Variation of COL7A1 gene in dystrophic epidermolysis bullosa pruriginosa / 中华医学遗传学杂志

OBJECTIVE@#To perform gene mutation analysis in a Chinese pedigree with dystrophic epidermolysis bullosa pruriginosa (DEB-Pr), and explore phetotype, genotype, and genotypes-phenotypes relationship of DEB-Pr.@*METHODS@#Potential variants of the COL7A1 gene were detected by skin targeted sequencing panel and verified by Sanger sequencing. The pathogenicity of the variation was analyzed.@*RESULTS@#Compound heterozygous variants, c.4128delT and c.8234G>A, were detected in the COL7A1 gene of the two patients. The c.4128delT(p.Pro1376fs) variant was derived from their mother and unreported previously. According to the American College of Medical Genetics and Genomics Standards and Guidelines, it was suggested to be a pathogenic mutation. The c.8234G>A(p.Arg2745Gln) variant was derived from their father, and possibly is a pathogenic variation.@*CONCLUSION@#In this study, the compound heterozygous variants of c.4128delT(p.Pro1376fs) and c.8234G>A(p.Arg2745Gln) of the COL7A1 gene probably underlies the disease in this patient and his sister. And our study expands the database on mutations of DEB-Pr.

Genetic analysis of a child with recessive dystrophic epidermolysis bullosa due to compound heterozygous variants of (COL7A1 gene / 中华医学遗传学杂志

OBJECTIVE@#To carry out genetic testing and prenatal diagnosis for a family affected with recessive dystrophic epidermolysis bullosa (RDEB).@*METHODS@#All exons of the COL7A1 gene and their flanking regions were subjected to PCR and Sanger sequencing. Suspected variant was validated in family members, based on which prenatal diagnosis was provided.@*RESULTS@#Sanger sequencing found that the proband has carried two variants of the COL7A1 gene, namely c.7289delC (p.Pro2430Glnfs*36) and c.7474C>T (p.Arg2492*), which were respectively derived from his mother and father. The same variants were not found among 100 healthy controls. By prenatal diagnosis, the fetus was found to have inherited the c.7474C>T (p.Arg2492*) variant from its father.@*CONCLUSION@#The pathogenic variants of the COL7A1 gene of the RDEB family were clarified, based on which prenatal diagnosis was provided.

Reproductive alternatives for patients with dystrophic epidermolysis bullosa / Opções reprodutivas para pacientes com epidermólise bolhosa distrófica

ABSTRACT Epidermolysis bullosa describes a group of skin conditions caused by mutations in genes encoding proteins related to dermal-epidermal adhesion. In the United States, 50 cases of epidermolysis bullosa per 1 million live births are estimated, 92% of which classified as simplex, 5% dystrophic, 1% junctional and 2% non-classified. Dystrophic epidermolysis bullosa is associated with autosomal, dominant and recessive inheritance. Epidermolysis bullosa causes severe psychological, economic and social impacts, and there is currently no curative therapy, only symptom control. Embryonic selection is available for epidermolysis bullosa patients in order to prevent perpetuation of the condition in their offspring.

RESUMO O termo "epidermólise bolhosa" descreve um grupo de afecções cutâneas causadas por mutações em genes que codificam proteínas relacionadas à aderência dermoepidérmica. Nos Estados Unidos, estima-se a ocorrência de 50 casos de epidermólise bolhosa por 1 milhão de nascidos vivos, sendo 92% deles da forma simples, 5% da forma distrófica, 1% da forma juncional e 2% não classificados. A epidermólise bolhosa do tipo distrófica foi associada a padrões autossômicos, dominante e recessivo. A epidermólise bolhosa causa sérios impactos psicológicos, econômicos e sociais, e não há tratamento curativo atualmente − apenas controle dos sintomas. A seleção embrionária é disponível para portadores de epidermólise bolhosa, a fim de evitar a perpetuação da condição em seus descendentes.

