IgA autoantibodies in the pemphigoids and linear IgA bullous dermatosis

Please cite this paper as: IgA autoantibodies in the pemphigoids and linear IgA bullous dermatosis. Experimental Dermatology 2010; 19: 648–653. Background: Patients with bullous pemphigoid (BP), mucous membrane pemphigoid (MMP) and pemphigoid gestationis (PG) have IgG antibodies against BP180 and BP230, components of the hemidesmosomes. Patients with linear IgA bullous dermatosis (LABD) have IgA autoantibodies against a 97/120‐kDa protein which is highly homologous to a shedded fragment of the BP180‐ectodomain. Objectives: The aim of our study was to determine the incidence of IgA autoantibodies directed against BP180/BP230 in the pemphigoids and LABD and to determine the antigenic regions that are targeted by IgA autoantibodies. Methods: Utilizing baculovirus‐expressed recombinant BP180 and BP230 proteins, we performed immunoblot analyses for IgA reactivity of sera from patients with BP (n = 30), MMP (n = 10), PG (n = 6), LABD (n = 6) and from control patients with non‐related pruritic dermatoses (n = 8). Results: IgA reactivity against BP180 and/or BP230 was detected in 19/30 of the BP, in 7/10 of the MMP, in 6/6 of the LABD and in 3/6 of the PG sera, respectively, but not in the control group. In all subgroups, the major antigenic site recognized by IgA antibodies was located within the NH₂‐terminus of the BP180‐ectodomain, but only a minority of the sera showed also IgA reactivity against the BP180‐NC16a‐domain. IgA reactivity against the central domain of BP180 was more frequently seen than against its COOH‐terminus. IgA against the COOH‐ and NH₂‐terminus of BP230, respectively, was detected in 6/30 of the BP, 1/10 of the MMP, 1/6 of the LABD and 0/8 control sera. Conclusion: IgA reactivity against BP180 and/or BP230 is a common finding in the pemphigoids.     

IgE autoantibodies against the intracellular domain of BP180

Background  Bullous pemphigoid (BP) is an autoimmune subepidermal blistering disease characterized by autoantibodies against the hemidesmosomal proteins BP180 (type XVII collagen) and BP230. BP not only involves IgG-mediated neutrophil activation, leading to blistering, but also IgE-dependent activation of mast cells and basophils. While IgG and IgE autoantibodies target the extracellular noncollagenous (NC) 16A domain of BP180, little is known whether other BP180 regions are targeted by these antibody classes.
Objectives  To characterize IgE and IgG autoantibody binding to antigenic sites on the intracellular domain (ICD) of BP180 compared with BP180 NC16A.
Methods  IgE/IgG autoreactivity against recombinant BP180 ICD and NC16A was determined by immunoblotting of sera from 18 patients with BP and 10 controls.
Results  Total serum IgE was elevated in 16 of 18 BP sera. Most BP sera tested positive (15 of 18) to NC16A with both immunoglobulin classes. Additionally, 14 of 18 sera showed IgE reactivity with an epitope mapped to the ICD of BP180 (amino acid residues 103–266). Mapping of ICD antigenic sites revealed similar IgE and IgG reactivities for most regions except for greater IgE reactivity to amino acid residues 234–398 (11 of 18 BP sera) than IgG (five of 18). Control sera failed to display IgE reactivity to these antigens.
Conclusions  The results indicate that BP180 NC16A is not the only antigenic determinant of IgE autoantibodies in BP and that additional, novel epitopes exist on different regions of the ICD of BP180. The heterogeneous autoimmune response against BP180 suggests intramolecular epitope spreading during disease progression.     

