Understanding the effects of per- and polyfluoroalkyl substances on early skin development: Role of ciliogenesis inhibition and altered microtubule dynamics.

Per- and polyfluoroalkyl substances (PFAS) are a class of highly stable chemicals, widely used in everyday products, and widespread in the environment, even in pregnant women. While epidemiological studies have linked prenatal exposure to PFAS with atopic dermatitis in children, little is known about their toxic effects on skin development, especially during the embryonic stage. In this study, we utilized human embryonic stem cells to generate non-neural ectoderm (NNE) cells and exposed them to six PFAS (perfluorooctanoic acid (PFOA), undecafluorohexanoic acid (PFHxA), heptafluorobutyric acid (PFBA), perfluorooctane sulfonate (PFOS), perfluorohexane sulfonate (PFHxS) and perfluorobutyric acid (PFBS)) during the differentiation process to assess their toxicity to early skin development. Our results showed that PFOS altered the spindle-like morphology of NNE cells to a pebble-like morphology, and disrupted several NNE markers, including KRT16, SMYD1, and WISP1. The six PFAS had a high potential to cause hypohidrotic ectodermal dysplasia (HED) by disrupting the expression levels of HED-relevant genes. Transcriptomic analysis revealed that PFOS treatment produced the highest number (1156) of differentially expressed genes (DEGs) among the six PFAS, including the keratinocyte-related genes KRT6A, KRT17, KRT18, KRT24, KRT40, and KRT81. Additionally, we found that PFOS treatment disturbed several signaling pathways that are involved in regulating skin cell fate decisions and differentiation, including TGF-ß, NOTCH, Hedgehog, and Hippo signaling pathways. Interestingly, we discovered that PFOS inhibited, by partially interfering with the expression of cytoskeleton-related genes, the ciliogenesis of NNE cells, which is crucial for the intercellular transduction of the above-mentioned signaling pathways. Overall, our study suggests that PFAS can inhibit ciliogenesis and hamper the transduction of important signaling pathways, leading potential congenital skin diseases. It sheds light on the underlying mechanisms of early embryonic skin developmental toxicity and provides an explanation for the epidemiological data on PFAS. ENVIRONMENTAL IMPLICATION: We employed a model based on human embryonic stem cells to demonstrate that PFOS has the potential to elevate the risk of hypohidrotic ectodermal dysplasia. This is achieved by targeting cilia, inhibiting ciliogenesis, and subsequently disrupting crucial signaling pathways like TGF-ß, NOTCH, Hedgehog, and Hippo, during the early phases of embryonic skin development. Our study highlights the dangers and potential impacts of six PFAS pollutants on human skin development. Additionally, we emphasize the importance of closely considering PFHxA, PFBA, PFHxS, and PFBS, as they have shown the capacity to modify gene expression levels, albeit to a lesser degree.

Compound heterozygous WNT10A missense variations exacerbated the tooth agenesis caused by hypohidrotic ectodermal dysplasia.

BACKGROUND: The aim of this study was to analyse the differences in the phenotypes of missing teeth between a pair of brothers with hypohidrotic ectodermal dysplasia (HED) and to investigate the underlying mechanism by comparing the mutated gene loci between the brothers with whole-exome sequencing. METHODS: The clinical data of the patients and their mother were collected, and genomic DNA was extracted from peripheral blood samples. By Whole-exome sequencing filtered for a minor allele frequency (MAF) ≤0.05 non-synonymous single-nucleotide variations and insertions/deletions variations in genes previously associated with tooth agenesis, and variations considered as potentially pathogenic were assessed by SIFT, Polyphen-2, CADD and ACMG. Sanger sequencing was performed to detect gene variations. The secondary and tertiary structures of the mutated proteins were predicted by PsiPred 4.0 and AlphaFold 2. RESULTS: Both brothers were clinically diagnosed with HED, but the younger brother had more teeth than the elder brother. An EDA variation (c.878 T > G) was identified in both brothers. Additionally, compound heterozygous variations of WNT10A (c.511C > T and c.637G > A) were identified in the elder brother. Digenic variations in EDA (c.878 T > G) and WNT10A (c.511C > T and c.637G > A) in the same patient have not been reported previously. The secondary structure of the variant WNT10A protein showed changes in the number and position of α-helices and ß-folds compared to the wild-type protein. The tertiary structure of the WNT10A variant and molecular simulation docking showed that the site and direction where WNT10A binds to FZD5 was changed. CONCLUSIONS: Compound heterozygous WNT10A missense variations may exacerbate the number of missing teeth in HED caused by EDA variation.

