Cold urticaria in association with cholinergic urticaria

The chronic blistering dermatosis, linear IgA disease (LAD), is characterized by subepidermal bullae and linear deposits of IgA in the basement membrane zone (BMZ) of skin. Circulating anti-BMZ antibodies are infrequently found. The immunogiobulin deposition is considered tobe pathogenetically significant. To investigate the clinicopathological correlation we studied 10 patients with LAD. All were evaluated clinically for evidence of mucosal involvement and immunopathologically for immunoglobulin deposition in skin, oral mucosa and conjunctiva. In addition, normal skin, oral mucosa (intact and chemically separated by incubation with IM NaCl) and conjunctiva were used as substrates in an indirect immunofluorescent technique (IIF) to detect circulating anti-BMZ antibodies in patients'sera.
All 10 patients had clinical evidence of oral involvement and all had linear deposits of IgA in oral mucosa. In one patient, oral mucosa gave a positive result when used as substrate for IIF. Six patients had clinical evidence of ocular involvement but none (of seven examined by conjunctival biopsy) had linear IgA deposits in conjunctiva. Five sera were positive for circulating IgA using conjunctiva as substrate, including one patient with no clinical evidence of ocular disease.
This study demonstrates that clinical or immunological assessment alone is unreliable in the evaluation of mucosal disease in LAD. In addition, we have shown that conjunctiva provides the most sensitive substrate for the detection of circulating IgA by IIF.     

Use of 1M NaCl split skin in the indirect immunofluorescence of the linear IgA bullous dermatoses

We compared 1M NaCl split skin with intact skin as substrates for detection of circulating IgA anti-basement membrane (BMZ) antibodies in linear IgA dermatosis (LAD). The sera of 63 patients with LAD including 27 adults and 36 with chronic bullous dermatosis of childhood (CBDC) were examined. 62% of patients overall had circulating IgA anti-BMZ antibodies detectable on intact skin. 73% of patients had circulating antibodies detectable on lM NaCl split skin as an additional 7 sera were positive. This was a statistically significant increase (p<0.01). The sera were mostly positive at a higher titre on the split skin when compared with intact skin. On routine indirect immunofluorescence (IIF) all positive sera produced linear fluorescence on the epidermal side of the split. Twenty serum samples were incubated with split skin overnight; 4 of these specimens exhibited linear fluorescence on the epidermal and dermal sides of the split after this prolonged incubation. These findings suggest that 1M NaCl split skin is a more sensitive substrate for detection of circulating IgA anti-BMZ antibodies in LAD, that these antibodies are heterogeneous and that the target antigen has an epidermal component.     

IgA basement membrane zone autoantibodies in bullous pemphigoid detect epidermal antigens of 270–280 kDa, 230 kDa, and 180 kDa molecular weight by immunoblotting

Bullous pemphigoid (BP) is an acquired subepidermal blistering disease characterized by circulating IgG autoantibodies binding to the 230 and 180 kDa hemidesmosomal proteins. Associated basement membrane zone (BMZ) autoantibodies of the IgA class have been reported in few BP patients. The incidence and clinical relevance of these IgA antibodies, as well as their target antigens are unknown. Sera of 26 patients with BP were analysed for circulating IgG- and IgA-anti-BMZ autoantibodies by indirect immunofluorescence on salt-split human skin. All of the patients had circulating IgG autoantibodies and, in addition, nine (35%) also had circulating anti-BMZ IgA antibodies, that bound to the epidermal side of salt-split skin. By immunoblotting, IgA antibodies in seven of nine sera recognized either the 180 kDa, the 230 kDa, or both BP antigens. Moreover, IgA anti-BMZ antibodies in seven sera also detected an epidermal protein of 270-280 kDa. IgA antibodies did not identify specific bands on immunoblots of dermal extracts. There was no clinical difference between BP patients with or without circulating anti-BMZ-IgA.     

