[The role of the innate immune system in atopic dermatitis]. / Zur Rolle des angeborenen Immunsystems bei atopischer Dermatitis.

The mechanisms how the innate immune system detects microbes and mounts a rapid immune response have been more and more elucidated in the past years. Subsequently it has been shown that innate immunity also shapes adaptive immune responses and determines their quality that can be either inflammatory or tolerogenic. As atopic dermatitis is characterized by disturbances of innate and adaptive immune responses, colonization with pathogens and defects in skin barrier function, insight into mechanisms of innate immunity has helped to understand the vicious circle of ongoing skin inflammation seen in atopic dermatitis patients. Elucidating general mechanisms of the innate immune system and its functions in atopic dermatitis paves the way for developing new therapies. Especially the novel insights into the human microbiome and potential functional consequences make the innate immune system a very fundamental and promising target. As a result atopic dermatitis manifestations can be attenuated or even resolved. These currently developed strategies will be introduced in the current review.

About the role and underlying mechanisms of cofactors in anaphylaxis.

Anaphylaxis is the systemic and most severe presentation of type I allergy. A number of conditions were identified that modulate the onset of anaphylaxis such as co- or augmentation factors, which significantly lower the allergen dose necessary for triggering anaphylaxis. Next to physical exercise or alcohol consumption, co-administration of nonsteroidal anti-inflammatory drugs (NSAID) or concomitant infectious diseases are well-documented cofactors of anaphylaxis. Registries for anaphylaxis document a role for cofactors in about 30% of anaphylactic reactions. Some disease entities such as 'wheat-dependent exercise-induced anaphylaxis' (WDEIA) are explicitly characterized by elicitation of anaphylaxis only in the presence of at least one such cofactor. Using WDEIA as a model disease, studies demonstrated that exercise increases skin prick test reactivity to and bioavailability of the allergen. Additional data indicate that alcohol consumption and NSAID administration display similar effects. Modulation of the cellular activation threshold is another mechanism underlying cofactor-induced anaphylaxis, most likely also functional when infectious diseases orchestrate elicitation of anaphylaxis. Cofactors are increasingly accepted to play a fundamental role in eliciting anaphylaxis. Consequently, to improve patient management modalities, a better understanding of the underlying mechanisms is warranted. This review aims to update clinicians and clinical scientists on recent developments.

The Staphylococcus aureus lipoprotein SitC colocalizes with Toll-like receptor 2 (TLR2) in murine keratinocytes and elicits intracellular TLR2 accumulation.

SitC is one of the predominant lipoproteins in Staphylococcus aureus. Recently, SitC was shown to be capable of stimulating Toll-like receptor 2 (TLR2), but the mechanism of TLR2 activation by SitC has not been analyzed in detail so far. In this study, we purified C-terminally His-tagged SitC (SitC-His) from Staphylococcus aureus. SitC-His induced interleukin-6 (IL-6) and tumor necrosis factor alpha (TNF-α) release in human monocytes and also NF-κB activation in TLR2-transfected HEK293 cells, indicating TLR2-specific activation. SitC not only induced a TLR2-dependent release of IL-6 in primary murine keratinocytes (MKs) but also induced intracellular accumulation of TLR2, which was time and concentration dependent. Cy2-labeled SitC-His colocalized specifically with TLR2 in MKs and was also internalized in TLR2 knockout MKs, suggesting a TLR2-independent uptake. Neither activation nor colocalization of SitC-His was observed with TLR4 or Nod2. The results show that the native lipoprotein SitC-His specifically colocalizes with TLR2, is internalized by host cells, induces proinflammatory cytokines, and triggers intracellular accumulation of TLR2.

The activin binding proteins follistatin and follistatin-related protein are differentially regulated in vitro and during cutaneous wound repair.

Follistatin is a secreted protein that binds activin in vitro and in vivo and thereby inhibits its biological functions. Recently, related human and murine genes, designated follistatin-related gene (FLRG), were identified, and their products were shown to bind activin with high affinity. In this study we further characterized the murine FLRG protein, and we analyzed its tissue-specific expression and regulation in comparison with those of follistatin. Transient expression of the mouse FLRG protein in COS-1 cells revealed that the FLRG cDNA encodes a secreted glycoprotein. FLRG mRNA was expressed at high levels in the lung, the testis, the uterus and, particularly, the skin. Immunohistochemistry revealed the presence of FLRG in the basement membrane between the dermis and the epidermis and around blood vessels. FLRG mRNA expression was induced in keratinocytes by keratinocyte growth factor, epidermal growth factor and transforming growth factor-beta 1, and in fibroblasts by platelet-derived growth factor and epidermal growth factor. The induction was more rapid, but weaker, than that of follistatin. Most interestingly, both follistatin and FLRG were expressed during the wound healing process, but their distribution within the wound was different. The different expression pattern of FLRG and follistatin and their differential regulation suggest different functions of these activin-binding proteins in vivo.

