Stefan Blunder

Role of PPAR, LXR, and PXR in epidermal homeostasis and inflammation

Epidermal lipid synthesis and metabolism are regulated by nuclear hormone receptors (NHR) and in turn epidermal lipid metabolites can serve as ligands to NHR. NHR form a large superfamily of receptors modulating gene transcription through DNA binding. A subgroup of these receptors is ligand-activated and heterodimerizes with the retinoid X receptor including peroxisome proliferator-activated receptor (PPAR), liver X receptor (LXR) and pregnane X receptor (PXR). Several isotypes of these receptors exist, all of which are expressed in skin. In keratinocytes, ligand activation of PPARs and LXRs stimulates differentiation, induces lipid accumulation, and accelerates epidermal barrier regeneration. In the cutaneous immune system, ligand activation of all three receptors, PPAR, LXR, and PXR, has inhibitory properties, partially mediated by downregulation of the NF-kappaB pathway. PXR also has antifibrotic effects in the skin correlating with TGF-beta inhibition. In summary, ligands of PPAR, LXR and PXR exert beneficial therapeutic effects in skin disease and represent promising targets for future therapeutic approaches in dermatology. This article is part of a Special Issue entitled The Important Role of Lipids in the Epidermis and their Role in the Formation and Maintenance of the Cutaneous Barrier. Guest Editors: Kenneth R. Feingold and Peter Elias.

Low Energy Shock Wave Therapy Induces Angiogenesis in Acute Hind-Limb Ischemia via VEGF Receptor 2 Phosphorylation

Objectives Low energy shock waves have been shown to induce angiogenesis, improve left ventricular ejection fraction and decrease angina symptoms in patients suffering from chronic ischemic heart disease. Whether there is as well an effect in acute ischemia was not yet investigated. Methods Hind-limb ischemia was induced in 10–12 weeks old male C57/Bl6 wild-type mice by excision of the left femoral artery. Animals were randomly divided in a treatment group (SWT, 300 shock waves at 0.1 mJ/mm2, 5 Hz) and untreated controls (CTR), n = 10 per group. The treatment group received shock wave therapy immediately after surgery. Results Higher gene expression and protein levels of angiogenic factors VEGF-A and PlGF, as well as their receptors Flt-1 and KDR have been found. This resulted in significantly more vessels per high-power field in SWT compared to controls. Improvement of blood perfusion in treatment animals was confirmed by laser Doppler perfusion imaging. Receptor tyrosine kinase profiler revealed significant phosphorylation of VEGF receptor 2 as an underlying mechanism of action. The effect of VEGF signaling was abolished upon incubation with a VEGFR2 inhibitor indicating that the effect is indeed VEGFR 2 dependent. Conclusions Low energy shock wave treatment induces angiogenesis in acute ischemia via VEGF receptor 2 stimulation and shows the same promising effects as known from chronic myocardial ischemia. It may therefore develop as an adjunct to the treatment armentarium of acute muscle ischemia in limbs and myocardium.

Alteration of inflammatory response by shock wave therapy leads to reduced calcification of decellularized aortic xenografts in mice

OBJECTIVES Tissue-engineered xenografts represent a promising treatment option in heart valve disease. However, inflammatory response leading to graft failure and incomplete in vitro repopulation with recipient cells remain challenging. Shock waves (SWs) were shown to modulate inflammation and to enhance re-epithelialization. We therefore aimed to investigate whether SWs could serve as a feasible adjunct to tissue engineering. METHODS Porcine aortic pieces were decellularized using sodium deoxycholate and sodium dodecylsulphate and implanted subcutaneously into C57BL/6 mice (n = 6 per group). The treatment (shock wave therapy, SWT) group received SWs (0.1 mJ/mm(2), 500 impulses, 5 Hz) for modulation of inflammatory response directly after implantation; control animals remained untreated (CTR). Grafts were harvested 72 h and 3 weeks after implantation and analysed for inflammatory cytokines, macrophage infiltration and polarization, osteoclastic activity and calcification. Transmission electron microscopy (TEM) was performed. Endothelial cells (ECs) were treated with SWs and analysed for macrophage regulatory cytokines. In an ex vivo experimental set-up, decellularized porcine aortic valve conduits were reseeded with ECs with and without SWT (0.1 mJ/mm(2), 300 impulses, 3 Hz), fibroblasts as well as peripheral blood mononuclear cells (all human) and tested in a pulsatile flow perfusion system for cell coverage. RESULTS Treated ECs showed an increase of macrophage migration inhibitory factor and macrophage inflammatory protein 1β, whereas CD40 ligand and complement component C5/C5a were decreased. Subcutaneously implanted grafts showed increased mRNA levels of tumour necrosis factor α and interleukin 6 in the treatment group. Enhanced repopulation with recipient cells could be observed after SWT. Augmented macrophage infiltration and increased polarization towards M2 macrophages was observed in treated animals. Enhanced recruitment of osteoclastic cells in proximity to calcified tissue was found after SWT. Consequently, SWT resulted in decreased areas of calcification in treated animals. The reseeding experiment revealed that fibroblasts showed the best coverage compared with other cell types. Moreover, SW-treated ECs exhibited enhanced repopulation compared with untreated controls. CONCLUSIONS SWs reduce the calcification of subcutaneously implanted decellularized xenografts via the modulation of the acute macrophage-mediated inflammatory response and improves the in vitro repopulation of decellularized grafts. It may therefore serve as a feasible adjunct to heart valve tissue engineering.

