Relevance of contact sensitizations in occupational dermatitis patients with special focus on patch testing of workplace materials.

BACKGROUND: Studies on patch testing with workplace materials and evaluation of current occupational relevance of positive patch test reactions are scarce in patients with occupational dermatitis (OD). OBJECTIVES: To identify frequent sensitizations with occupational relevance and to determine the value of patch testing with workplace materials in OD patients. PATIENTS AND METHODS: Results and clinical data of 654 patients with suspected OD patch tested between 2013 and 2017 were analysed. RESULTS: Occupational allergic contact dermatitis was diagnosed in 113 (17.3%) patients. Mechanics had the widest range of occupational sensitizations. Sensitization to epoxy resin was rated occupationally relevant in almost all handicraft trades. Among positive patch test reactions to workplace products, those to water-based metal working fluids and leave-on cosmetic products were most frequent. Despite frequent testing, protective gloves only rarely elicited positive reactions. Preservatives and rubber compounds were most frequently identified as currently occupationally relevant. CONCLUSIONS: Rubber allergy is occupationally relevant especially in healthcare workers and cleaners. Generally, preservatives including formaldehyde releasers are important allergens in OD patients. Leave-on cosmetic products must not be forgotten as allergen sources. Patch testing both workplace materials and standardized test preparations has a complementary value and is beneficial for the diagnostic work-up of OD patients.

Health education decreases incidence of hand eczema in metal work apprentices: Results of a controlled intervention study.

BACKGROUND: Metal work apprentices (MWAs) frequently develop work-related hand eczema (HE). OBJECTIVES: To evaluate the effect of health education on incidence of work-related HE in MWAs and to assess confounding factors. MATERIALS/METHODS: In a prospective controlled intervention study, 131 MWAs received educational training on prevention of HE, whereas 172 MWAs and 118 office work apprentices served as controls. At baseline and during three yearly follow-ups, questionnaires were completed and hands were examined. Saliva samples were collected for assessment of filaggrin (FLG) null mutations and an explorative genome-wide association study (GWAS), and levels of various cytokines were assessed from stratum corneum samples. RESULTS: The 2-year and 3-year incidence of HE in the metalwork control group was 20.9% and 32.6%, respectively, which was significantly higher than in the intervention group (odds ratio [OR] 2.63, 95% confidence interval [CI] 1.31 to 5.28, P < .01 and OR 3.47, 95% CI 1.88 to 6.40, P < .0001). The knowledge score was higher in unaffected MWAs (P < .05). Other factors significantly associated with developing HE in MWAs were smoking cigarettes (P < .01) and FLG mutations (P < .001). No significant associations were found regarding epidermal cytokine levels and GWAS. CONCLUSIONS: Health education is effective in primary prevention of HE in MWAs. Individual factors should be considered in targeted counseling.

Transient epidermal barrier deficiency and lowered allergic threshold in filaggrin-hornerin (FlgHrnr-/- ) double-deficient mice.

BACKGROUND: Filaggrin (Flg) and hornerin (Hrnr) share similar structural and functional features. Both proteins have been implicated as essential proteins for skin barrier maintenance. Loss-of-function mutations of these genes constitute a risk factor for atopic dermatitis and eczema-related asthma. Furthermore, both FLG and HRNR protein levels are downregulated in patients with atopic dermatitis. Thus, mice deficient for Flg and Hrnr provide a novel model to study skin barrier impairment and the susceptibility for cutaneous inflammation. METHODS: By using appropriate targeting vectors and breeding strategies, we established a homozygous FlgHrnr double-deficient (FlgHrnr-/- ) mouse model lacking both genes including the intergenomic sequence. RESULTS: Neonates appeared normal, but developed a transient scaly phenotype with overall flaky appearance, but no overt skin phenotype in adulthood, thereby reflecting a subclinical barrier defect seen in humans. Structurally, FlgHrnr-/- mice displayed a markedly reduced granular layer and a condensed cornified layer. Functionally, FlgHrnr-/- mice showed permeability abnormalities and metabolic aberrations regarding the production of natural moisturizing factors (NMFs) in the stratum corneum. Surprisingly, although the immune system revealed no aberrations under steady-state conditions, FlgHrnr-/- mice are predisposed to mount an allergic contact dermatitis, especially at hapten threshold levels eliciting allergic reactions. CONCLUSIONS: Together, our FlgHrnr-/- mouse model nicely reflects the epicutaneous sensitization susceptibilities and inflammatory reactions to environmental insults in humans with impaired skin barrier functions.

Targeting IgE in Severe Atopic Dermatitis with a Combination of Immunoadsorption and Omalizumab.