A missense mutation in the highly conserved TNF-like domain of Ectodysplasin A is the candidate causative variant for X-linked hypohidrotic ectodermal dysplasia in Limousin cattle: Clinical, histological, and molecular analyses.

Ectodysplasin A related hypohidrotic ectodermal dysplasia (XLHED) is a well-studied fetal developmental disorder in mammals that mainly affects ectodermal structures. It has been identified in a variety of species, including mice, rats, dogs, cattle, and humans. Here, we report the clinical, histological, and molecular biological analyses of a case of XLHED in Limousin cattle. An affected Limousin calf showed pathognomonic signs of ectodermal dysplasia, i.e. sparse hair and characteristic dental aplasia. Histopathologic comparison of hairy and glabrous skin and computed tomography of the mandible confirmed the phenotypic diagnosis. In addition, a keratoconjunctivitis sicca was noted in one eye, which was also confirmed histopathologically. To identify the causative variant, we resequenced the bovine X-chromosomal ectodysplasin A gene (EDA) of the affected calf and compared the sequences to the bovine reference genome. A single missense variant (rs439722471) at position X:g.80411716T>C (ARS-UCD1.3) was identified. The variant resulted in an amino acid substitution from glutamic acid to glycine within the highly conserved TNF-like domain. To rule out the possibility that the variant was relatively common in the cattle population we genotyped 2,016 individuals including 40% Limousin cattle by fluorescence resonance energy transfer analysis. We also tested 5,116 multibreed samples from Run9 of the 1000 Bull Genomes Project for the said variant. The variant was not detected in any of the cattle tested, confirming the assumption that it was the causative variant. This is the first report of Ectodysplasin A related hypohidrotic ectodermal dysplasia in Limousin cattle and the description of a novel causal variant in cattle.

Hypohidrotic Ectodermal Dysplasia Milia Treatment With Fractional Carbon Dioxide Laser and Laser-Assisted Drug Delivery of Triamcinolone.

Hypohidrotic ectodermal dysplasia (HED) is a genetic disorder characterized by hypohidrosis, hypodontia, and hypotrichosis. Skin manifestations, including dyspigmentation and milia-like papules that coalesce into plaques, are difficult to treat. There is no cure for HED, therefore treatment is focused on managing symptoms and improving quality of life. There is limited evidence in the literature for safe and effective treatments improving HED-related facial skin aesthetics. The facial skin rashes caused by HED demonstrate an unmet clinical need in dermatology. Current therapies are limited to prevention methods such as keeping the skin cool by avoiding heat and applying topical moisturizers to help treat dry, pruritic skin. Herein we present a method for successful treatment of a 34-year-old African American male using fractional carbon dioxide CO2 ablative laser with laser-assisted drug delivery of triamcinolone 0.1% ointment that resulted in decreased milia-like papules, improved dyspigmentation, smoother skin tone, and high patient satisfaction. J Drugs Dermatol. 2023;22(11):1130-1132    doi:10.36849/JDD.7650.

X-linked genodermatoses from diagnosis to tailored therapy.

Background: Genodermatoses are rare heterogeneous genetic skin diseases with multiorgan involvement. They severely impair an individual's well-being and can also lead to early death. Methods: During the progress of this review, we have implemented a targeted research approach, diligently choosing the most relevant and exemplary articles within the subject matter. Our method entailed a systematic exploration of the scientific literature to ensure a compre-hensive and accurate compilation of the available sources. Results: Among genodermatoses, X-linked ones are of particular importance and should always be considered when pediatric males are affected. Regardless of other syndromic forms without prevalence of skin symptoms, X-linked genodermatoses can be classified in three main groups: keratinization defects, pigmentation defects, and inflammatory skin diseases. Typical examples are dyskeratosis congenita, keratosis follicularis spinulosa decalvans, hypohidrotic ectodermal dysplasia, chondrodysplasia punctata, hypohidrotic ectodermal dysplasia, incontinentia pigmenti, chronic granulomatous disease, CHILD syndrome and ichthyosis. In this field, genetic diagnosis of the specific disease is important, also considering that numerous clinical trials of orphan drugs and genetic therapies are being proposed for these rare genetic diseases. Conclusions: Thus, this chapter starts from clinical to molecular testing and ends with a review of all clinical trials on orphan drugs and gene therapy for genodermatoses.

