Functional and proteomic analysis of a full thickness filaggrin-deficient skin organoid model

Wellcome Open Research

article

Martina S. Elias¹  Sheila C. Wright¹  William V. Nicholson¹  Kimberley D. Morrison¹  Alan R. Prescott²  Sara Ten Have³  Phillip D. Whitfield⁴  Angus I. Lamond³  Sara J. Brown¹ 

[1] Skin Research Group, Division of Molecular and Clinical Medicine, School of Medicine, University of Dundee;Dundee Imaging Facility, School of Life Sciences, University of Dundee;Centre for Gene Regulation and Expression, School of Life Sciences, University of Dundee;Lipidomics Research Facility, Division of Biomedical Sciences, University of the Highlands and Islands;Department of Dermatology, Ninewells Hospital

关键词: Axon guidance;    atopic dermatitis;    eczema;    filaggrin;    gene ontology;    keratinocyte-immune crosstalk;    organoid;    proteomics;   

DOI  :  10.12688/wellcomeopenres.15405.2

学科分类：内科医学

来源: Wellcome

【 摘 要 】

Background: Atopic eczema is an itchy inflammatory disorder characterised by skin barrier dysfunction. Loss-of-function mutations in the gene encoding filaggrin (FLG) are a major risk factor, but the mechanisms by which filaggrin haploinsufficiency leads to atopic inflammation remain incompletely understood. Skin as an organ that can be modelled using primary cellsin vitro provides the opportunity for selected genetic effects to be investigated in detail.Methods: Primary human keratinocytes and donor-matched primary fibroblasts from healthy individuals were used to create skin organoid models with and without siRNA-mediated knockdown ofFLG. Biological replicate sets of organoids were assessed using histological, functional and biochemical measurements.Results:FLG knockdown leads to subtle changes in histology and ultrastructure including a reduction in thickness of the stratum corneum and smaller, less numerous keratohyalin granules. Immature organoids showed some limited evidence of barrier impairment withFLG 8000 proteins per sample. Gene ontology and pathway analyses identified an increase in transcriptional and translational activity but a reduction in proteins contributing to terminal differentiation, including caspase 14, dermokine, AKT1 and TGF-beta-1. Aspects of innate and adaptive immunity were represented in both the up-regulated and down-regulated protein groups, as was the term ‘axon guidance’.     Conclusions: This work provides further evidence for keratinocyte-specific mechanisms contributing to immune and neurological, as well as structural, aspects of skin barrier dysfunction. Individuals with filaggrin deficiency may derive benefit from future therapies targeting keratinocyte-immune crosstalk and neurogenic pruritus.

【 授权许可】

CC BY