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Identification of novel gene products involved in epidermal keratinocyte differentiation and pathologic skin barrier repair
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences. (Dermatology and Venerology)ORCID iD: 0000-0001-9280-9832
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences. (Dermatology and Venerology)
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences. (Dermatology and Venerology)
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences. (Dermatology and Venerology)
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(English)Manuscript (preprint) (Other academic)
Abstract [en]

A functional skin barrier relies on a correct structural organization of the outermost epidermal layer, stratum corneum, as the final stage of keratinocyte terminal differentiation. However, the factors involved in keratinocyte differentiation, especially those which are important for the skin barrier function and barrier repair, are still largely unknown. In this study we re-analysed two sets of microarrays obtained from cultured differentiated keratinocytes or skin biopsies of autosomal recessive congenital ichthyosis (ARCI) patients with hyperkeratosis and defect skin barrier due to TGM1 mutations. When comparing these arrays to those of proliferating keratinocytes and healthy control epidermis respectively, we identified 24 augmented genes not previously associated with keratinocyte differentiation and skin barrier repair. Increased expression of 9 of these genes, i.e. AKR1B10, BLNK, ENDOU, GCNT4, GLTP, RHCG, SLC15A1, TMEM86A, VSNL1, was verified by qPCR of differentiated keratinocytes and patients skin extracts. In a separate experiment, the regulation of the genes by two well-known transcription factors, retinoic acid receptors (RARs) and peroxisome proliferator activating receptors (PPARs), was studied in cell cultures by adding the pan-RAR agonist all-trans retinoic acid (atRA) or PPARd agonist GW501516. In line with the anti-keratinizing effects of retinoids, atRA reduced the mRNA expressions of some of these genes both in monolayer and organotypic keratinocyte cultures. Confirming the importance of PPARd for keratinocyte differentiation, PPARd-agonist GW501516 induced all 9 novel genes associated with differentiation and skin barrier repair. The results increase the understanding about genes involved in human epidermal keratinocyte differentiation, with potential repercussions on the development of new ways to restore the barrier function in conditions with a defect skin barrier.

National Category
Dermatology and Venereal Diseases
Research subject
Dermatology and Venerology
Identifiers
URN: urn:nbn:se:uu:diva-377555OAI: oai:DiVA.org:uu-377555DiVA, id: diva2:1290860
Funder
Swedish Research Council, K2013-57X-22309-3Available from: 2019-02-21 Created: 2019-02-21 Last updated: 2019-02-21
In thesis
1. Congenital Recessive Ichthyosis: Studies of Gene Expressions Related to Keratinocyte Differentiation and Skin Barrier Repair
Open this publication in new window or tab >>Congenital Recessive Ichthyosis: Studies of Gene Expressions Related to Keratinocyte Differentiation and Skin Barrier Repair
2019 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Autosomal recessive congenital ichthyosis (ARCI) is a rare monogenetic disorder characterized by a defective skin barrier, hyperkeratosis, and dry, scaly skin. It affects keratinocyte differentiation and is caused by mutations in any of at least 12 genes believed to control the formation of ω-O-acylceramide and the corneocyte lipid envelope (CLE): ABCA12, ALOXE3, ALOX12B, CERS3, CYP4F22, ELOVL4, LIPN, NIPAL4, PNPLA1, SDR9C7, SLC27A4, and TGM1.

Studies of keratinocyte differentiation and gene expression in ARCI may help us better understand the pathobiology of skin barrier formation. One way to verify that ARCI-related gene products are operating in a chain of events essential for lipid barrier formation is to use immunofluorescence and in situ proximity ligation assays to demonstrate the proteins’ colocalizations in the epidermis. In paper I, a new method for the objective quantitative image analysis of protein expression and colocalization in different epidermal layers of skin sections was developed using free, open-source software, CellProfiler. Using this method and microarray analyses of skin biopsies from ARCI patients with TGM1 mutations (n = 5) compared with those of healthy controls (n = 4), many ARCI-related genes were found to be upregulated in patient epidermis (paper II). Because many other genes involved in keratinocyte differentiation and immune/inflammatory response, including PPARD, were also induced in the patients’ microarrays, the effects of a ligand-dependent transcription factor, PPARδ, encoded by PPARD, were studied on ARCI-related gene expression in cultured keratinocytes, usually showing the pronounced upregulation by PPARδ agonists (paper III). Furthermore, using previous array data obtained from cultured differentiated keratinocytes and from skin biopsies of patients with TGM1 mutations, nine novel candidate markers of differentiation were identified, and the upregulation was verified by qPCR of mRNA from cultured keratinocytes and skin biopsies. These transcripts were also induced by PPARδ agonists in cultured proliferating keratinocytes (paper IV).

To conclude, the upregulation of other ARCI-related genes in patients with TGM1 mutations might reflect a feedback mechanism in ω-O-acylceramide biosynthesis, which, however, is unable to restore the patients’ skin barrier. In theory, substitution therapy with ω-O-acylceramide and recombinant TGm-1 may be required. Because PPARδ activation appears involved in upregulating ARCI-related genes and nine novel differentiation marker genes, all potentially important for barrier repair, this approach could become a treatment option for several types of ichthyosis and wound healing.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2019. p. 58
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 1541
Keywords
genodermatoses, oligoarray, trancriptomics, transglutaminase-1, cornified envelope, peroxisome proliferator-activated receptor δ, all-trans retinoic acids.
National Category
Dermatology and Venereal Diseases
Research subject
Dermatology and Venerology
Identifiers
urn:nbn:se:uu:diva-377557 (URN)978-91-513-0578-3 (ISBN)
Public defence
2019-04-11, Fåhraeussalen, Rudbeck Laboratory, C5, entréplan, Uppsala, 13:15 (English)
Opponent
Supervisors
Available from: 2019-03-20 Created: 2019-02-21 Last updated: 2019-05-07

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