A Case of Epidermolysis Bullosa Acquisita Associated with Psoriasis / 대한피부과학회지

Epidermolysis bullosa acquisita (EBA) is an autoimmune subepidermal blistering disease characterized by the presence of circulating IgG autoantibodies to type VII collagen. Various types of autoimmune blistering disease have been reported in association with psoriasis. A 58-year-old man with a 5-year history of psoriasis vulgaris presented with painful and mildly pruritic erythematous multiple bullae and vesicles. Histopathologically, there was a subepidermal bulla with infiltration of inflammatory cells composed of neutrophils and eosinophils. The salt-split skin indirect immunofluorescence test showed IgG binding to the dermal side of the separation, and immunoblotting using normal human dermal extract revealed antibodies directed against a 290-kDa polypeptide. He was diagnosed with EBA and started medication of oral prednisolone and mycophenolate mofetil. Skin lesions were continuously regressed. Of all the autoimmune blistering diseases coexisting with psoriasis, bullous pemphigoid is the most frequent. However, a few cases of EBA associated with psoriasis have been reported in the literature. We report a rare case of EBA coexisting with psoriasis vulgaris.

Scanning electron microscopy of a blister roof in dystrophic epidermolysis bullosa / Microscopia eletronica de varredura do teto de uma bolha de epidermolise bolhosa distrofica

In dystrophic epidermolysis bullosa the genetic defect of anchoring fibrils leads to cleavage beneath the basement membrane, with its consequent loss. We performed scanning electron microscopy of an inverted blister roof of a case of dystrophic epidermolysis bullosa, confirmed by immunomapping and gene sequencing. With a magnification of 2000 times a net attached to the blister roof could be easily identified. This net was composed of intertwined flat fibers. With higher magnifications, different fiber sizes could be observed, some thin fibers measuring around 80 nm and thicker ones measuring between 200 and 300 nm.

Na epidermólise bolhosa distrófica, o defeito genético das fibrilas ancorantes leva à clivagem abaixo da membrana basal, com sua consequente perda. Realizamos microscopia eletrônica de varredura do teto invertido de uma bolha de um caso de epidermólise bolhosa distrófica, cujo diagnóstico foi confirmado com imunomapeamento e com sequenciamento gênico. Com uma ampliação de 2.000 vezes, pôde ser facilmente identificada uma rede ligada ao teto da bolha. Essa rede era composta por fibras achatadas e entrelaçadas. Com grandes aumentos, fibras de diferentes tamanhos puderam ser observadas: algumas finas, medindo cerca de 80 nm, e outras mais largas, medindo entre 200 nm e 300 nm.

Transient Bullous Dermolysis of the Newborn / 대한피부과학회지

Transient bullous dermolysis of the newborn (TBDN) is a rare subtype of the dystrophic epidermolysis bullosa characterized by blistering at birth which improves spontaneously during early life. Electron microscopy showed sublamina densa separation with dilated rough endoplasmic reticulum and electron dense inclusions. Immunofluorescence mapping using anti-type VII collagen antibody showed widespread intraepidermal type VII collagens which are a characteristic finding of TBDN. Here, we report two cases of TBDN presenting typical clinical manifestations, electron microscopy findings, and immunofluorescence mapping results. The skin lesions of both patients healed spontaneously 2~3 months later.

Type VII Collagen Gene Mutations (c.8569G>T and c.4879G>A) Result in the Moderately Severe Phenotype of Recessive Dystrophic Epidermolysis Bullosa in a Korean Patient

