HLA haplotypes and class II molecular alleles in Sardinian and Italian patients with pemphigus vulgaris

HLA class II antigens and DRB1, DQA1, DQB1 alleles were studied in 16 Italian and in 16 Sardinian patients with pemphigus vulgaris (PV). In the last group the complete HLA A-DQ haplotypes, including the complotypes, were defined by family studies. As in other populations, two PV susceptibility haplotypes were found: HLA-DRB 1*0402, DQA1*0301, DQB1*0302 and HLA-DRB 1*1401, DQA1*0104, DQB 1*0503. The first haplotype was largely prevalent in the Sardinian patients and was a part of the extended haplotype HLA-A2, Cw4, B35, S31, DR4, DQ8. The strength of the allele associations to PV is in agreement with the view that the main PV susceptibility genes are the DRB 1*0402 and DQB 1*0503 alleles. A genetic resistance to PV seems to be conferred by the HLA-DR3, DQ2 haplotype in the Sardinian population.     

Distribution of HLA class II alleles among Spanish patients with pemphigus vulgaris

Abstract: Twenty-six unrelated Spanish Caucasian individuals affected by pemphigus vulgaris (PV) were HLA typed and frequencies compared with those of 200 ethnically matched healthy controls. Twenty-three out of 26 patients were HLA-DR4. The frequency of HLA-DR14 was also increased (31%; controls: 4%). Of the 23 patients positive for HLA-DR4, 21 carried the DRB1*0402 allele. Therefore, the frequency of HLA-DRB1*0402 among patients was 81% (4% in controls; P =4.7times 10^-27, OR=100.8). Interestingly, HLA-DR13, a frequent HLA-DR specificity in the Spanish general population (27%), was absent among the PV patients ( P =0.009; P _c=0.1; OR=0.05). Taking together these data, we can conclude that, in the Spanish population, PV is preferentially and strongly associated with HLA-DRB1*0402, whereas DRB1*13 seems to confer a protective effect in our population.     

Serological and molecular analysis of HLA class I and II alleles in Thai patients with psoriasis vulgaris

Abstract: The HLA class I and class II alleles in 67 patients with type I psoriasis vulgaris, 23 patients with type II psoriasis vulgaris and 140 healthy individuals were analyzed. The frequencies of HLA-A2, -B46, -B57 and DQB1*0303 were significantly increased in type I psoriasis compared to the controls (Pc<0.05). Molecular analysis of HLA-A2 alleles showed an increase in HLA-A*0207 and a decrease in HLA-A*0203 in type I psoriasis. HLA-DQBl*0301 was significantly decreased in type I psoriasis compared to the normal controls (Pc<0.05). No association of any alleles with type II psoriasis was observed. This data demonstrated two susceptible haplo-types: HLA-A1-B57-DRB1*0701-DQA1*0201-DQB1*0303 (AH57.1) and HLA-A2-B46-DRB1*0901-DQA1*0301-DQB1*0303 (AH46.1) for type I psoriasis in the Thai population. Besides, the haplotype AH46.1 was also associated with type II psoriasis.     

HLA-DRB1*0402 haplotypes without DQB1*0302 in Venezuelan patients with pemphigus vulgaris

The two basic forms of autoimmune intraepidermal blistering diseases, pemphigus vulgaris (PV) and pemphigus foliaceus (PF), affect different layers of the skin, have different symptoms and target different antigens. We have defined human leukocyte antigen (HLA)-DRB1-DQB1 alleles and haplotypes in a case-control study of 66 non-Jewish patients attending a public reference Hospital over the past 10 years. The control group consisted of 101 matched individuals tested also by medium to high-resolution polymerase chain reaction-sequence-specific oligonucleotide with primers and probes from the 12th and 13th International Histocompatibility Workshop. Patients and controls were descendants of three-generation individuals born in the country. Among the patients, 49 had PV, 50% showed predominantly mucosal involvement, 50% showed predominantly the cutaneous clinical phenotype and 17 had PF. Statistically significant HLA-DR frequency differences between patients with PV and controls were found only for DRB1*0402 and DRB1*1401 [odds ratio (OR) = 27.22, confidence interval (CI) 94.7-7.82, P= 1.1 x 10(-14) and OR = 46.56, CI 801.4-2.70 P= 7.5 x 10(-6), respectively]. Both alleles were also increased in the patients with PF compared with the controls (OR = 7.0, P= 0.038 and OR = 21.64, P= 0.009, respectively), but the significance of the difference did not resist Bonferroni correction. Haplotype analysis showed that DRB1*0402 was always present with DQB1*0302 and DRB1*1401 with DQB1*0503, but no independent effect of the DQB1*0302 in the former haplotype was evident. Our results support the hypothesis that the DRB1*0402 without DQB1*0302 is the most relevant HLA-DRB1 allele responsible for the pathogenesis of pemphigus in Venezuelan patients with PV and discard the DQB1*0302 influence observed in other populations.     

