BP180/type XVII collagen: its role in acquired and inherited disorders or the dermal-epidermal junction

Abstract BP180 is a member of the collagen protein family and is also referred to as type XVII collagen or BP antigen 2. It is a transmembrane protein constituent of the dermal-epidermal anchoring complex. The long-held hypothesis that BP180 functions as a cell-matrix adhesion molecule has been supported by recent investigations of human disorders of the dermal-epidermal junction in which BP180 is either genetically defective or targeted by the immune system. In generalized atrophic benign epidermolysis bullosa, mutations of BP180 result in an inherited subepidermal blistering disease. In bullous pemphigoid, herpes/ pemphigoid gestationis, cicatricial pemphigoid, lichen planus pemphigoides and linear IgA disease, autoantibodies are directed to different epitopes on the BP180 ectodomain. Recent molecular investigations have provided new insights, not only into the mechanism of autoantibody-mediated subepidermal blistering, but also into the biochemical structure and cell biological functions of BP180 and other components of the dermal-epidermal anchoring complex. These findings have suggested new directions for the development of diagnostic and therapeutic tools for these autoimmune and genetic diseases. Received: 4 September 1998 / Accepted: 27 October 1998     

Immunofluorescent analysis of the basement membrane zone in lichen planus suggests destruction of the lamina lucida in bullous lesions

Lichen planus is an inflammatory dermatosis which is characterized histologically by an intense lymphocytic infiltrate at the dermal epidermal junction. This frequently results in disruption of the basement membrane zone, occasionally causing clinical blisters. In order to better understand the specific portion of the basement membrane zone which is disrupted by the lymphocytic infiltrate, we examined 7 cases of lichen planus with antibodies directed against anchoring filaments (GB3), the bullous pemphigoid antigen, anchoring fibrils (type VII collagen) and type IV collagen. In lesions without separation at the BMZ, all antibodies were strongly expressed, as in normal skin. In lesions with early separation, there was a focal decrease in GB3 staining, but types VII and IV collagen labelled normally. In lesions resulting in blisters, GB3 staining was essentially absent, and anti-types IV and VII collagen remained, but stained in a disrupted, less discrete pattern. The bullous pemphigoid antigen showed only slight deviation from the normal staining pattern. These findings suggest that the basement membrane zone in lichen planus is disrupted in the lamina lucida region. The lamina densa and sub-lamina clensa zones remain intact even in bullous lesions of lichen planus.     

Evidence for a functional defect of the lamina lucida in recessive dystrophic epidermolysis bullosa demonstrated by suction blisters

Suction blisters were raised in non-lesional skin of sixteen patients with various types of epidermolysis bullosa (EB). The ultrastructural level of separation was found in each type to be within the lamina lucida of the epidermal basement membrane zone. Suction blister times were normal in EB simplex, dominant dystrophic and localized recessive dystrophic EB, but were markedly reduced in both junctional EB and severe generalized recessive dystrophic EB. These data indicate an unpredicted abnormality of adhesion within the lamina lucida in severe recessive dystrophic EB, in addition to the well-recognized defect at a deeper level, beneath the lamina densa, in this condition. The use of suction blister times may prove valuable in the diagnosis of EB.     

Bullous pemphigoid: role of complement and mechanisms for blister formation within the lamina lucida

Bullous pemphigoid (BP), an autoimmune subepidermal blistering skin disease, demonstrates tense blisters with or without widespread erythema, blistering along the lamina lucida, immunoglobulin G and/or complement deposits at the basement membrane zone, and the presence of circulating autoantibodies against hemidesmosomal molecules. These autoantibodies usually react against 180‐kDa and/or 230‐kDa proteins, designated as BP180 and BP230, respectively. The precise blistering mechanisms after autoantibodies bind to antigens are not fully understood. Immune complexes are thought to initially activate the complement cascade, which may induce activation of proteases and/or cytokines and cause dermal–epidermal separation. However, why does separation run specifically within the lamina lucida in a space as narrow as 500 nm wide? This review mainly focuses on the possible mechanisms of BP‐specific blistering and how separation occurs along the lamina lucida, based on existing evidence.     

