Oliver Seifert

Keloid scarring: bench and bedside

Wound healing is a fundamental complex-tissue reaction leading to skin reconstitution and thereby ensuring survival. While, fetal wounds heal without scarring, a normal “fine line” scar is the clinical outcome of an undisturbed wound healing in adults. Alterations in the orchestrated wound healing process result in hypertrophic or keloid scarring. Research in the past decades attempted to identify genetic, cellular, and molecular factors responsible for these alterations. These attempts lead to several new developments in treatments for keloids, such as, imiquimod, inhibition of transforming growth factor beta, and recombinant interleukin-10. The urgent need for better therapeutics is underlined by recent data substantiating an impaired quality of life in keloid and hypertrophic scar patients. Despite the increasing knowledge about the molecular regulation of scar formation no unifying theory explaining keloid development has been put forward until today. This review aims to give an overview about the genetic and molecular background of keloids and focus of the current research on keloid scarring with special emphasis on new forthcoming treatments. Clinical aspects and the spectrum of scarring are summarized.

Identification of unique gene expression patterns within different lesional sites of keloids

Keloid disease is a significant clinical problem, especially in black populations, with an estimated incidence of 4–16%. Keloids are fibroproliferative dermal tumors developing as a result of deregulated wound healing. The dynamic nature of keloids is illustrated by clinical regression in the center, while the margin remains active growing into the surrounding healthy skin. Therefore, the gene expression profiles of fibroblasts from different sites of the keloids were characterized using Affymetrix microarrays covering the whole human genome. This study revealed 105 genes that were differentially regulated (79 genes were up‐regulated and 26 down‐regulated) in a unique gene expression profile in different sites of keloids where progression or regression of the process was in progress. The apoptosis inhibitor AVEN was found to be up‐regulated at the active margin of keloids, while apoptosis‐inducing genes such as ADAM12 and genes inducing extracellular matrix (ECM) degradation such as matrix metalloproteinase‐19 were up‐regulated in the regressing keloid center. We identified genes previously not described in the development of keloids. Activating proapoptotic genes or inhibiting ECM‐inducing genes as INHBA or monocyte chemoattractant protein‐1 might be possible target genes for new treatment strategies for keloid disease.

The skin pathergy test: innately useful?

Pathergy is the term used to describe hyper-reactivity of the skin that occurs in response to minimal trauma. A positive skin pathergy test (SPT), characterised by erythematous induration at the site of the needle stick with a small pustule containing sterile pus at its centre, is among the criteria required for a diagnosis of Behçet’s disease (BD) and in certain population has been shown to be highly specific for this condition. Problems with standardising the induction manoeuvre for the SPT as well as the method of assessment of the response have limited the usefulness of the SPT in the clinical setting. Extensive investigation into histopathological and immunological aspects of pathergy has led to a number of hypotheses relating to the aetiology of the skin pathergy reaction and the disease itself, but the cause is considered to be unknown. Pathergy lesions, the development of new skin lesions or the aggravation of existing ones following trivial trauma, are also reported in pyoderma gangrenosum and has been noted in other neutrophilic dermatoses such as Sweet’s syndrome. The response of such patient groups to the systematic application of the SPT has not been described. We propose that a new way of considering the pathergy reaction is to see it as an aberration of the skin’s innate reactivity from a homeostatic reactive mode closely coupled to tissue healing to an abnormal destructive/inflammatory mode. Our understanding of BD and other similar conditions would profit by more detailed mechanistic knowledge of skin homeostasis to minimal trauma in both health and disease through a more structured and systematic use of the SPT.

Ultraviolet exposure of melanoma cells induces fibroblast activation protein- α in fibroblasts: Implications for melanoma invasion