Dystrophic epidermolysis bullosa (DEB) are caused by mutations in the COL7A1 gene, which encodes type VII collagen. Even though more than 500 different COL7A1 mutations have been identified in DEB, it still remains to be under-investigated. To investigate the mutation of COL7A1 in moderately severe phenotype of recessive DEB (RDEB) in a Korean patient, the mutation detection strategy was consisted of polymerase chain reaction (PCR) amplification of genomic DNA, followed by heteroduplex analysis, nucleotide sequencing of the PCR products demonstrating altered mobility. In this study, we found that one mutation (c.8569G>T) was detected within exon 116. The mutation of c.8569G>T in exon 116 changed the GAG (Glu) to TAG, eventually resulted in premature termination of type VII collagen polypeptide. Furthermore the mother did not have the mutation c.8569G>T in exon 116. The other novel mutation (c.4879G>A) was detected within exon 51 of both patient and mother, thereby resulting in changing valine (Val) to isoleucine (Ile) in type VII collagen polypeptide. Taken together, in this study we identified compound heterozygosity for COL7A1 mutations (c.8569G>T and c.4879G>A) in moderately severe RDEB in a Korean patient. We hope that this data contribute to the expanding database on COL7A1 mutations in DEB.

Epidermólisis ampollar distrófica dominante generalizada severa / Dominant dystrophic epidermolysis bullosa severe generalized

La epidermólisis ampollar distrófica (EAD) es una genodermatosis que se caracteriza por la aparición de ampollas en respuesta al trauma friccional, que luego dejan cicatrices extensas y milia. En ella, la formación de ampollas ocurre por alteraciones morfológicas y cuantitativas en las fibrillas de anclaje en la sublámina densa, cuyo principal componente es el colágeno tipo VII. Comunicamos un caso de EAD en un niño de 7 años que presentaba múltiples cicatrices atróficas, erosiones sobre base eritematosa, algunas con costras en su superficie y numerosas ampollas con contenido serohemorrágico en todo el tegumento. En la cavidad oral se evidenciaban múltiples erosiones, caries y microstomía. Además, el niño presentaba pérdida completa de las uñas y pseudosindactilia de manos y pies. El estudio histopatológico de una lesión ampollar mostró piel desprovista de revestimiento epidérmico. El resultado del inmunomapeo evidenció fluorescencia en el lado epidérmico para todos los anticuerpos testeados: antígeno del penfigoide ampollar, laminina, colágeno IV y colágeno VII; patrón sugestivo de EAD dominante.

Dystrophic epidermolysis bullosa (DEB) is an inherited disease characterized by blistering of the skin following minor trauma that heals with extensive scars and milia. In this disease there are morphologic and quantitative alterations in the anchoring fibrils composed mostly by collagen type VII. We report a case of a 7-year-old boy with multiple atrophic scars, erosions covered by crusts and several serum-hemorrhagic blisters affecting all of tegument. There were multiple oral erosions, caries and microstomia. All nails were missing and presented pseudosyndactyly on the hands and feet. The histopathological exam of a blister revealed skin with absence of the epidermis. The immunomapping study showed positive fluorescence of antibodies against bullous pemphigoid antigen, laminin, collagen IV and collagen VII in the top of the blister (epidermis); suggestive of dominant DEB.

Epidermolysis bullosa pruriginosa: a rare autosomal dominant variant.

A 35 years female presented with extremely pruritic, violaceous, small vesiculopapular lesions over both shins since 11/12 years of age. The intensity of pruritus slightly descreased following oozing of fluid. History of similar incidence in her mother and maternal grandfather was present. There was no toe-nail dystrophy. Histopathology report showed the lesions had hyperkeratotic, mild acanthosis, dermal lymphohistiocytic infiltrate and subepidermal cleft. The case was diagnosed to be a case of epidermolysis bullosa pruriginosa.

Histologic characteristics of in vitro constructed skin basement membrane / 中华烧伤杂志