Key role of heparan sulfate chains in assembly of anchoring complex at the dermal-epidermal junction

Epidermal basement membrane forms anchoring complex composed of hemidesmosomes, anchoring filaments, lamina densa and anchoring fibrils to link epidermis to dermis. However, the anchoring complex is rarely formed in skin equivalent models, probably because of degradation of extracellular matrix (ECM) proteins and heparan sulfate chains by matrix metalloproteinases (MMPs) and heparanase, respectively. To explore the roles of ECM proteins and heparan sulfate in anchoring complex assembly, we used specific inhibitors of MMPs and heparanase, and the formation of anchoring complex was analysed in terms of polarized deposition of collagen VII, BP180 and β4 integrin at the dermal-epidermal junction (DEJ) by means of immunohistochemistry and transmission electron microscopy (TEM). The deposition of collagen VII was polarized to the basal side by the addition of MMP inhibitor, and the staining intensity was increased by combined treatment with MMP inhibitor and heparanase inhibitor, which enhanced anchoring fibril formation as observed by TEM. BP180 was polarized to the basal side by heparanase inhibitor, which protects HS chains, but not by MMP inhibitor. MMP inhibitor improved the polarization of β4 integrin. Hemidesmosomes were formed in the presence of each inhibitor, as observed by TEM, and formation was greatly enhanced by the combined treatment. These findings suggest that heparan sulfate chains, in addition to ECM proteins at the DEJ, play an important role in the assembly of anchoring complex, especially hemidesmosomes and anchoring fibrils.     

Influence of heparan sulfate chains in proteoglycan at the dermal-epidermal junction on epidermal homeostasis

Basement membrane (BM) plays important roles in skin morphogenesis and homeostasis by controlling dermal-epidermal interactions. However, it remains unclear whether heparan sulfate (HS) chains of proteoglycan in epidermal BM contribute to epidermal homeostasis. To explore the function of HS chains at the dermal-epidermal junction (DEJ), we used a skin equivalent (SE) model. This model lacked HS at the DEJ and showed abnormal expression of the differentiation markers filaggrin and loricrin; similar changes were seen in ultraviolet B-irradiated human skin. Perlecan (core-protein of HS proteoglycan) remained localized at the DEJ in both SE and UV-irradiated human skin. Heparanase, which degrades HS, was increased in epidermis of UV-irradiated skin, compared with unirradiated skin. We found that deposition of HS at the DEJ in the SE model was markedly augmented by a synthetic heparanase inhibitor, and release of HS into conditioned medium was suppressed. The inhibitor also increased filaggrin and loricrin expression. Moreover, the recovery of HS was associated with an increase of Ki67-positive basal cells, compared with control SE cultured without inhibitor. Comparative gene expression analysis in epidermis of SE cultured in the presence and absence of heparanase inhibitor, using DNA microarrays, showed that recovery of HS was associated with increased expression of differentiation-related genes and down-regulation of degradation-enzyme-related genes. These results indicate that degradation of HS at the DEJ by heparanase impairs epidermal homeostasis in SE, leading to abnormal differentiation and proliferation behaviour. Thus, HS chains in epidermal BM appear to play an important role in epidermal homeostasis.     