Autoantibodies to basement membrane proteins BP180 and BP230 are commonly detected in normal subjects by immunoblotting

Autoantibodies to basement membrane proteins BP180 and BP230 are characteristic of bullous pemphigoid and other subepidermal immunobullous disorders. These antibodies are, however, reported in other pruritic dermatoses, non-bullous disorders and non-cutaneous disease. Few studies have assessed basement membrane antibodies in normal subjects; antibody prevalence in this population is not clear. This study aims to examine basement membrane zone antibodies in normal middle-aged to elderly subjects. Sera from 61 healthy subjects (majority age 50–70 years) were assessed by immunoblot, indirect immunofluorescence and enzyme-linked immunosorbent assay. Ninety-one bullous pemphigoid patients acted as positive controls. Antigenic target, antibody class and titre were examined; sera binding BP180 were assessed for reactivity to the non-collagenous 16A (NC16A) domain. Thirty-six normal subjects (59%) had antibodies to either BP180 or BP230 on immunoblot analysis. BP180 was the commonest target antigen, detected in 35 subjects; binding to the immunodominant NC16A domain was not detected. Immunofluorescence was positive in three subjects. Of the bullous pemphigoid sera, 88% were positive on immunoblot or immunofluorescence; a higher frequency had antibodies against BP230. In conclusion, significant numbers of normal healthy subjects have circulating autoantibodies to basement membrane proteins, chiefly BP180 detectable by immunoblot, but these do not bind the NC16A domain.     

Antiplectin autoantibodies in subepidermal blistering diseases

Background  Hemidesmosomal proteins may become targets of autoimmunity in subepidermal blistering diseases. Well-known recognized autoantigens are the intracellular plaque protein BP230, the transmembrane BP180 and its shed ectodomain LAD-1.
Objectives  To establish the prevalence of autoimmunity against plectin, another intracellular plaque protein, and to investigate its antigenic sites.
Methods  Two hundred and eighty-two patients with subepidermal blistering diseases, investigated by routine immunoblot analysis for possible antiplectin antibodies, were included in the study. Epitope mapping was performed using recombinantly produced overlapping plectin domains from the actin-binding domain to the rod domain. The COOH-terminal region of plectin was not included in the study.
Results  In 11 of 282 (3·9%) patients an immunoblot staining pattern identical to that of antiplectin monoclonal antibody HD121 was found. Affinity-purified antibodies bound back to normal human skin in a pattern typical for plectin, i.e. to the epidermal basement membrane zone as well as to keratinocytes in the epidermis, and to myocytes. No binding was seen to plectin-deficient skin of a patient with epidermolysis bullosa simplex with muscular dystrophy. Epitope mapping of the plectin molecule showed that the central coiled-coil rod domain is an immunodominant hotspot as 92% of the sera with antiplectin antibodies reacted with it. Most patients with antiplectin antibodies also had antibodies to other pemphigoid antigens.
Conclusions  Plectin is a minor pemphigoid antigen with an immunodominant epitope located on the central rod domain.     

IgG, IgA and IgE autoantibodies against the ectodomain of BP180 in patients with bullous and cicatricial pemphigoid and linear IgA bullous dermatosis

BACKGROUND: Bullous pemphigoid (BP), linear IgA bullous dermatosis (LABD) and cicatricial pemphigoid (CP) are clinically distinct autoimmune bullous skin diseases characterized by autoantibodies against components of the epidermal basement membrane. Like most patients with BP, a significant subgroup of patients with CP has circulating IgG specific for BP180, a transmembraneous protein of hemidesmosomes. Moreover, sera of patients with LABD contain IgA autoantibodies reactive with a 97/120-kDa protein, LABD antigen 1, which is highly homologous to the extracellular portion of BP180. OBJECTIVES: We aimed to determine whether, in these diseases, autoantibody reactivity to BP180 is restricted to distinct immunoglobulin subtypes. METHODS: Utilizing a baculovirus-encoded form of the ectodomain of BP180, sera from patients with BP (n = 10), CP (n = 9), LABD (n = 10) and normal human control sera (n = 10) were analysed by immunoblot for IgG, IgA and IgE reactivity against BP180. RESULTS: All of 10 BP sera displayed IgG, IgA and IgE reactivity with BP180. Six and seven of nine CP sera, respectively, contained IgG and IgA autoantibodies reactive with BP180, but none of nine sera contained BP180-specific IgE. Nine of 10 LABD sera contained IgA, and six of 10 IgG, which was reactive with BP180, but none of 10 sera showed IgE reactivity to BP180. CONCLUSIONS: The presence of IgG and IgA autoantibody responses to BP180 in patients with three clinically distinct autoimmune bullous diseases indicates that an autoimmune response to the same distinct adhesion protein may lead to different clinical manifestations. It is therefore conceivable that variable epitopes of BP180 are targeted by the different autoantibody isotypes, resulting in the distinct clinical pictures.     