A new variant of the ectodysplasin A receptor death domain gene associated with anhidrotic ectodermal dysplasia in a Turkish family and its simple diagnosis by restriction fragment length polymorphism.

Ectodermal dysplasia (ED), which exhibits a wide range of clinical symptoms, may be classified into three major types: hypohidrotic, anhidrotic, and hidrotic. A male child (proband) showing anhidrotic dysplasia was used as the subject of this study. The biopsy of the big toe revealed that the male child had no sweat glands. Genetic analysis of the patient revealed a mutation caused by a homozygous nucleotide substitution in the EDAR-associated death domain (EDARADD) (rs114632254) gene c.439G>A (p.Gly147Arg). Phenotypically, his teeth were sharp, but eight teeth were missing (oligodontia). The patient had normal nails with dry skin, sparse hair, everted lower lip vermilion, hyperpigmented eyelids, and abnormal nasal bridge morphology around the eyes. There is also a homozygous dominant (healthy) female and a heterozygous male in this family, who are cousins (aunt children) to the heterozygous parents. The daughter of the patient was also heterozygous. This mutation represents homozygous recessive inheritance, which we describe for the first time. Furthermore, we demonstrated that this genetic disorder can be readily diagnosed using the restriction fragment length polymorphism (RFLP) method after digestion with MnII restriction endonuclease.

A Causal Treatment for X-Linked Hypohidrotic Ectodermal Dysplasia: Long-Term Results of Short-Term Perinatal Ectodysplasin A1 Replacement.

X-linked hypohidrotic ectodermal dysplasia (XLHED), caused by a genetic deficiency of ectodysplasin A1 (EDA1), is a rare developmental disorder of ectodermal derivatives such as hair, sweat glands, and teeth. The absence of sweat glands and perspiration can evoke life-threatening hyperthermia. As molecular genetic findings are not always conclusive, the concentrations of circulating EDA1 may help to distinguish between total and partial EDA1 deficiencies. We previously treated nine male patients with obvious signs of XLHED with a recombinant EDA1 replacement protein, Fc-EDA, either shortly after birth (n = 3) or by prenatal administration in gestational week 26 and beyond (n = 6). Here, we present the long-term follow-up for up to six years. In patients who had received Fc-EDA after birth, neither sweat glands nor sweating ability were detected at the age of 12-60 months. In contrast, prenatal EDA1 replacement resulted in ample sweat gland development and pilocarpine-inducible sweating in all treated subjects, who also attained more permanent teeth than their untreated affected relatives. Normal perspiration has persisted for six years in the two oldest boys treated repeatedly with Fc-EDA in utero. When they had a sauna, adequate thermoregulation was evidenced. Lower sweat production after single prenatal dosing may indicate a dose-response relationship. The absence of circulating EDA1 in five prenatally treated subjects proved that these children would have been unable to perspire if they had been left untreated. The sixth infant was shown to produce an EDA1 molecule that, albeit interacting with its cognate receptor, cannot activate EDA1 signaling. In conclusion, a causal treatment of XLHED before birth is feasible.

Next generation sequencing panel target genes: possible diagnostic tool for ectodermal dysplasia related diseases.