Bullous pemphigoid: a correlative study of autoantibodies, circulating immune complexes and dermo-epidermal deposits

Twenty bullous pemphigoid (BP) patients were studied to establish any correlation between free anti-basement membrane zone (BMZ) antibodies, circulating immune complexes (CIC) and dermo-epidermal junction deposits. CIC levels were evaluated by 2% polyethylene glycol (PEG) precipitation. The twenty patients were found to have IgG and/or C3 deposited in the BMZ. Eight of the twelve patients who had no free anti-BMZ antibodies displayed a positive in vivo C4 and/or C_Iq staining and high levels of CIC. Moreover, CIC were detected in only one patient with positive circulating free anti-BMZ antibodies. The presence of free anti-BMZ antibodies was generally found to correlate with the absence of cutaneous deposits of C1q and/or C4 and with negative CIC; on the other hand, the absence of free anti-BMZ antibodies was generally found to correlate with high levels of CIC and with deposits of C3 and C_Iq and/or C4. The absence of circulating free anti-BMZ antibodies in BP patients, could be explained by the formation of CIC. It is possible that BMZ antigens released from damaged tissue could combine with free antibodies and form complexes in the blood. The release could involve locally formed immune complexes. Elevated CIC levels were generally found to correlate with the presence of active disease.     

Purification of bullous pemphigoid IgG subclasses and their capability for complement fixation.

The antibody reactivity and complement fixing capability of circulating IgG subclass antibodies of patients with bullous pemphigoid (BP) were investigated. Four subclasses of polyclonal IgG were purified from the sera of BP patients. The first stage of purification was a combination of chromatographies on DEAE Affi-Gel Blue and protein A-Cellulofine columns. The four polyclonal IgG subclasses were then isolated from the above-mentioned semi-purified subclass fractions using IgG subclass-specific immuno-affinity chromatographies. Using an immunohistopathological technique, IgG deposits were detected at the basement membrane zone (BMZ) in the normal skin incubated with IgG subclasses other than IgG3. C3 deposits were detected at BMZ in the skin incubated with IgG1. Neither IgG nor C3 deposits were in skin incubated with IgG3. These findings suggest that the subclasses of circulating BP antibodies are IgG1, IgG2 and IgG4, but not IgG3. Moreover, only the BP IgG1 subclass appears to fix complement.     

IgE and its related phenomena in bullous pemphigoid

This study was designed to analyse IgE and its related phenomena in bullous pemphigoid (BP). We analysed 17 BP sera by indirect immunofluorescenee (IIF) and immunoblotting (IB) using a monoclonal antibody to IgE. In addition, inflammatory cells in lesional skin from 11 patients with BP were analysed by the alkaline phosphatase-anti-alkaline phosphatase (APAAP) technique using monoclonal antibodies to IgE and Fc?RII/CD23. IgE class anti-basement membrane zone (BMZ) autoantibody was detected in nine of 17 sera (52.9%) by IIF. IgG class anti-BMZ antibody could block the BMZ-binding reactivity of IgE class antibody. Titres of IgE class autoantibody in the sera ranged from 1:40 to 1:320, and statistically correlated with serum IgF levels. Two of 11 sera contained an IgE class autoantibody which recognized a 230-kDa BP antigen by IB. By radio-allergosorbent test (RAST), IgE-specific antibodies to an extended series of common inhalant and food allergens were detectable in six sera with higb concentrations of total IgF(over 3300 IU/ml). IgE-bearing and Fc?RII-expressing cells were demonstrated in the upper dermis and along the BMZ in seven of 11 biopsy specimens by tbe APAAP technique. The distribution and number of IgE-bearing cells in the lesions were similar to those of the FcEERII-expressing cells. Tbese results suggest that botb IgE-mediated immune responses and autoimmunity characterize BP as distinctive features.     

