Summary
The main proteinaceous component of the keratohyalin granules within the granular layer keratinocytes of the epidermis is the giant, repetitive polyprotein profilaggrin. When granular layer ...cells commit to terminal differentiation to form the flattened squames of the stratum corneum, profilaggrin is rapidly cleaved into multiple copies of the 37 kDa filaggrin monomer, which binds to and condenses the keratin cytoskeleton, thereby facilitating cellular compression. Within the stratum corneum, filaggrin is broken down to form natural moisturising factor, a pool of amino acids and derivatives thereof that exerts multiple effects. Filaggrin is therefore essential for normal stratum corneum biogenesis and physiology. In 2006, the McLean group identified the first loss‐of‐function mutations in the filaggrin gene (FLG) as the cause of the common monogenic genodermatosis ichthyosis vulgaris (IV). In parallel, they showed by multiple methods that these mutations, carried by up to 10% of various human populations are the major genetic predisposing factor for atopic dermatitis (eczema) and all of the associated allergic phenotypes that constitute the atopic diathesis. This paradigm‐shifting work showed that skin barrier deficiency is a major early event in the pathophysiology of eczema and allergy.
What is already known?
Filaggrin is a critically important, multifunctional protein required for the normal biogenesis and physiology of the stratum corneum.
What does (recent) work tell us?
Loss‐of‐function mutations in the filaggrin gene are common in the human population and cause clinically dry skin, ichthyosis vulgaris and importantly, are a major genetic predisposing factor for atopic dermatitis and its associated allergic conditions. This work revealed that skin barrier deficiency is the major pathomechanism underlying atopic disease.
CRISPR/Cas9-based therapeutics hold the possibility for permanent treatment of genetic disease. The potency and specificity of this system has been used to target dominantly inherited conditions ...caused by heterozygous missense mutations through inclusion of the mutated base in the short-guide RNA (sgRNA) sequence. This research evaluates a novel approach for targeting heterozygous single-nucleotide polymorphisms (SNPs) using CRISPR/Cas9. We determined that a mutation within KRT12, which causes Meesmann's epithelial corneal dystrophy (MECD), leads to the occurrence of a novel protospacer adjacent motif (PAM). We designed an sgRNA complementary to the sequence adjacent to this SNP-derived PAM and evaluated its potency and allele specificity both in vitro and in vivo. This sgRNA was found to be highly effective at reducing the expression of mutant KRT12 mRNA and protein in vitro. To assess its activity in vivo we injected a combined Cas9/sgRNA expression construct into the corneal stroma of a humanized MECD mouse model. Sequence analysis of corneal genomic DNA revealed non-homologous end-joining repair resulting in frame-shifting deletions within the mutant KRT12 allele. This study is the first to demonstrate in vivo gene editing of a heterozygous disease-causing SNP that results in a novel PAM, further highlighting the potential for CRISPR/Cas9-based therapeutics.
Summary
Background Filaggrin loss‐of‐function (FLG) mutations are associated with eczema and skin barrier impairment, but it is unclear whether skin barrier impairment precedes phenotypic eczema in ...FLG mutation carriers.
Objectives To study the association between FLG mutations, skin barrier impairment and clinical eczema at 3 months of age.
Methods A total of 88 infants were examined for eczema. Disease severity was determined by the SCORAD eczema severity score. Transepidermal water loss (TEWL) was measured on unaffected forearm skin. Venous blood samples were screened for the four most common FLG mutations found in the U.K. white population (R501X, 2282del4, R2447X and S3247X). Median SCORAD and TEWL measurements in children with and without eczema and FLG mutations were compared.
Results Thirty‐three per cent (29/88) of children had clinical eczema. Median SCORAD was 10·6 (range 3·5–31·0). TEWL (g m−2 h−1) was higher in children with eczema compared with unaffected infants (median TEWL 14·24 vs. 11·24, P < 0·001). Higher TEWL was associated with more severe disease (r = 0·59, P < 0·001, median TEWL, SCORAD < 15, 13·1 vs. 29·6, SCORAD ≥ 15, P = 0·029). Clinically dry skin was associated with higher TEWL, even in the absence of eczema (median TEWL 17·55 vs. 11·08, P = 0·008). Seventeen per cent (15/88) of children carried at least one FLG mutation. FLG mutation carriers were significantly more likely to have clinically dry skin, even in the absence of eczema odds ratio (OR) 8·50, 95% confidence interval (CI) 1·09–66·58, P = 0·042. FLG mutation carriers were also more likely to have eczema by 3 months of age (OR 4·26, 95% CI 1·34–13·57, P = 0·014). FLG mutations were significantly associated with higher median TEWL (all children, FLG‘yes’ 21·59 vs. FLG‘no’ 11·24, P < 0·001), even without clinical eczema (FLG‘yes’ 15·99 vs. FLG‘no’ 10·82, P = 0·01).