The Ca2+-binding proteins S100A8 and S100A9 are encoded by novel injury-regulated genes.

To gain insight into the molecular mechanisms underlying cutaneous wound repair, we performed a large scale screen to identify novel injury-regulated genes. Here we show a strong up-regulation of the RNA and protein levels of the two Ca(2+)-binding proteins S100A8 and S100A9 in the hyperthickened epidermis of acute murine and human wounds and of human ulcers. Furthermore, both genes were expressed by inflammatory cells in the wound. The increased expression of S100A8 and S100A9 in wound keratinocytes is most likely related to the activated state of the keratinocytes and not secondary to the inflammation of the skin, since we also found up-regulation of S100A8 and S100A9 in the epidermis of activin-overexpressing mice, which develop a hyperproliferative and abnormally differentiated epidermis in the absence of inflammation. Furthermore, S100A8 and S100A9 expression was found to be associated with partially differentiated keratinocytes in vitro. Using confocal microscopy, both proteins were shown to be at least partially associated with the keratin cytoskeleton. In addition, cultured keratinocytes efficiently secreted the S100A8/A9 dimer. These results together with previously published data suggest that S100A8 and S100A9 are novel players in wound repair, where they might be involved in the reorganization of the keratin cytoskeleton in the wounded epidermis, in the chemoattraction of inflammatory cells, and/or in the defense against microorganisms.

Transcriptional activity of GLI1 is negatively regulated by protein kinase A.

The transcription factor GLI1 is one of three vertebrate members of the GLI family, which is characterized by a highly conserved DNA-binding domain of five zinc fingers. We have analyzed whether human GLI1 is a target of PKA regulation. It was found that PKA inhibits GLI1 transcriptional activity. However, no evidence for proteolytic processing or for alteration in the subcellular distribution of GLI1 was obtained. The responsive PKA site (aa333-336) was localized to the second zinc finger of GLI1. Mutation of Ser336 revealed that PKA could also stimulate GLI1 transcriptional activity. Thus, our data demonstrate both negative and positive regulation of human GLI1 by PKA.

1,25 dihydroxyvitamin-D3 attenuates the confluence-dependent differences in the osteoblast characteristic proteins alkaline phosphatase, procollagen I peptide, and osteocalcin.

In the present study a cell culture model of primary human osteoblasts based on degrees of confluence was investigated by measuring basal and 1,25(OH)2D3stimulated levels of the osteoblast characteristic proteins alkaline phosphatase (AP), procollagen I-peptide (PICP), and osteocalcin (OC), as well as the corresponding gene expression. Primary osteoblast-like cell cultures from seven donors were treated in the second passage with 1,25(OH)2D3 (5 x 10(-8) M for 48 hours) and investigated at four stages of confluence (stage I 50%, stage II 75%, stage III 100%, and stage IV 7 days postconfluence). In untreated cultures passing through the different stages of confluence, we saw a 1.8-fold increase of AP activity, a 2. 3-fold increase of OC secretion, but a decrease of PICP levels to 0. 36-fold. Gene expression showed only minor variation between the different confluence stages. 1,25(OH)2D3 did not significantly affect PICP production. Alkaline phosphatase protein was stimulated during proliferation until confluence, with no effect thereafter. Surprisingly, OC secretion and mRNA expression were stimulated in all four stages to the same absolute level independent of basal values. We conclude that our results correspond to other studies showing differentiation-stage dependent changes of basal levels of osteoblast-specific proteins. However, 1,25(OH)2D3 stimulation decreased the confluence-dependent difference for AP and abolished it for osteocalcin, thus leading to a more differentiated phenotype of the osteoblast. Therefore, 1,25(OH)2D3 stimulation might improve the reproducibility of results obtained at different confluence stages from cultures of clinical samples.

Gli genes and limb development.

During development of the limb Shh plays a key role as a mediator of zone of polarizing activity (ZPA). However, the molecular mechanisms by which Shh directs anterior/posterior patterning in the limb remain unknown. Members of the Gli gene family encode zinc-finger transcription factors and represent likely candidates for being regulators of Shh target genes. In this review we would like to summarize the current knowledge on expression and function of Gli genes in limb development.