Patients with atopic dermatitis (AD) tend to have greatly elevated levels of serum immunoglobulin E (IgE). However, the role of IgE in the pathogenesis of AD is debated. This investigator-initiated open-label pilot study evaluates an anti-IgE-treatment approach by combining extracorporeal immunoadsorption and anti-IgE antibody omalizumab in 10 patients with severe, therapy-refractory AD. IgE levels decreased after immunoadsorption and decreased continuously in all patients during anti-IgE therapy. The reverse trend was observed during 6 months follow-up without treatment. In parallel with these observations, an improvement in AD was observed during the treatment period, with aggravation during follow-up. Further research is needed, based on the principle of reducing IgE levels in order to improve clinical symptoms, using a combination anti-IgE treatment approach, adjusted according to IgE levels.

Internalization routes of cell-penetrating melanoma antigen peptides into human dendritic cells.

Optimized delivery of antigens combined with sustainable maturation of dendritic cells (DCs) is crucial for generation of effective antitumoral immune responses. Multiple approaches for ex vivo antigen loading and improvement in immunogenicity have been described. We have recently established a single-step protocol consisting of a fusion peptide (a sequence of the melanoma antigen Melan-A and a cationic cell-penetrating HIV TAT domain) bound in complexes with a toll-like receptor agonist. As the exact cellular uptake mechanisms of TAT-coupled antigens have been a matter of considerable debate and significantly depend on cell type, cargo and concentrations, we evaluated internalization routes into human immature DCs in comparison with non-phagocytic cell lines. We found that Melan-A-TAT fusion peptide uptake by DCs is mainly energy dependent, superior compared with polylysine-coupled Melan-A and significantly higher in DCs as compared with Jurkat cells or HUVECs. Furthermore, we could track the uptake of the fusion peptide exclusively through early endosomes to lysosome compartments after 90 min by fluorescence microscopy and immunoelectron microscopy. Specific endocytosis inhibitors revealed major internalization of the fusion peptide by DCs via clathrin-mediated endocytosis, whereas uptake by non-phagocytic HUVECs differed significantly, involving macropinocytosis as well as clathrin-mediated endocytosis. As our understanding of the processes involved in internalization of TAT-coupled cargos by human DCs broadens, and DC activation becomes available by single-step procedures as described, further development of simultaneous DC maturation and intra-cellular peptide targeting is warranted.

S1 guideline diagnosis and therapy of cutaneous larva migrans (creeping disease).

GUIDELINE OBJECTIVES: These guidelines aim to enhance patient care by optimizing the diagnosis and treatment of infections due to creeping disease (cutaneous larva migrans) and to raise awareness among doctors of current treatment options. METHODS: S1 guideline, non-systematic literature search, consensus process using a circular letter.

Immunostimulatory activity of murine keratinocyte-derived exosomes.

It has long been known that keratinocytes influence cutaneous immunity through secretion of soluble factors. Exosomes, small membrane vesicles of endocytotic origin, have been implicated in intercellular communication processes such as the transfer of tumor cell antigens and the activation of recipient dendritic cells (DC). However, little is known about immunomodulatory functions of keratinocyte-derived exosomes. To address this question, we analysed exosome secretion of the murine keratinocyte cell line MPEK under steady state as well as inflammatory conditions (+/- IFNγ). These exosomes were readily taken up by bone marrow-derived DC (BMDC) in vitro resulting in a matured phenotype, as evidenced by increased CD40 expression as well as by the production of large amounts of IL-6, IL-10 and IL-12. When the transfer of antigen-specific information through exosomes was investigated, it was found that keratinocytes took up antigen (ovalbumin) and transferred it to their exosomes. However, these antigen-harbouring exosomes failed to induce antigen-specific T cell responses via BMDC. Together, this novel biological function suggests that keratinocytes are able to direct unspecific immune processes but do not elicit specific immune responses.

Atypical location of lymphocytoma cutis in a child.

Lyme borreliosis is a common infectious disease that can affect myocardial muscle or the central nervous system if not treated at an early stage. Here we report a unique case of an atypical location of lymphocytoma cutis in a 3-year-old boy. Histologic and immunohistochemical analysis revealed the correct diagnosis.

Requirement of the RNA-editing enzyme ADAR2 for normal physiology in mice.

