A mouse organotypic tissue culture model for autosomal recessive congenital ichthyosis

Rosenberger, Sabine; Dick, Angela; Latzko, Susanne; Haußer, Ingrid; Stark, Hans Jürgen; Rauh, Manfred; Schneider, Holm; Krieg, Peter

doi:10.1111/bjd.13308

Cited by 9 publications

(9 citation statements)

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“…After isolation of both keratinocytes and fibroblasts from skin biopsies of individuals P5 and P6, cells (up to four passages) were seeded at final concentrations of 1 3 10 5 fibroblasts and 0.7 3 10 6 keratinocytes and cocultured for up to 14 days, as previously described. 26,27 To achieve equivalents that are as similar as possible to in vivo skin of diseased persons, the keratinocytes were cocultured on a dermal layer consisting of the fibroblasts from the affected individual and collagen Type I isolated from tendons of rat tails, serving as a scaffold for the epidermis. After 7 and 14 days of exposition to air (lifting), the models were harvested and prepared for downstream functional analyses.…”

Section: D Organotypic Tissue Culture Modelmentioning

confidence: 99%

“…Paraffin sections of biopsies were prepared as outlined above. 8 mm cryo sections of organotypic models were fixed according to Rosenberger et al 27 Double immunofluorescence staining was performed as described in Radner et al with the following minor modifications. 10 Antigen retrieval of paraffin sections was performed in a pressure cooker at pH 9.0 (Tris EDTA buffer).…”

Section: Immunofluorescence Analysismentioning

confidence: 99%

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Mutations in SULT2B1 Cause Autosomal-Recessive Congenital Ichthyosis in Humans

Heinz

Kim

Marrakchi

et al. 2017

The American Journal of Human Genetics

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Ichthyoses are a clinically and genetically heterogeneous group of genodermatoses associated with abnormal scaling of the skin over the whole body. Mutations in nine genes are known to cause non-syndromic forms of autosomal-recessive congenital ichthyosis (ARCI). However, not all genetic causes for ARCI have been discovered to date. Using whole-exome sequencing (WES) and multigene panel screening, we identified 6 ARCI-affected individuals from three unrelated families with mutations in Sulfotransferase family 2B member 1 (SULT2B1), showing their causative association with ARCI. Cytosolic sulfotransferases form a large family of enzymes that are involved in the synthesis and metabolism of several steroids in humans. We identified four distinct mutations including missense, nonsense, and splice site mutations. We demonstrated the loss of SULT2B1 expression at RNA and protein levels in keratinocytes from individuals with ARCI by functional analyses. Furthermore, we succeeded in reconstructing the morphologic skin alterations in a 3D organotypic tissue culture model with SULT2B1-deficient keratinocytes and fibroblasts. By thin layer chromatography (TLC) of extracts from these organotypic cultures, we could show the absence of cholesterol sulfate, the metabolite of SULT2B1, and an increased level of cholesterol, indicating a disturbed cholesterol metabolism of the skin upon loss-of-function mutation in SULT2B1. In conclusion, our study reveals an essential role for SULT2B1 in the proper development of healthy human skin. Mutation in SULT2B1 leads to an ARCI phenotype via increased proliferation of human keratinocytes, thickening of epithelial layers, and altered epidermal cholesterol metabolism.

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Section: D Organotypic Tissue Culture Modelmentioning

confidence: 99%

Section: Immunofluorescence Analysismentioning

confidence: 99%

Mutations in SULT2B1 Cause Autosomal-Recessive Congenital Ichthyosis in Humans

Heinz

Kim

Marrakchi

et al. 2017

The American Journal of Human Genetics

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“…The conjunction of cornified envelope proteins and sphingolipids or fatty acids forms the corneocyte lipid envelope, which is essential for a properly functioning epidermal barrier (Fig. a) …”

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confidence: 99%

Diminished protein-bound ω-hydroxylated ceramides in the skin of patients with ichthyosis with 12R-lipoxygenase (LOX) or eLOX-3 deficiency

et al. 2017

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DEAR EDITOR, Autosomal recessive congenital ichthyoses (ARCIs) are a group of hereditary skin disorders. The disease phenotype is associated with an impaired epidermal barrier leading to increased transepidermal water loss, temperature instability and hypernatraemic dehydration in infancy. 1 ARCIs are known to be caused by mutations in at least nine different genes, which encode proteins involved in the formation of the epidermal barrier. 2 Two of these genes, ALOX12B and ALOXE3, code for the epidermal lipoxygenases 12R-LOX and eLOX-3, which are known to be indispensable for barrier function and lipid metabolism in the skin. 3 Both eLOX-3-and 12R-LOX-deficient mice die shortly after birth due to transepidermal water loss, and show dramatically