Conclusions By the age of 3 months, FLG mutations are associated with an eczema phenotype, dry skin and TEWL. The observation that TEWL is elevated in unaffected FLG mutation carriers suggests that skin barrier impairment precedes clinical eczema.
Summary
Background
Pachyonychia congenita (PC) is a rare autosomal dominant keratinizing disorder characterized by severe, painful, palmoplantar keratoderma and nail dystrophy, often accompanied by ...oral leucokeratosis, cysts and follicular keratosis. It is caused by mutations in one of five keratin genes: KRT6A, KRT6B, KRT6C, KRT16 or KRT17.
Objectives
To identify mutations in 84 new families with a clinical diagnosis of PC, recruited by the International Pachyonychia Congenita Research Registry during the last few years.
Methods
Genomic DNA isolated from saliva or peripheral blood leucocytes was amplified using primers specific for the PC‐associated keratin genes and polymerase chain reaction products were directly sequenced.
Results
Mutations were identified in 84 families in the PC‐associated keratin genes, comprising 46 distinct keratin mutations. Fourteen were previously unreported mutations, bringing the total number of different keratin mutations associated with PC to 105.
Conclusions
By identifying mutations in KRT6A, KRT6B, KRT6C, KRT16 or KRT17, this study has confirmed, at the molecular level, the clinical diagnosis of PC in these families.
What's already known about this topic?
Pachyonychia congenita (PC) is caused by autosomal dominant mutations in KRT6A, KRT6B, KRT6C, KRT16 or KRT17.
Plantar pain is the main symptom.
Palmoplantar keratoderma and nail dystrophy are the predominant characteristics, often accompanied by oral leucokeratosis, cysts and follicular keratosis.
What does this study add?
This study identifies PC‐associated keratin mutations in 84 new families with PC recruited by the International Pachyonychia Congenita Research Registry.
Fourteen of the 46 distinct keratin mutations were previously unreported.
This study expands the large well‐phenotyped and genotyped case series of patients with PC, which is an invaluable resource for the development of mutation‐specific and/or gene‐specific therapies and for future clinical trials.
Summary
Background Atopic dermatitis (AD) and loss‐of‐function mutations in the filaggrin gene (FLG) are both associated with chronic irritant contact dermatitis (ICD). As FLG mutations also are a ...major risk factor for AD, it is not clear whether FLG mutations are an independent risk factor for ICD or whether the risk is mediated by AD.
Objectives To investigate the relative contribution and interaction of FLG mutations and AD in German patients with occupational ICD and controls (vocational school apprentices).
Methods A total of 634 patients and 393 controls were genotyped for R501X, 2282del4, R2447X and S3247X. Current or past flexural eczema was used as an indicator of AD.
Results FLG mutations were found in 15·9% of the patients with ICD and 8·3% of the controls, with a crude odds ratio (OR) of 2·09 95% confidence interval (CI) 1·33–3·28 for the combined genotype. The adjusted OR for FLG mutations, corrected for AD, was 1·62 (95% CI 1·01–2·58). Subjects with AD were at approximately three times higher risk of developing ICD than controls (OR 2·89; 95% CI 2·09–3·99). There was no evidence of an interaction between these two risk factors.
Conclusions Our results indicate that both FLG mutations and AD increase the risk of ICD. Individuals with concurrent FLG mutations and AD are at the highest risk of developing ICD.
What’s already known about this topic?
•Filaggrin gene (FLG) loss‐of‐function mutations increase the risk of developing atopic dermatitis (AD).
It has been reported that FLG mutations are associated with irritant contact dermatitis (ICD); however, it is unclear whether they are an independent risk factor or work through AD.
What does this study add?
•FLG mutations are associated with ICD even when adjusted for AD.
• Individuals with concurrent FLG mutations odds ratio (OR) 1·61 and AD (OR 2·89) are at the highest risk of developing ICD (combined OR 4·7).
See also the Commentary by Thys sen
Summary
Background Null mutations in the filaggrin gene (FLG) cause ichthyosis vulgaris (IV) and predispose to atopic dermatitis (AD). Cohort studies in Europe and Japan have reported an FLG ...mutation carrier frequency of between 14% and 56%, but the prevalent European FLG mutations are rare or absent in Chinese patients with IV and AD.
