Proteomic biomarkers for psoriasis and psoriasis arthritis.

UNLABELLED: Although several new biomarkers have been recently proposed for psoriasis (Ps) and psoriasis arthritis (PsA), nothing is known about their diagnostic sensitivity and specificity, and their routine use. We therefore searched in-depth for new biomarker candidates using a biobank with EDTA-plasma from 158 individuals, patients and healthy controls. Samples from 6 selected pairs (patients against healthy controls) were searched proteomically using a workflow of extensive and precise design that is highly comprehensive. Subsequent verification was performed using ELISA and the entire biobank. By proteomic methods, 208 altered proteins were identified. Of these, 15 biomarker candidates were selected for verification. Of these 15, 4 individual parameters and 11 combinations significantly discriminated between patient and control groups. These individual parameters were Zn-α2-glycoprotein, complement C3, polymeric immunoglobulin receptor, and plasma kallikrein. Significant discrimination was obtained by combinations of 2 or 3 parameters. One combination seemed suitable for diagnosing PsA. Moreover, several candidates desmoplakin, complement C3, polymeric immunoglobulin receptor, and cytokeratin 17, correlated with PASI in all patients. This first comprehensive proteomic study on non-depleted plasma identified several biomarker candidates that have not been described before as well as some known from previous studies. BIOLOGICAL SIGNIFICANCE: Our non-gel proteomic analysis is based on the highly comprehensive and significantly optimized chromatographic protein pre-fractionation. The method allows a biomarker search in non-depleted plasma. The subsequent verification by ELISA identifies several biomarker-candidates for the unbiased diagnosis of psoriasis and psoriasis arthritis. Four of the identified candidate markers might be used individually. Combinations of several parameters improve the diagnostic sensitivity and specificity. The still not validated candidates form a reserve for further evaluation. Moreover, mass spectrometric data uncover several biomarker-candidates which show diverse protein species of the same protein with opposing changes in the same sample.

Beta-defensin-2 genomic copy number variation and chronic periodontitis.

Chronic periodontitis (ChP) is a multifactorial disease influenced by microbial and host genetic variability; however, the role of beta-defensin-2 genomic (DEFB4) copy number (CN) variation (V) in ChP remains unknown. The association of the occurrence and severity of ChP and DEFB4 CNV was analyzed. Our study included 227 unrelated Caucasians, that is, 136 ChP patients (combined ChP) and 91 control individuals. The combined ChP group was subdivided into the severe ChP and slight-to-moderate ChP subgroups. To determine DEFB4 CNV, we isolated genomic DNA samples and analyzed them by relative quantitation using the comparative CT method. The serum beta-defensin-2 (hBD-2) level was determined via ELISA. The distribution pattern and mean DEFB4 CN did not differ significantly in combined ChP cases vs. the controls; however, the mean DEFB4 CN in the severe ChP group differed significantly from those for the control and slight-to-moderate ChP groups. Low DEFB4 CN increased the risk of severe ChP by about 3-fold. DEFB4 CN was inversely associated with average attachment loss. Mean serum hBD-2 levels were highest in the controls, followed by the slight-to-moderate ChP group and the severe ChP group. The results suggested an association between decreased DEFB4 CN and serum hBD-2 levels and periodontitis severity.

Histamine modulates γδ-T lymphocyte migration and cytotoxicity, via Gi and Gs protein-coupled signalling pathways.

BACKGROUND AND PURPOSE: The biogenic amine, histamine plays a pathophysiological regulatory role in cellular processes of a variety of immune cells. This work analyses the actions of histamine on γδ-T lymphocytes, isolated from human peripheral blood, which are critically involved in immunological surveillance of tumours. EXPERIMENTAL APPROACH: We have analysed effects of histamine on the intracellular calcium, actin reorganization, migratory response and the interaction of human γδ T cells with tumour cells such as the A2058 human melanoma cell line, the human Burkitt's Non-Hodgkin lymphoma cell line Raji, the T-lymphoblastic lymphoma cell line Jurkat and the natural killer cell-sensitive erythroleukaemia cell line, K562. KEY RESULTS: γδ T lymphocytes express mRNA for different histamine receptor subtypes. In human peripheral blood γδ T cells, histamine stimulated Pertussis toxin-sensitive intracellular calcium increase, actin polymerization and chemotaxis. However, histamine inhibited the spontaneous cytolytic activity of γδ T cells towards several tumour cell lines in a cholera toxin-sensitive manner. A histamine H(4) receptor antagonist abolished the histamine induced γδ T cell migratory response. A histamine H(2) receptor agonist inhibited γδ T cell-mediated cytotoxicity. CONCLUSIONS AND IMPLICATIONS: Histamine activated signalling pathways typical of chemotaxis (G(i) protein-dependent actin reorganization, increase of intracellular calcium) and induced migratory responses in γδ T lymphocytes, via the H(4) receptor, whereas it down-regulated γδ T cell mediated cytotoxicity through H(2) receptors and G(s) protein-coupled signalling. Our data suggest that histamine activated γδ T cells could modulate immunological surveillance of tumour tissue.