Bullous pemphigoid antigen II (BP180) and its soluble extracellular domains are major autoantigens in mucous membrane pemphigoid: the pathogenic relevance to HLA class II alleles and disease severity

BACKGROUND: Mucous membrane pemphigoid (MMP), a chronic autoimmune subepithelial blistering disease, is associated with circulating IgG and/or IgA autoantibodies against several basement membrane zone antigens. The heterogeneity of clinical presentation and diversity of target autoantigens have contributed to difficulties in characterizing this condition immunologically. OBJECTIVES: To analyse serum autoantibody profile and HLA class II alleles in MMP patients and to correlate this with the clinical presentation of disease. METHODS: Well-defined subgroups consisting of 124 patients with MMP were examined for IgG and IgA reactivity with immunoblotting using human epidermal, dermal and placental amnion proteins. The results were further analysed on the basis of detailed clinical (sites of involvement and disease severity) and immunopathological criteria (immunofluorescence study and HLA class II alleles). RESULTS: Immunoblot assay revealed that the majority of MMP patients had IgG (93 of 124, 75%) and/or IgA autoantibodies (63 of 124, 51%) to BP180 (including its soluble ectodomains, 120-kDa LAD-1 and 97-kDa LABD97 antigens). Other antigens targeted predominantly by IgG autoantibodies included: BP230 in 34 (27%), beta4 integrin in 26 (21%), and laminin 5 in three (2%). All the BP230+ sera and 23 (88%) beta4 integrin+ sera also reacted with at least one of the BP180 antigens. Over 85% of patients with reactivity to beta4 integrin had ocular involvement. In most cases of MMP, more severe clinical features were associated with antibody reactivity to multiple basement membrane zone antigens, as well as reactivity to multiple BP180 component antigens. Dual BP180/LAD-1 reactivity with IgG and IgA was associated with a more severe phenotype. In addition, the subset-dependent autoantibody reactivity correlated well with specific HLA class II alleles, DQB1*0301, DRB1*04 and DRB1*11. CONCLUSIONS: Our results confirmed that BP180 is a major autoantigen targeted by the sera of patients with MMP. The disease-prevalent HLA class II alleles and humoral autoimmune response against the particular subsets of antigenic epitope(s) within BP180 ectodomain may contribute to the clinicopathological significance and disease severity of MMP.     

Evaluation of a BP180-NC16a enzyme-linked immunosorbent assay in the initial diagnosis of bullous pemphigoid

BACKGROUND: Bullous pemphigoid (BP) is the most common subepidermal immunobullous disease, characterized by circulating IgG autoantibodies targeting BP180 and BP230 hemidesmosomal proteins. Several immunological studies have demonstrated that the membrane proximal noncollagenous domain NC16a of BP180 is the immunodominant region targeted by BP autoantibodies. Recently, a commercial BP180 NC16a-specific enzyme-linked immunosorbent assay (ELISA) has become available for detecting pathogenic anti-BP180 autoantibodies in BP sera. However, it remains unclear whether the diagnostic potential of the ELISA is equivalent to that of the 'gold-standard' diagnostic technique of immunofluorescence (IF). OBJECTIVES: To examine the usefulness of a commercially available BP180-NC16a ELISA in the initial serodiagnosis of BP. METHODS: Sera from a large cohort of patients with BP (n = 102) and control subjects (age- and sex-matched normal volunteers, n = 60; pemphigus foliaceus, n = 18; pemphigus vulgaris, n = 16) were assayed by BP180-NC16a ELISA. All BP sera were obtained at presentation before initiation of systemic immunosuppressive therapy. The values of IgG antibody levels measured by ELISA were compared with those measured by indirect IF on salt-split skin. Results Receiver operating characteristic analysis was used to calculate the cut-off value for the ELISA in the diagnosis of BP which maximizes both sensitivity and specificity, and to estimate the diagnostic accuracy of the ELISA as represented by the area under the curve (AUC = 0.965). A cut-off value of 9 was associated with a sensitivity of 89% (91 of 102 BP sera showed a positive result) and a specificity of 98%. Fifty-eight of 60 normal controls and all the pemphigus sera showed a negative result. There was a correlation between the mean ELISA values and indirect IF titres (Spearman rank correlation 0.286; P = 0.004). CONCLUSIONS: Our results suggest that the BP180-NC16a ELISA is a useful tool for the detection of pathogenic anti-BP180 IgG autoantibodies at the initial disease stage of BP. Because it is not only highly sensitive and specific, but is also easy to perform, is objective, and semiquantitative, the ELISA may provide valuable information for the accurate and reliable serodiagnosis of BP.     