The predominance of IgG4 in prodromal bullous pemphigoid

Background Prodromal bullous pemphigoid (PBP) and bullous pemphigoid (BP) demonstrate immunoglobulin G (IgG) and/or C3 deposition at the basement membrane zone (BMZ) on direct immunofluorescence. BP‐180‐specific IgG1, IgG4, and IgE antibodies have been detected in BP. However, the distribution of IgG subclasses is unknown in PBP. Objectives We will describe the role of anti‐BMZ IgG subclasses in PBP and we will correlate these findings to better understand the pathogenesis of PBP. Methods Skin biopsies and serum samples were obtained from 45 patients who had PBP. The skin tissue was processed for direct immunofluorescence studies. Sera were analyzed by indirect immunofluorescence for the presence of circulating anti‐BMZ IgG antibodies (by standard IIF) and IgG subclasses antibodies (by sandwich double antibody immunofluorescence [SDAI]). Sera were also analyzed for antibodies against BP‐180 and BP‐230 antigens by enzyme‐linked immunosorbent assay (ELISA). Results Thirty‐two patients (71%) had IgG and C3 staining at the BMZ, while 13 patients (29%) had isolated C3 staining at the BMZ on direct immunofluorescence. All patients demonstrated staining on the epidermal side of the salt‐split skin. Of the seven skin specimens that were available for C5‐9 SDAI testing, all were found to be positive along BMZ area. Standard IIF studies demonstrated the presence of circulating BMZ antibodies in 11 of the 30 patients (36.6%). When SDAI for IgG subclass differentiation was utilized, 17 of 30 (56.6%) patients were found to have circulating anti‐BMZ antibodies. All of these 17 patients had IgG4 subclass antibodies. Thirteen patients did not have detectable IgG subclass anti‐BMZ antibody on SDAI. Sixteen of 30 patients had detectable anti‐BP‐180 or anti‐BP‐230 antibodies, while 12 (40%) did not have detectable antibody against BP antigens on ELISA. Conclusions IgG4 is the initial and predominant anti‐BMZ antibody subclass detected in PBP. Demonstration of linear C5‐9 at the BMZ enhances the early diagnosis of PBP. Predominance of IgG4 and the initial presence of IgG4 on skin lesions as well as the presence of only IgG4 subclass anti‐BMZ antibody suggest that IgG4 subclass antibody could be the initial immunologic event encountered in patients with PBP.     

Evidence that anti-basement membrane zone antibodies in bullous eruption of systemic lupus erythematosus recognize epidermolysis bullosa acquisita autoantigen

Circulating and tissue-deposited IgG antibodies to the cutaneous basement membrane zone (BMZ) were detected in 3 patients with the clinical, pathologic, and immunologic features of bullous eruption of systemic lupus erythematosus (SLE). The antibodies were present in sera and IgG fractions in all cases and in eluates of cutaneous immune deposits from one of the cases. The antibodies were easily detected in sera by indirect immunofluorescence on adult human thigh skin separated through the lamina lucida by incubation in 1.0 M NaCl but were less easily detected on intact neonatal foreskin. The antibodies had features of epidermolysis bullosa acquisita (EBA) anti-BMZ antibodies including binding to the dermal side of the BMZ in separated skin, binding to the cutaneous but not vascular or glomerular basement membranes, binding to and just below the lamina densa, and binding to 290 or 290 and 145 kD dermal proteins previously identified as components of the EBA autoantigen. The antibodies were relatively specific for SLE patients with features of bullous eruption of SLE since they were detected in 3 of 4 of those cases and in only 1 of 20 SLE patients without blisters. These results show anti-BMZ antibodies with features of EBA antibodies are present in patients with bullous eruption of SLE and suggest there may be a close relationship between that disease and EBA. The results also suggest that EBA antibodies may be part of the autoantibody spectrum of SLE and that separated skin is more sensitive than intact skin for their detection.     

免疫即迹法与分离人皮肤IIF洁检测类天疱疮抗体的比较研究

采用１ｍｏｌ／ＬＮａＣｌ分离正常人皮肤为底物的ＩＩＦ法和免疫印迹技术检测１２２例类天疱疮患者血清中抗基底膜抗体，比较研究上述两种方法对检测类天疱疮抗体的灵敏度及价值。结果分离皮肤ＩＩＦ法阳性８９例（７２．９５％）。免疫印迹法检测，与２３０ＫＤ、１８０ＫＤ和１６０ＫＤ分子量蛋白抗原结合阳性９２例（７５．４１％）。两法合用总阳性率为８３．６１％。分离皮肤ＩＩＦ法适合于常规采用，而免疫印迹法是前者的进一步补充。     

A case of linear IgA disease presenting initially with IgG immune deposits

We describe a case of linear IgA bullous disease initially presenting with histopathologic and immunofluorescent findings consistent with bullous pemphigoid. Initial immunofluorescent studies demonstrated a predominance of linear IgG at the basement membrane zone (BMZ) of perilesional skin and a low titer circulating IgG anti-BMZ antibody. Repeat studies 3 years later revealed a predominance of linear IgA immune deposits at the BMZ and no circulating anti-BMZ antibody. Dapsone therapy was initiated at this time with a good therapeutic response noted. Suction blister studies, immunoelectron microscopy, split skin immunofluorescent studies and Western immune blot were performed and provided indirect evidence that the BMZ antigen in this case is distinct from the bullous pemphigoid antigen component of the BMZ.     