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“…A generally accepted alternative approach is to use cells from a healthy control and mimic the patient's disease with gene knock-down. 8 Rosenberger et al, 9 from the German Cancer Research Centre in Heidelberg present, in this issue, a completely new approach for the generation of a 3D skin model for congenital ichthyosis. The team used freshly isolated neonatal keratinocytes and fibroblasts from the Alox12b À/À knockout mouse to establish 3D, full-thickness organotypic models to mimic ARCI.…”

Section: Original Article P 1347mentioning

confidence: 99%

“…These results are very similar to those seen in the knockout mouse and are comparable with the typical features of ALOX12B-deficient patients with ARCI. Unexpectedly, using a dye penetration assay, Rosenberger et al 9 saw huge differences in the epidermal barrier function of the murine models compared with patient skin and human 3D skin models. These differences might have been caused by the nature of the keratinocytes used, which are, in contrast to all thus far known human systems, neonatal and proliferate extensively, resulting in an extremely pronounced stratum corneum with marked hyperkeratosis.…”

Section: Original Article P 1347mentioning

confidence: 99%

Update: advanced methods in three-dimensional organotypic tissue engineering for congenital ichthyosis and other rare keratinization disorders

Eckl

2014

Br J Dermatol

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The biology of melanoma is challenging and the exact mechanisms by which ultraviolet (UV) radiation affects the malignant transformation of melanocytes to melanoma are not well defined. UV radiation has been implicated as the main aetiological factor in the development of all types of skin cancer, including melanoma located at sun-exposed sites. 1 Specifically, intermittent sun exposure seems to be the most important risk factor for melanoma. 2 Therefore, research efforts have placed an emphasis on understanding how solar UV wavelengths, mainly UVA and UVB, alter specific protein targets and the corresponding biological mechanisms in the context of melanoma development and/or progression. In this issue, W€ aster et al. 3 demonstrate that activator protein-1 induces proapoptotic signaling and that nuclear factor kappa B is a key antiapoptotic/ prosurvival factor following both UVA-and UVB-induced cellular damage in human melanocytes. In addition, in an attempt to reconcile some of the discrepancies reported from observations following irradiations with a single spectral fraction, W€ aster et al. 3 observed significant increases in phosphorylated extracellular regulated kinase and c-Jun N-terminal kinase after melanocytes were exposed either to UVA or UVB radiation.This work contributes to an expanding field of research demonstrating that p53, p21, p16 and retinoblastoma protein in UV-irradiated human melanocytes are important in the maintenance of genomic stability and prevention of malignant transformation. 4,5 Most recently, in vivo studies by Viros et al. 6 have shown that exposure to UV radiation causes accelerated melanomagenesis in mice expressing BRAF(V600E), the most common mutation in melanoma, through an increase in mutations of the tumor suppressor p53. These studies have provided insight into the molecular mechanisms of melanoma development and transformation. In addition, these studies have shown the importance of understanding the phenotype and genetic features of the melanocytes selected, as these cells will react differently from other cells to the stimuli utilized.One key aspect of understanding the biological functions of epidermal melanocytes is the central role that these cells play in determining the responses of the skin to UV exposure. Increased melanogenesis and pigment darkening is evident after UV exposure and constitutes an important defence mechanism in guarding against the damaging effects of chronic UV exposure. 7 As important as melanogenesis is in protecting the skin from UV radiation, this process can only occur with the assistance of factors secreted by neighbouring dermal fibroblasts and epidermal keratinocytes. 8 As we continue to understand the biological aspects of each epidermal cell type, specifically during gene-environment interactions, it will shed light on how the transformation of epidermal cells to cancer occurs. Given the difficulty in curing melanoma, owing to its heterogeneity and plasticity, ultimately resulting in chemoresistance, there is great emphasis on melano...

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A mouse organotypic tissue culture model for autosomal recessive congenital ichthyosis

Cited by 9 publications

References 42 publications

Mutations in SULT2B1 Cause Autosomal-Recessive Congenital Ichthyosis in Humans

Mutations in SULT2B1 Cause Autosomal-Recessive Congenital Ichthyosis in Humans

Diminished protein-bound ω-hydroxylated ceramides in the skin of patients with ichthyosis with 12R-lipoxygenase (LOX) or eLOX-3 deficiency

Update: advanced methods in three-dimensional organotypic tissue engineering for congenital ichthyosis and other rare keratinization disorders

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