Autoantibodies to the extracellular and intracellular domain of bullous pemphigoid 180, the putative key autoantigen in bullous pemphigoid, belong predominantly to the IgG1 and IgG4 subclasses

BACKGROUND: Autoantibodies to the extracellular domain (ECD) of bullous pemphigoid (BP) antigen 180 (BP180) are thought to play a crucial part in the pathophysiology of BP. OBJECTIVES: As the various IgG subclasses have different biological properties, we have sought to assess the relative isotype distribution of IgG to BP180 and their reactivity against the ECD and intracellular domain (ICD) of BP180. METHODS: The reactivity of 27 sera from patients with BP was assayed by immunoblotting against recombinant proteins covering the ECD and ICD of BP180. RESULTS: Twenty-seven (100%) and 21 (77%) of 27 BP sera, respectively, contained IgG1 and IgG4 autoantibodies binding to the ECD of BP180. Fourteen (82%) and six (35%) of the 17 BP sera that were reactive with the ICD of BP180 had autoantibodies of the IgG1 and IgG4 subclass, respectively. The profile of the isotype restriction appeared to be similar when the response to the ECD vs. that to the ICD was compared. IgG2 and IgG3 reactivity with BP180 was found less frequently. Patients with BP of longer duration showed a tendency to have, in addition to IgG1, an IgG4 response. CONCLUSIONS: Consistent with prior evidence indicating that subepidermal blister formation in BP is dependent upon complement activation, the frequent finding of complement-fixing IgG1 autoantibodies to both the ECD and ICD of BP180 might have pathogenic relevance in BP. These findings provide new insights relevant for our understanding of the immune response to BP180, the putative key autoantigen in BP.     

Role of BP230 autoantibodies in bullous pemphigoid

Bullous pemphigoid (BP) is an autoimmune disease associated with subepidermal blistering due to autoantibodies directed against BP180 and BP230. BP180 is currently considered as the major pathogenic autoantigen. However, previous clinical findings suggested that anti-BP230 autoantibodies alone can cause skin lesions in animal models and many BP patients. The characteristics of BP230 and the pathogenic roles of anti-BP230 antibodies have been proposed. First, at the molecular level, BP230 mediates the attachment of keratin intermediate filaments to the hemidesmosomal plaque and interacts with other constituents of hemidesmosomes. Second, the presence of BP230 autoantibodies may correlate with specific clinical features of BP. The immunoglobulin (Ig)G autoantibodies from BP patients react mainly against the C-terminus of BP230, while the IgE autoantibodies are still inconclusive. Third, in vivo, autoantibodies against BP230 involved in the disease may not only induce the inflammatory response but also impair the structural stability of hemidesmosomes. This article reviews recently published work about the role of BP230 and its antibodies, including IgG and IgE, aiming to find clues of its clinical association and lay the foundation for the research on the pathogenicity of antibodies against BP230.     