Iloprost has potent anti-inflammatory properties on human monocyte-derived dendritic cells.

BACKGROUND: The stable prostaglandin I2 analogue (iloprost) iloprost has been shown to inhibit allergic airway inflammation in mice by modulating the function of myeloid dendritic cells (DCs). OBJECTIVE: The aim of the current study was to investigate the biological activity of iloprost on human monocyte-derived DCs. METHODS: I prostanoid (IP) receptor expression was analysed by RT-PCR. Cytokine secretion by DCs and CD4+ T cells was measured by ELISA. The expression of the transcription factor FoxP3 after co-culture of DCs with CD4+ CD45RA+ T cells was analysed by flow cytometry. RESULTS: Human monocyte-derived DCs were found to express mRNA specific for the PGI2 receptor IP, and stimulation with iloprost resulted in increased cyclic AMP levels in both immature DCs (iDCs) and mature DCs (mDCs). Moreover, iloprost dose dependently inhibited the secretion of TNF-alpha, IL-6, IL-8 and IL-12p70 in mDCs, while it enhanced IL-10 production. Changes in cytokine secretion were paralleled by an altered T-cell priming capacity of DCs: in co-culture experiments of iloprost-treated mDC and naïve CD45RA+ T cells, an induction of regulatory T cells could be observed, as demonstrated by increased intracellular FoxP3 expression and IL-10 production. Additionally, iloprost inhibited the MIP-3beta-induced migration of mDCs. CONCLUSION: In summary, our results provide evidence that iloprost profoundly affects the function of human myeloid DCs. Therefore, iloprost might also be a new therapeutical option for the treatment of asthma in humans.

Both copy number and sequence variations affect expression of human DEFB4.

Copy number variations (CNVs) were found to contribute massively to the variability of genomes. One of the best studied CNV region is the beta-defensin cluster (DEFB) on 8p23.1. Individual DEFFB copy numbers (CNs) between 2 and 12 were found, whereas low CNs predispose for Crohn's disease. A further level of complexity is represented by sequence variations between copies (multisite variations, MSVs). To address the relation of DEFB CN and MSV to the expression of beta-defensin genes, we analyzed DEFB4 expression in B-lymphoblastoid cell lines (LCLs) and primary keratinocytes (normal human epidermal keratinocyte, NHEK) before and after stimulation with lipopolysaccharide, tumor necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma). Moreover, we quantified one DEFB4 MSV in DNA and mRNA as a marker for variant-specific expression (VSE) and resequenced a region of approximately 2 kb upstream of DEFB4 in LCLs. We found a strong correlation of DEFB CN and DEFB4 expression in 16 LCLs, although several LCLs with very different CNs exhibit similar expression levels. Quantification of the MSV revealed VSE with consistently lower expression of one variant. Costimulation of NHEKs with TNF-alpha/IFN-gamma leads to a synergistic increase in total DEFB4 expression and suppresses VSE. Analysis of the DEFB4 promoter region showed remarkably high density of sequence variabilities (approximately 1 MSV/41 bp).

[Economic aspects of standardization of modern wound care]. / Okonomische Effekte der Standardisierung in der modernen Wundversorgung.