ADAR2, an RNA editing enzyme that converts specific adenosines to inosines in certain pre-mRNAs, often leading to amino acid substitutions in the encoded proteins, is mainly expressed in brain. Of all ADAR2-mediated edits, a single one in the pre-mRNA of the AMPA receptor subunit GluA2 is essential for survival. Hence, early postnatal death of mice lacking ADAR2 is averted when the critical edit is engineered into both GluA2 encoding Gria2 alleles. Adar2(-/-)/Gria2(R/R) mice display normal appearance and life span, but the general phenotypic effects of global lack of ADAR2 have remained unexplored. Here we have employed the Adar2(-/-)/Gria2(R/R) mouse line, and Gria2(R/R) mice as controls, to study the phenotypic consequences of loss of all ADAR2-mediated edits except the critical one in GluA2. Our extended phenotypic analysis covering ∼320 parameters identified significant changes related to absence of ADAR2 in behavior, hearing ability, allergy parameters and transcript profiles of brain.

IL-22 and TNF-α represent a key cytokine combination for epidermal integrity during infection with Candida albicans.

T cells exercise their full impact on target cells through a combination of secreted cytokines. The recently described T helper cell subset Th22 is characterized by a combinatorial secretion of IL-22 and TNF-α. Here, we demonstrate that IL-22 increases the TNF-α-dependent induction and secretion of several immune-modulatory molecules such as initial complement factors C1r and C1s, antimicrobial peptides S100A7 and HBD-2 (human ß defensin 2), and antimicrobial chemokines CXCL-9/-10/-11 in primary human keratinocytes. The synergism of IL-22 and TNF-α is transmitted intracellularly by MAP kinases and downstream by transcription factors of the AP-1 family. The induction of innate immunity is relevant in an in vitro infection model, where keratinocytes stimulated with Th22 supernatants or recombinant IL-22 plus TNF-α effectively inhibit the growth of Candida albicans and maintain survival of epithelia. Accordingly, the combinatorial stimulation of keratinocytes with IL-22 and TNF-α most efficiently conserves the integrity of the epidermal barrier in a three-dimensional skin infection model as compared with IFN-γ, IL-17, IL-22 or TNF-α alone. In summary, we demonstrate that IL-22 and TNF-α represent a potent, synergistic cytokine combination for cutaneous immunity.

Mouse phenotyping.

Model organisms like the mouse are important tools to learn more about gene function in man. Within the last 20 years many mutant mouse lines have been generated by different methods such as ENU mutagenesis, constitutive and conditional knock-out approaches, knock-down, introduction of human genes, and knock-in techniques, thus creating models which mimic human conditions. Due to pleiotropic effects, one gene may have different functions in different organ systems or time points during development. Therefore mutant mouse lines have to be phenotyped comprehensively in a highly standardized manner to enable the detection of phenotypes which might otherwise remain hidden. The German Mouse Clinic (GMC) has been established at the Helmholtz Zentrum München as a phenotyping platform with open access to the scientific community (www.mousclinic.de; [1]). The GMC is a member of the EUMODIC consortium which created the European standard workflow EMPReSSslim for the systemic phenotyping of mouse models (http://www.eumodic.org/[2]).

Efficacy and safety of omalizumab in patients with chronic urticaria who exhibit IgE against thyroperoxidase.

BACKGROUND: A subgroup of patients with chronic spontaneous urticaria (CU) exhibits IgE antibodies directed against autoantigens, such as thyroperoxidase (TPO). We conducted this study to investigate whether such patients with CU with IgE against TPO benefit from treatment with omalizumab, a humanized anti-IgE mAb licensed for the treatment of severe persistent allergic (IgE-mediated) asthma. OBJECTIVES: We sought to assess the efficacy of omalizumab treatment in patients with CU with IgE autoantibodies against TPO. METHODS: In this multicenter, randomized, double-blind, placebo-controlled study patients with CU (male/female, 18-70 years of age) with IgE autoantibodies against TPO who had persistent symptoms (wheals and pruritus) despite standard antihistamine therapy were randomized to receive either omalizumab (75-375 mg, dose determined by using the approved asthma dosing table) or placebo subcutaneously once every 2 or 4 weeks for 24 weeks. The primary end point was the change from baseline in mean weekly urticaria activity score after 24 weeks of treatment, as calculated from patients' diaries. The safety and tolerability of omalizumab were also assessed. RESULTS: Of the 49 randomized patients (omalizumab, n = 27; placebo, n = 22), 42 completed the study. At week 24, patients demonstrated a mean reduction in the weekly urticaria activity score from baseline of 17.8 with omalizumab and 7.9 with placebo (P = .0089). Complete protection from wheal development was observed in 19 (70.4%) patients in the omalizumab group compared with only 1 (4.5%) patient in the placebo group. The rate of adverse events was similar in both groups. CONCLUSIONS: The results of this study indicate that omalizumab is an effective treatment option for patients with CU with IgE autoantibodies against TPO who are refractory to conventional treatment.