Correlation of autoantibodies against BP180/BP230 in response to topical corticosteroids in patients with bullous pemphigoid

Topical steroids are effective in treating bullous pemphigoid (BP). Autoantibodies against BP180 are related to disease activity, but correlation of these autoantibodies with response to topical steroid therapy has not yet been clearly evaluated. We investigate the usefulness of close and early monitoring of autoantibodies against BP180 and BP230 for assessment of response to therapy and early detection of therapeutic failure in BP patients treated topically. In eight BP patients under treatment with topical or systemic steroid therapy we retrospectively evaluated clinical course and autoantibodies against BP180 and BP230 as well as indirect immunofluorescence titres (IIF). Data were included at diagnosis, during hospitalization and follow‐ups. Autoantibodies against BP180 parallel disease activity in all topically and as well as systemically treated patients. Autoantibodies against BP230 correlated in five out of eight patients. Autoantibodies directed against BP180 and, to a lesser degree, against BP230 correlate with the clinical course of topically treated BP patients. Monitoring autoantibodies against BP180 is a useful tool to evaluate the efficacy of topical therapy in BP.     

大疱性类天疱疮患者血清抗BP180分子不同表位自身抗体定量分析及亲和力分析

目的 探讨大疱性类天疱疮（BP）患者血清抗BP180分子不同表位的自身抗体量。方法 制备BP180分子NC16A片段的不同抗原表位区NC16A-1、NC16A-2和NC16A-3,利用柱亲和层析的方法从10例BP患者血清中分别纯化抗不同表位区的自身抗体,定量,并用硫氰酸盐洗脱法测定抗NC16A-1、抗NC16A-2和抗NC16A-3自身抗体的相对亲和力。结果 经亲和层析纯化获得针对BP180-NC16A不同表位区的自身抗体,从20 mg总IgG中纯化抗NC16A-1、NC16A-2和NC16A-3自身抗体的产量分别为（49.0 ± 20.7） μg、（117.7 ± 22.4） μg和（39.5 ± 18.9） μg。抗NC16A-2自身抗体的量和亲和力水平均明显高于其他两个表位区的自身抗体。结论 BP致病性自身抗体所识别的主要抗原表位可能位于BP180分子的NC16A-2表位区（aa507-aa520）。     

Antibody-Binding to the 180-kD Bullous Pemphigoid Antigens at the Lateral Cell Surface Causes Their Internalization and Inhibits Their Assembly at the Basal Cell Surface in Cultured Keratinocytes

We demonstrated the effects of monoclonal antibodies to the 180-kD and 230-kD BP antigens (BPA) and of BP sera on Ca⁺⁺-induced formation of hemidesmosomes in cultured human keratinocytes (a cell line, DJM-1) by immunofluorescence microscopy. Under low Ca⁺⁺ (0.07 mM) conditions, the 180-kD and 230-kD BPAs were distributed homogeneously on the basal plasma membrane, while they formed a peculiar concentric ring or arch (ring/arch) arrangement in high-Ca⁺⁺ (1.87 mM) medium. On the other hand, the apical-lateral cell membrane was stained homogeneously with antibodies to the 180-kD BPA, but not to the 230-kD BPA, both in low and high Ca⁺⁺ media. The low-high Ca⁺⁺ switch at first caused disappearance of the antigen from the basal plasma membrane and then formed the high-Ca⁺⁺ ring/arch pattern within 3 hrs. In this system, monoclonal antibodies to the 180-kD and 230-kD BPAs and the sera from 5 BP patients, 2 pemphigus vulgaris (PV) patients, and 4 normal volunteers were added into the culture media. The addition of anti-180-kD BPA antibodies or any BP serum caused the internalization of the 180-kD BPA from the apical-lateral cell membrane and inhibited the Ca⁺⁺-induced formation of the ring/arch pattern on the basal membrane, possibly by inhibiting the movement of the antigen from the lateral to the basal membrane to form hemidesmosomes. The internalized fluorescence dots were shown to be composed of the 180-kD BPA and patient's IgG, but not of the 230-kD BPA by double-immunostaining, suggesting the internalized 180-kD BPA was from the apical-lateral membrane, but not from hemidesmosomes. Monoclonal antibodies to the 230-kD BPA and normal and PV sera did not cause these effects. These results suggest that autoantibodies to the 180-kD BPA, but not to the 230-kD BPA, may directly bind the antigen on the cell surface and disturb the formation of hemidesmosomes.     