<p><b>OBJECTIVE</b>To observe the histological characteristics of constructed basement membrane in tissue-engineered skin.</p><p><b>METHODS</b>Forskins from circumcision in normal children were obtained with informed consent of the parents, and then the epidermal keratinocytes (KC) and dermal fibroblasts (Fb) were isolated with trypsin and collagenase D digestion in sequence. Tissue engineered skin with composite chitosan was maintained in a submerged state for 3 days, and then at the air-liquid interface. The tissue-engineered skins were fixed in neutral formalin and then embedded in paraffin after culture for 7, 10 and 15 days, respectively for immunohistological examination of the basement membrane component,including the condition of collagen type IV (COL-IV), collagen type VII (COL-VII), and laminin (LN).</p><p><b>RESULTS</b>HE staining showed that the keratinocytes formed a fine stratified squamous epithelium with the presence of basal, spinous, granular and corneous cell layers, and there was various amount of cells in flat and fusiform shape in each layer. It was found that a regular red staining strip situated at the dermal epidermal junction. Positive staining of collagen IV, collagen VII as well as LN was observed by immunohistological examination.</p><p><b>CONCLUSION</b>The results suggest that the composite chitosan tissue engineered skin has a good prospect for clinical use because it presents a perfect reconstruction of basement membrane.</p>

Las epidermolisis bullosas distróficas en México: 2470insG representa la mutación mas común en 21 familias / 2470insG, represents the commonest mutation in Mexican patients with dystrophic bullous epidermolysis: a study of 21 families

Antecedentes: Las epidermolisis ampollosas congénitas son enfermedades caracterizadas por ampollas en piel y mucosas al mínimo traumatismo. Son tres tipos: simple, unión y distrófica. Las epidermolisis ampollosas distróficas (EAD) son causadas por mutaciones en el gen COL 7Al que codifica la producción del colágeno tipo VII localizado en las fibrillas de anclaje de la unión dermoepidérmica. Objetivo: Determinar las bases moleculares de las EAD en México. Material y métodos: se analizaron ADN de 21 familias mexicanas con EAD. Se realizó reacción en cadena de la polimerasa, estudios de heteroduplex secuenciación de nucleótidos en ADN de los pacientes. Resultados: Se detectó 59 de 67 mutaciones en 36 pacientes. Se encontraron seis mutaciones de tipo codón de terminación prematuro, substitución de glicina, remoción de intrones de novo y depleción interna. La mutación comúnmente más encontrada fue la 2470insG, en 21 (58.35%) de 36 pacientes. Conclusiones: En pacientes con EAD, la mutación 2470insG es la más frecuente en México. Recomendamos analizar esta mutación a Mexicanos con EAD como primera opción. Estos resultados son útiles para clasificar los subtipos de EAD, dar asesoramiento genético, así como para entender un poco más la fisiopatología de esta enfermedad mecano ampollosa.

BACKGROUND: Type VII collagen gene (COL 7 Al) mutations are the cause of dystrophic epidermolysis bullosa (DEB), but most mutations are specific to individual families and there is limited data on the nature of COL 7 Al mutations in certain ethnic populations. OBJECTIVE: To determine the molecular basis of DEB in Mexican patients and describe the most frequent mutation among this ethnic population. METHODS: Most subjects were approached at FUNDACION DEBRA MEXICO AC. Molecular analysis was performed by polymerase chain reaction (PCR) of genomic DNA using COL 7 A l-specific primers, heteroduplex analysis, and direct nucleotide sequencing. RESULTS: Fifty nine of 67 COL 7 Al possible mutations (88%) were identified; 36 individuals (31 recessive, five dominant) from 21 families. Recessive mutations included six frameshift mutations, four silent glycine substitutions and two splice site mutations. CONCLUSIONS: The present study informs a different kind of mutation observed in our patient population. Only two mutations informed in this study had been described earlier among another ethnic group. The most frequent mutation was 2470insG, affecting 21 (58.3%) out of 36 patients with DEB. These new data will be helpful in facilitating the accurate diagnosis of an DEB subtype, and will add further insight into the pathophysiology of this mechanobullous disease.