A comprehensive analysis of the COL29A1 gene does not support a role in eczema.

BACKGROUND: Based on a recent positional cloning approach, it was claimed that the collagen 29A1 gene (COL29A1), which encodes an epidermal collagen, represents a major risk gene for eczema underlying a previously reported linkage to chromosome 3q21. However, thus far, not a single replication attempt has been published, and no definitive functional data have been provided. OBJECTIVES: We aimed to determine whether COL29A1 polymorphisms contribute to eczema susceptibility and whether COL29A1 expression is altered in eczema. METHODS: We investigated the reported association of COL29A1 variants with eczema, subtypes of eczema, and eczema-related traits in 5 independent and large study populations comprehensively phenotyped for allergic diseases: a set of 1687 German patients with eczema and 2387 population control subjects, a collection of 274 German families with eczema-diseases children, a cross-sectional population of German children (n = 3099), the Swedish population-based birth cohort Children Allergy and Milieu in Stockholm, an Epidemiologic Study (BAMSE) (n = 2033), and the European cross-sectional Prevention of Allergy-Risk Factors for Sensitization Related to Farming and Anthroposophic Lifestyle (PARSIFAL) study (n = 3113). An additional set of 19 COL29A1 coding single nucleotide polymorphisms was analyzed in BAMSE and PARSIFAL. COL29A1 expression was investigated by using in situ hybridization. RESULTS: We found no evidence for a relationship between COL29A1 polymorphisms and eczema. The equivalence test rejected the hypothesis of association even excluding small effects. In situ hybridization carried out on biopsy specimens from lesional and nonlesional skin of patients with eczema and from healthy control subjects did not show any differences in the cellular distribution pattern of COL29A1 expression at the mRNA level. CONCLUSIONS: Our results suggest that COL29A1 is unlikely to contain genetic variants that have a major effect on eczema or atopy susceptibility.

TNF receptor I on human keratinocytes is a binding partner for staphylococcal protein A resulting in the activation of NF kappa B, AP-1, and downstream gene transcription.

Primary human keratinocytes and immortalized HaCaT cells were analysed for their capacity to bind purified staphylococcal protein A (SpA). Co-incubation with FITC-labelled SpA led to a dose-depending attachment. Pull-down experiments with cellular extracts revealed the TNFα receptor I (TNF RI) as binding partner on keratinocytes. Thus, we next looked for expression of this receptor in human epidermis and cultured keratinocytes. TNF RI is strongly expressed on all keratinocytes analysed, both at the mRNA and protein level and activation by SpA at optimal doses of 50-100 µg/ml resulted in the phosphorylation of the TNF RI downstream kinases MEK1/2, JNK1/2, and p38 subsequently leading to translocation of the p65 NF kappa B subunit and AP-1 into the nucleus. This translocation was then followed by increased expression of IL-8 and COX-2, two known NF kappa B-induced pro-inflammatory genes. To further test the relevance of our findings, we analysed in vitro production of over 100 strains isolated from atopic eczema showing that more than 85% of the tested strains produced extracellular SpA in substantial amounts. Thus, besides superantigens, haemolysins, and other cell wall components, Staphylococcus aureus exerts pro-inflammatory stimuli on human keratinocytes through the production of SpA signalling through TNF RI.

Quantitation of serum IgE by using chimeras of human IgE receptor and avian immunoglobulin domains.

Anti-IgE therapeutics represent an efficient approach in the management of IgE-mediated allergic asthma. However, monitoring the reduction of IgE levels into a therapeutically efficient range requires the determination of residual serum IgE. We established an analytical approach to distinguish free and anti-IgE complexed serum IgE based on soluble derivatives of the human high-affinity IgE receptor. Soluble receptor derivatives represent an ideal means to analyze receptor antagonism by any ligand or blocking antibody. Therefore, the FcεRI ectodomain was fused with avian IgY constant domains that circumvent susceptibility to interference phenomena and improve assay performance. After production in HEK293 cells, subsequent characterization by enzyme-linked immunosorbent assay and immunoblotting confirmed the suitability of avian IgY constant domains for immobilization and detection purposes. To provide further insights into the different IgE reactivities, free allergen-specific IgE was also determined. Monitoring of sera from omalizumab-treated patients during the course of therapy revealed the applicability for assessment of omalizumab-complexed versus noncomplexed serum IgE. These parameters may allow correlation to clinical responses during anti-IgE therapy with the perspective of biomonitoring.

Specific CD8 T cells in IgE-mediated allergy correlate with allergen dose and allergic phenotype.