Detection of autoantibodies against bullous pemphigoid and pemphigus antigens by an enzyme-linked immunosorbent assay using the bacterial recombinant proteins

Abstract To develop a new method to evaluate autoantibodies in various autoimmune bullous skin diseases, we examined reactivity of bullous pemphigoid (BP) and pemphigus vulgaris (PV) patients' sera with partial bacterial fusion proteins of the 230 kD BP antigen (BPAG1) and PV antigen, respectively, by an enzyme-linked immunosorbent assay (ELISA), and compared the results with those of immunoblotting. We used two fusion proteins derived from the mouse BPAG1 and a fusion protein derived from the amino-terminus (EC 1-2) of PV antigen. For both BP and PV sera, the ELISA scores were well correlated with the reactivities on immunoblot assays. The present study indicates that ELISA using recombinant antigen proteins in various autoimmune bullous skin diseases will be a new useful technique to detect the autoantibodies. However, development of recombinant proteins with the entire molecule and correct conformation will be necessary to establish a perfect ELISA system in the future.     

Absence of anti-BP180 antibodies in mothers of infants with bullous pemphigoid

BACKGROUND: It is not clear whether bullous pemphigoid (BP) of infancy is linked to maternal transmission of pathogenic autoantibodies. Objectives To search for anti-BP180 antibodies in the sera of infants with BP and their mothers, using sensitive and specific methods. METHODS: Four infants (<6 months) with BP and their mothers were tested for anti-BP180 antibodies by indirect immunofluorescence, immunoblotting and enzyme-linked immunosorbent assay (ELISA). RESULTS: We found anti-BP180 antibodies in the sera of the four infants with all methods. These antibodies reacted with the extracellular domain NC16A. In the serum of their mothers we found 180 and 160 kDa proteins, each in one case, but indirect immunofluorescence and ELISA were negative, suggesting the absence of anti-BP180 autoantibodies reacting with the extracellular domain NC16A. CONCLUSIONS: BP of infants is not due to maternofetal transmission of pathogenic autoantibodies. Other hypotheses for the pathophysiology of BP are discussed.     

The 120-kDa soluble ectodomain of type XVII collagen is recognized by autoantibodies in patients with pemphigoid and linear IgA dermatosis

BACKGROUND: Type XVII collagen promotes adhesion of basal keratinocytes to epidermal basement membrane, and is the target of disease in patients with certain inherited or acquired blistering diseases. Two forms of type XVII collagen are produced by cultured human keratinocytes: a 180-kDa full-length, transmembrane protein, and a recently identified 120-kDa soluble fragment that corresponds to its collagenous ectodomain. OBJECTIVES: We aimed to determine the incidence and pattern of reactivity of autoantibodies against the 180- and 120-kDa forms of type XVII collagen in sera from 40 patients with bullous pemphigoid (BP), pemphigoid gestationis or cicatricial pemphigoid (CP), as well as six patients with linear IgA dermatosis (LAD). METHODS: Various immunochemical techniques were used. RESULTS: These studies found that the 120-kDa fragment of type XVII collagen was bound by circulating autoantibodies in 13 of 38 patients with BP or CP and all six patients with LAD. While many pemphigoid sera had specific reactivity against one but not both forms of this protein, autoantibodies from patients with LAD bound only the soluble ectodomain. CONCLUSIONS: These findings are consistent with the presence of both neoepitopes and cross-reactive epitopes on the ectodomain of type XVII collagen. The finding that sera from patients with LAD showed specific reactivity to epidermal basement membrane suggests that such neoepitopes are present in human skin and that their targeting by autoantibodies may contribute to disease pathogenesis.     