Autosomal Recessive Keratoderma-Ichthyosis- Deafness (ARKID) Syndrome Is Caused by VPS33B Mutations Affecting Rab Protein Interaction and Collagen Modification

In this paper, we report three patients with severe palmoplantar keratoderma associated with ichthyosis and sensorineural deafness. Biallelic mutations were found in VPS33B, encoding VPS33B, a Sec1/Munc18 family protein that interacts with Rab11a and Rab25 proteins and is involved in trafficking of the collagen-modifying enzyme LH3. Two patients were homozygous for the missense variant p.Gly131Glu, whereas one patient was compound heterozygous for p.Gly131Glu and the splice site mutation c.240-1G>C, previously reported in patients with arthrogryposis renal dysfunction and cholestasis syndrome. We demonstrated the pathogenicity of variant p.Gly131Glu by assessing the interactions of the mutant VPS33B construct and its ability to traffic LH3. Compared with wild-type VPS33B, the p.Gly131Glu mutant VPS33B had reduced coimmunoprecipitation and colocalization with Rab11a and Rab25 and did not rescue LH3 trafficking. Confirming the cell-based experiments, we found deficient LH3-specific collagen lysine modifications in patients’ urine and skin fibroblasts. Additionally, the epidermal ultrastructure of the p.Gly131Glu patients mirrored defects in tamoxifen-inducible VPS33B-deficient Vps33bfl/fl-ERT2 mice. Both patients and murine models revealed an impaired epidermal structure, ascribed to aberrant secretion of lamellar bodies, which are essential for epidermal barrier formation. Our results demonstrate that p.Gly131Glu mutant VPS33B causes an autosomal recessive keratoderma-ichthyosis-deafness syndrome.

Alterations in Epidermal Eicosanoid Metabolism Contribute to Inflammation and Impaired Late Differentiation in FLG-Mutated Atopic Dermatitis.

Loss-of-function mutations in the FLG gene cause ichthyosis vulgaris (IV) and represent the major predisposing genetic risk factor for atopic dermatitis (AD). Although both conditions are characterized by epidermal barrier impairment, AD also exhibits signs of inflammation. This work was aimed at delineating the role of FLG loss-of-function mutations on eicosanoid metabolism in IV and AD. Using human epidermal equivalents (HEEs) generated with keratinocytes isolated from nonlesional skin of patients with FLG wild-type AD (WT/WT), FLG-mutated AD (FLG/WT), IV (FLG/FLG), or FLG WT control skin, we assessed the potential autocrine role of epidermal-derived eicosanoids in FLG-associated versus FLG-WT AD pathogenesis. Ultrastructural analyses demonstrated abnormal stratum corneum lipid architecture in AD and IV HEEs, independent of FLG genotype. Both AD (FLG/WT) and IV (FLG/FLG) HEEs showed impaired late epidermal differentiation. Only AD (FLG/WT) HEEs exhibited significantly increased levels of inflammatory cytokines. Analyses of lipid mediators revealed increased arachidonic acid and 12-lipoxygenase metabolites. Whereas treatment of control HEEs with arachidonic acid increased expression of inflammatory cytokines, 12-hydroxy-eicosatetraenoic acid attenuated expression of late differentiation markers. Thus, FLG mutations lead to alterations in epidermal eicosanoid metabolism that could serve as an autocrine trigger of inflammation and impaired late epidermal differentiation in AD.

