Study On Gene Mutation Of Two Dystrophic Epidermolysis Bullosa Pedigrees

Dystrophic epidermolysis bullosa (DEB) is a subtype of inherited epidermolysis bullosa characterized by its blister formation located below the lamina densa. DEB can be autosomal dominant or autosomal recessive, based on its inheritance pattern. With the development of molecular biology, it becomes evident that DEB is caused by mutations in typeVII collagen gene which encodes one of the important components of the anchoring fibrils.Objective: Study on gene mutation of two dystrophic epidermolysis bullosa pedigrees.Method: The dominant DEB of the two pedigrees was diagnosed by using pathology, electron microscopy and immunofluorescence examinations. PCR and DNA sequencing were performed for the identification of mutations in typeVDcollagen gene. Mutations detected were also confirmed by using the restriction enzyme reaction and five polymorphic microsatellite sequences at loci D3S1358, D20S161, D5S818, D17S1293, CSFIPO analysis.Result:(1) Ultrastructural analyses revealed that pathological cleft was located below the lamina densa, but anchoring fibrils was normal.(2) Bright linear deposition along dermal-epidermal junction was shown by immunochemistry using FITC-conjugated monoclonal antibody LH7:2 .(3) Two mutations were found in typeVDcollagen gene in the two pedigrees by using PCR and direct DNA sequencing: G6240A in exon 73 resulting in amino acid codon change of G2043R in the first pedigree, and G6376A in exon 75 resulting in amino acid codon change of G2088E in the second pedigree.(4) G6240A and G6376A mutations cause changes of restriction enzyme sites. By using the restriction enzyme reaction and microsatellite analysis, G6240A in the first pedigree was proved to be a de novo mutation. In the second pedigree, the patient and his father brought the same G6376E mutation, but other members in this pedigree were unaffected.Conclusion: Both G6240A and G6376A are pathogenic mutations. In the first pedigree, G6240A was proved to be a de novo mutation. In the second pedigree, G6376A mutation of the patient was apparently inherited from his father.