BACKGROUND: Despite the advantages of modern wound management, it is still employed too rarely because of higher costs and lack of information available to the medical staff. At the University Hospital Jena, the assortment of hydroactive wound dressings has been optimized, in order to increase acceptance and attain cost advantages. MATERIAL AND METHODS: The defined target sizes were differentiated according to economic and qualitative parameters. The economic parameters included the reduction in the total costs for wound management, adjusted for the case mix and the number of cases. Among the qualitative parameters were reducing the number of suppliers and the number of orders outside the standard offerings. RESULTS: Via standardization, the average expenses per case for hydroactive wound applications could be reduced by 10.3% and the average expenses per weighted German DRG by 15.9%. As a result of the reorganization of the assortment, the number of suppliers was reduced by 57.1% and two main and one secondary supplier were designated. The number of orders outside the standard assortment was reduced from 173 to 21 (87.8%). CONCLUSION: The standardization of the assortment of hydroactive wound applications has led to an increase in procedural efficiency and effectiveness at the University Hospital Jena, in addition to a reduction in the cost of material and in the number of suppliers.

Microtubules regulate expression of ICAM-1 in epidermoid cells (KB cells).

The intercellular adhesion molecule-1/CD54 (ICAM-1) functions as a counterreceptor for other adhesion molecules (e.g. the lymphocyte function-associated antigen-1/CD11a/CD18) required for the interaction of a large variety of cells with leucocytes. Constitutive expression of ICAM-1 in human epidermoid cells (KB cells) is low, but inducible by interferon-gamma (IFN-gamma). Treatment of KB cells with microtubule-disrupting agents, like colchicine, nocodazole and vinblastine, potentiated the constitutive and cytokine-induced ICAM-1 expression on the cell surface. Actinomycin D inhibited microtubule-disrupting agent-induced ICAM-1 surface expression. Increased steady-state levels of ICAM-1 transcripts were found after treatment of KB cells with microtubule-disrupting agents. However, microtubule-disrupting agents neither altered the glyceraldehyde-3-phosphate dehydrogenase mRNA levels nor the amount of expressed alpha(2)-, alpha(3)-and beta(1)-integrins at the cell surface. In addition, they did not change the ICAM-1 mRNA half-life. These studies indicate a control function of the microtubule network on the expression of ICAM-1.

Additive impairment of the barrier function by mechanical irritation, occlusion and sodium lauryl sulphate in vivo.

BACKGROUND: The interaction between potential irritants in the workplace might be important because workers are not usually exposed to a single irritant, but to multiple potentially harmful substances. Physical irritant contact dermatitis caused by friction or mechanical abrasion is a common occupational dermatosis. Prolonged water exposure by occlusion is also common in the workplace. Several studies have revealed the negative effect of the common anionic detergent sodium lauryl sulphate (SLS) on permeability barrier function. OBJECTIVES: To study the additive impairment of permeability barrier function by mechanical irritation combined with 0.5% SLS or prolonged water exposure by occlusion, as models of mild irritation. METHODS: The volar forearms of 20 healthy volunteers were exposed to mechanical irritation and occlusion with water or 0.5% SLS for four consecutive days in a combined tandem repeated irritation test (TRIT). Permeability barrier function was measured with a Tewameter TM 210. Irritation was assessed with a Chromameter CR 300 and a visual score. RESULTS: Barrier disruption in our model was rated as follows: occlusion with SLS and mechanical irritation > occlusion with SLS > occlusion with water and mechanical irritation > mechanical irritation and occlusion with water > occlusion with a glove and mechanical irritation > mechanical irritation > occlusion with water. Barrier disruption caused by occlusion or mechanical irritation was enhanced by the tandem application. The choice of irritant under occlusion, time of occlusion and order of tandem application all affected the degree of barrier disruption. Evaporimetry was able to detect early stages in the development of an irritant reaction before it became visible. Chromametry was not able to detect this early response. CONCLUSIONS: Physical irritants (friction, abrasive grains, occlusion) and detergents such as SLS represent a significant irritation risk and should be minimized, especially when acting together, as shown in our TRIT model.

Sequential application of cold and sodium lauryl sulphate decreases irritation and barrier disruption in vivo in humans.