Enhanced expression of genes related to xenobiotic metabolism in the skin of patients with atopic dermatitis but not with ichthyosis vulgaris

Previous transcriptome analyses underscored the importance of immunological and skin barrier abnormalities in atopic dermatitis (AD). We sought to identify pathogenic pathways involved in AD by comparing the transcriptomes of AD patients stratified for filaggrin (FLG)-null mutations to those of both healthy donors and patients with ichthyosis vulgaris. We applied RNA sequencing to analyze the whole transcriptome of nonlesional skin. We found that 607 genes (476 up-regulated and 131 down-regulated by >2-fold) and 193 genes (172 up-regulated and 21 down-regulated by >2-fold) were differentially expressed when all AD or ichthyosis vulgaris patients were compared with healthy donors, respectively. Expression of genes involved in RNA/protein turnover and adenosine triphosphate synthesis, as well as genes involved in cell death, response to oxidative stress, DNA damage/repair, and autophagy, were significantly enriched in AD skin and, to a lesser extent, in ichthyosis vulgaris skin. FLG-null mutations appear to hardly interfere with current observations. Genes related to xenobiotic metabolism were up-regulated in AD skin only, as were genes related to arachidonic, linoleic, and α-linolenic acid metabolism. Thus, this work newly links AD pathogenesis to aberrant expression of genes related to xenobiotic metabolism.

Aggregated neutrophil extracellular traps resolve inflammation by proteolysis of cytokines and chemokines and protection from antiproteases

Papillon‐Lefèvre syndrome (PLS) is characterized by nonfunctional neutrophil serine proteases (NSPs) and fulminant periodontal inflammation of unknown cause. Here we investigated neutrophil extracellular trap (NET)‐associated aggregation and cytokine/chemokine‐release/degradation by normal and NSP‐deficient human and mouse granulocytes. Stimulated with solid or soluble NET inducers, normal neutrophils formed aggregates and both released and degraded cytokines/chemokines. With increasing cell density, proteolytic degradation outweighed release. Maximum output of cytokines/chemokines occurred mostly at densities between 2 × 107 and 4 × 107 neutrophils/cm3. Assessment of neutrophil density in vivo showed that these concentrations are surpassed during inflammation. Association with aggregated NETs conferred protection of neutrophil elastase against αl‐antitrypsin. In contrast, eosinophils did not influence cytokine/chemokine concentrations. The proteolytic degradation of inflammatory mediators seen in NETs was abrogated in Papillon–Lèfevre syndrome (PLS) neutrophils. In summary, neutrophil‐driven proteolysis of inflammatory mediators works as a built‐in safeguard for inflammation. The absence of this negative feedback mechanism might be responsible for the nonresolving periodontitis seen in PLS.—Hahn, J., Schauer, C., Czegley, C., Kling, L., Petru, L., Schmid, B., Weidner, D., Reinwald, C., Biermann, M. H. C., Blunder, S., Ernst, J., Lesner, A., Bäuerle, T., Palmisano, R., Christiansen, S., Herrmann, M., Bozec, A., Gruber, R., Schett, G., Hoffmann, M. H. Aggregated neutrophil extracellular traps resolve inflammation by proteolysis of cytokines and chemokines and protection from antiproteases. FASEB J. 33, 1401–1414 (2019). www.fasebj.org

ÖGDV Preisträger stellen sich vor: Der Österreichische Dermatologenpreis - Unilever Preis 2017 ging an Dr. Stefan Blunder aus Innsbruck