An Ultrastructural Examination of the Epitope Expression at Each Domain of Type VII Collagen in Patients with Epidermolysis Bullosa Acquisita / 대한피부과학회지

BACKGROUND: Epidermolysis bullosa acquisita (EBA) is an autoimmune disease characterized by circulating IgG autoantibodies which bind to the type VII collagen (C-VII). The major antigenic epitopes in C-VII, to which most EBA autoantibodies react, have been considered to be present in the N-terminal noncollagenous (NC1) domain. However, a novel EBA subgroup was recently identified with circulating antibodies, which target domain(s) other than or along with the NC1 domain of C-VII. These data suggest that there might be some heterogeneity in the autoantibody specificity to bind the domain-oriented epitopes in EBA. OBJECTIVE: The purpose of this study was to determine whether additional or independent epitopes exist in the C- terminal noncollagenous (NC2) and/or collagenous triple-helical (CTH) domain among Korean patients with EBA. METHODS: For this investigation, postembedding, indirect, immunogold electron microscopy was performed with the sera from 10 cases of EBA, having circulating autoantibodies against C-VII. The identification of the epitope and the relevant domain in each case were determined by ultrastructural localization of the immunogold particles. RESULTS: From 10 sera examined, all 10 cases showed deposits of gold particles confined to the area along the lamina densa (LD), without any other pattern of deposition. There was no case which revealed any independent/distinct deposits of the gold particles in the dermis below the LD. The ultrastructural locations of each domain (NC1, on the LD; NC2, 300~360 nm below the LD; CTH, between the area of NC1 and NC2) indicated that the epitopes recognized in all 10 Korean cases of EBA were expressed at the NC1 domain of C-VII. We did not find any additional or independent epitope in other domain. CONCLUSION: The results suggest that there may not be a wide heterogeneity in the domain-oriented topographic expression of antigenic epitopes in EBA; it is highly likely that the major epitopes present in Korean EBA cases reside within the NC1 domain of C7, similar to those observed with white population.

Epidermólise bolhosa distrófica recessiva mitis: relato de caso clínico / Recessive dystrophic epidermolysis bullosa mitis: case report

As epidermólises bolhosas são dermatoses bolhosas congênitas que levam à formação de bolhas espontaneamente ou após trauma. São reconhecidos três grupos de da doença, de acordo com o segundo consenso internacional: simples, juncional e distrófica. Nas formas distróficas, o defeito genético deve-se à mutação no gene COL7A1, responsável pela codificação do colágeno VII, principal constituinte das fibrilas de ancoragem, que participam na aderência da lâmina densa à derme. Os autores relatam o caso de paciente do sexo feminino, de 15 anos, apresentando ulcerações nas pernas, bolhas serosas e lesões atrófico-acastanhadas nos braços e tronco. Foram observadas distrofias ungueais e alterações dentárias, iniciadas a partir do nascimento. O exame histopatológico da bolha revelou quadro compatével com epidermólise bolhosa, que, associado aos dados clínicos, permitiram a classificação do caso na forma distrófica recessiva mitis.

A Case of Cicatricial Pemphigoid with Autoantibodies Against the BP180 and BP230 Antigens / 대한피부과학회지

Cicatricial pemphigoid is a chronic autoimmune subepithelial blistering disease that predominantly involves mucous membrane with resultant scar formation. It may involve oral, ocular, nasal, pharyngeal, laryngeal, esophageal, and anogenital mucous membranes. Cicatricial pemphigoid is a heterogenous group of diseases with respect to the autoimmune target antigens including BP180(type XVII collagen), BP230, laminin 5(epiligrin), laminin 6, type VII collagen and other newly described antigen. We describe a patient with cicatricial pemphigoid in whom circulating IgA and IgG autoantobodies against BP180 and BP230 antigens were detected simunltaneously.

The Effects of Basic Fibroblast Growth Factor(bFGF)on Type I and VII Collagen Gene Expression in Cultured Dermal Fibroblast