BACKGROUND: Ectodermal dysplasias (EDs) are a large and complex group of disorders affecting the ectoderm-derived organs; the clinical and genetic heterogeneity of these conditions renders an accurate diagnosis more challenging. The aim of this study is to demonstrate the clinical utility of a targeted resequencing panel through enhancing the molecular and clinical diagnosis of EDs. Given the recent developments in gene and protein-based therapies for X-linked hypohidrotic ectodermal dysplasia, there is a re-emerging interest in identifying the genetic basis of EDs and the respective phenotypic presentations, in an aim to facilitate potential treatments for affected families. METHODS: We assessed seventeen individuals, from three unrelated families, who presented with diverse phenotypes suggestive of ED. An extensive multidisciplinary clinical evaluation was performed followed by a targeted exome resequencing panel (including genes that are known to cause EDs). MiSeqTM data software was used, variants with Qscore >30 were accepted. RESULTS: Three different previously reported hemizygous EDA mutations were found in the families. However, a complete genotype-phenotype correlation could not be established, neither in our patients nor in the previously reported patients. CONCLUSIONS: Targeted exome resequencing can provide a rapid and accurate diagnosis of EDs, while further contributing to the existing ED genetic data. Moreover, the identification of the disease-causing mutation in an affected family is crucial for proper genetic counseling and the establishment of a genotype-phenotype correlation which will subsequently provide the affected individuals with a more suitable treatment plan.

A novel deletion of exon 4 in the Ectodysplasin A gene associated with X-linked hypohidrotic ectodermal dysplasia.

OBJECTIVE: Identify the disease-causing mutation in a patient with features of X-linked hypohidrotic ectodermal dysplasia, which is a genetic disorder characterized by hypodontia, hypohidrosis and hypotrichosis. It is caused by mutations in Ectodysplasin A gene, which encodes ectodysplasin A, a member of the tumor necrosis factor superfamily. DESIGN: Genetic analysis, was performed using chromosomal microarray analysis, whole exome sequencing and multiplex ligation-dependent probe amplification analysis in a 4-year-old boy with hypohidrotic ectodermal dysplasia features. Moreover, the boy's parents were tested for clinically significant findings identified in order to elucidate the pattern of inheritance of the finding detected in the proband. RESULTS: A novel deletion of entire exon 4 in Ectodysplasin A gene identified in the 4-year-old patient. This deletion was found in heterozygous state in the mother of the proband and was not detected in his father. RNA analysis revealed an in-frame deletion r.527_706del, p.(176_236del) in exon 4 of the Ectodysplasin A gene. CONCLUSION: We identified a novel gross deletion in the Ectodysplasin A gene in a male patient with X-linked hypohidrotic ectodermal dysplasia. Clinical and molecular genetic analysis are crucial to set an accurate diagnosis in patients with hypohidrotic ectodermal dysplasia. These results highlight the importance of the collagen domain of Ectodysplasin A, encoded by exon 4, for its function in vivo.

Structural insights into pathogenic mechanism of hypohidrotic ectodermal dysplasia caused by ectodysplasin A variants.

EDA is a tumor necrosis factor (TNF) family member, which functions together with its cognate receptor EDAR during ectodermal organ development. Mutations of EDA have long been known to cause X-linked hypohidrotic dysplasia in humans characterized by primary defects in teeth, hair and sweat glands. However, the structural information of EDA interaction with EDAR is lacking and the pathogenic mechanism of EDA variants is poorly understood. Here, we report the crystal structure of EDA C-terminal TNF homology domain bound to the N-terminal cysteine-rich domains of EDAR. Together with biochemical, cellular and mouse genetic studies, we show that different EDA mutations lead to varying degrees of ectodermal developmental defects in mice, which is consistent with the clinical observations on human patients. Our work extends the understanding of the EDA signaling mechanism, and provides important insights into the molecular pathogenesis of disease-causing EDA variants.

Protocol for the Phase 2 EDELIFE Trial Investigating the Efficacy and Safety of Intra-Amniotic ER004 Administration to Male Subjects with X-Linked Hypohidrotic Ectodermal Dysplasia.

