Lentiviral expression of wild-type LAMA3A restores cell adhesion in airway basal cells from children with epidermolysis bullosa.

The hallmark of epidermolysis bullosa (EB) is fragile attachment of epithelia due to genetic variants in cell adhesion genes. We describe 16 EB patients treated in the ear, nose, and throat department of a tertiary pediatric hospital linked to the United Kingdom's national EB unit between 1992 and 2023. Patients suffered a high degree of morbidity and mortality from laryngotracheal stenosis. Variants in laminin subunit alpha-3 (LAMA3) were found in 10/15 patients where genotype was available. LAMA3 encodes a subunit of the laminin-332 heterotrimeric extracellular matrix protein complex and is expressed by airway epithelial basal stem cells. We investigated the benefit of restoring wild-type LAMA3 expression in primary EB patient-derived basal cell cultures. EB basal cells demonstrated weak adhesion to cell culture substrates, but could otherwise be expanded similarly to non-EB basal cells. In vitro lentiviral overexpression of LAMA3A in EB basal cells enabled them to differentiate in air-liquid interface cultures, producing cilia with normal ciliary beat frequency. Moreover, transduction restored cell adhesion to levels comparable to a non-EB donor culture. These data provide proof of concept for a combined cell and gene therapy approach to treat airway disease in LAMA3-affected EB.

Management of acute sloughing of the esophageal lining in patients with dystrophic epidermolysis bullosa-A series of six pediatric patients.

BACKGROUND: Dystrophic epidermolysis bullosa (DEB) is a subtype of an inherited skin disorder characterized by skin and mucosal fragility due to collagen VII (COL7A1) gene mutations. Esophageal strictures leading to chronic dysphagia and acute episodes are well recognized complications within this subtype. Sloughing of esophageal mucosa and the treatment of this emergency have heretofore received limited attention in the EB literature. METHODS: We retrospectively reviewed the electronic medical records of the patients who had an acute episode of sloughing of the esophageal lining between 2008 and 2021 and extracted the information regarding their clinical presentation and management. RESULTS: Six patients out of 210 with recessive DEB severe (RDEB-S) (n = 4), RDEB intermediate (RDEB-I) (n = 1) and dominant DEB (DDEB) (n = 1) were identified. The mean age at the time of the episode was 2.7 years. All patients had early-onset severe gastroesophageal reflux. Clinically, they presented with a coughing episode (n = 6), hematemesis (n = 6), vomiting (n = 6), and choking (n = 3), followed by coughing up a string like tissue of variable length, part of the esophageal mucosal lining. Four patients recovered with medical management only, two patients required gastrostomy insertions for feeding due to severe persistent dysphagia and one also required a Nissen's fundoplication to manage severe reflux. One patient had aspiration pneumonia. CONCLUSIONS: Sloughing of varying lengths of segments of the esophagus is an emergency. The lining coughed up needs to be cut at the mouth not pulled and the emergency services called immediately for urgent assessment and management. Expert multidisciplinary care is needed to manage this rare but serious condition.

Biallelic TUFT1 variants cause woolly hair, superficial skin fragility and desmosomal defects.

BACKGROUND: Desmosomes are complex cell junction structures that connect intermediate filaments providing strong cell-to-cell adhesion in tissues exposed to mechanical stress. OBJECTIVES: To identify causal variants in individuals with woolly hair and skin fragility of unknown genetic cause. METHODS: This research was conducted using whole-genome sequencing, whole-exome sequencing, clinical phenotyping, haplotype analysis, single-cell RNA sequencing data analysis, immunofluorescence microscopy and transmission electron microscopy. RESULTS: We identified homozygous predicted loss-of-function tuftelin-1 (TUFT1) variants in nine individuals, from three families, with woolly hair and skin fragility. One donor splice-site variant, c.60+1G>A, was present in two families, while a frameshift variant, p.Gln189Asnfs*49, was found in the third family. Haplotype analysis showed the c.60+1G>A substitution to be a founder variant in the Irish population that likely arose approximately 20 generations ago. Human and mouse single-cell RNA sequencing data showed TUFT1 expression to be enriched in the hair dermal sheath and keratinocytes. TUFT1 expression was highly correlated with genes encoding desmosomal components implicated in diseases with phenotypes that overlap with the cohort presented here. Immunofluorescence showed tuftelin-1 to be mainly localized to the peripheral cell membranes of keratinocytes in normal skin. Skin samples from individuals with TUFT1 variants showed markedly reduced immunoreactivity for tuftelin-1, with a loss of the keratinocyte cell membrane labelling. Light microscopy revealed keratinocyte adhesion, mild hyperkeratosis and areas of superficial peeling. Transmission electron microscopy showed panepidermal acantholysis with widening of intercellular spaces throughout the epidermis and desmosomal detachment through the inner plaques. CONCLUSIONS: Biallelic loss-of-function TUFT1 variants cause a new autosomal recessive skin/hair disorder characterized by woolly hair texture and early-onset skin fragility. Tuftelin-1 has a role in desmosomal integrity and function.

The impact of using a multistrain probiotic supplement on gastrointestinal function in children and adolescents with severe recessive dystrophic epidermolysis bullosa.

BACKGROUND: Children and adolescents with severe recessive dystrophic epidermolysis bullosa (RDEB-S) often have severe constipation in addition to gastrointestinal dysbiosis, due to frequent antibiotic use and reduced oral diet. Constipation is treated with long-term use of high daily doses of macrogol gel (Movicol Paediatric PlainTM or LaxidoTM). Constipation is refractory to increases in fibre and fluids, and impacts severely on quality of life. AIM: To study the initial impact and efficacy of using a multistrain probiotic supplement daily for 12 weeks in patients with RDEB-S. The authors sought to determine the impact of such a supplement on  gastrointestinal symptoms, stool consistency and the use of macrogol gel to treat constipation, as well as understanding patient reaction, palability and ease of use. METHODS: Patients were identified through the epidermolysis bullosa tertiary multidisciplinary team clinic in July 2021. Patients were included if they had a diagnosis of RDEB-S, prescribed at least one sachet of macrogol gel and provided written consent to take part. Patients were provided, proprietary liquid multistrain probiotic supplement (Symprove™) with a high bacterial count, at a dose of 1 mL kg-1 once a day. Each patient completed an anonymous, nine-question, electronic survey to document symptoms and report overall findings at the start and end of a 12-week trial period. RESULTS: Four patients with RDEB-S (two boys and two girls; age range 7-14 years) who met the inclusion criteria were approached to take part. All patients had chronic constipation requiring daily macrogol gel use (range 2-5 sachets per day). Three out of four (75%) completed the 12-week course. At baseline (before supplementation commenced), all three (100%) patients reported poor oral appetite, constipation, flatulence, abdominal bloating and pain, and frequent skin infections requiring oral antibiotics, with two of the three (66%) patients also having nausea. After 12 weeks of supplementation, all three patients (100%) reported a significant improvement in abdominal pain and bloating, nausea, stool consistency, stool frequency, flatulence and increased appetite. Two of the three patients (66%) were able to reduce their macrogol gel usage and the third patient (33%) was able to stop macrogol gel usage altogether during the study period. All three patients said they would choose to continue using the supplement if it was available. CONCLUSION: We have shown in this case series that giving a multistrain probiotic supplement in patients with RDEB-S has the potential to improve stool consistency and reduce or prevent the need for chronic macrogol gel use. Future larger-scale, placebo-controlled trials are needed to confirm these results.