A COL7A1 Variant in a Litter of Neonatal Basset Hounds with Dystrophic Epidermolysis Bullosa.

We investigated three neonatal Basset Hound littermates with lesions consistent with epidermolysis bullosa (EB), a group of genetic blistering diseases. A clinically normal bitch was bred to her grandfather by artificial insemination. Out of a litter of seven puppies, two affected puppies died and one was euthanized, with these puppies being submitted for diagnostic necropsy. All had multiple bullae and ulcers involving the nasal planum and paw pads, as well as sloughing claws; one puppy also had oral and esophageal ulcers. The complete genome of one affected puppy was sequenced, and 37 known EB candidate genes were assessed. We found a candidate causative variant in COL7A1, which encodes the collagen VII alpha 1 chain. The variant is a complex rearrangement involving duplication of a 107 bp region harboring a frameshift deletion of 7 bp. The variant is predicted to truncate more than 75% of the open reading frame, p.(Val677Serfs*11). Targeted genotyping of this duplication confirmed that all three affected puppies were homozygous for the duplication, whereas 12 unaffected Basset Hounds did not carry the duplication. This variant was also not seen in the genomes of more than 600 dogs of other breeds. COL7A1 variants have been identified in humans and dogs with dystrophic epidermolysis bullosa (DEB). The identified COL7A1 variant therefore most likely represents the causative variant and allows the refinement of the preliminary EB diagnosis to DEB.

Nonsense variant in COL7A1 causes recessive dystrophic epidermolysis bullosa in Central Asian Shepherd dogs.

A rare hereditary mechanobullous disorder called epidermolysis bullosa (EB) causes blistering in the skin and the mucosal membranes. To date, nineteen EB-related genes have been discovered in human and other species. We describe here a novel EB variant in dogs. Two newborn littermates of Central Asian Shepherd dogs with severe signs of skin blistering were brought to a veterinary clinic and euthanized due to poor prognosis. In post-mortem examination, the puppies were shown to have findings in the skin and the mucosal membranes characteristic of EB. A whole-genome sequencing of one of the affected puppies was performed to identify the genetic cause. The resequencing data were filtered under a recessive model against variants from 31 other dog genomes, revealing a homozygous case-specific nonsense variant in one of the known EB-causing genes, COL7A1 (c.4579C>T, p.R1527*). The variant results in a premature stop codon and likely absence of the functional protein in the basement membrane of the skin in the affected dogs. This was confirmed by immunohistochemistry using a COL7A1 antibody. Additional screening of the variant indicated full penetrance and breed specificity at ~28% carrier frequency. In summary, this study reveals a novel COL7A1 variant causing recessive dystrophic EB and provides a genetic test for the eradication of the disease from the breed.

Congenital dystrophic epidermolysis bullosa (DEB) in Sprague Dawley rats: a case series.

BACKGROUND: Epidermolysis bullosa is a rare skin disease caused by defects in the basement membrane and/or other dermoepidermal junction components. HYPOTHESIS/OBJECTIVES: We describe a series of spontaneous cases of dystrophic epidermolysis bullosa (DEB) in a colony of Sprague Dawley rats investigated with histopathology, transmission electron microscopy (TEM) and inheritance pattern. ANIMALS: Four, 4-day-old pups from a litter of Sprague Dawley rats developed blistering, haemorrhagic skin lesions and were euthanized. Age-matched controls from the same litter were normal. Several months later two more litters presented with identical findings. All three litters had the same sire, suggesting a genetic component. METHODS: Skin from affected and control animals was evaluated histologically and with TEM. Unaffected sibling pairs from affected litters were bred in order to potentially reproduce the disease and determine the mode of inheritance. RESULTS: Histologically, there was significant dermoepidermal clefting below the basement membrane with variable amounts of haemorrhage and cellular debris within the clefts. Ultrastructurally, clefting occurred below the basement membrane with an intact lamina densa and normal hemidesmosomes. Anchoring filaments were strikingly absent. Litters produced from phenotypically unaffected sibling pairs resulted in a total of four more litters with approximately a quarter of pups affected. CONCLUSIONS AND CLINICAL IMPORTANCE: Based on the gross lesions, histopathological features and TEM determination of separation below the lamina densa and lack of normal anchoring fibrils, these cases are most consistent with DEB. This is the first report of naturally occurring, localized and reproducible recessive DEB in Sprague Dawley rats.

Epidermolysis bullosa in animals: a review.

Epidermolysis bullosa (EB) is a hereditary mechanobullous disease of animals and humans, characterized by an extreme fragility of the skin and mucous membranes. The main feature of EB in humans and animals is the formation of blisters and erosions in response to minor mechanical trauma. Epidermolysis bullosa is caused by mutations in the genes that code for structural proteins of the cytoskeleton of the basal keratinocytes or of the basement membrane zone. Based on the ultrastructural levels of tissue separation, EB is divided into the following three broad categories: epidermolysis bullosa simplex, junctional epidermolysis bullosa and dystrophic epidermolysis bullosa. Human types of EB are divided into several subtypes based on their ultrastructural changes and the mode of inheritance; subtypes are not fully established in animals. In humans, it is estimated that EB affects one in 17,000 live births; the frequency of EB in different animals species is not known. In all animal species, except in buffalo with epidermolysis bullosa simplex, multifocal ulcers are observed on the gums, hard and soft palates, mucosa of the lips, cheek mucosa and dorsum of the tongue. Dystrophic or absent nails, a frequent sign seen in human patients with EB, corresponds to the deformities and sloughing of the hooves in ungulates and to dystrophy or atrophy of the claws in dogs and cats. This review covers aspects of the molecular biology, diagnosis, classification, clinical signs and pathology of EB reported in animals.

