified as a risk factor for psoriasis. Expression of 16 LCE genes of LCE groups 1, 2, 3, 5, and 6 was examined in vivo and in vitro. Quantitative PCR demonstrated that moderate to high LCE expression was largely confined to skin and a few oropharyngeal tissues. Genes of the LCE3 group demonstrated increased expression in lesional psoriatic epidermis and were induced after superficial injury of normal skin, whereas expression of members of other LCE groups was down-regulated under these conditions. Immunohistochemistry and immunoelectron microscopy demonstrated that LCE2 protein expression was restricted to the uppermost granular layer and the stratum corneum. Stimulation of in vitro reconstructed skin by several psoriasis-associated cytokines resulted in induction of LCE3 members. The data suggest that LCE proteins of groups 1, 2, 5, and 6 are involved in normal skin barrier function, whereas LCE3 genes encode proteins involved in barrier repair after injury or inflammation. These findings may provide clues to the mechanistic role of LCE3B/C deletion in psoriasis.

Molecular diagnostics of psoriasis, atopic dermatitis, allergic contact dermatitis and irritant contact dermatitis.

Background  Microarray studies on the epidermal transcriptome in psoriasis and atopic dermatitis (AD) have revealed genes with disease‐specific expression in keratinocytes of lesional epidermis. These genes are possible candidates for disease‐specific pathogenetic changes, but could also provide a tool for molecular diagnostics of inflammatory skin conditions in general.

Expression of the Vanin Gene Family in Normal and Inflamed Human Skin: Induction by Proinflammatory Cytokines

The vanin gene family encodes secreted and membrane-bound ectoenzymes that convert pantetheine into pantothenic acid and cysteamine. Recent studies in a mouse colitis model indicated that vanin-1 has proinflammatory activity and suggest that pantetheinases are potential therapeutic targets in inflammatory diseases. In a microarray analysis of epidermal gene expression of psoriasis and atopic dermatitis lesions, we identified vanin-3 as the gene showing the highest differential expression of all annotated genes that we studied (19-fold upregulation in psoriasis). Quantitative real-time PCR analysis confirmed the microarray data on vanin-3 and showed similar induction of vanin-1, but not of vanin-2, in psoriatic epidermis. Immunohistochemistry showed that vanin-3 is expressed in the differentiated epidermal layers. Using submerged and organotypic keratinocyte cultures, we found that vanin-1 and vanin-3 are induced at the mRNA and protein level by psoriasis-associated proinflammatory cytokines (Th17/Th1) but not by Th2 cytokines. We hypothesize that increased levels of pantetheinase activity are part of the inflammatory-regenerative epidermal differentiation program, and may contribute to the phenotype observed in psoriasis.

A Comprehensive Analysis of Pattern Recognition Receptors in Normal and Inflamed Human Epidermis: Upregulation of Dectin-1 in Psoriasis

Human epidermis plays an important role in host defense by acting as a physical barrier and signaling interface between the environment and the immune system. Pattern recognition receptors (PRRs) are crucial to maintain homeostasis and provide protection during infection, but are also causally involved in monogenic auto-inflammatory diseases. This study aimed to investigate the epidermal expression of PRRs and several associated host defense molecules in healthy human skin, psoriasis, and atopic dermatitis (AD). Using microarray analysis and real-time quantitative PCR, we found that many of these genes are transcribed in normal human epidermis. Only a few genes were differentially induced in psoriasis (CLEC7A (dectin-1), Toll-like receptor (TLR) 4, and mannose receptor C type 1 (MRC1)) or AD (MRC1, IL1RN, and IL1β) compared with normal epidermis. A remarkably high expression of dectin-1 mRNA was observed in psoriatic epidermis and this was corroborated by immunohistochemistry. In cultured primary human keratinocytes, dectin-1 expression was induced by IFN-γ, IFN-α, and Th17 cytokines. Keratinocytes were unresponsive, however, to dectin-1 ligands such as β-glucan or heat-killed Candida albicans, nor did we observe synergy with TLR2/TLR5 ligands. In conclusion, upregulation of dectin-1 in psoriatic lesions seems to be under control of psoriasis-associated cytokines. Its role in the biology of skin inflammation and infection remains to be explored.

