Publications by authors named "Patrick L J M Zeeuwen"

66 Publications

Skin microbiota in health and disease: From sequencing to biology.

J Dermatol 2020 Oct 17;47(10):1110-1118. Epub 2020 Aug 17.

Department of Dermatology, RIMLS, Radboudumc, Nijmegen, The Netherlands.

Microbiota live in a closely regulated interaction with their environment, and vice versa. The presence and absence of microbial entities is greatly influenced by features of the niche in which they thrive. Characteristic of this phenomenon is that different human skin sites harbor niche-specific communities of microbes. Microbial diversity is considerable, and the current challenge lies in determining which microbes and (corresponding) functionality are of importance to a given ecological niche. Furthermore, as there is increasing evidence of microbial involvement in health and disease, the need arises to fundamentally understand microbiome processes for application in health care, nutrition and personal care products (e.g. diet, cosmetics, probiotics). This review provides a current overview of state-of-the-art sequencing-based techniques and corresponding data analysis methodology for profiling of complex microbial communities. Furthermore, we also summarize the existing knowledge regarding cutaneous microbiota and their human host for a wide range of skin diseases.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1111/1346-8138.15536DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7589227PMC
October 2020

Terminal keratinocyte differentiation in vitro is associated with a stable DNA methylome.

Exp Dermatol 2020 Jul 17. Epub 2020 Jul 17.

Department of Dermatology, Radboud Institute for Molecular Life Sciences (RIMLS), Radboud University Medical Center (Radboudumc), Nijmegen, The Netherlands.

The epidermal compartment of the skin is regenerated constantly by proliferation of epidermal keratinocytes. Differentiation of a subset of these keratinocytes allows the epidermis to retain its barrier properties. Regulation of keratinocyte fate-whether to remain proliferative or terminally differentiate-is complex and not fully understood. The objective of our study was to assess if DNA methylation changes contribute to the regulation of keratinocyte fate. We employed genome-wide MethylationEPIC beadchip array measuring approximately 850 000 probes combined with RNA sequencing of in vitro cultured non-differentiated and terminally differentiated adult human primary keratinocytes. We did not observe a correlation between methylation status and transcriptome changes. Moreover, only two differentially methylated probes were detected, of which one was located in the TRIM29 gene. Although TRIM29 knock-down resulted in lower expression levels of terminal differentiation genes, these changes were minor. From these results, we conclude that-in our in vitro experimental setup-it is unlikely that changes in DNA methylation have an important regulatory role in terminal keratinocyte differentiation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1111/exd.14153DOI Listing
July 2020

Know your enemy: Unexpected, pervasive and persistent viral and bacterial contamination of primary cell cultures.

Exp Dermatol 2020 Jun 7. Epub 2020 Jun 7.

Department of Dermatology, Radboud University Medical Center (Radboudumc), Radboud Institute for Molecular Life Sciences (RIMLS), Nijmegen, The Netherlands.

In biomedical research, cell culture contamination is one of the main culprits of experimental failure. Contamination sources and concomitant remedies are numerous and challenging to manage. We herein describe two cases of uncommon contamination of cell cultures that we encountered, and the successful determination and eradication strategies. The first case describes the infection with human adenovirus C that originated from pharyngeal tonsils used for isolation of primary tonsillar epithelial cells. It is known that viral contamination of in vitro cell cultures can occur symptomless and is therefore difficult to identify. The contamination was pervasive and persistent, as it was widely spread in flow cabinets and apparatus, and has caused a serious delay to our research projects and the inevitable loss of valuable (patient-derived) cell sources. Eradication was successful by formalin gas sterilization of the flow cabinet and elimination of all infected cell lines from our biobank after PCR-guided determination. Secondly, we encountered a spore-forming bacterium, namely Brevibacillus brevis, in our cell culture facility. This bacterium originated from contaminated tap water pipes and spread via regular aseptic culture techniques due to survival of the bacterial spores in 70% ethanol. B brevis overgrew the cultures within a few days after seeding of the primary cells. Chlorine solution effectively killed this spore-forming bacterium. Both cases of contamination were identified using DNA sequencing which enabled the deployment of targeted aseptic techniques for the elimination of the persistent contamination.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1111/exd.14126DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7496648PMC
June 2020

A generic workflow for Single Locus Sequence Typing (SLST) design and subspecies characterization of microbiota.

Sci Rep 2019 12 27;9(1):19834. Epub 2019 Dec 27.

Center for Molecular and Biomolecular Informatics (CMBI), Radboud Institute for Molecular Life Sciences (RIMLS), Radboud University Medical Center (Radboudumc), Nijmegen, The Netherlands.