BACKGROUND: Irritant contact dermatitis (ICD) is one of the most frequent types of occupational dermatitis. Different factors are involved in the development of contact dermatitis. In the food-processing industry, the combined exposure to different irritants may be involved in the development of ICD. Few data have been published regarding the irritant potential of sodium lauryl sulphate (SLS) in combination with cold. OBJECTIVES: The present study was intended to analyse whether cold exposure and low skin temperature influence the development of ICD. METHODS: Twenty (part I) and 12 (part II) healthy volunteers were exposed twice daily for 4 days to SLS alone, different low temperatures alone (4 degrees C six times for 90 s with an interval of 20 s or 15 degrees C for 10 min) or a combination of cold and SLS (19.6 microL SLS 1% cm(-2), part I; or 52.6 microL SLS 0.5% cm(-2), part II) using the tandem repetitive irritation test. Irritant cutaneous reactions were measured by noninvasive biophysical methods with transepidermal water loss as a parameter for permeability barrier function and skin colour reflectance together with visual scoring as parameters for inflammatory reactions. RESULTS: Cold alone caused no significant skin reaction compared with untreated control. Exposure to SLS alone and SLS together with cold (independent of the applied temperature of 4 or 15 degrees C) twice daily induced a clear irritant reaction and barrier disturbance. Reactions did not differ whether SLS was applied before or after cold. Furthermore, 'tandem application' of cold and SLS diminished the barrier disruption and irritant reaction compared with SLS alone. CONCLUSIONS: We conclude that the application of cold may have a protective effect on the development of ICD, at least in our short-term model.

The quality of human skin xenografts on SCID mice: a noninvasive bioengineering approach.

BACKGROUND: Animal models are important tools for studies in skin physiology and pathophysiology. Due to substantial differences in skin characteristics such as thickness and number of adnexa, the results of animal studies cannot always be directly transferred to the human situation. Therefore, transplantation of human skin on to SCID (severe combined immunodeficiency) mice might offer a promising tool to perform studies in viable human skin without the direct need for human volunteers. OBJECTIVES: To characterize the physiological and anatomical changes of a human skin transplant on a SCID animal host. METHODS: In this study human skin was transplanted on to 32 SCID mice and followed for 6 months. Barrier function was assessed by transepidermal water loss (TEWL; tewametry) and moisture content of the stratum corneum was studied by measurement of electrical capacitance (corneometry). RESULTS: The results showed considerable deviations of TEWL values and skin hydration between the grafts and human skin in vivo. The human skin showed epidermal hyperkeratosis and moderate sclerosis of the corium 4 and 6 months after transplantation on to SCID mice. CONCLUSIONS: Our results indicate that human skin does not completely preserve its physiological and morphological properties after transplantation on to SCID mice. Therefore, results from experiments using this model system need to be discussed cautiously.

Fruit acids and sodium hydroxide in the food industry and their combined effect with sodium lauryl sulphate: controlled in vivo tandem irritation study.

BACKGROUND: Cutaneous exposure to a variety of irritants has been extensively studied in recent years. Nevertheless, knowledge of the induction of irritant dermatitis, especially by mild irritants at low doses and for a short duration of exposure, is still incomplete. OBJECTIVES: To quantify the irritant effects and barrier disruption properties of ascorbic acid (ASC), acetic acid (ACA) and sodium hydroxide (NaOH), particularly in combination with an anionic detergent, sodium lauryl sulphate (SLS). METHODS: In a tandem repeated irritation test, the irritants were applied for 30 min twice daily for 4 days to the skin of the mid-back of 19 healthy volunteers of both sexes. We used bioengineering techniques for measurement of transepidermal water loss (TEWL) and skin colour reflectance, as well as visual scoring. RESULTS: Repeated application of ASC and ACA caused a moderate increase in TEWL and erythema. The sequential application of ASC or ACA and SLS enhanced these effects. NaOH induced a strong reaction when applied both occlusively and nonocclusively as well as in combination with SLS, with an early onset of the inflammatory signs, leading to discontinuation of the application on the third day in most of the test fields. Notably, the irritant effect of NaOH was not as marked when applied sequentially with SLS. CONCLUSIONS: Our results demonstrate that concurrent application of an anionic detergent and a mild acidic irritant can lead to disruption of the barrier function which, although not additive, is still considerable. The combined application of SLS and mild acids does not prevent SLS-induced irritation. Furthermore, we showed that NaOH in low concentrations may also act as a potent irritant but that its effect is not enhanced by SLS. The necessity of adequate skin protection and reduction of contact with substances that are potentially barrier disruptive and irritant, e.g. in the food industry, is emphasized, not only when handling detergents, but also when processing food products.

Surveillance of nosocomial infections in dermatology patients in a German university hospital.