Differentiating anti-lamina lucida and anti-sublamina densa anti-BMZ antibodies by indirect immunofluorescence on 1.0 M sodium chloride-separated skin

Sixty-one bullous disease sera containing IgG anti-BMZ antibodies were examined by indirect immunofluorescence on intact skin and skin separated through the lamina lucida by incubation in 1.0 M NaCl. All sera produced an indistinguishable pattern of linear immunofluorescence on intact skin at dilutions of 1:10 or higher. On separated skin, antibodies bound to either the epidermal (epidermal pattern), dermal (dermal pattern), or epidermal and dermal (combined pattern) sides of the separation. The binding patterns were consistent on separated skin from several donors and titers of anti-basement membrane zone antibodies on separated skin were comparable to those on intact skin. Sera from 3 patients with herpes gestationis (HG), 36 patients with bullous pemphigoid (BP), and 1 patient with clinical and histologic features of epidermolysis bullosa acquisita (EBA) showed an epidermal pattern. Sera from 9 patients with BP showed a combined pattern and sera from 6 patients with EBA and 6 patients with clinical and histologic features of BP showed a dermal pattern. Indirect immunoelectron microscopy of selected sera showed antibodies producing the epidermal and combined patterns were anti-lamina lucida antibodies and those producing the dermal pattern were anti-sublamina densa antibodies. These results show indirect immunofluorescence on separated skin is a dependable method for differentiating bullous disease anti-lamina lucida and anti-sublamina densa antibodies and that differentiating between the antibodies is essential for accurate diagnosis in some patients. The results also suggest BP anti-lamina lucida antibodies may have more than one antigenic specificity.     

Localized pemphigoid: a case report showing in situ deposition as well as presence in serum of IgG and IgA anti-basement membrane zone antibodies

In this paper, we describe a 69-year-old Japanese woman with localized pemphigoid. Direct immunofluorescence study showed linear IgG, IgA and C3 deposits at the basement membrane zone (BMZ). In the serum, circulating anti-BMZ antibodies of both IgG and IgA classes were found. This is the first report of a patient with localized pemphigoid in whom circulating anti-BMZ antibodies of not only the IgG class, but also of the IgA class were present.     

Erosive lichen planus of the vulva: weak circulating basement membrane zone antibodies are present

The objective of this study was to investigate whether circulating basement membrane zone (BMZ) antibodies are present in erosive lichen planus (LP) of the vulva. In total, 56 consecutive women with biopsy-confirmed erosive LP of the vulva were recruited from a vulval clinic in a district general hospital and teaching hospital in Oxfordshire. Indirect immunofluorescence (IgG and IgA) was performed on 56 sera, and 15 were tested to IgG subclasses (1-4). Immunoblotting was carried out on salt-split and urea-extracted epidermal skin extracts on 11. The main outcome measure was the presence or absence of staining at the BMZ. Of the 56 sera, 34 (61%) had weak (neat or 1 : 5) epidermal-binding BMZ antibodies (25 had IgG, 5 had IgA, 4 had both IgG and IgA). All 15 sera tested to IgG showed epidermal binding to one or more IgG subclasses: IgG1 (7 sera), IgG2 (7), IgG3 (7) and IgG4 (0). Immunoblotting identified IgG antibodies to bullous pemphigoid (BP)180 (10/11) and BP230 (2/11). The majority (61%) of patients with vulval erosive LP had circulating serum IgG BMZ antibodies, chiefly reacting with BP180. There was subclass restriction of the IgG response to IgG1, 2 and 3. The significance of these antibodies is uncertain, but they may be a marker for the disease.     

