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Cicatricial pemphigoid is a chronic subepithelial autoimmune blistering disease of mucous membranes and skin. Recently, a subtype of cicatricial pemphigoid with autoantibodies to epiligrin was identified. We describe a Taiwanese patient who presented with ocular, oral, and cutaneous involvement. Direct immunofluorescence showed IgG and C3 deposition in epidermal basement membrane; indirect immunofluorescence showed circulating IgG autoantibodies reactive with the dermal side of 1 mol/L sodium chloride-split skin. Immunoblotting of laminin 5 isolated from the extracellular matrix of cultured human keratinocytes showed no specific reactivity. In contrast, with immunoprecipitation of the conditioned culture media from biosynthetically radiolabeled human keratinocytes, this patient's serum clearly reacted with a series of disulfide-linked polypeptides that correspond to laminin 5(alpha3beta3gamma2) and laminin 6(alpha3beta1gamma1). This is the first confirmed case of a patient of Chinese ancestry with this disease entity.

Patients with one form of cicatricial pemphigoid have IgG antibasement membrane autoantibodies against laminin 5 (alpha3beta3gamma2). Although passive transfer of rabbit anti-laminin 5 IgG to neonatal mice has been shown to induce subepidermal blisters that mimic those in patients, it has not been possible to directly assess the pathogenic activity of human autoantibodies in this animal model because the latter do not bind murine skin. To address this question, a disease model in adult mice as well as SCID mice bearing human skin grafts was developed. Adult BALB/C mice challenged with rabbit anti-laminin 5 IgG developed, in a concentration-related fashion, erythema, erosions, and crusts surrounding injection sites, histologic evidence of noninflammatory, subepidermal blisters, and deposits of rabbit IgG and murine C3 in epidermal basement membranes. Anti-laminin 5 IgG also induced subepidermal blisters in: adult complement-, mast cell-, and immuno-deficient mice; adult BALB/C mice pretreated with dexamethasone; and human skin grafts on SCID mice. Alterations did not develop in matching controls challenged with identical amounts of purified normal rabbit IgG or bovine serum albumin. Using this adult mouse model, human skin grafts on SCID mice were challenged with purified IgG from patients with alpha subunit-specific, anti-laminin 5 autoantibodies, or normal controls. Patient (but not control) IgG induced epidermal fragility as well as noninflammatory, subepidermal blisters in grafted human (but not adjacent murine) skin. Moreover, whereas all mice that received patient autoantibodies had anti-laminin 5 IgG in their circulation, deposits of human IgG were present only in the epidermal basement membranes of grafts. Interestingly, these in situ and circulating autoantibodies were predominately of the IgG4 subclass. These studies demonstrate that human anti-laminin 5 autoantibodies are pathogenic in vivo and describe an animal model that can be used to define disease pathomechanisms and biologically important domains within this autoantigen.

Sera from 20 patients with antiepiligrin cicatricial pemphigoid were studied to define the specific reactivity of their IgG autoantibodies. IgG from all patients bound exclusively to the dermal side of 1 mol/L NaCl split skin and immunoprecipitated laminin 5 (alpha3beta3gamma2) from extracts of human keratinocytes (HKs). Immunoblot studies on purified laminin 5 subunits demonstrated that patient IgG bound alpha3 alone in 16 patients. In two patients, IgG autoantibodies were directed predominantly to the gamma2 subunit, yet showed trace reactivity to alpha3 as well. Sera from two patients did not immunoblot any laminin 5 subunits, their IgG presumably immunoprecipitating laminin 5 via a conformational epitope. Sera from patients with alpha3 subunit-specific IgG immunoprecipitated all subunits of laminin 5 as well as polypeptides of 190 and 200 kDa from the conditioned media of HKs. Preclearance studies and experiments utilizing affinity-purified patient IgG demonstrated that the latter signified laminin 6 (alpha3beta1gamma1) that was bound by cross-reactive alpha3 subunit-specific patient IgG. Sera from patients with gamma2 subunit-specific IgG showed no reactivity to laminin 6, except for faint reactivity provided by low levels of their alpha3 subunit-specific IgG. Taken together, these findings indicate that antiepiligrin cicatricial pemphigoid signifies an autoimmune response to subunits present in laminin 5.

A 48-year-old woman with anti-epiligrin cicatricial pemphigoid (CP) who showed clinical features resembling epidermolysis bullosa acquisita was found to have adenocarcinoma of the stomach. Histological examination of lesional skin demonstrated a subepidermal blister. Direct immunofluorescence microscopy of perilesional skin revealed linear deposits of IgG and C3 at the basement membrane zone. The patient's serum contained IgG autoantibodies that bound to the dermal side of 1 mol/L NaCl-split normal human skin as determined by indirect immunofluorescence microscopy, and the lamina lucida as determined by indirect immunoelectron microscopy. The patient's serum immunoprecipitated laminin-5 from extracts and media of biosynthetically radiolabelled human keratinocytes. Immunoblot studies showed that the patient's autoantibodies specifically bound the alpha3 subunit of this laminin isoform. Fragility of the skin and bullous lesions disappeared after total gastrectomy, but soon reappeared possibly in association with metastatic disease in a lymph node. The possibility that anti-epiligrin CP may develop paraneoplastically in some patients is discussed.