Heterogeneity of Brunsting-Perry type pemphigoid: a case showing blister formation at the lamina lucida, immune deposition beneath the lamina densa and autoantibodies against the 290-kD polypeptide along the lamina densa

An otherwise healthy 31-year-old man presented with multiple, vesicular, subepidermal blistering on the head, face, chest and oral cavity, leaving shallow scar formation, typical of Brunsting-Perry type pemphigoid. Direct immunofluorescence showed linear deposition of immunoglobulin (Ig)G and C3 along the basement membrane zone (BMZ), and indirect showed anti-BMZ autoantibodies (IgG, >40×) reacting with the dermal side under the salt-split study. Immunofluorescence staining for type IV collagen and laminins, as well as routine electron microscopy, demonstrated that the cleavage level of the blister was intra-lamina lucida. The immunoperoxidase method applied to lesional skin demonstrated IgG deposits along the lamina densa. The post-embedding immunogold method demonstrated that the autoantibodies against BMZ reacted with the lamina densa and the dermis just beneath it. Immunoblot studies demonstrated that the autoantibodies reacted with the 290-kD polypeptide (suggesting type VII collagen) when dermal extract was used as the substrate. The patient was treated with combination therapy consisting of 30 mg prednisolone, 900 mg nicotinamide and 750 mg tetracycline, and the number of newly forming blisters decreased. We concluded that Brunsting-Perry type pemphigoid, a rare autoimmune blistering disease, includes cases showing characteristics of epidermolysis bullosa acquisita as well as bullous pemphigoid. This case showed discrepancy between the blistering level (intra-lamina lucida) and location of antigen (lamina densa and sub-lamina densa areas).     

Ultrastructure of the epidermal-dermal junction and cutaneous basal lamina in Bowen's disease

Ten tissue sections from 10 examples of Bowen's disease were excised from paraffin blocks, rehydrated, and incubated in 90% formic acid at 45°C for 18 h. The epidermis was gently removed with the aid of a dissecting microscope, and the remaining dermis with attached basal lamina was processed for scanning electron microscopy. This surface showed a well-preserved basal lamina. The dermal papillae in the areas of Bowen's disease were elongated and had frequent secondary protrusions. The normal 0.5 μ, interconnecting corrugations were often replaced by either broad, coarse corrugations or by large areas of smooth-to-undulating basal lamina. This study demonstrates marked alterations in spatial interactions between neoplastic epidermis and underlying dermis in Bowen's disease.     

A direct method to determine the strength of the dermal-epidermal junction in a mouse model for epidermolysis bullosa

Epidermolysis bullosa (EB) describes a spectrum of rare, incurable, inherited mechanobullous disorders unified by the fact that they are caused by structural defects in the basement membrane zone which disrupt adhesion between the epidermis and dermis. Mouse models provide valuable tools to define the molecular basis of these diseases and to test novel therapeutic approaches. There is need for rapid, quantitative tests that measure the integrity of dermal-epidermal adhesions in such models. To address this need, we describe a novel quantitative method to determine the mechanical strength of the adhesion between tail skin epidermis and dermis. We show that this test reliably measures the force required to cause dermal-epidermal separation in tails of mice that are genetically predisposed to a form of non-Herlitz Junctional EB which develops as the result of a hypomorphic mutation in the laminin gamma 2 gene (Lamc2(jeb) ). This simple, quantitative method of directly measuring the tensile strength of dermal-epidermal adhesion provides a novel dimension to the pathophysiological screening, evaluation, and therapeutic treatment of mice that may develop progressive forms of EB and potentially other disorders that compromise cutaneous integrity.     

Salt split technique: a useful tool in the diagnosis of subepidermal bullous disorders

Background:

Direct immunofluorescence (DIF) is the gold standard in the diagnosis of immunobullous diseases. However, it cannot reliably differentiate various subtypes of subepidermal immune- bullous diseases (SIBD). Salt split technique (SST) could be used under such circumstances to differentiate them. There is paucity of reports in the Indian literature regarding the SST.

Aim:

This study was designed to evaluate the utility of direct SST in subepidermal blistering diseases.

Materials and Methods:

Fourteen clinically diagnosed cases of subepidermal blistering diseases were included in the study. Two perilesional punch biopsies were taken one each for DIF and salt split study.

Results:

Linear basement membrane zone band with IgG and/or C₃ was seen in 14 cases of patients BP. Salt split study showed epidermal or mixed pattern of deposits in 12 patients and exclusive floor pattern in two patients. The diagnosis was revised in these two patients to epidermolysis bullosa acquisita.