X-linked hypohidrotic ectodermal dysplasia (XLHED) is a rare genetic disorder characte-rised by abnormal development of the skin and its appendages, such as hair and sweat glands, the teeth, and mucous glands of the airways, resulting in serious, sometimes life-threatening complications like hyperthermia or recurrent respiratory infections. It is caused by pathogenic variants of the ectodysplasin A gene (EDA). Most affected males are hemizygous for EDA null mutations that lead to the absence or inactivity of the signalling protein ectodysplasin A1 (EDA1) and, thus, to the full-blown phenotype with inability to perspire and few if any teeth. There are currently no long-term treatment options for XLHED. ER004 represents a first-in-class protein replacement molecule designed for specific, high-affinity binding to the endogenous EDA1 receptor (EDAR). Its proposed mechanism of action is the replacement of missing EDA1 in yet unborn patients with XLHED. Once bound to EDAR, ER004 activates the EDA/NFκB signalling pathway, which triggers the transcription of genes involved in the normal development of multiple tissues. Following preclinical studies, named-patient use cases demonstrated significant potential of ER004 in affected males treated in utero during the late second and third trimesters of pregnancy. In order to confirm these results, we started the EDELIFE trial, a prospective, open-label, genotype-match controlled, multicentre clinical study to investigate the efficacy and safety of intra-amniotic ER004 administration as a prenatal treatment for male subjects with XLHED. This article summarises the rationale, the study protocol, ethical issues of the trial, and potential pitfalls.

Different degree of loss-of-function among four missense mutations in the EDAR gene responsible for autosomal recessive hypohidrotic ectodermal dysplasia may be associated with the phenotypic severity.

Hypohidrotic ectodermal dysplasia is a rare condition characterized by hypohidrosis, hypodontia, and hypotrichosis. The disease can show X-linked recessive, autosomal dominant or autosomal recessive inheritance trait. Of these, the autosomal forms are caused by mutations in either EDAR or EDARADD. To date, the underlying pathomechanisms or genotype-phenotype correlations for autosomal forms have not completely been disclosed. In this study, we performed a series of in vitro studies for four missense mutations in the death domain of EDAR protein: p.R358Q, p.G382S, p.I388T, and p.T403M. The results revealed that p.R358Q- and p.T403M-mutant EDAR showed different expression patterns from wild-type EDAR in both western blots and immunostainings. NF-κB reporter assays demonstrated that all the mutant EDAR showed reduced activation of NF-κB, but the reduction by p.G382S- and p.I388T-mutant EDAR was moderate. Co-immunoprecipitation assays showed that p.R358Q- and p.T403M-mutant EDAR did not bind with EDARADD at all, whereas p.G382S- and p.I388T-mutant EDAR maintained the affinity to some extent. Furthermore, we demonstrated that all the mutant EDAR proteins analyzed aberrantly bound with TRAF6. Sum of the data suggest that the degree of loss-of-function is different among the mutant EDAR proteins, which may be associated with the severity of the disease.

A novel mutation in the collagen domain of EDA results in hypohidrotic ectodermal dysplasia by impacting the receptor-binding capability.

BACKGROUND: Hypohidrotic ectodermal dysplasia (HED) mainly results from gene mutations in the EDA/EDAR/NF-κB pathway. Function analysis of the mutations in the collagen domain of ectodysplasin A (EDA)result in HED has been rarely studied. This study aimed at determining the mechanism by which the novel collagen domain mutation of EDA results in HED. METHODS: We analyzed the DNAs from a Chinese family with HED and performed bioinformatics analysis. A new three-dimensional structure model of the EDA trimer was built and used to predict the effect of the mutations on EDA. We performed a western blot to detect EDA1 proteins in cell lysates and supernatants. We then performed coimmunoprecipitation to determine whether the mutation would affect the interaction of EDA1 with the EDA receptor (EDAR). Dual luciferase reporter assay and immunofluorescence were performed to detect the effect of the mutant EDA1 protein on nuclear factor kappa B (NF-κB) activation. RESULTS: A novel missense mutation (c.593G > A, p. Gly198Glu) in the collagen domain of EDA was detected. The mutation was predicted to be disease-causing. A three-dimensional structure model of the EDA trimer was first built in this study, in which the mutation site is located around the receptor binding domain. Functional studies showed that there was no difference in the secretion activity between the mutant EDA1 and the wild-type EDA1. However, the receptor-binding activity and the transcription activation of NF-κB were impaired by the mutation. CONCLUSION: We identified a novel mutation (c.593G > A, p. Gly198Glu) in the collagen domain of EDA. Bioinformatics analysis and functional studies showed this mutation was damaging, indicating that mutations in the collagen domain of EDA could result in HED by affecting the receptor-binding activity of EDA and the transcriptional activity of NF-κB.