IgA linear dermatosis (author's transl)]

Besides the typical forms of dermatitis herpetiformis (DH) and bullous pemphigoid (BP) of adults and children, there are cases combining clinical, histological and electronmicroscopic features of both. Linear continuous IgA deposits along basement membrane zone (BMZ) are a most characteristic finding. They differ from the granular IgA deposits in DH, even if these are also distributed along the BMZ (however, preserving as a rule their granular pattern). IgG circulating anti-BMZ antibodies are absent, whereas in some cases IgA anti-BMZ antibodies may be found. In contrast to DH, there is no gluten-sensitive enteropathy, and the gluten-free diet is ineffective. The recognition of this bullous disease as a distinct entity is of practical significance because these cases respond well to combined treatment with sulfones and corticosteroids, all in small doses. Because of diagnostic importance of linear IgA deposits at BMZ we have proposed the name IgA linear dermatosis. In children a counterpart of IgA linear dermatosis of adults is chronic bullous disease of childhood (CBDC), which we propose to call IgA linear dermatosis of childhood.     

An Analysis of 24 Patients with IgA Deposition at the BMZ

A study of 24 patients with IgA deposition at the BMZ of the skin showed that five conditions could be recognized: 1) linear IgA bullous dermatosis in adults (LAD, 7 cases); 2) linear IgA and IgG bullous dermatosis in adults (LAGD, 10 cases); 3) chronic bullous disease of childhood (CBDC, 3 cases); 4) dermatitis herpetiformis (DH, 1 case), and 5) systemic lupus erythematosus (SLE, 3 cases). Histopathologically, 5 of 7 patients with LAD were similar to the DH group, but 7 of 10 patients with LAGD were similar to the BP group. Half the patients with LAD and LAGD had oral lesions, and most of them had excellent responses to dapsone and Tripterygium Wilfordii, but the patients with CBDC did not respond to these treatments. In the patients with LAD and LAGD, the positivity rates of IgA anti-BMZ antibodies examined by indirect immunofluorescence (IIF) on intact skin and NaCl split skin were 41% and 64%, respectively. The heterogeneity of the histopathologic pictures of LAD and LAGD, the incidence of DH, and the value of using NaCl split skin for IIF are discussed.     

抗核抗体阴性的大疱性系统性红斑狼疮1例

报告１例抗核抗体阴性的大疱性系统性红斑狼疮。在整个病程中共查过７次抗核抗体均阴性。ＤＩＦ、ＩＩＦ，免疫印迹均无阳性发现。肾脏穿刺活检免疫荧光见ＩｇＧ颗粒状沉积。并讨论其诊断依据和大疱性系统性红斑狼疮抗核抗体为阴性的可能原因。     

Blister fluid for the diagnosis of subepidermal immunobullous diseases: a comparative study of basement membrane zone autoantibodies detected in blister fluid and serum

The subepidermal immunobullous diseases bullous pemphigoid (BP), cicatricial pemphigoid (CP), pemphigoid gestationis (PG) and linear IgA disease (LAD) are characterized by circulating and in vivo deposition of antibodies to antigens in the cutaneous basement membrane zone (BMZ). Indirect immunofluorescence (IMF) of serum is a routine diagnostic test to detect circulating BMZ antibodies in these diseases. We have compared the titres of IgG and IgA and their subclasses, also of IgM and IgE BMZ antibodies in serum and aspirated blister fluid in 35 adult patients with subepidermal immunobullous diseases: BP ( n  = 30), PG ( n  = 2), CP ( n  = 1), and LAD ( n  = 2), by indirect IMF on intact and salt-split skin. The antibody titre in blister fluid was the same or one dilution less than serum in most cases and there was no significant difference between these results ( P  > 0.05). IgG1 and IgG4 were the predominant subclasses in both blister fluid and serum in BP. Indirect IMF of serum and blister fluid was also carried out on trypsinized epidermal cells in a subgroup of patients with BP ( n  = 19). Typical polar fluorescence was obtained in all 14 cases which had positive indirect IMF on intact and split skin. Our findings demonstrate that blister fluid can be used as an alternative to serum for indirect IMF in subepidermal immunobullous diseases. This avoids the need for venesection and has a practical application in children and those with poor venous access.     