BACKGROUND: Nosocomial infections (NIs) are a growing problem in healthcare today. Thus, surveillance of NIs is an important aspect of modern infection control, which aims to improve the quality of care. OBJECTIVES: To identify overall and site-specific NI rates in dermatology patients in a German university hospital. METHODS: In a prospective study, 1450 patients were surveyed for NIs according to criteria laid down by the Centers for Disease Control and Prevention. Case records were reviewed twice a week, microbiology reports were assessed and the ward staff was consulted. RESULTS: Altogether, 37 NIs were identified in 35 patients, of whom two had two NIs. The overall incidence was 2.5 NIs per 100 patients, and the incidence density was 1.9 NIs per 1000 patient days. Twenty-one patients developed superficial surgical site infections (SSIs). Thirteen of the 21 SSIs occurred after surgical removal of basal cell carcinoma (BCC; 172 in total). This represents an infection rate of 7.6% after surgery for BCC. CONCLUSIONS: Our data suggest that routine surveillance in dermatological wards is not accorded a high priority. However, surveillance of SSIs, especially following surgery for BCC, may be indicated.

Elevated expression of extracellular matrix metalloproteinase inducer (CD147) and membrane-type matrix metalloproteinases in venous leg ulcers.

BACKGROUND: Matrix metalloproteinases (MMPs) contribute to matrix remodelling in venous leg ulcers. Extracellular MMP inducer (EMMPRIN; CD147) has been reported to increase MMP expression, and membrane type 1 MMP (MT1-MMP) has been implicated in the activation of MMPs. OBJECTIVES: To examine whether and to what degree EMMPRIN, MMP-2, MT1-MMP and membrane type 2 MMP (MT2-MMP) are expressed in venous leg ulcers as well as the association with MMP activity. METHODS: EMMPRIN, MMP-2, MT1-MMP and MT2-MMP were analysed by zymography and immunohistochemistry in biopsies from healthy skin and lesional tissue from venous leg ulcers. RESULTS: Zymography provided direct evidence of increased proteolytic activity of MMP-2 in lesional skin in comparison with healthy controls. Immunostaining showed intense expression of EMMPRIN, MMP-2, MT1-MMP and MT2-MMP in dermal structures of venous leg ulcers, whereas only EMMPRIN and MMP-2 showed elevated expression in perivascular regions. Our findings indicate that venous leg ulcers are characterized by elevated expression of EMMPRIN, MMP-2, MT1-MMP and MT2-MMP. The immunohistological findings of skin alterations reflect the dynamic process of activation of soluble and membrane-bound MMPs, which may be highly induced by EMMPRIN. CONCLUSIONS: These data suggest for the first time that membrane-bound MMPs may favour enhanced turnover of the extracellular matrix and support unrestrained MMP activity in venous leg ulcers.

Expression and function of adenosine receptors in human dendritic cells.

Dendritic cells (DCs) are specialized antigen-presenting cells characterized by their ability to migrate into target sites, process antigens, and activate naive T cells. In this study, we analyzed the biological activity and intracellular signaling of adenosine by using reverse transcriptase-polymerase chain reaction assays to investigate mRNA expression of A(1), A(2a) and A(3) adenosine receptors in immature and mature human DCs. Functional experiments on adenosine stimulation showed chemotaxis, intracellular calcium transients, and actin polymerization, but no activation of adenylate cyclase in immature DCs. Experiments with receptor isotype-selective agonists and antagonists as well as pertussis toxin revealed that chemotaxis, calcium transients, and actin polymerization were mediated via G(i-) or G(0-)protein-coupled A(1) and A(3) receptors. Maturation of DCs induced by lipopolysaccharide (LPS) resulted in down-regulation of A(1) and A(3) receptor mRNAs, although A(2a) receptor mRNA was still expressed. However, in LPS-differentiated DCs, adenosine and an A(2a) receptor agonist stimulated adenylate cyclase activity, enhanced intracellular cAMP levels, and inhibited interleukin 12 (IL-12) production. These effects could be completely prevented by pretreatment with A(2) receptor antagonist. These findings strongly suggest that adenosine has important but distinct biological effects in DCs activity as a chemotaxin for immature DCs and as a modulator of IL-12 production in mature DCs. These effects can be explained by differential expression of adenosine receptor subtypes.