Fibroblast activation protein-α (FAP-α) promotes tumor growth and cell invasiveness through extracellular matrix degradation. How ultraviolet radiation (UVR), the major risk factor for malignant melanoma, influences the expression of FAP-α is unknown. We examined the effect of UVR on FAP-α expression in melanocytes, keratinocytes and fibroblasts from the skin and in melanoma cells. UVR induces upregulation of FAP-α in fibroblasts, melanocytes and primary melanoma cells (PM) whereas keratinocytes and metastatic melanoma cells remained FAP-α negative. UVA and UVB stimulated FAP-α-driven migration and invasion in fibroblasts, melanocytes and PM. In co-culture systems UVR of melanocytes, PM and cells from regional metastases upregulated FAP-α in fibroblasts but only supernatants from non-irradiated PM were able to induce FAP-α in fibroblasts. Further, UV-radiated melanocytes and PM significantly increased FAP-α expression in fibroblasts through secretory crosstalk via Wnt5a, PDGF-BB and TGF-β1. Moreover, UV radiated melanocytes and PM increased collagen I invasion and migration of fibroblasts. The FAP-α/DPPIV inhibitor Gly-ProP(OPh)2 significantly decreased this response implicating FAP-α/DPPIV as an important protein complex in cell migration and invasion. These experiments suggest a functional association between UVR and FAP-α expression in fibroblasts, melanocytes and melanoma cells implicating that UVR of malignant melanoma converts fibroblasts into FAP-α expressing and ECM degrading fibroblasts thus facilitating invasion and migration. The secretory crosstalk between melanoma and tumor surrounding fibroblasts is mediated via PDGF-BB, TGF-β1 and Wnt5a and these factors should be evaluated as targets to reduce FAP-α activity and prevent early melanoma dissemination.

Gene expression profiling of macrophages: implications for an immunosuppressive effect of dissolucytotic gold ions

BackgroundGold salts has previously been used in the treatment of rheumatoid arthritis but have been replaced by biologicals such as TNF-α inhibitors. The mechanisms behind the anti-inflammatory effect of metallic gold ions are still unknown, however, recent data showed that charged gold atoms are released from pure metallic gold implants by macrophages via a dissolucytosis membrane, and that gold ions are taken up by local macrophages, mast cells and to some extent fibroblasts. These findings open the question of possible immunomodulatory effects of metallic gold and motivate efforts on a deeper understanding of the effect of metallic gold on key inflammatory cells as macrophages.MethodsHuman macrophage cells (cell line THP-1) were grown on gold foils and intracellular uptake was analysed by autometallography. The impact of phagocytised gold ions on viability of THP-1 cells was investigated by trypan blue staining and TUNEL assay. The global gene expression profile of THP-1 cells after incorporation of gold ions was studied using microarray analysis comprising approximately 20,000 genes. The gene expression data was confirmed by measurement of secreted proteins.ResultsAutometallography showed intracellular uptake of gold ions into THP-1 cells. No significant effect on viability of THP-1 cells was demonstrated. Our data revealed a unique gene expression signature of dissolucytotic THP-1 cells that had taken up gold ions. A large number of regulated genes were functionally related to immunomodulation. Gold ion uptake induced downregulation of genes involved in rheumatoid arthritis such as hepatocyte growth factor, tenascin-C, inhibitor of DNA binding 1 and 3 and matrix metalloproteinase 13.ConclusionThe data obtained in this study offer new insights into the mode of action of gold ions and suggest for the investigation of effects on other key cells and a possible future role of metallic gold as implants in rheumatoid arthritis or other inflammatory conditions.

UV radiation promotes melanoma dissemination mediated by the sequential reaction axis of cathepsins–TGF- β 1–FAP- α

Background:Ultraviolet radiation (UVR) is the major risk factor for development of malignant melanoma. Fibroblast activation protein (FAP)-α is a serine protease expressed on the surface of activated fibroblasts, promoting tumour invasion through extracellular matrix (ECM) degradation. The signalling mechanism behind the upregulation of FAP-α is not yet completely revealed.Methods:Expression of FAP-α was analysed after UVR exposure in in vitro co-culture systems, gene expression arrays and artificial skin constructs. Cell migration and invasion was studied in relation to cathepsin activity and secretion of transforming growth factor (TGF)-β1.Results:Fibroblast activation protein-α expression was induced by UVR in melanocytes of human skin. The FAP-α expression was regulated by UVR-induced release of TGF-β1 and cathepsin inhibitors prevented such secretion. In melanoma cell culture models and in a xenograft tumour model of zebrafish embryos, FAP-α mediated ECM degradation and facilitated tumour cell dissemination.Conclusions:Our results provide evidence for a sequential reaction axis from UVR via cathepsins, TGF-β1 and FAP-α expression, promoting cancer cell dissemination and melanoma metastatic spread.