Dystrophic epidermolysis bullosa in goats.

Clinical, histopathological and ultrastructural findings of caprine dystrophic epidermolysis bullosa (DEB) with autosomal recessive inheritance are reported. The goats presented with exungulation, erosions, crusts and scars on the skin and ulcers in the oral cavity. Microscopically, the skin showed subepidermal separation with clefts filled occasionally with clear eosinophilic fluid, cellular debris or neutrophils. Ultrastructurally, the site of blister formation was the sublamina densa in the epidermal basement membrane zone. In skin with blister formation and in clinically uninvolved skin, the basal lamina was preserved, but the anchoring fibrils were sparse and rudimentary. A twin brother of an affected kid was mated over 5 years with his mother; three out of the 10 kids born presented with epidermolysis bullosa, indicating that the disease has an autosomal recessive mode of inheritance. It is suggested that the disease is similar to human severe generalized recessive DEB.

Dystrophic epidermolysis bullosa in Assaf lambs.

Dystrophic epidermolysis bullosa (DEB) was diagnosed in five newborn Assaf lambs from a flock in which around 1-2% of the lambs were lame. Affected animals had marked erythema of the coronary band and subsequent detachment of the hooves. Blisters were present on the lips and in the oral and oesophageal mucosae. Erosions and crusts were noted in the axillary and inguinal areas. Microscopically, there was detachment of the epidermis from the dermis. The roof of these bullous spaces was formed by the intact epidermis with its basement membrane. The bullae were generally filled with pale eosinophilic proteinaceous fluid and erythrocytes. Immunohistochemical analysis revealed absence of collagen VII in the basement membrane zone. The disease in these lambs appears similar to the severe recessive form of human DEB.

Genetic correction of canine dystrophic epidermolysis bullosa mediated by retroviral vectors.

We have assessed the suitability of retroviral vectors for gene therapy of recessive dystrophic epidermolysis bullosa (RDEB) in dogs expressing a mutated collagen type VII. Isolation and analysis of the 9 kb dog collagen type VII cDNA identified the causative genetic mutation G1906S and disclosed the interspecies conservation of collagen type VII. Highly efficient transfer of the wild-type collagen type VII cDNA to both dog RDEB and human primary RDEB collagen type VII-null keratinocytes using recombinant vectors derived from LZRS-Ires-zeo and MSCV retroviruses achieved sustained and permanent expression of the transgene product. The expression and post-translational modification profile of the recombinant collagen type VII was comparable to that of the wild-type counterpart. The recombinant canine collagen type VII in human RDEB keratinocytes and dog cells corrected the observable defects caused by RDEB keratinocytes in cell cultures and in vitro reconstructed skin. Hypermotility was fully reverted in human RDEB keratinocytes, and strongly reduced in the dog RDEB cells. This observation suggests that not only infection efficiency but also high expression levels are required to ensure therapeutic efficacy in the presence of mutated gene products. Our results set the basis for preclinical gene therapy assays in the first immune-competent large animal model for an inherited skin disease and broaden the spectrum of preclinical and clinical applications of retroviral vectors in the transfer of large recombinant genes in epithelial cells.

Reduced anchoring fibril formation and collagen VII immunoreactivity in feline dystrophic epidermolysis bullosa.

Dystrophic epidermolysis bullosa was diagnosed in a cat with juvenile-onset epithelial sloughing of the oral mucosa, footpads, and haired skin. Dermoepidermal separation occurred in the absence of inflammation or cytolysis of basal epidermal cells. Collagen IV-specific immunostaining corroborated the fact that clefting took place below the epidermal basement membrane. Ultrastructural examination revealed that the proband's anchoring fibrils exhibited a filamentous morphology and were decreased in number compared with those in a normal cat. Finally, the attenuated immunoreactivity for collagen VII in our patient led us to suspect that its encoding gene, COL7A1, could be mutated in this case of feline dystrophic epidermolysis bullosa.

Dystrophic form of inherited epidermolysis bullosa in a dog (Akita Inu).

We report a dog with dystrophic epidermolysis bullosa. This 4-year-old female Akita Inu, a species of Canis familiaris var. japonicus Temminck, had a 3-year-history of ulcers and scars over the pressure areas on the limbs, and dystrophic nails, since the age of 1 year, which corresponds to early adulthood in humans. Electron microscopy of a blister revealed separation beneath the lamina densa, and a reduction in the number of anchoring fibrils. The NC-1 domain of type VII collagen was positively stained with monoclonal antibody LH7.2 at the basement membrane zone. These findings indicate that humans and dogs have a similar response to antibody LH7.2, which may aid the development of an animal model for this disease.