Conclusion:

SST is a simple, inexpensive procedure and should be routinely employed in the diagnosis of subepidermal bullous diseases.     

Papillomavirus-induced changes in the topography of the epidermal-dermal junction: a scanning electron microscopic study of basal lamina

Papillomavirus infects human epidermal cells and causes verruca vulgaris, which is characterized by altered epidermal growth rates and differentiation patterns. There is also a prominent induced dermal proliferative response. Ten formalin-fixed skin specimens from 10 patients, each with histologically characteristic verruca vulgaris, were excised from paraffin blocks, deparaffinized in xylene, and rehydrated in graded ethanol solutions. Hydrated specimens were incubated in 90% formic acid at 45 degrees C for 18 h. Using a dissecting microscope the epidermis was gently separated from the dermis, which then was fixed in 3% glutaraldehyde and processed for scanning electron microscopy. Scanning electron microscopy showed a well-preserved epidermal-dermal junction (EDJ) covered by basal lamina. In comparison with normal EDJ, the EDJ of verruca vulgaris showed markedly elongated dermal papillae. Some papillae were broad and thin, and others were broad and thick. Irregularly shaped and branching secondary papillae were common. Also, there were changes in the basal lamina: the normal corrugation pattern was replaced by relatively smooth undulations or coarse, vertically oriented ridges on papillae. Papillary tips were often smooth. This study shows that there were prominent alterations in the topography of the EDJ and basal lamina in verruca vulgaris. This technique should be useful in evaluating epithelial-connective tissue morphologic interactions in formalin fixed archival tissue in other diseases characterized by alterations in epidermal growth rates and differentiation patterns.     

Ontogeny of structural components at the dermal-epidermal junction in human embryonic and fetal skin: the appearance of anchoring fibrils and type VII collagen

The ontogeny and composition of the dermal-epidermal junction (DEJ) in developing human embryonic and fetal skin was studied at progressive stages of gestation by immunofluorescence microscopy and immunocytochemistry using transmission electron microscopy (TEM). The DEJ of embryonic skin at 5 weeks estimated gestational age (EGA) was a simple basement membrane zone limited to the basal cell plasma membrane, lamina lucida, and lamina densa. A network of reticular collagen fibrils (reticular lamina) was deposited beneath the lamina densa by 6 weeks. Coincident with the onset of increased complexity in epidermal and dermal structure, at the time of the embryonic to fetal transition, the DEJ displayed additional components that were markers of maturation. At 7-8 weeks EGA, fine filamentous structures extended from the DEJ into the reticular lamina. By 9 weeks EGA hemidesmosomes and banded anchoring fibrils were recognizable, although distributed sparsely at the DEJ. With increasing gestational age, these structures displayed greater electron density and structural completeness. By the end of the first trimester, the DEJ appeared ultrastructurally similar to that of mature skin. Weak immunofluorescent labeling demonstrated the presence of type VII collagen at the DEJ by 8 weeks EGA. From 10-12 weeks EGA immunofluorescent labeling of the DEJ for type VII collagen was distinctly punctate, while immunoperoxidase labeling observed by TEM was linear, continuous, and sublamina densa in position. With ongoing gestation the immunofluorescent labeling became increasingly stronger at the DEJ. Thus, type VII collagen was present at the DEJ in the zone immediately beneath the lamina densa, before the appearance of mature anchoring fibrils but coordinate with the appearance of fine filamentous, unbanded structures, and appeared to increase with the development and accumulation of anchoring fibrils.     

炎性细胞在大疱性类天疱疮发病机制中的作用研究进展

大疱性类天疱疮(BP)是由抗半桥粒抗原BPl80和BP230抗体所导致的自身免疫性水疱性疾病.补体活化、肥大细胞脱颗粒、中性粒细胞聚集等都在BP的发病机制中起着重要的作用.该文就炎性细胞在大疱性类天疱疮发病机制中的作用作一综述.