Im Rahmen der Jahrestagung der Österreichischen Gesellschaft für Dermatologie und Venerologie 2017 in Salzburg wurde Dr. Stefan Blunder mit dem Österreichischen Dermatologen-Preis ausgezeichnet. Dieser von UNILEVER Austria GmbH gestiftete Preis wird für Arbeiten mit Grundlagen orientierten experimentellen Fragestellungen verliehen. Die Pathogenese der Atopischen Dermatitis ist Gegenstand intensiver Forschungsbemühungen. Dies ist der Fall obwohl die Rolle des Immunsystems sowie der epidermalen Barriere in der Entstehung dieser Erkrankung gemeinhin anerkannt sind. Insbesondere der Stellenwert des Gens Filaggrin , welches für ein zentrales epidermales Strukturprotein kodiert, wird diskutiert. Mutationen im Gen Filaggrin führen zur Beeinträchtigung der epidermalen Barriere und sind nicht nur kausal für die nicht-entzündliche Hauterkrankung Ichthyosis Vulgaris, sondern stellen auch den wichtigsten, bisher bekannten genetischen Risikofaktor für die Entstehung der Atopischen Dermatitis dar. Das Ziel der prämierten Arbeit war es, die molekulare Signatur der Haut von Patienten mit Atopischer Dermatitis mit und ohne Filaggrin Mutation sowie Ichthyosis vulgaris zu entschlüsseln, um somit die Rolle von Filaggrin in der Entstehung der Atopischen Dermatitis versus Ichthyosis vulgaris genauer defi nieren zu können. Zu diesem Zweck verwendeten die Autoren „Next Generation Sequencing“ um die Transkriptome von Hautproben von Kontrollpersonen und Patienten mit (i) Atopischer Dermatitis ohne Filaggrin Mutationen, (ii) Atopischer Dermatitis mit Filaggrin Mutationen, (iii) Ichthyosis Vulgaris sowie von Patienten mit (iiii) Ichthyosis Vulgaris und Atopischer Dermatitis zu vergleichen [ 1 ] . Dabei wurde die überraschende Entdeckung gemacht, dass in der Atopischen Dermatitis – unabhängig vom Filaggrin Mutationsstatus – Gene, welche im Metabolismus von Xenobiotika eine entscheidende Rolle spielen, verstärkt exprimiert werden. Xenobiotika sind nicht-natürliche Umweltsubstanzen, mit denen die Haut konfrontiert ist. Die Ergebnisse lassen den unerwarteten Schluss zu, dass vermehrte Exposition gegenüber „körperfremden“ Stoffen, wie zum Beispiel pharmazeutischen Wirkstoffen, Pestiziden, Toxinen, endokrinen Disruptoren, Umweltgiften und anderen chemischen Verbindungen, eine bisher unbekannte Rolle in der Entstehung der Atopischen Dermatitis spielt. Somit weist die Arbeit auf einen bis dato kaum beachteten Mechanismus in der Entstehung der Atopischen Dermatitis hin, welcher zukünftig einen möglichen Ansatz in der Behandlung und der Prävention der Atopischen Dermatitis darstellen könnte. Dr. Blunder absolvierte sein Medizinstudium an der Medizinischen Universität Innsbruck und der Universität Heidelberg. Während seines Medizinstudiums verbrachte er acht Monate am Sidney Kimmel Comprehensive Cancer Center der School of Medicine der Johns Hopkins University bei Herrn Prof. Dr. Drew Pardoll. Nach Abschluss seines Medizinstudiums arbeitete er in der Forschungsgruppe von Herrn Prof. Dr. Christoph Höller an der Abteilung für Dermatologie der Medizinischen Universität Wien. Daran anschließend absolvierte er ein Ph.D. Studium bei Frau Prof. Dr. Sandrine Dubrac und Herrn Prof. Dr. Matthias Schmuth an der Klinik für Dermatologie, Venerologie und Allergologie der Medizinischen Universität Innsbruck. Ebendort steht er zurzeit in Ausbildung zum Facharzt. Sein derzeitiger Forschungsschwerpunkt liegt auf dem Gebiet der epidermalen Barriere.

Epidermale Barriere bei hereditären Ichthyosen, atopischer Dermatitis und Psoriasis

Zahlreiche Hautkrankheiten gehen mit Störungen der Hautbarriere einher. Man unterscheidet primäre und sekundäre Barrierestörungen. Erstere sind monogen durch Defekte in Schlüsselkomponenten der Epidermis bedingt (z.B. Ichthyosen). Sekundäre Barrierestörungen treten bei entzündlichen Hautkrankheiten auf, bei denen die epidermale Homöostase beeinträchtigt ist (z.B. Ekzeme, Psoriasis u.v.a.m.). Bei diesen Erkrankungen hemmt die Entzündung den Aufbau oder Erhalt intakter Barrierstrukturen. Neue Forschungsergebnisse sprechen dafür, dass bei der atopischen Dermatitis und in geringerem Maß bei der Psoriasis eine Kombination primärer und sekundärer Barrierestörungen vorliegt. Voraussichtlich lassen sich zukünftig Subtypen dieser Erkrankungen definieren, bei denen die eine oder die andere Komponente im Vordergrund steht.

Slow growth of hair and nails, craniofacial abnormalities and brachyphalangy.

Prominent forehead, bulbous tip of the nose, long philtrum with slightly developed philtrum ridges, thin vermilion of the upper lip and slight rarification of the lateral eyebrows (Figure 1). Fine, thin, blonde hair. Fair complexion, dry skin. The nails were thin and short, particularly on the thumbs and great toes, but normal nail structure. Limbs: Brachydactyly, ulnar deviation in the proximal interphalangeal joint of the index finger, thickening of the proximal phalanges of the index, middle and ring fingers, clinodactyly of the fifth fingers, broad and short thumbs and great toes. Hypermobility of the joints; skeletal system otherwise normal. No palmoplantar hyperhidrosis or hyperlinearity. No dental anomalies.