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.
Basement Membrane ; Collagen Type III* ; Collagen Type IV ; Collagen Type VII ; Collagen* ; Cornea* ; Corneal Stroma ; Epithelium ; Immunohistochemistry ; Lasers, Excimer* ; Photorefractive Keratectomy

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.
Antibodies ; Collagen Type VII* ; Cornea* ; Epithelial Cells ; Fluorescence ; Fluorescent Antibody Technique, Indirect ; Graft Rejection ; Humans ; Immunohistochemistry

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.
Collagen Type VII/genetics* ; Epidermolysis Bullosa Dystrophica/genetics* ; Female ; Humans ; Male ; Mutation ; Pedigree ; Phenotype

OBJECTIVETo observe the histological characteristics of constructed basement membrane in tissue-engineered skin.

METHODSForskins 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).

RESULTSHE 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.

CONCLUSIONThe 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.

Basement Membrane ; cytology ; Cells, Cultured ; Child ; Chitosan ; metabolism ; Collagen Type IV ; metabolism ; Collagen Type VII ; metabolism ; Humans ; Laminin ; metabolism ; Organ Culture Techniques ; Skin, Artificial ; Tissue Engineering ; methods

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.
Autoantibodies* ; Blister ; Cicatrix ; Collagen Type VII ; Humans ; Immunoglobulin A ; Immunoglobulin G ; Laminin ; Mucous Membrane ; Pemphigoid, Benign Mucous Membrane*

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.
Antibodies ; Autoantibodies ; Autoimmune Diseases ; Collagen ; Collagen Type VII* ; Dermis ; Epidermolysis Bullosa Acquisita* ; Epidermolysis Bullosa* ; Epitopes ; Humans ; Immunoglobulin G ; Microscopy, Electron ; Population Characteristics ; Sensitivity and Specificity

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.
Antibodies ; Collagen ; Collagen Type VII* ; Dermis ; DNA, Complementary ; Extracellular Matrix ; Fibroblasts* ; Gene Expression ; Humans ; In Situ Hybridization ; Polymerase Chain Reaction ; Reverse Transcription ; RNA, Messenger* ; Skin

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.
Blister ; Collagen ; Collagen Type VII ; Electrons ; Endoplasmic Reticulum, Rough ; Epidermolysis Bullosa Dystrophica ; Fluorescent Antibody Technique ; Humans ; Infant, Newborn ; Microscopy, Electron ; Parturition ; Skin

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.
Blotting, Northern ; Cell Proliferation ; Collagen Type I ; Collagen Type VII ; Collagen* ; Cytokines ; Extracellular Matrix ; Fibroblast Growth Factors ; Fibroblasts* ; Gene Expression* ; Homeostasis ; Humans ; Methods ; Muscle, Smooth, Vascular ; RNA, Messenger ; Skin ; Wound Healing

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.
Antibodies ; Autoantibodies ; Blister ; Collagen Type VII ; Eosinophils ; Epidermolysis Bullosa Acquisita* ; Epidermolysis Bullosa* ; Fluorescent Antibody Technique, Indirect ; Humans ; Immunoblotting ; Immunoglobulin G ; Middle Aged ; Neutrophils ; Pemphigoid, Bullous ; Prednisolone ; Psoriasis* ; Skin