Genetically Programmed Differences in Epidermal Host Defense between Psoriasis and Atopic Dermatitis Patients

In the past decades, chronic inflammatory diseases such as psoriasis, atopic dermatitis, asthma, Crohn’s disease and celiac disease were generally regarded as immune-mediated conditions involving activated T-cells and proinflammatory cytokines produced by these cells. This paradigm has recently been challenged by the finding that mutations and polymorphisms in epithelium-expressed genes involved in physical barrier function or innate immunity, are risk factors of these conditions. We used a functional genomics approach to analyze cultured keratinocytes from patients with psoriasis or atopic dermatitis and healthy controls. First passage primary cells derived from non-lesional skin were stimulated with pro-inflammatory cytokines, and expression of a panel of 55 genes associated with epidermal differentiation and cutaneous inflammation was measured by quantitative PCR. A subset of these genes was analyzed at the protein level. Using cluster analysis and multivariate analysis of variance we identified groups of genes that were differentially expressed, and could, depending on the stimulus, provide a disease-specific gene expression signature. We found particularly large differences in expression levels of innate immunity genes between keratinocytes from psoriasis patients and atopic dermatitis patients. Our findings indicate that cell-autonomous differences exist between cultured keratinocytes of psoriasis and atopic dermatitis patients, which we interpret to be genetically determined. We hypothesize that polymorphisms of innate immunity genes both with signaling and effector functions are coadapted, each with balancing advantages and disadvantages. In the case of psoriasis, high expression levels of antimicrobial proteins genes putatively confer increased protection against microbial infection, but the biological cost could be a beneficial system gone awry, leading to overt inflammatory disease.

Strong induction of AIM2 expression in human epidermis in acute and chronic inflammatory skin conditions

Absent in melanoma 2 (AIM2) is a double‐stranded DNA receptor, and its activation initiates an interleukin‐1 beta processing inflammasome. AIM2 is implicated in host defense against several pathogens, but could hypothetically also contribute to autoinflammatory or autoimmune diseases, such as is the case for NLRP3. Using thoroughly characterised antibodies, we analysed AIM2 expression in human tissues and primary cells. A strong epidermal upregulation of AIM2 protein expression was observed in several acute and chronic inflammatory skin disorders, such as psoriasis, atopic dermatitis, venous ulcera, contact dermatitis, and experimental wounds. We also found AIM2 induction by interferon‐gamma in submerged and three‐dimensional in vitro models of human epidermis. Our data highlight the dynamics of epidermal AIM2 expression, showing Langerhans cell and melanocyte‐restricted expression in normal epidermis but a pronounced induction in subpopulations of epidermal keratinocytes under inflammatory conditions.

The cystatin M/E-cathepsin L balance is essential for tissue homeostasis in epidermis, hair follicles, and cornea.

Cystatin M/E (CST6) is a nonredundant, epithelium‐specific protease inhibitor with a presumed role in epidermal differentiation and tumor suppression. We have previously reported that cystatin M/E deficiency in Cst6−/− mice causes neonatal lethality because of excessive transepidermal water loss. Biochemical evidence suggests that cystatin M/E controls the activity of legumain, cathepsin L, cathepsin V, and transglutaminase‐3. Using a genetic approach we sought to define the role of cystatin M/E in epithelial biology by identification of its target proteases and their downstream functions. Ablation of cathepsin L in a Cst6−/− background (Cst6−/− Ctsl−/− double‐knockout mice) restored viability and resulted in normalization of stratum corneum morphology. Ablation of legumain or transglutaminase‐3 in Cst6−/− mice, however, did not rescue the lethal phenotype. Intriguingly, both Cst6+ ∼Ctsl−/− and Cst6−/− Ctsl+/− mice were viable, but the absence of cystatin M/E caused scarring alopecia in adult animals. In the cornea of Cst6−/− Ctsl+/− mice, we observed keratitis, hyperplasia, and transition to a cornified epithelium. Evidence is provided that activation of cathepsin D and transglutaminase‐1 are downstream events, dependent of cathepsin L activity. We conclude that a tightly regulated balance between cathepsin L and cystatin M/E is essential for tissue integrity in epidermis, hair follicles, and corneal epithelium.—Zeeuwen, P. L. J. M., van Vlijmen‐Willems, I. M. J. J., Cheng, T., Rodijk‐Olthuis, D., Hitomi, K., Hara‐Nishimura, I., John, S., Smyth, N., Reinheckel, T., Hendriks, W. J. A. J., Schalkwijk, J. The cystatin M/E‐cathepsin L balance is essential for tissue homeostasis in epidermis, hair follicles and cornea. FASEB J. 24, 3744–3755 (2010). www.fasebj.org

Gram-positive anaerobe cocci are underrepresented in the microbiome of filaggrin-deficient human skin

Mutations in the filaggrin gene, which cause the skin disease ichthyosis vulgaris and are a genetic risk factor for atopic dermatitis, alter the cutaneous microbiome thereby affecting keratinocyte host defense responses following skin barrier disruption