Bullous pemphigoid: serum antibody titre and antigen specificity

Abstract 2 antigens have been identified as possible targets for autoantibody depositions in bullous pemphigoid: a 230-kD protein (BP230) and a 180-kD protein (BP180). We studied the relationship of these 2 antigens with the immunofluorescence determined serum antibody titre. 2 groups of bullous pemphigoid patients were selected on the basis of immunoblot-determined antibody specificity. One group (13 patients) had antibody specificity for BP230 and not for BP180, while the other group (9 patients) had antibody specificity for BP180 and not for BP230. The immunofluorescence titres of the circulating antibodies determined on monkey oesophagus substrate displayed, for the BP230-specific group, a mean of 1:1102. The maximal observed titre was 1:5120. The mean titre in the BP180-specific group was only 1:29, with a highest titre of only 1:160. This result suggests that in routine indirect immunofluorescence of bullous pemphigoid sera, the contribution of the BP180-specific antibodies to the total anti-epidermal basement membrane zone antibody titre is relatively much lower than that of the BP230-specilic antibodies. Thus, at high dilutions, only the BP230-speeific antibodies contribute to the overall indirect immunfluorescence titre.     

Development of a novel ELISA system for detection of anti-BP180 IgG and characterization of autoantibody profile in bullous pemphigoid patients

BACKGROUND: The NC16A immunodominant region of the bullous pemphigoid (BP) antigen BP180 has been used to develop several enzyme-linked immunosorbent assays (ELISAs) as diagnostic tools for BP autoantibody detection. OBJECTIVES: Because BP180 autoantibody reactivity is not restricted to NC16A, we have investigated the possibility of developing an ELISA based on selected epitopes additional to this immunodominant region. METHODS: Initially 78 BP sera were tested using an NC16A ELISA and IgG reactivity was detected in 64 BP sera (82%). The 14 NC16A-negative BP sera were then analysed by immunological screening against seven BP180-specific epitopes. Recombinant phages displaying BP180 epitopes were grown as plaques, blotted onto a nitrocellulose filter and incubated with BP sera. RESULTS: Three and five NC16A-negative BP sera reacted with epitopes AA 1080-1107 and AA 1331-1404 of the BP180 ectodomain, respectively. Thus, a novel ELISA with GST-1080 and GST-1331 (GST-1080/1331) was developed: 32 of 78 BP sera (41%) proved positive by this assay. The combined use of ELISAs with GST-NC16A and GST-1080/1331 detected IgG reactivity in 72 of 78 BP sera, increasing the sensitivity from 82% to 92%. In addition, autoreactivity against the three extracellular epitopes appeared to be related to the presence of both skin and mucosal involvement as assessed by Fisher's exact probability test. CONCLUSIONS: Our findings further characterize the autoimmune response in BP by identifying a subgroup of NC16A-negative patients who react with different BP180 extracellular epitopes. The developed ELISA system appears more sensitive than the ELISA based on NC16A alone and also informative about the epitope profile of BP patients.     

Omalizumab as an alternative therapeutic tool in the treatment of bullous pemphigoid: A case report

Bullous pemphigoid (BP) is an acquired autoimmune bullous disease characterized by autoantibodies against the hemidesmosomal proteins found in the basal keratinocytes of the basement membrane zone (BMZ): a 180 kDa protein (type XVII collagen) mainly and the 230 kDa antigen. There is such evidence that the antibodies against the BMZ components are not only of IgG type, but also this bullous disease may have IgE antibodies directed to the BMZ that contribute to the pathogenesis of the disorder. IgE is not only thought to contribute to the pathogenesis of BP, it has also been suggested that eosinophils play a role in the development of the first signs associated with BP. A humanized monoclonal antibody directed to IgE, omalizumab, is approved for the treatment of severe asthma and chronic spontaneous urticaria, and it may be useful in the treatment of BP in the first stages of the disease.     

大疱性类天疱疮230000和160000抗原的区域性分布

利用免疫印迹技术筛选出2份大疱性类天疱疮(BP)血清,一份只和230 000分子结合,一份只和160 000分子结合,分别和人体22处正常皮肤作间接免疫荧光(ⅡF),发现230 000抗原和160 000抗原的表达存在明显的区域性差异.230 000抗原在胸腹和大腿屈侧含量最高,头皮和足跖最低.160 000抗原在胸腹含量最高,头皮和颈部含量最低.230 000抗原和160 000抗原在同一部位的表达基本一致,但在腋窝、大腿伸侧和大腿屈侧三个部位明显高于后者.上述差异与临床上BP的好发部位有一定关联,靶抗原表达的区域性差异是造成自身免疫性皮肤病皮疹特异分布形式的一个重要原因.