Objectives To investigate further the spectrum of FLG‐null mutations in Chinese patients and to compare it with that in other populations.
Methods We conducted comprehensive FLG genetic analysis in a discovery cohort of 92 Singaporean Chinese individuals with IV and/or moderate‐to‐severe AD. All detected FLG mutations were then screened in a cohort of 425 patients with AD and 440 normal controls.
Results In total, 22 FLG‐null mutations, of which 14 are novel, were identified in this study; the combined null FLG genotype of 17 mutations detected in cases and controls showed strong association with AD Fisher’s exact test; P = 5·3 × 10−9; odds ratio (OR) 3·3, palmar hyperlinearity (Fisher’s exact test; P = 9·0 × 10−15; OR 5·8), keratosis pilaris (Fisher’s exact test; P = 0·001; OR 4·7) and with increased severity of AD (permutation test; P = 0·0063).
Conclusions This study emphasizes the wider genetic landscape of FLG‐null mutations in Asia that is slowly emerging.
Summary
Background
Filaggrin is central to the pathogenesis of atopic dermatitis (AD). The cheeks are a common initiation site of infantile AD. Regional and temporal expression of levels of filaggrin ...degradation products natural moisturizing factors (NMFs), activities of filaggrin‐processing enzymes bleomycin hydrolase (BH) and calpain‐1 (C‐1) and plasmin, and corneocyte envelope (CE) maturity in early life are largely unknown.
Objectives
We conducted a cross‐sectional, observational study investigating regional and age‐dependent variations in NMF levels, activity of proteases and CE maturity in stratum corneum (SC) from infants to determine whether these factors could explain the observed predilection sites for AD in early life.
Methods
We measured NMF using a tape‐stripping method at seven sites in the SC of 129 children (aged < 12 months to 72 months) and in three sites in 56 neonates and infants (< 48 h to 3 months). In 37 of these neonates and infants, corneocyte size, maturity, BH, C‐1 and plasmin activities were determined.
Results
NMF levels are low at birth and increase with age. Cheek SC, compared with elbow flexure and nasal tip, has the lowest NMF in the first year of life and is the slowest to reach stable levels. Cheek corneocytes remain immature. Plasmin, BH and C‐1 activities are all elevated by 1 month of age in exposed cheek skin, but not in elbow skin.
Conclusions
Regional and temporal differences in NMF levels, CE maturity and protease activities may explain the predilection for AD to affect the cheeks initially and are supportive of this site as key for allergen priming in early childhood. These observations will help design early intervention and treatment strategies for AD.
What's already known about this topic?
Atopic dermatitis (AD) frequently starts in early infancy, and the first eczematous lesions emerge on the cheeks.
Filaggrin is a major structural protein in the stratum corneum (SC).
Filaggrin deficiency is associated with the development of AD and, in the context of AD, food allergies and asthma.
Filaggrin is metabolized into natural moisturizing factors (NMFs), which can be measured in the SC.
What does this study add?
Regional differences in NMF levels, corneocyte envelope immaturity and protease activities may help explain why infantile AD most often initially affects the cheeks.
Filaggrin processing, corneocyte maturity, and protease activities show regional and temporal differences in infant skin.
These findings may explain disease patterns in early‐life AD.
What is the translational message?
Cheek skin may be highly relevant for allergen priming.
Emollient therapy at the vulnerable cheek site might help to prevent AD and/or food sensitization.
Linked Editorial: Thyssen. Br J Dermatol 2018; 179:235–236.
Plain language summary available online
Respond to this article
Summary
Background
Atopic dermatitis (AD) is the most common inflammatory skin disease. It is highly heterogeneous in clinical presentation, treatment response, disease trajectory and associated ...atopic comorbidities. Immune biomarkers are dysregulated in skin and peripheral blood.
Aims
We used noninvasive skin and peripheral biomarkers to observe the effects of real‐world topical corticosteroid (TCS) treatment in infants with AD, by measuring skin and blood biomarkers before and after therapy.
Methods
Seventy‐four treatment‐naïve infants with AD underwent 6 weeks of TCS treatment. Stratum corneum (SC) and plasma blood biomarkers as well as SC natural moisturizing factor (NMF) were measured before and after TCS therapy. Immune markers included innate, T helper (Th)1 and Th2 immunity, angiogenesis, and vascular factors. AD severity was assessed by the Scoring Atopic Dermatitis index, and skin barrier function by transepidermal water loss (TEWL). Twenty healthy infants were recruited as controls.