A Missense Mutation in the Collagen Triple Helix of EDA Is Associated with X-Linked Recessive Hypohidrotic Ectodermal Dysplasia in Fleckvieh Cattle.

Mutations within the ectodysplasin A (EDA) gene have been associated with congenital hypotrichosis and anodontia (HAD/XHED) in humans, mice, dogs and cattle. We identified a three-generation family of Fleckvieh cattle with male calves exhibiting clinical and histopathological signs consistent with an X-linked recessive HAD (XHED). Whole genome and Sanger sequencing of cDNA showed a perfect association of the missense mutation g.85716041G>A (ss2019497443, rs1114816375) within the EDA gene with all three cases following an X-linked recessive inheritance, but normal EDAR and EDARADD. This mutation causes an exchange of glycine (G) with arginine (R) at amino acid position 227 (p.227G>R) in the second collagen triple helix repeat domain of EDA. The EDA variant was associated with a significant reduction and underdevelopment of hair follicles along with a reduced outgrowth of hairs, a complete loss of seromucous nasolabial and mucous tracheal and bronchial glands and a malformation of and reduction in number of teeth. Thermostability of EDA G227R was reduced, consistent with a relatively mild hair and tooth phenotype. However, incisors and canines were more severely affected in one of the calves, which correlated with the presence of a homozygous missense mutation of RNF111 (g.51306765T>G), a putative candidate gene possibly associated with tooth number in EDA-deficient Fleckvieh calves.

Appearance Says It All; A Rare Case Of Hypohidrotic Ectodermal Dysplasia.

Ectodermal Dysplasia (ED) is a rare genetic condition characterized by the involvement of ectoderm derivatives such as hair, nail, sweat glands, and teeth. It has many variants, but the two most common ones are hypohidrotic/anhidrotic ectodermal dysplasia and hidrotic ectodermal dysplasia. Herein, we present a case of a 20-year-old female with hypohidrotic ectodermal dysplasia who had anodontia, hypohidrosis, and hypotrichosis, and her condition went unrecognized until she was seen for gastroenteritis at a tertiary care center. This case report will help spread education and awareness regarding such a rare and under-recognized condition. Early diagnosis and intervention help improve the quality of life.

Orthodontic and dentofacial orthopedic treatments in patients with ectodermal dysplasia: a systematic review.

OBJECTIVE: The objective of this systematic review was to determine the orthodontic and dentofacial orthopedic treatments carried out in patients with ectodermal dysplasia to facilitate functional and aesthetic rehabilitation. METHODS: The systematic review was conducted in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-analysis statement. We systematically searched PubMed, Web of Science, Scopus, Scielo, LILACS, EBSCOhost and Embase databases up to 6 January 2022. We included articles describing patients with any type of ectodermal dysplasia who received orthodontic or dentofacial orthopedic treatment to facilitate functional and aesthetic oral rehabilitation. The search was not restricted by language or year of publication. The quality of the studies was assessed using the Joanna Briggs Institute Quality Assessment Scale of the University of Adelaide for case series and case reports. The review was registered at the University of York Centre for reviews (CRD42021288030). RESULTS: Of the initial 403 studies found, 29 met the inclusion criteria. After applying the quality scale, 23 were left for review-21 case reports and 2 case series. The initial age of patients ranged from 34 months to 24 years. Thirteen studies were on hypohidrotic and/or anhidrotic ectodermal dysplasia, of which two were X-chromosome linked. In one study, the patient had Wiktop syndrome, and in nine the type of ectodermal dysplasia was not specified. The duration of treatment was 7 weeks to 10 years. The treatments described were: fixed orthodontic appliances or simple acrylic plates designed for tooth movement, including leveling and aligning, closing of diastemata, retraction of impacted teeth in the dental arch; clear aligners; fixed and/or removable appliances for the correction of skeletal and/or dentoalveolar relationships; palatal expanders in combination with face masks for orthopedic traction of the maxilla; and orthognathic surgery. Only three studies provided cephalometric data. CONCLUSION: The level of evidence of the articles reviewed was low and most orthopedic and dentofacial orthodontic treatments described were focused on correcting dental malpositioning and jaw asymmetries and not on stimulating growth from an early age. Studies with greater scientific evidence are needed to determine the best treatment for these patients.