We present TaxPhlAn, a new method and bioinformatics pipeline for design and analysis of single-locus sequence typing (SLST) markers to type and profile bacteria beyond the species-level in a complex microbial community background. TaxPhlAn can be applied to any group of phylogenetically-related bacteria, provided reference genomes are available. As TaxPhlAn requires the SLST targets identified to fit the phylogenetic pattern as determined through comprehensive evolutionary reconstruction of input genomes, TaxPhlAn allows for the identification and phylogenetic inference of new biodiversity. Here, we present a clinically relevant case study of high-resolution Staphylococcus profiling on skin of atopic dermatitis (AD) patients. We demonstrate that SLST enables profiling of cutaneous Staphylococcus members at (sub)species level and provides higher resolution than current 16S-based techniques. With the higher discriminative ability provided by our approach, we further show that the presence of Staphylococcus capitis on the skin together with Staphylococcus aureus associates with AD disease.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-019-56065-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6934516PMC
December 2019

Microbe-host interplay in atopic dermatitis and psoriasis.

Nat Commun 2019 10 16;10(1):4703. Epub 2019 Oct 16.

Department of Dermatology, University Hospital Schleswig-Holstein, Kiel, 24105, Germany.

Despite recent advances in understanding microbial diversity in skin homeostasis, the relevance of microbial dysbiosis in inflammatory disease is poorly understood. Here we perform a comparative analysis of skin microbial communities coupled to global patterns of cutaneous gene expression in patients with atopic dermatitis or psoriasis. The skin microbiota is analysed by 16S amplicon or whole genome sequencing and the skin transcriptome by microarrays, followed by integration of the data layers. We find that atopic dermatitis and psoriasis can be classified by distinct microbes, which differ from healthy volunteers microbiome composition. Atopic dermatitis is dominated by a single microbe (Staphylococcus aureus), and associated with a disease relevant host transcriptomic signature enriched for skin barrier function, tryptophan metabolism and immune activation. In contrast, psoriasis is characterized by co-occurring communities of microbes with weak associations with disease related gene expression. Our work provides a basis for biomarker discovery and targeted therapies in skin dysbiosis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41467-019-12253-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6795799PMC
October 2019

Targeting the Cutaneous Microbiota in Atopic Dermatitis by Coal Tar via AHR-Dependent Induction of Antimicrobial Peptides.

J Invest Dermatol 2020 02 22;140(2):415-424.e10. Epub 2019 Jul 22.

Department of Dermatology, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, Nijmegen, The Netherlands. Electronic address:

Skin colonization by Staphylococcus aureus and its relative abundance is associated with atopic dermatitis (AD) disease severity and treatment response. Low levels of antimicrobial peptides in AD skin may be related to the microbial dysbiosis. Therapeutic targeting of the skin microbiome and antimicrobial peptide expression can, therefore, restore skin homeostasis and combat AD. In this study, we analyzed the cutaneous microbiome composition in 7 patients with AD and 10 healthy volunteers upon topical coal tar or vehicle treatment. We implemented and validated a Staphylococcus-specific single-locus sequence typing approach combined with classic 16S ribosomal RNA marker gene sequencing to study the bacterial composition. During coal tar treatment, Staphylococcus abundance decreased, and Propionibacterium abundance increased, suggesting a shift of the microbiota composition toward that of healthy controls. We, furthermore, identified a hitherto unknown therapeutic mode of action of coal tar, namely the induction of keratinocyte-derived antimicrobial peptides via activation of the aryl hydrocarbon receptor. Restoring antimicrobial peptide levels in AD skin via aryl hydrocarbon receptor-dependent transcription regulation can be beneficial by creating a (anti)microbial milieu that is less prone to infection and inflammation. This underscores the importance of coal tar in the therapeutic aryl hydrocarbon receptor armamentarium and highlights the aryl hydrocarbon receptor as a target for drug development.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.jid.2019.06.142DOI Listing
February 2020

Stable pantothenamide bioisosteres: novel antibiotics for Gram-positive bacteria.

J Antibiot (Tokyo) 2019 09 6;72(9):682-692. Epub 2019 Jun 6.

Department of Dermatology, Radboud University Medical Center, Nijmegen, The Netherlands.