A 71-year-old female patient presented with erosions of the oral, genital and ocular mucosa, but without skin lesions. One year prior to the onset of lesions, a metastatic adenocarcinoma of the endometrium had been diagnosed. In perilesional skin, direct immunofluorescence showed linear deposits of C3 and IgG along the basement membrane zone. Indirect immunofluorescence demonstrated anti-basement membrane zone antibodies which bound exclusively to the dermal side of 1M NaCl-split skin. In immunoprecipitation studies, the autoantibodies bound a set of proteins characteristic of epiligrin/laminin-5, and they specifically immunoblotted the alpha-subunit of this protein. These results confirmed our presumed diagnosis of anti-epiligrin cicatricial pemphigoid. Combined treatment with methylprednisolone and dapsone resulted in prompt remission, despite progression of her malignancy. So far, only eight patients with anti-epiligrin cicatricial pemphigoid have been reported worldwide. Our patient represents the first case from Austria and is clinically and immunopathologically similar to previously reported cases.

This study characterized the specific reactivity, IgG subclass, and complement fixing ability of anti-laminin-5 IgG from 12 patients with anti-epiligrin cicatricial pemphigoid. Circulating IgG from all patients bound the dermal side of 1 M NaCl split skin, immunoprecipitated laminin-5 produced by biosynthetically radiolabeled human keratinocytes, and (in 10 of 12 cases) immunoblotted the laminin-alpha3 subunit. Analysis of the distribution of IgG subclasses in these patients' circulating anti-laminin-5 autoantibodies by semiquantitative indirect immunofluorescence microscopy using the HP series of subclass-specific monoclonal antibodies revealed: (i) IgG4 predominant autoantibodies in seven of 11 sera; (ii) IgG1 and IgG2 at substantially lower levels in a smaller number of sera; and (iii) no specific IgG3 anti-laminin-5 autoantibodies in any patients. The same IgG4-dominant profile of anti-laminin-5 autoantibodies was found in enzyme-linked immunosorbent assay studies of purified human laminin 5. Direct immunofluorescence microscopy of six skin biopsies from three patients found that IgG4 was also the predominant subclass of IgG in epidermal basement membranes in situ. Consistent with these findings, sera from 11 of 11 patients with anti-laminin-5 IgG autoantibodies did not fix C3 to epidermal basement membranes in vitro. These immunochemical studies suggest that complement activation does not play a major role in the pathophysiology of this disease and that subepidermal blisters in these patients may develop via a direct effect of anti-laminin-5 IgG itself.

Inflammatory bowel diseases are uncommon in the Chinese, but the incidence is rising. Their differentiation from infective colitis is often not clear-cut and diagnosing inflammatory bowel diseases can be difficult in Asia. We have studied Chinese patients with ulcerative colitis (N = 19) and Crohn's disease (N = 12) for anti-neutrophil cytoplasmic antibodies (ANCA) by indirect immunofluorescence (IIF) and enzyme-linked immunosorbent assays (ELISA). Patients with enteric fever (N = 29) and irritable bowel syndrome (N = 24) were recruited as controls. Seventy-three percent of ulcerative colitis patients exhibited either p-ANCA (31%) or c-ANCA (42%) by IIF. Twenty-five percent of Crohn's disease patients were found to be p-ANCA positive. However, these ANCA were nonreactive to anti-alpha granule, antiproteinase 3, antimyeloperoxidase, or antilactoferrin. All positive patients had extensive colitis. Sera collected from patients suffering from enteric fever and irritable bowel syndrome were negative for ANCA by IIF and ELISA. We concluded that the detection of ANCA is helpful in diagnosing inflammatory bowel diseases. Further attempts to characterize these autoantibodies are needed.

Inflammatory bowel diseases are known to be rare among the Chinese. The diagnosis of ulcerative colitis has been difficult in some of the Asian countries where infective colitis is more prevalent. Twenty-three Hong Kong Chinese patients diagnosed to have ulcerative colitis were reviewed. The symptoms were relatively mild and extraintestinal manifestation had been rare. Patients responded well to steroid therapy and sulfasalazine. Three patients in this series were found to have cyst and/or trophozoites of Entamoeba histolytica in stool. In this series, 19 patients were tested for antineutrophil cytoplasmic antibody (ANCA). Fourteen patients (73.5%) were positive, of which six (31.5%) showed a perinuclear staining pattern and eight (42%) demonstrated a cytoplasmic pattern. Five patients (26.5%) were negative for any ANCA, and none was positive for both. Sera of these patients were also tested for anti-alpha granules, anti-myeloperoxidase, and anti-lactoferrin activities. None was positive. Control sera collected from 16 patients with irritable bowel syndrome were all negative for the tests. In conclusion, testing of ANCAs may help in making the diagnosis of idiopathic inflammatory bowel disease in difficult situations.