BACKGROUND: bFGF, a member of the fibroblast growth factor family, potently induces vascular smooth muscle cell proliferation and decreased synthesis of the collagens. OBJECTIVE: For further investigation of the effect of bFGF on extracellular matrix homeostasis in the skin, we evaluated the expression of type I and type VII collagen gene at the transcriptional levels. METHOD: We examined that recombinant human bFGF affects the expression of genes involved in ECM synthesis and remodeling in human dermal fibroblasts cultures as judged by Northern blot analysis. RESULTS: The steady state levels of type I and VII collagen gene mRNA were decreased with age dependent pattern up to 0.13 and 0.44 folds respectively. The transcriptional levels of type I collagen mRNA were increased by TGF-B, treatment but markedly decreased by bFGF as well as TNF-a. But there were no synergistic effects bFGF and TNF-a on type I collagen gene expression. The levels of type VII collagen gene expression were increased by both bFGF and TGF-B,. The TNF-a showed slightly antagnostic effects on type VII collagen gene expression. CONCLUSION: The type I and VII collagen gene expression in dermal fibroblasts is clearly subjected to modulation by the cytokines including bFGF with uncoordinate regulatory pathway. In addition to its function of vascular proliferation, bFGF also may play a major role in physiologic skin condition and in repair process such as formation of a stable dermoepidermal junction during skin wound healing.

Immunogold Localization of Type VII Collagen in the Adhesion Complex of Pathologic Cornea

Type VII collagen is one of the major structural components of the corneal epithelial adhesion complex. Using the immunogold technique combined with indirect immunofluorescence analysis, the fine structural distribution of type VII collagen was studied in the corneas obtained from 5 enucleated hyman eyes (age range, 1-77 years) including one pathologic cornea from graft rejection. The findings on normal cornea corroborated the results from previous studies. In pathologic cornea from graft rejection, type VII collagen antibodies generated linear and irregular patchy fluorescence staining along the epithelial-stromal interface and immunogold binding to type VII collagen mainly occurred within the undulating lamina densa, more densealy distributed anchoring plaques and anchoring fibrils. The distribution of type VII collagen in pathologic human cornea from graft rejection is similar to normal human cornea. But, in pathologic cornea, type VII collagen is more densely distributed in superficial stroma and forms more extended anchoring network, which may be derived from the increased secretion of the type VII collagen due to the activated basal epithelial cell during healing process.

A Case of Multiple Histiocytomas in a Healthy Person / 대한피부과학회지

Epidermolysis bullosa acquisita(EBA) is a rare chronic subepidermal bullous disease wit,h autoantibodies to type VII collagen. Clinically, EBA usually begins after the age of 50 with pruritic vesicle and blister formation often on traumatized skin and leaves atrophic scars and milia. A 48-year-old woman visited our department with a 5 month history of a generalized pruritic blistering eruption that began on the scalp and spread to the face, trunk, extremities, and lip. The biopsy specimen revealed dermoepidermal separation, infiltration of perivascular mononuclear cells. Direct immunofluorescence(DIF) showed positive linear deposition of IgG at the BMZ. Bullous pemphigoid was diagnosed and treated with prednisolone 20-40mg daily. The skin lesions were improved after 2 week. During follow-up, trauma-induced vesicles occurred frequently and healed with remaining malia and scars. The biopsy specimen of trauma induced vesicles revealed subepidermal blisters and inflammatory cell infiltration in the dermis. DIF of perilesional skin showed positive linear deposition of IgG at the BMZ. DIF of salt split skin showed linear IgG deposits on the dermal floor. EBA was confirmed.

Immunohistochemical Changes of Rabbit Cornea After Excimer Laser Surface Ablation: collagen type III, IV, VI, VII

We performed photorefractive keratectomy(PRK) on 10 rabbit eyes and determined the distribution of collagen type III, IV VI, VII at postoperative 2, 4 and 6 months to examine immunohistochemical changes after PRK. Type III collagen was not found in the normal cornea but strongly detected in the regenerated corneal stroma at all intervals. It was most prominent at 2 months after surgery and then decreased. Type IV collagen was detected in basement membrane in both normal and ablated corneas at all intervals and the staining was more intense in ablatd corneas than in normal cornea. There was no difference of staining intensity among the groups of different intervals. Type IV collagen was found in both normal and healed corneal stroma at all intervals and there was no difference of staining intensity between normal and ablated corneas and among the groups of different intervals. Type VII collagen was observed as a linear continuous band along the basal surface of epithelium in normal cornea. At 2 months after surgery, type VII collagen staining in basement membrane zone became denser than normal cornea, but segmented. At 4 months after surgery, continuous band of collagen type VII staining was observed, but it was less intense than in normal cornea. At 6 months after surgery, the intensity of continuous band of collagen type VII was the same as in normal cornea. This results suggest that the presence of type III collagen in the regenerated cornea may be related to the development of postoperative subepithelial opacity after PRK and the normalization of collagen type IV and VII at postoperative 6 months may mean the complete reestablished of the adhesion of regenerated epithelium and stroma.