The emergence of multidrug resistant bacteria has prioritized the development of new antibiotics. N-substituted pantothenamides, analogs of the natural compound pantetheine, were reported to target bacterial coenzyme A biosynthesis, but these compounds have never reached the clinic due to their instability in biological fluids. Plasma-stable pantothenamide analogs could overcome these issues. We first synthesized a number of bioisosteres of the prototypic pantothenamide N7-Pan. A compound with an inverted amide bond (CXP18.6-012) was found to provide plasma-stability with minimal loss of activity compared to the parent compound N7-Pan. Next, we synthesized inverted pantothenamides with a large variety of side chains. Among these we identified a number of novel stable inverted pantothenamides with selective activity against Gram-positive bacteria such as staphylococci and streptococci, at low micromolar concentrations. These data provide future direction for the development of pantothenamides with clinical potential.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41429-019-0196-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6760626PMC
September 2019

STAT1 gain-of-function compromises skin host defense in the context of IFN-γ signaling.

J Allergy Clin Immunol 2019 04 18;143(4):1626-1629.e5. Epub 2018 Dec 18.

Department of Internal Medicine, Radboud University Medical Center, Radboud Center for Infectious Diseases, Nijmegen, The Netherlands. Electronic address:

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.jaci.2018.11.033DOI Listing
April 2019

Deficiency of the human cysteine protease inhibitor cystatin M/E causes hypotrichosis and dry skin.

Genet Med 2019 07 14;21(7):1559-1567. Epub 2018 Nov 14.

Department of Dermatology, Radboud Institute for Molecular Life Sciences (RIMLS), Radboud University Nijmegen Medical Center (Radboudumc), Nijmegen, The Netherlands.

Purpose: We aimed to assess the biological and clinical significance of the human cysteine protease inhibitor cystatin M/E, encoded by the CTS6 gene, in diseases of human hair and skin.

Methods: Exome and Sanger sequencing was performed to reveal the genetic cause in two related patients with hypotrichosis. Immunohistochemical, biophysical, and biochemical measurements were performed on patient skin and 3D-reconstructed skin from patient-derived keratinocytes.

Results: We identified a homozygous variant c.361C>T (p.Gln121*), resulting in a premature stop codon in exon 2 of CST6 associated with hypotrichosis, eczema, blepharitis, photophobia and impaired sweating. Enzyme assays using recombinant mutant cystatin M/E protein, generated by site-directed mutagenesis, revealed that this p.Gln121* variant was unable to inhibit any of its three target proteases (legumain and cathepsins L and V). Three-dimensional protein structure prediction confirmed the disturbance of the protease/inhibitor binding sites of legumain and cathepsins L and V in the p.Gln121* variant.

Conclusion: The herein characterized autosomal recessive hypotrichosis syndrome indicates an important role of human cystatin M/E in epidermal homeostasis and hair follicle morphogenesis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41436-018-0355-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6752276PMC
July 2019

The skin barrier: Epidermis vs environment.

Exp Dermatol 2018 08;27(8):805-806

Department of Dermatology, Radboud Institute for Molecular Life Sciences (RIMLS), Radboud University Nijmegen Medical Center (Radboudumc), Nijmegen, The Netherlands.

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1111/exd.13731DOI Listing
August 2018

3D skin models for 3R research: The potential of 3D reconstructed skin models to study skin barrier function.

Exp Dermatol 2018 05 6;27(5):501-511. Epub 2018 Apr 6.

Department of Dermatology, Radboud university medical center (Radboudumc), Radboud Institute for Molecular Life Sciences (RIMLS), Nijmegen, The Netherlands.

The skin barrier is an important shield regulating the outside-in as well as inside-out penetration of water, nutrients, ions and environmental stimuli. We can distinguish four different barrier compartments: the physical, chemical, immunological and microbial skin barrier. Well-functioning of those is needed to protect our body from the environment. To better understand the function and the contribution of barrier dysfunction in skin diseases, 3D skin or epidermal models are a valuable tool for in vitro studies. In this review, we summarize the development and application of different skin models in skin barrier research. During the last years, enormous effort was made on optimizing these models to better mimic the in vivo composition of the skin, by fine-tuning cell culture media, culture conditions and including additional cells and tissue components. Thereby, in vitro barrier formation and function has been improved significantly. Moreover, in this review we point towards changes and chances for in vitro 3D skin models to be used for skin barrier research in the nearby future.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1111/exd.13531DOI Listing
May 2018

Immortalized N/TERT keratinocytes as an alternative cell source in 3D human epidermal models.

Sci Rep 2017 09 19;7(1):11838. Epub 2017 Sep 19.

Department of Dermatology, Radboud Institute for Molecular Life Sciences (RIMLS), Radboud University Medical Center (Radboudumc), PO BOX 9101, 6500 HB, Nijmegen, The Netherlands.