Significant Changes in the Skin Microbiome in Patients with Chronic Plaque Psoriasis after Treatment with Narrowband Ultraviolet B

Changes in the skin microbiome have been shown to promote cutaneous inflammation. The skin microbiome of patients with chronic plaque type psoriasis was analysed before and after treatment with narrowband ultraviolet B (UVB). Swab samples of the microbiome were taken from lesional and non-lesional skin of 26 patients. Microbiotas were characterized by sequencing 16S rRNA bacterial genes on the Illumina MiSeq platform. Lesional skin microbiome diversity correlated with psoriasis severity (measured with the Psoriasis Area and Severity Index; PASI). There was a significantly lower abundance of the phylum Firmicutes and the genus Staphylococcus in lesional skin compared with non-lesional skin before UVB treatment. Responders (> 75% target Psoriasis Severity Index (PSI) improvement) had significantly lower abundance of the phyla Firmicutes in lesional and non-lesional skin and lower abundance of the genera Staphylococcus, Finegoldia, Anaerococcus, Peptoniphilus, Gardnerella, Prevotella and Clostridium in lesional skin after UVB treatment. Pseudomonas significantly decreased in lesional and non-lesional skin of treatment responders. These results suggest that skin microbiome alterations after UVB treatment could be related to treatment and treatment response.

Bullous Pemphigoid: Validation of the National Patient Register in Two Counties in Sweden, 2001 to 2012.

Bullous pemphigoid (BP) is the most common autoimmune blistering disease. Since 2001, data from all specialized outpatient and inpatient care institutions in Sweden, have been registered with the National Patient Register (NPR), based on a unique personal identification number. Previous validations of the register have shown high accuracy for various non-dermatological autoimmune diseases. In order to validate the diagnosis of BP, all residents aged < 20 years in 2 counties in Sweden (539,000 inhabitants) diagnosed with bullous pemphigoid (ICD-10; L12.0, L12.8, L12.9) in the period 2001 to 2012 were identified in the NPR. Medical records, as well as immuno- and histopathological data, were reviewed for this study. A total of 323 patients with BP were identified in the NPR. Of these, 178 patients had a directly confirmed diagnosis of BP from immuno- and histopathological data, reviewed by a dermatopathologist. For the remaining 145 patients medical records were retrieved and further reviewed by 2 dermatologists. Of these, 105 patients had a confirmed diagnosis of BP. The medical records of 16 patients were missing, and 24 patients were not classified as having BP. Overall, a positive predictive value of 92% (283/307) was found for BP in the NPR. In conclusion, the present validation of medical records and immuno- and histopathological data showed high validity for the diagnosis of BP in the Swedish NPR.

Transforming Growth Factor Beta Gene Signatures are Spatially Enriched in Keloid Tissue Biopsies and Ex vivo-Cultured Keloid Fibroblasts

The keloid lesion is recognised as a spatially heterogeneous mass both in cellular and acellular composition and biological activity. Here, we have utilised a bioinformatic approach to determine whether this spatial heterogeneity is also evident at the molecular level and to identify key upstream regulators of signalling pathways enriched in the lesion in a spatially-restricted manner. Differentially expressed genes (20% change, p < 0.05) obtained from microarray datasets derived from whole keloid biopsies and ex vivo-cultured keloid fibroblasts, both from distinct regions of the keloid lesion (leading edge, centre, and top) have been analysed to show that the TGFβ family plays a significant but spatially dependent role in regulation of keloid gene expression. Furthermore, we have identified additional upstream signalling molecules involved in driving keloid biology and provide information on therapeutic targets whose modulation might be expected to lead to significant therapeutic efficacy.

Expression of low-density lipoprotein-related receptors 5 and 6 (LRP5/6) in psoriasis skin

Low‐density lipoprotein‐related receptors 5 and 6 (LRP5/6) are transmembrane receptors with key functions in canonical Wnt signalling. Wnt ligands are thought to play an important role in innate immunity and psoriasis, and recent studies assigned LRP5/6 anti‐inflammatory properties. The objective of this study was to investigate the expression of LRP5 and LRP6 in lesional and non‐lesional skin in peripheral blood and in mononuclear cells of patients with chronic plaque type psoriasis compared with control individuals. To investigate the effect of UV‐B radiation, LRP5/6 skin gene expression was analysed before and after narrowband UV‐B treatment. Our results showed significantly decreased gene expression of LRP5 and LRP6 in lesional skin and in peripheral blood from patients with psoriasis compared with non‐lesional skin and healthy control skin. Immunohistochemistry did not reveal differences in protein expression of LRP5/6. Narrowband UV‐B treatment induced a significant increase in LRP5 and LRP6 gene expression in lesional skin. Decreased gene expression of LRP5/6 in lesional skin and upregulation after nb UV‐B treatment suggest a possible role for LRP5/6 in psoriasis.