Results
TCS therapy predictably led to improvement in disease severity. Levels of immune markers in the skin and in the peripheral blood showed significant change from baseline, though most did not reach healthy control levels. The most prominent change from baseline in the SC was in markers of innate immune activation, interleukin (IL)‐18, IL‐8 and IL‐1α, and the Th2 chemokines C‐C motif chemokine (CCL)17 and CCL22. In blood, the largest changes were in Th2‐skewed biomarkers: CCL17, IL‐13, CCL22, IL‐5, and CCL26. TEWL decreased after therapy; no significant changes from baseline were found for NMF.
Conclusions
The profound impact of topical therapy on systemic biomarkers suggests that the skin compartment generates a major component of dysregulated systemic cytokines in infant AD. There may be long‐term beneficial effects of correcting systemic immune dysregulation through topical therapy.
What is already known about this topic?
Atopic dermatitis (AD) is a clinically heterogeneous condition with multiple clinical manifestations and a complex pathogenesis.
Noninvasive sampling of the stratum corneum has yielded significant insights into cytokine profiles of this compartment in AD.
Peripheral blood signatures have distinct cytokine profiles in childhood AD.
What does this study add?
We show the effects of topical corticosteroid (TCS) therapy on both skin‐derived and peripheral blood biomarkers.
TCS therapy had an effect on the cutaneous compartment but also substantially normalized the peripheral blood compartment cytokine signatures.
The long‐term effects of normalizing peripheral immune signatures are unknown but could potentially be beneficial.
What is the translational message?
TCS therapy can normalize systemic immune dysregulation in infant AD.
Summary
Background Filaggrin is a key protein involved in maintaining skin barrier function and hydration. Mutations in the filaggrin gene (FLG) cause ichthyosis vulgaris (IV) and are a major ...predisposing factor for atopic dermatitis (AD) in individuals of European and Asian descent. It has been proposed that FLG mutations are population specific and a difference in the spectra of mutations between different ancestral groups has been described. However, it is unknown whether FLG mutations in the African population are a causative genetic factor for IV and predispose to AD, or whether other mechanisms are more prominent.
Objectives The present aim was to investigate the role of FLG mutations as predisposing factors for IV or AD among individuals from Ethiopia.
Methods A case series of Ethiopian patients with AD (n = 103) and IV (n = 7) together with controls (n = 103; subjects without past or present history of AD, dry skin or atopic manifestations) was collected at the outpatient dermatology clinics at ALERT Dermatology Hospital, Tikur Anbessa Hospital and Gondar University Hospital, Ethiopia. AD was diagnosed by a dermatologist using the U.K. Working Party’s diagnostic criteria. The IV diagnosis was based on clinical examination and genetic testing of the steroid sulphatase gene to exclude X‐linked recessive ichthyosis. Patients were studied with direct sequencing (n = 40) and/or allelic discrimination (n = 110). Immunohistochemical analysis was performed for filaggrin expression in the skin of patients (n = 7) and controls (n = 2).
Results The Ethiopian patients and controls were genotyped for the four previously described common European FLG null mutations (R501X, 2282del4, S3247X, R2447X) and no carriers were found. In one patient with AD a novel heterozygous 2‐bp deletion, 632del2, leading to a premature stop codon was revealed by direct sequencing. No additional carrier of this deletion or other mutations was found. In addition, no difference in filaggrin expression was detected in AD or IV skin compared with healthy control skin.
Conclusions Our results indicate that FLG loss‐of‐function‐variants are less common in patients with IV and AD in the Ethiopian population, suggesting that other factors may be of importance in the pathogenesis in this ethnic group.
See also the Commentary by Carlsen
Mutations in FLG coding profilaggrin cause ichthyosis vulgaris and are an important predisposing factor for atopic dermatitis. Until now, most case–control studies and population‐based screenings ...have been performed only for prevalent mutations. In this study, we established a high‐throughput FLG mutation detection system by real‐time PCR with a set of two double‐dye probes and conducted comprehensive screening for almost all of the Japanese‐population‐specific FLG mutations (ten FLG mutations). The present comprehensive screening for all ten FLG mutations provided a more precise prevalence rate for FLG mutations (11.1%, n = 820), which seemed high compared with data of previous reports based on screening for limited numbers of FLG mutations. Our comprehensive screening suggested that population‐specific FLG mutations may be a significant predisposing factor for hay fever (odds ratio = 2.01 95% CI: 1.027–3.936, P < 0.05), although the sample sizes of this study were too small for reliable subphenotype analysis on the association between FLG mutations and hay fever in the eczema patients and the noneczema individuals, and it is not clear whether the association between FLG mutations and hay fever is due to the close association between FLG mutations and hay fever patients with eczema.
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