The strong societal urge to reduce the use of experimental animals, and the biological differences between rodent and human skin, have led to the development of alternative models for healthy and diseased human skin. However, the limited availability of primary keratinocytes to generate such models hampers large-scale implementation of skin models in biomedical, toxicological, and pharmaceutical research. Immortalized cell lines may overcome these issues, however, few immortalized human keratinocyte cell lines are available and most do not form a fully stratified epithelium. In this study we compared two immortalized keratinocyte cell lines (N/TERT1, N/TERT2G) to human primary keratinocytes based on epidermal differentiation, response to inflammatory mediators, and the development of normal and inflammatory human epidermal equivalents (HEEs). Stratum corneum permeability, epidermal morphology, and expression of epidermal differentiation and host defence genes and proteins in N/TERT-HEE cultures was similar to that of primary human keratinocytes. We successfully generated N/TERT-HEEs with psoriasis or atopic dermatitis features and validated these models for drug-screening purposes. We conclude that the N/TERT keratinocyte cell lines are useful substitutes for primary human keratinocytes thereby providing a biologically relevant, unlimited cell source for in vitro studies on epidermal biology, inflammatory skin disease pathogenesis and therapeutics.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-017-12041-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5605545PMC
September 2017

The Effects of Human Beta-Defensins on Skin Cells in vitro.

Dermatology 2017 8;233(2-3):155-163. Epub 2017 Jul 8.

Department of Dermatology, Radboud Institute for Molecular Life Sciences (RIMLS), Radboud University Medical Centre (Radboudumc), Nijmegen, The Netherlands.

Background: Defensins are antimicrobial peptides that exert immunomodulatory and chemotactic functions. Based on these properties and their high expression levels in the skin, they are likely to affect skin inflammation, infection, and wound healing. This may lead to therapeutic applications in (burn) wound healing.

Objective: We aimed to investigate the effects of human β-defensins (hBDs) on keratinocytes and fibroblasts, 2 major skin cell types involved in skin regeneration.

Methods: Monolayer keratinocyte and fibroblast cultures were exposed to recombinant hBDs, and we overexpressed hBD2 and hBD3 in keratinocytes of reconstructed epidermal equivalents by lentiviral transduction. The effects were measured by immunohistochemistry, quantitative real-time PCR, and migration assays. Kinome analyses were performed on cultured keratinocytes to investigate the signal transduction events elicited by hBD stimulation.

Results: We found that hBD3 induced the expression of cytokines and chemokines in keratinocytes, which was not observed in fibroblasts. hBD2, however, stimulated cell migration only in fibroblasts, which was not found for hBD3. Both defensins are likely to exert receptor-mediated effects in keratinocytes, as witnessed by changes in protein kinase activation following stimulation by hBD2 and hBD3. Kinome analysis suggested that protein kinase C activation was a common event for both defensins. We observed, however, considerable differences in keratinocyte responses between stimulation by exogenous recombinant defensins and endogenous defensins expressed following lentiviral transduction.

Conclusion: Defensins exert modest biological effects on skin cells that are potentially beneficial in wound healing, but many questions regarding the biological mechanisms of action and relevance for the in vivo situation are still remaining.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1159/000477346DOI Listing
July 2018

Psoriasis-Associated Late Cornified Envelope (LCE) Proteins Have Antibacterial Activity.

J Invest Dermatol 2017 11 17;137(11):2380-2388. Epub 2017 Jun 17.

Department of Dermatology, Radboud University Medical Center, Radboud Institute for Molecular Life Sciences, Nijmegen, The Netherlands. Electronic address:

Terminally differentiating epidermal keratinocytes express a large number of structural and antimicrobial proteins that are involved in the physical barrier function of the stratum corneum and provide innate cutaneous host defense. Late cornified envelope (LCE) genes, located in the epidermal differentiation complex on chromosome 1, encode a family of 18 proteins of unknown function, whose expression is largely restricted to epidermis. Deletion of two members, LCE3B and LCE3C (LCE3B/C-del), is a widely-replicated psoriasis risk factor that interacts with the major psoriasis-psoriasis risk gene HLA-C*06. Here we performed quantitative trait locus analysis, utilizing RNA-seq data from human skin and found that LCE3B/C-del was associated with a markedly increased expression of LCE3A, a gene directly adjacent to LCE3B/C-del. We confirmed these findings in a 3-dimensional skin model using primary keratinocytes from LCE3B/C-del genotyped donors. Functional analysis revealed that LCE3 proteins, and LCE3A in particular, have defensin-like antimicrobial activity against a variety of bacterial taxa at low micromolar concentrations. No genotype-dependent effect was observed for the inside-out or outside-in physical skin barrier function. Our findings identify an unknown biological function for LCE3 proteins and suggest a role in epidermal host defense and LCE3B/C-del-mediated psoriasis risk.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.jid.2017.06.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5651197PMC
November 2017

Cathepsin B as a potential cystatin M/E target in the mouse hair follicle.