Basement membrane antibodies in sera of haematopoietic cell recipients are associated with graft‐versus‐host disease

Background Graft‐versus‐host disease (GvHD) occurs frequently after haematopoietic cell transplantation (HCT). Mucocutaneous lesions of GvHD may mimic bullous autoimmune dermatoses, and 10 cases of concurrent GvHD and a bullous autoimmune disease have been reported in the literature. Objective To determine the frequency of circulating antibodies to the cutaneous basement membrane zone (BMZ) in HCT patients with GvHD in comparison with HCT patients without GvHD, psoriasis patients and healthy controls. Subjects and methods We examined 42 patients with chronic GvHD, 18 HCT patients without GvHD, 11 psoriasis patients and 40 healthy controls, prospectively. Sera were tested by indirect immunofluorescence (IIF) on salt‐split skin, NC16a‐ELISA and immunoblot using keratinocyte extracts. Univariate statistical analyses and logistic regression were performed to assess possible correlations of graft and patient characteristics with the presence of BMZ antibodies. Results Circulating basement membrane zone (BMZ) antibodies were detected in 10/42 (24%) GvHD sera by immunoblot, but not in any of the HCT sera from patients without GvHD (0/18; 0%). The antibodies targeted collagen VII, BP230, collagen XVII/BP180 or p200/laminin γ1. Clinically manifest bullous autoimmune dermatoses (bullous pemphigoid or epidermolysis bullosa acquisita) were found in two GvHD patients. 1/11 (9%) psoriasis sera and 1/40 (2.4%) healthy control sera reacted with collagen XVII or BP230, respectively. Conclusions Circulating BMZ antibodies are significantly associated with chronic GvHD in contrast to uncomplicated HCT. Recurrent mucocutaneous lesions in chronic inflammatory skin disorders may liberate antigens, which may lead to production of BMZ antibodies, particularly in the context of GvHD‐mediated reduced self‐tolerance.     

Correlation of serum anti-basement membrane zone antibody and malignancy in bullous pemphigoid

73 patients with bullous pemphigoid were studied to examine the association between serum anti-basement membrane zone (anti-BMZ) antibody and malignancy. Indirect immunofluorescence (IIF) was performed on the sera. 37 patients had positive IIF for anti-BMZ antibodies, ranging in titer from 1:20 to 1:10,230. On a follow-up of 7-41 months (mean 22 months), only 3 patients (8%) had malignancies (endometrium, bladder, squamous cell carcinoma of the skin). 36 patients had negative IIF for anti-BMZ antibodies. On a follow-up of 5-43 months (mean 16 months), 3 patients (8%) had malignancies (breast, colon, endometrium). No correlation between the presence or absence of anti-BMZ antibody in the serum and the development of malignant disease was observed.     

Absence of specific histologic changes in guinea pig skin treated with bullous pemphigoid antibodies

Previous studies have reported that intradermal injections of bullous pemphigoid antibodies into guinea pigs can reproduce the histologic and immunohistologic features of bullous pemphigoid lesions. In this study we examined this model to determine its reproducibility and suitability for testing other types of anti-BMZ antibodies. Twenty guinea pigs were injected intradermally with 0.1, 0.3, or 0.5 ml of either bullous pemphigoid serum or IgG fraction containing high-titer complement-binding anti-BMZ antibodies or an equivalent volume of normal human serum or IgG fraction as control. Sites were biopsied at intervals after injection and were examined by routine histology and direct immunofluorescence. The results showed (a) no difference in the incidence of dermal epidermal separation or type of inflammation in experimental and control sites; (b) no evidence of an eosinophil-rich inflammatory reaction typical of bullous pemphigoid; (c) an absence of linear BMZ deposits of IgG and complement in the majority of sites injected with bullous pemphigoid antibodies; and (d) no correlation between dermal-epidermal separation and deposition of immune reactants at the BMZ. These results suggest the histologic changes seen in guinea pigs that are administered intradermal injections of bullous pemphigoid antibodies are nonspecific and that the model is not suitable for testing the pathogenicity of anti-BMZ antibodies in sera or IgG fractions.