A Novel Subepidermal Blistering Disease with Autoantibodies to a 200-kDa Antigen of the Basement Membrane Zone

Abstract

Several components of the basement membrane zone (BMZ) have been identified as antigenic targets in autoimmune bullous diseases. We report a novel disease with autoantibodies to a BMZ antigen that is different from the targets described so far. The patient suffering from this disorder showed tense bullae and severe mucous membrane involvement rapidly responding to oral tetracyclines and colchicine. Histopathologic findings resembled those of dermatitis herpetiformis. Direct immunofluorescence microscopy showed linear deposits of IgG and C3 at the BMZ. By indirect immunofluorescence studies on split human skin, using both 1 M NaCl and suction blistering for dermal-epidermal separation, IgG antibodies localized exclusively to the dermal side of the split. The antibodies were mainly of the IgG4 sub-class. By Western blot analysis of epidermal and dermal extracts, the patient's serum unequivocally reacted with a dermal antigen of 200 kDa. It did not recognize bullous pemphigoid antigens (the autoantigen of epidermolysis bullosa acquisita), purified preparations of laminin-1 and laminin-5, or the recently described 105-kDa BMZ antigen. By immunoblotting of concentrated conditioned SCC-25 medium, the patient's antibodies reacted with a band of 200 kDa and several bands of lower molecular weight. No reactivity was seen with extracts of cultured human fibroblasts. By indirect immunogold electron microscopy, immunoreactants localized to the lower lamina lucida. After clearance of skin lesions, both indirect immunofluorescence and Western blot analysis became negative. This patient suffers from a novel autoimmune bullous disease with autoantibodies to a 200-kDa antigen of the BMZ.

Profibrotic Phenotype of Conjunctival Fibroblasts from Mucous Membrane Pemphigoid

“…We have previously observed that normal conjunctival fibroblasts do not contract collagen in response to TNF␣. 22 In contrast, the addition of IL-13 to the collagen lattice conditioned medium resulted in less contraction by pemphigoid fibroblasts compared with normal fibroblasts, relative to the amount of contraction in the SFM negative control ( Figure 7D). …”

“…10 The diagnosis of MMP was based on clinical presentation with progressive conjunctival cicatrization and positive direct immunofluorescence microscopy of the conjunctiva or other tissues (oral mucosa, genital mucosa, and skin) showing linear deposition of IgG and/or IgA and/or C3 at the BMZ (Figure 1), and/or positive circulating anti-epithelial BMZ autoantibodies. 19 Serological testing for anti-epithelial BMZ autoantibodies was carried out by i) indirect immunofluorescence microscopy on human salt-split skin (for IgG and IgA), ii) enzyme-linked immunosorbent assay using the immunodominant NC16A domain of BP180 (type XVII collagen) as target antigen (MBL, Nagoya, Japan), iii) immunoblotting with the linear IgA dermatoses antigen 1 (LAD-1) representing the soluble ectodomain of BP180 generated from the concentrated supernatant of cultured human keratinocytes, 20 iv) immunoblotting with the extracellular matrix of cultured human keratinocytes for anti-laminin 332 reactivity, 21 v) immunblotting with extract from human dermis for antibodies to the p200 antigens and type VII collagen, 22 and vi) immunoblotting with the recombinant NC1 domain of type VII collagen, the immunodominant region in epidermolysis bullosa acquisita. Compared with indirect immunofluorescence microscopy, which detects circulating autoantibodies in about half the MMP sera, the combined testing for anti-NC16A and anti-LAD-1 autoantibodies has been shown previously to increase sensitivity to more than 80% in MMP sera.…”

B Cells, Dendritic Cells, and Macrophages Are Required To Induce an Autoreactive CD4 Helper T Cell Response in Experimental Epidermolysis Bullosa Acquisita

“…Sixmicrometer frozen skin sections were incubated with biotin-conjugated Ly-6G (1:200 dilution) Ab at 37˚C for 1 h. Subsequently, sections were incubated with Ultra-Sensitive ABC Peroxidase Staining Kit (Thermo Scientific Pierce Protein Research Products) at room temperature (RT) for 0.5 h and were developed by 3,39-diaminobenzidine (Merck, Darmstadt, Germany). Dermal extracts of murine skin were prepared as described (34).…”

Clinical features and diagnosis of epidermolysis bullosa acquisita

“…By indirect IF microscopy of human salt-split skin, antitype VII collagen antibodies label the floor of the artificial blister ( Figure 7) [138]. However, antibodies against laminin 332 and the p200 protein/ laminin γ1 also bind to the floor of the blister [3,127,139,140]. In latter two diseases, however, direct IF microscopy reveals an n-serrated pattern.…”