FASEB J 2017 10 8;31(10):4286-4294. Epub 2017 Jun 8.

Department of Dermatology, Radboud Institute for Molecular Life Sciences, Radboud University Nijmegen Medical Center, Nijmegen, The Netherlands;

Deficiency of the cysteine protease inhibitor cystatin M/E (Cst6) in mice leads to disturbed epidermal cornification, impaired barrier function, and neonatal lethality. We report the rescue of the lethal skin phenotype of (Cst6-deficient; ) mice by transgenic, epidermis-specific, reexpression of Cst6 under control of the human involucrin (INV) promoter. Rescued Tg(INV-) mice survive the neonatal phase, but display severe eye pathology and alopecia after 4 mo. We observed keratitis and squamous metaplasia of the corneal epithelium, comparable to mice, as we have reported in other studies. We found the INV promoter to be active in the hair follicle infundibulum; however, we did not observe Cst6 protein expression in the lower regions of the hair follicle in Tg(INV-) mice. This result suggests that unrestricted activity of proteases is involved in disturbance of hair follicle biology, eventually leading to baldness. Using quenched activity-based probes, we identified mouse cathepsin B (CtsB), which is expressed in the lower regions of the hair follicle, as an additional target of mouse Cst6. These data suggest that Cst6 is necessary to control CtsB activity in hair follicle morphogenesis and highlight Cst6-controlled proteolytic pathways as targets for preventing hair loss.-Oortveld, M. A. W., van Vlijmen-Willems, I. M. J. J., Kersten, F. F. J., Cheng, T., Verdoes, M., van Erp, P. E. J., Verbeek, S., Reinheckel, T., Hendriks, W. J. A. J., Schalkwijk, J., Zeeuwen, P. L. J. M. Cathepsin B as a potential cystatin M/E target in the mouse hair follicle.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1096/fj.201700267RDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5602906PMC
October 2017

Reply to Meisel et al.

J Invest Dermatol 2017 04 22;137(4):961-962. Epub 2016 Nov 22.

NIZO Food Research B.V., Ede, The Netherlands.

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.jid.2016.11.013DOI Listing
April 2017

Epidermal equivalents of filaggrin null keratinocytes do not show impaired skin barrier function.

J Allergy Clin Immunol 2017 06 11;139(6):1979-1981.e13. Epub 2016 Oct 11.

Department of Dermatology, Radboud University Medical Center, Radboud Institute for Molecular Life Sciences, Nijmegen, The Netherlands. Electronic address:

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.jaci.2016.09.016DOI Listing
June 2017

An In vitro Model for Bacterial Growth on Human Stratum Corneum.

Acta Derm Venereol 2016 Nov;96(7):873-879

Department of Dermatology, Radboud university medical center, Nijmegen, The Netherlands.

The diversity and dynamics of the skin microbiome in health and disease have been studied recently, but adequate model systems to study skin microbiotas in vitro are largely lacking. We developed an in vitro system that mimics human stratum corneum, using human callus as substrate and nutrient source for bacterial growth. The growth of several commensal and pathogenic bacterial strains was measured for up to one week by counting colony-forming units or by quantitative PCR with strain-specific primers. Human skin pathogens were found to survive amidst a minimal microbiome consisting of 2 major skin commensals: Staphylococcus epidermidis and Propionibacterium acnes. In addition, complete microbiomes, taken from the backs of healthy volunteers, were inoculated and maintained using this system. This model may enable the modulation of skin microbiomes in vitro and allow testing of pathogens, biological agents and antibiotics in a medium-throughput format.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.2340/00015555-2401DOI Listing
November 2016

Perfusion Intensity Correlates with Expression Levels of Psoriasis-Related Genes and Proteins.

Skin Pharmacol Physiol 2015 12;28(6):296-306. Epub 2015 Sep 12.

Department of Dermatology, Radboudumc, Nijmegen, The Netherlands.

Background: Previous research revealed heterogeneity in the perfusion intensity within clinically homogenous-appearing plaques, without differences in erythema. In addition, an increased perfusion was found within the perilesional skin. This raises the question whether the heterogeneity in perfusion found both inside and outside a lesion influences the expression levels of genes and proteins involved in the pathogenesis of psoriasis.

Objectives: To correlate the perfusion intensity to mRNA and protein expression of genes associated with the pathogenesis of psoriasis and to visualize the dynamics of the perfusion intensity over time using laser Doppler perfusion imaging.