RATIONALE: Studies in humans and rodents have indicated a causative role for CD8(+) T cells in IgE-mediated allergic inflammation, but their function is still controversial. OBJECTIVES: To analyze the role of allergen-specific CD8(+) T cells during the development of allergic airway inflammation in two parallel but diverging outcome models. METHODS: We used H2-Kb SIINFEKL (OVA(257-264)) multimers to analyze induction, natural distribution, and phenotype of allergen-specific CD8(+) T cells in a murine C57BL/6 model of ovalbumin (OVA)-induced allergic airway inflammation using low-dose or high-dose OVA sensitization. MEASUREMENTS AND MAIN RESULTS: The low-dose protocol was characterized by a significant induction of total and OVA-specific IgE, eosinophilic airway inflammation, IL-4 levels in bronchoalveolar lavage fluid. And significant alterations in lung function. The high dose protocol was characterized by a significant reduction of the allergic phenotype. Using OVA(257-264) H2-Kb multimers, we observed lung and airway infiltrating OVA-specific CD8(+) T cells showing an effector/effector-memory phenotype. The high-dose protocol caused significantly higher infiltration of allergen-specific CD8(+) cells to the airways and enhanced their cytotoxicity. Adoptive transfer with CD8(+) T cells from transgenic OT-I mice to TAP1(-/-) or wild-type mice showed their migration to the lungs and TAP1-dependent proliferation after OVA-aerosol exposure. TAP1(-/-) mice defective in CD8(+) T cells showed exacerbated symptoms in the low-dose sensitization model. CONCLUSIONS: Allergen-specific CD8(+) T cells seem to protect from allergic inflammation in the lungs. Their number, which is dependent on the sensitization dose, appears to be a critical predictor for the severity of the allergic phenotype.

Genome-wide scan on total serum IgE levels identifies FCER1A as novel susceptibility locus.

High levels of serum IgE are considered markers of parasite and helminth exposure. In addition, they are associated with allergic disorders, play a key role in anti-tumoral defence, and are crucial mediators of autoimmune diseases. Total IgE is a strongly heritable trait. In a genome-wide association study (GWAS), we tested 353,569 SNPs for association with serum IgE levels in 1,530 individuals from the population-based KORA S3/F3 study. Replication was performed in four independent population-based study samples (total n = 9,769 individuals). Functional variants in the gene encoding the alpha chain of the high affinity receptor for IgE (FCER1A) on chromosome 1q23 (rs2251746 and rs2427837) were strongly associated with total IgE levels in all cohorts with P values of 1.85 x 10(-20) and 7.08 x 10(-19) in a combined analysis, and in a post-hoc analysis showed additional associations with allergic sensitization (P = 7.78 x 10(-4) and P = 1.95 x 10(-3)). The "top" SNP significantly influenced the cell surface expression of FCER1A on basophils, and genome-wide expression profiles indicated an interesting novel regulatory mechanism of FCER1A expression via GATA-2. Polymorphisms within the RAD50 gene on chromosome 5q31 were consistently associated with IgE levels (P values 6.28 x 10(-7)-4.46 x 10(-8)) and increased the risk for atopic eczema and asthma. Furthermore, STAT6 was confirmed as susceptibility locus modulating IgE levels. In this first GWAS on total IgE FCER1A was identified and replicated as new susceptibility locus at which common genetic variation influences serum IgE levels. In addition, variants within the RAD50 gene might represent additional factors within cytokine gene cluster on chromosome 5q31, emphasizing the need for further investigations in this intriguing region. Our data furthermore confirm association of STAT6 variation with serum IgE levels.

The role of toll-like receptors in host defenses and their relevance to dermatologic diseases.

The family of toll-like receptors (TLRs) plays a central role in the cutaneous immune defense system. To date, different TLRs have been found on several major cell populations of the skin, such as keratinocytes, fibroblasts, antigen-presenting cells, and melanocytes. Activation of TLRs leads, via different intracellular signaling pathways, to the production of pro-inflammatory stimuli, and is considered a danger signal that should transform the skin in to the functional state of defense. However, TLRs have also been implicated in tissue homeostasis and renewal. Within the group of TLRs, two types have been identified: surface-expressed TLRs, which are predominantly active against bacterial cell wall compounds; and intracellular receptors, which preferentially recognize virus-associated pattern molecules. In addition, surface-expressed receptors trigger phagocytotic and maturation signals, while the intracellular TLRs lead to the induction of antiviral genes. Our review aims to outline the importance of TLRs in the pathogenesis of numerous skin diseases and the potential of TLR agonists as a treatment option for various skin diseases.