Ectodysplasin A1 Deficiency Leads to Osteopetrosis-like Changes in Bones of the Skull Associated with Diminished Osteoclastic Activity.

Pathogenic variants of the gene Eda cause X-linked hypohidrotic ectodermal dysplasia (XLHED), which is characterized by structural abnormalities or lack of ectodermal appendages. Signs of dysplasia are not restricted to derivatives of the ectodermal layer, but mesodermal abnormalities, such as craniofacial dysmorphism, are also frequently observed, suggesting close reciprocal interactions between the ectoderm and mesoderm; however, a causal link has remained unsubstantiated. We investigated the functional impact of defective ectodysplasin A1 (Eda1) signaling on postnatal bone homeostasis in Eda1-deficient Tabby mice. Interestingly, Eda1 was detected in wild-type mouse calvariae throughout postnatal lifetime. In calvariae, bone-lining Osterix (Osx)+ osteoblasts stained positive for Eda1, and osteoclasts were revealed as Eda receptor (Edar)-positive. Moreover, adult Eda1-deficient calvarial bone showed osteopetrosis-like changes with significantly diminished marrow space, which was maintained during adulthood. Concomitantly with osteopetrosis-like changes, Tabby calvarial bone and Tabby bone marrow-derived osteoclasts had far less osteoclastic activity-associated co-enzymes including cathepsin K, Mmp9, Trap, and Tcirg1 (V-type proton ATPase a3 subunit) compared with wild-type calvariae in vivo or osteoclasts in vitro, indicating that Eda1 deficiency may affect the activity of osteoclasts. Finally, we confirmed that nuclear Nfatc1-positive osteoclasts were strongly diminished during mature osteoclastic differentiation under M-CSF and RANKL in the Tabby model, while Fc-EDA treatment of Tabby-derived osteoclasts significantly increased nuclear translocation of Nfatc1. Furthermore, we identified enhanced Nfatc1 and NF-κB transcriptional activity following Fc-EDA treatment in vitro using luciferase assays. Overall, the results indicate that diminished expressions of osteoclastic activity-associated co-enzymes may lead to disturbed bone homeostasis in Tabby calvariae postnatally.

A large deletion encompassing exon 2 of the ectodysplasin A (EDA) gene in a British blue crossbred calf with hypohidrotic ectodermal dysplasia.

BACKGROUND: Hypohidrotic ectodermal dysplasia (HED) is a congenital syndrome of mammals affecting organs and tissues of ectodermal origin characterized by absence or hypoplasia of hair, teeth, and eccrine glands. The disorder has been reported in several species, including humans, mice, dogs and cattle, associated with variants in genes affecting the ectodysplasin pathway, including the X-linked ectodysplasin A (EDA) gene. Until now, nine pathogenic variants have been found in the bovine EDA gene. Here we report a novel variant in EDA in a crossbreed male Belgian Blue calf with HED, and provide an overview of the phenotypic and allelic heterogeneity of EDA-related forms of HED in cattle. CASE PRESENTATION: A 45-day-old male crossbreed British Blue calf was referred with congenital hypotrichosis, oligodontia and omphalitis. On histopathological examination of the nasal planum, nasolabial glands and ducts were not observed. The density of hair follicles was low, and they were small, with a predominance of telogen-phase hairs, and some serocellular crusts. The phenotype of the calf resembled that of HED. Whole-genome sequencing (WGS) was performed and revealed a 21,899 base-pair deletion encompassing the coding exon 2 of EDA, predicted to result in an altered transcript and aberrant protein. CONCLUSIONS: The clinicopathological and genetic findings were consistent with a case of X-linked HED. A very similar EDA deletion has been previously reported in a family of Holstein cattle with HED. The newly identified hemizygous EDA loss-of-function variant is certainly pathogenic and therefore is the genetic cause for the observed phenotype. This case report provides an additional example of the potential of WGS-based precise diagnostics in livestock species such as cattle to increase the diagnostic yield in rare diseases.