Methods: Fourteen patients with plaque psoriasis were included. The superficial microcirculation and clinical local scores (single usability metric, SUM, scores) were analysed in one representative lesion every 2 weeks. After 8 weeks 4 biopsies were taken, one from a highly perfused area (hotspot) and one from a low perfusion area (coldspot) of the lesional skin, one biopsy from the highly perfused perilesional skin and one from the distant uninvolved skin.

Results: Statistically significant differences in mRNA and protein expression, including IL-17 and TBX21/T-Bet, were found between hotspots and coldspots, and between the highly perfused perilesional and the uninvolved skin. Hotspots tend to remain on the same location during 8 weeks of follow-up.

Conclusions: Within homogenous-appearing psoriatic plaques, there are remarkable differences in mRNA and protein levels, which are correlated with the perfusion intensity and can be detected by using laser Doppler perfusion imaging. In addition, differences in mRNA and protein expression between the highly perfused perilesional skin and the uninvolved skin were found, indicating that several biological changes occur well before clinical changes become manifest.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1159/000381628DOI Listing
August 2016

Absent in Melanoma 2 is predominantly present in primary melanoma and primary squamous cell carcinoma, but largely absent in metastases of both tumors.

J Am Acad Dermatol 2014 Nov 15;71(5):1012-5. Epub 2014 Oct 15.

Department of Dermatology, Radboud University Medical Center, Nijmegen, The Netherlands; Radboud Institute for Molecular Life Sciences, Nijmegen, The Netherlands.

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.jaad.2014.06.012DOI Listing
November 2014

Myeloid lineage-restricted somatic mosaicism of NLRP3 mutations in patients with variant Schnitzler syndrome.

J Allergy Clin Immunol 2015 Feb 16;135(2):561-4. Epub 2014 Sep 16.

Department of Internal Medicine, Radboud University Medical Center, Nijmegen, The Netherlands; Radboud Institute for Molecular Life Sciences (RIMLS), Nijmegen, The Netherlands; Nijmegen Center for Immunodeficiency and Autoinflammation, Nijmegen, The Netherlands.

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.jaci.2014.07.050DOI Listing
February 2015

Analysis of protein-protein interaction between late cornified envelope proteins and corneodesmosin.

Exp Dermatol 2014 Oct;23(10):769-71

Department of Dermatology, Radboud University Medical Center, Nijmegen, The Netherlands.

Deletion of two members of the late cornified envelope (LCE) family, LCE3B and LCE3C (LCE3C_LCE3B-del), has been identified as risk factor for psoriasis with a possible role in skin barrier function. Moreover, genetic interaction between LCE3C_LCE3B-del and HLA-C*06, located in the psoriasis susceptibility regions 4 and 1 (PSORS4 and 1), has been reported in several populations. Because of high linkage disequilibrium between the PSORS1 genes HLA-C*06 and corneodesmosin (CDSN), both genes are potentially involved in psoriasis. As corneodesmosin and LCE proteins are both constituents of the stratum corneum, we investigated potential direct protein-protein interactions between six LCE proteins and two corneodesmosin sequence variants. Partial colocalization of LCE2 and CDSN was observed in normal and psoriasis skin using immunofluorescence microscopy. Co-expression of eCFP-LCE and mRFP-CDSN proteins in COS-1 cells and human adult keratinocytes, and GST pull-down results did not provide evidence for direct interactions between LCE proteins and CDSN variants.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1111/exd.12524DOI Listing
October 2014

Genotype-phenotype correlations in a prospective cohort study of paediatric plaque psoriasis: lack of correlation between HLA-C*06 and family history of psoriasis.

Acta Derm Venereol 2014 Nov;94(6):667-71

Department of Dermatology, Radboud University Nijmegen Medical Centre, PO Box 9101, 6500 HB Nijmegen, The Netherlands.

This study aims to investigate associations between observed clinical parameters and known genetic risk factors of psoriasis in a well-defined prospective cohort of paediatric patients with plaque psoriasis (n = 151). Significant associations were found for paediatric-onset psoriasis with ERAP1 (p = 0.002), IL23R (p = 0.01), LCE3C_LCE3B-del (p = 0.00049) and HLA-C*06 (p = 3.15 × 10(-30)). Psoriasis severity was associated with the single nucleotide polymorphisms tagging IFIH1 and ERAP1 (p < 0.05). An onset before 10 years of age was associated with IL12B (p = 0.02). Nail psoriasis was more often seen in HLA-C*06-negative patients (p = 0.008). Remarkably, family history is clearly not associated with HLA-C*06 in this specific group. The large proportion of patients with a positive family history in HLA-C*06 negative patients (and the lack of correlation between the two) indicates that other genes, either alone or interaction between two or more genes, may have significant effects on heritability.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.2340/00015555-1810DOI Listing
November 2014

Microbiome and skin diseases.