Clinical Features of Epidermolysis Bullosa Acquisita

Epidermolysis bullosa acquisita (EBA) is an autoimmune blistering disease of the skin occurring mostly in middle-aged persons with characteristic skin lesions of inflammatory in a vesiculobullae and mechanobullous lesions. Separation of the skin occurs at the dermoepidermal junction (DEJ) initiated by an immune process involving the anchoring fibrils (AF) 764, which have a role in the normal adherence of the epidermis and the dermis. Patients with EBA have autoantibodies of IgG to type VII collagen which is the main component of AF. An electron microscopic. picture of normal DEJ is shown in figure 1, and the antigen site 2011s of this disease (AF) is noted at the upper-most part of the dermis. In EBA, a biopsy specimen shows subepidermal bulla with a variable degree of dermal in-filtrates. Immunofluorescence (IF) demonstrates a linear deposit of IgG. The pattern of immune deposits along the DEJ is similar to that of bullous pemphigoid. However, the linear iing. fashion is thicker and coarser. When examined by the indirect method with a semi-horizontal section of normal human skin substrates the same patterns can be observed: a fine linear deposit with bullous pemphigoid antibodies and a slightly coarser linear pattern with EBA antibodies. With salt-split skin substrates, the serum autoantibodies of IgG are found to be bound only to the dermal side, the AF zone (Fig. 2). This immunopathologic study can provide a diagnostic finding. Transmission electron microscopic examination reveals blister to be localized just beneath the lamina densa, the site of the immune deposit. In immunoblot analysis of the patient's serum against the. dermal extracts, serum antibodies are found to recognize type VII collagen of 290/145 kD (Fig. 3). This is a confirmatory technique (with antibody-positive sera) in the diagnosis of EBA.

Detection of Type VII Collagen mRNA in Scleroderma Fibroblasts / 대한피부과학회지

BACKGROUND: Type VII collagen is a relatively low abundance extracellular matrix protein among the collagenous molecules. Among the minor collagens. type VII collagen has been demon strated by a immunolocalization studies to be component of anchoring fibrils and structures extending perpendicularly from the lamina densa to the upperpapillary dermis. OBJECTIVE: The purpose of his study is to determine the expression of the type VII collagengene in a group of scleroderma patients as compared to normal skin. METHODS: We have examined the levels of type VII collagen mRNA using quantitative reverse transcription PCR and in sit gybridization in scleroderma skin fibroblasts. Immunofluorescent staining with anti-type VII collan antibody was performed in vitro and in vivo to evaluate the expression of type VII collagen at protein level. RESULTS: 1. the ratio of type VII collagen/GAPDH RT-PCR product were 63.3+/-15.3 in scleroderma and 21.7+7.6 in normal fibroblasts by RT-PCR. 2. The expression of type VII collagen mRNA was considerably lower than type I in scleroderma. A few positive signals by in situ hybridization with type VII collagen cDNA were shown in the dermis. 3. The staining was markedly enhanced in scleroderma fibroblasts and tissues compaired with normal subjects in imunofluorescent staining with anti-Type VII collagn antibody. CONCLUSION: RT-PCR and immunofluorescent staining with antibodies to type VII collagen shows enhanced gene expression in scleroderma skin fibroblasts These data suggest that type VII collagen may be the main soruce of the sclerotic change of skin in scleroderma.