Curr Opin Allergy Clin Immunol 2013 Oct;13(5):514-20

Department of Dermatology, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands.

Purpose Of Review: This article reviews recent findings on the skin microbiome. It provides an update on the current understanding of the role of microbiota in healthy skin and in inflammatory and allergic skin diseases.

Recent Findings: Advances in computing and high-throughput sequencing technology have enabled in-depth analysis of microbiota composition and functionality of human skin. Most data generated to date are related to the skin microbiome of healthy volunteers, but recent studies have also addressed the dynamics of the microbiome in diseased and injured skin. Currently, reports are emerging that evaluate the strategies to manipulate the skin microbiome, intending to modulate diseases and/or their symptoms.

Summary: The microbiome of normal human skin was found to have a high diversity and high interpersonal variation. Microbiota compositions of diseased lesional skin (in atopic dermatitis and psoriasis) showed distinct differences compared with healthy skin. The function of microbial colonization in establishing immune system homeostasis has been reported, whereas host-microbe interactions and genetically determined variation of stratum corneum properties might be linked to skin dysbiosis. Both are relevant for cutaneous disorders with aberrant immune responses and/or disturbed skin barrier function. Modulation of skin microbiota composition to restore host-microbiota homeostasis could be future strategies to treat or prevent disease.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1097/ACI.0b013e328364ebebDOI Listing
October 2013

Combination of pantothenamides with vanin inhibitors as a novel antibiotic strategy against gram-positive bacteria.

Antimicrob Agents Chemother 2013 Oct 22;57(10):4794-800. Epub 2013 Jul 22.

Department of Dermatology and Nijmegen Centre for Molecular Life Sciences, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands.

The emergence of resistance against current antibiotics calls for the development of new compounds to treat infectious diseases. Synthetic pantothenamides are pantothenate analogs that possess broad-spectrum antibacterial activity in vitro in minimal media. Pantothenamides were shown to be substrates of the bacterial coenzyme A (CoA) biosynthetic pathway, causing cellular CoA depletion and interference with fatty acid synthesis. In spite of their potential use and selectivity for bacterial metabolic routes, these compounds have never made it to the clinic. In the present study, we show that pantothenamides are not active as antibiotics in the presence of serum, and we found that they were hydrolyzed by ubiquitous pantetheinases of the vanin family. To address this further, we synthesized a series of pantetheinase inhibitors based on a pantothenate scaffold that inhibited serum pantetheinase activity in the nanomolar range. Mass spectrometric analysis showed that addition of these pantetheinase inhibitors prevented hydrolysis of pantothenamides by serum. We found that combinations of these novel pantetheinase inhibitors and prototypic pantothenamides like N5-Pan and N7-Pan exerted antimicrobial activity in vitro, particularly against Gram-positive bacteria (Staphylococcus aureus, Staphylococcus epidermidis, Streptococcus pneumoniae, and Streptococcus pyogenes) even in the presence of serum. These results indicate that pantothenamides, when protected against degradation by host pantetheinases, are potentially useful antimicrobial agents.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/AAC.00603-13DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3811400PMC
October 2013

Skin microbiome imbalance in patients with STAT1/STAT3 defects impairs innate host defense responses.

J Innate Immun 2014 22;6(3):253-62. Epub 2013 Jun 22.

Department of Medicine, Radboud University Nijmegen Medical Center, Nijmegen, The Netherlands.

Background: Chronic mucocutaneous candidiasis (CMC) and hyper-IgE syndrome (HIES) are primary immunodeficiencies mainly caused by mutations in STAT1 and STAT3, respectively. CMC and HIES patients have an increased risk for skin and mucosal infections with fungal pathogens and Staphylococcus aureus. However, it is unknown whether the genetic defects in these patients also affect the skin and mucosal microbiome, which in turn may influence host defense mechanisms.

Methods: The skin and oral microbiome of CMC and HIES patients was compared to that of healthy controls at five body sites using 16S rRNA sequencing. The influence of skin colonizers on the immune response was investigated using in vitro experiments.

Results: The microbiome of CMC and HIES patients contained more Gram-negative bacteria, especially Acinetobacter spp., and less of the normal Corynebacterium spp. compared to healthy controls. Exposure of human primary leukocytes to Acinetobacter suppressed the cytokine response to Candida albicans and S. aureus, while the normal corynebacteria did not suppress cytokine responses.

Discussion: These results demonstrate that central mediators of immune responses like STAT1 and STAT3 not only directly influence immune responses, but also result in changes in the skin microbiome that in turn can amplify the defective immune response against fungal and microbial pathogens.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1159/000351912DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4045018PMC
May 2015

Coal tar induces AHR-dependent skin barrier repair in atopic dermatitis.

J Clin Invest 2013 Feb 25;123(2):917-27. Epub 2013 Jan 25.

Department of Dermatology, Radboud University Nijmegen Medical Centre, Nijmegen Centre for Molecular Life Sciences, P.O. Box 9101, Nijmegen 6500 HB, The Netherlands.

Topical application of coal tar is one of the oldest therapies for atopic dermatitis (AD), a T helper 2 (Th2) lymphocyte-mediated skin disease associated with loss-of-function mutations in the skin barrier gene, filaggrin (FLG). Despite its longstanding clinical use and efficacy, the molecular mechanism of coal tar therapy is unknown. Using organotypic skin models with primary keratinocytes from AD patients and controls, we found that coal tar activated the aryl hydrocarbon receptor (AHR), resulting in induction of epidermal differentiation. AHR knockdown by siRNA completely abrogated this effect. Coal tar restored filaggrin expression in FLG-haploinsufficient keratinocytes to wild-type levels, and counteracted Th2 cytokine-mediated downregulation of skin barrier proteins. In AD patients, coal tar completely restored expression of major skin barrier proteins, including filaggrin. Using organotypic skin models stimulated with Th2 cytokines IL-4 and IL-13, we found coal tar to diminish spongiosis, apoptosis, and CCL26 expression, all AD hallmarks. Coal tar interfered with Th2 cytokine signaling via dephosphorylation of STAT6, most likely due to AHR-regulated activation of the NRF2 antioxidative stress pathway. The therapeutic effect of AHR activation herein described opens a new avenue to reconsider AHR as a pharmacological target and could lead to the development of mechanism-based drugs for AD.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1172/JCI65642DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3561798PMC
February 2013

Discovery of small molecule vanin inhibitors: new tools to study metabolism and disease.

ACS Chem Biol 2013 Mar 2;8(3):530-4. Epub 2013 Jan 2.

Nijmegen Centre for Molecular Life Sciences, Radboud University Nijmegen Medical Centre, The Netherlands.

Vanins are enzymes with pantetheinase activity and are presumed to play a role in the recycling of pantothenic acid (vitamin B5) from pantetheine. Pantothenic acid is an essential nutrient required to synthesize coenzyme A, a cofactor involved in many biological processes such as fatty acid synthesis and oxidation of pyruvate to fuel the citric acid cycle. Hydrolysis of pantetheine also liberates cysteamine, a known antioxidant. Vanin-1 is highly expressed in liver and is under transcriptional control of PPAR-α and nutritional status, suggesting a role in energy metabolism. The lack of potent and specific inhibitors of vanins has hampered detailed investigation of their function. We hereby report the design, synthesis, and characterization of a novel pantetheine analogue, RR6, that acts as a selective, reversible, and competitive vanin inhibitor at nanomolar concentration. Oral administration of RR6 in rats completely inhibited plasma vanin activity and caused alterations of plasma lipid concentrations upon fasting, thereby illustrating its potential use in chemical biology research.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/cb3006424DOI Listing
March 2013

Strong induction of AIM2 expression in human epidermis in acute and chronic inflammatory skin conditions.

Exp Dermatol 2012 Dec;21(12):961-4

Absent in melanoma 2 (AIM2) is a double-stranded DNA receptor, and its activation initiates an interleukin-1 beta processing inflammasome. AIM2 is implicated in host defense against several pathogens, but could hypothetically also contribute to autoinflammatory or autoimmune diseases, such as is the case for NLRP3. Using thoroughly characterised antibodies, we analysed AIM2 expression in human tissues and primary cells. A strong epidermal upregulation of AIM2 protein expression was observed in several acute and chronic inflammatory skin disorders, such as psoriasis, atopic dermatitis, venous ulcera, contact dermatitis, and experimental wounds. We also found AIM2 induction by interferon-gamma in submerged and three-dimensional in vitro models of human epidermis. Our data highlight the dynamics of epidermal AIM2 expression, showing Langerhans cell and melanocyte-restricted expression in normal epidermis but a pronounced induction in subpopulations of epidermal keratinocytes under inflammatory conditions.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1111/exd.12037DOI Listing
December 2012