HLA-DRB1、DQB1基因与汉族人群寻常型天疱疮的相关性研究

目的　探讨 HL A- DRB1、DQB1位点基因在汉族人群寻常型天疱疮易感性中的作用。方法用序列特异性引物 -聚合酶链反应方法 ,对 6 1例寻常型天疱疮 (pemphigus vulgaris,PV)患者和 5 7名正常对照进行了 HL A- DRB1、DQB1等位基因的分型 ,并分析了 DRB1、DQB1基因在两组中的分布。结果　与正常对照组比较 ,PV组 DR4、DRB1* 14 (* 14 0 1、* 14 0 4、* 14 0 5 )基因频率明显增高 (Pc分别 <0 .0 5及P<0 .0 1) ,差异有显著性 ;PV组 DQB1* 0 5 0 3、DQB1* 0 30 2基因频率明显增高 (Pc均 <0 .0 5 ) ,差异有显著性。对 DR4阳性样本的组内基因亚型分型结果发现 ,PV组中 DRB1* 0 4 0 3、DRB1* 0 4 0 6频率显著增高(Pc<0 .0 5 ) ,差异有显著性。 PV患者组单倍型 HL A- DRB1* 0 4 ,DQB1* 0 30 2和 HL A- DRB1* 14 ,DQB1* 0 5 0 3频率明显增高 (P<0 .0 5 )。结论　HL A- DRB1* 0 4 ,DQB1* 0 30 2和 HL A- DRB1* 14 ,DQB1* 0 5 0 3可能是汉族人 PV推测的易感单倍型。     

MHC class II genes in Mexican patients with idiopathic dilated cardiomyopathy

The purpose of the present study was to evaluate the relationship between class II major histocompatibility complex (MHC) genes (HLA-DR and HLA-DQB) and the genetic susceptibility to idiopathic dilated cardiomyopathy (IDC) in Mexican patients. The HLA-DR and DQB alleles were analyzed in 53 patients with IDC and 99 ethnically matched healthy controls using the polymerase chain reaction-sequence specific oligonucleotides (PCR-SSO) technique. IDC patients showed increased frequencies of HLA-DR4 (pC=0.02, OR=1.87), HLA-DQB1*0301 (pC=0.02, OR=1.92) and HLA-DQB1*0302 (pC=0.02, OR=1.87) when compared to healthy controls. On the other hand, IDC patients also showed decreased frequencies of HLA-DR11 allele (pC=0.03, OR=0.26) and HLA-DQB1*0201 (pC=0.04, OR=0.41). These data suggest that variation in class II HLA alleles could be a genetic factor involved in the susceptibility to IDC of the Mexican Mestizo population.     

The imbalance of gut microbiota and its correlation with plasma inflammatory cytokines in pemphigus vulgaris patients

Pemphigus vulgaris (PV) is an autoimmune disease characterized by the production of IgG autoantibodies owing to an imbalance in the Th1/Th2 and Th17/Tregs cell pathways. The role of gut microbiota in the development of immune system and autoimmune diseases has been unraveled in the last two decades. However, data pertaining to gut microbiota of PV patients is largely lacking. We aimed to compare the gut microbiota of PV patients and healthy controls and assessed potential correlation with circulating cytokines of Th1/Th2/Th17 cell. Faecal bacterial diversity was analysed in 18 PV patients and 14 age‐ and gender‐matched healthy individuals using hypervariable tag sequencing of the V3‐V4 region of the 16S rRNA gene. Plasma levels of 20 inflammatory cytokines were assessed using the Luminex screening system. As a result, we identified 10 differentially abundant taxa between patients and controls. At the genera level, Lachnospiracea_incertae_sedis and Coprococcus decreased, while Granulicatella, Flavonifractor enriched in PV. Plasma levels of C5a, interleukin (IL)‐2R, IL‐6, IL‐8, IL‐7, IL‐1β, IL17A, IL‐5 and IL‐21 were significantly increased in PV Flavonifractor exhibited a positive correlation with C5a, IL‐6, IL‐8, IL‐7, IL‐1β, IL17A and IL‐21. Lachnospiracea_incertae_sedis and Coprococcus showed a negative correlation with IL‐17A. Our results are consistent with the hypothesis that PV patients have gut microbial dysbiosis which might contribute to the immune disorder and the development of PV.     

Diagnostic features of pemphigus vulgaris in patients with pemphigus foliaceus: detection of both autoantibodies, long-term follow-up and treatment responses

There are several studies that describe the simultaneous presence and conversion of pemphigus foliaceus into pemphigus vulgaris and vice versa. We describe eight patients with clinical, histological and immunopathological features of pemphigus foliaceus, at the time of the initial diagnosis. After a mean period of 2.5 years, additional serological features of pemphigus vulgaris were observed. During a long-term follow-up, systemic therapies, their durations and treatment outcomes were recorded. These patients did not respond to conventional systemic therapy and developed multiple side-effects from these drugs. Hence, they were treated with intravenous immunoglobulin therapy (IVIg). Prior to the initiation of IVIg therapy, different assays were performed to detect the presence of autoantibodies, including indirect immunofluorescence (IIF), immunoblot assay using bovine gingival lysate, and ELISA. Twenty-five healthy normal individuals, 12 patients with pemphigus vulgaris, and eight patients with pemphigus foliaceus served as controls for comparison of serological studies. At the time of initial diagnosis, the sera of all eight study patients also demonstrated binding on an immunoblot assay to a 160-kDa protein (desmoglein 1) only. This is typically observed in pemphigus foliaceus. Prior to staring IVIg therapy, binding was observed to both the 160 kDa and 130 kDa (desmoglein 3) proteins on an immunoblot assay which was characteristic of pemphigus vulgaris. The antidesmogleins, 1 and 3 autoantibodies, were predominantly of the IgG4 subclass in all eight patients studied. IVIg therapy induced remission in four patients and control in four of the eight patients. The total follow-up period ranged from 2.6 to 9.5 years (mean 5.3 years). It is difficult to determine the exact time at which these patients with pemphigus foliaceus developed pemphigus vulgaris. It is possible that the disease was nonresponsive to conventional immunosuppressive therapy owing to the simultaneous presence of two autoantibodies.     

Common ancestral origin of pemphigus vulgaris in Jews and Spaniards: a study using microsatellite markers

Pemphigus is a group of autoimmune blistering diseases of the skin and mucous membranes. The association of pemphigus with human leukocyte antigen (HLA) is widely accepted. It was described in many ethnic groups and in most countries of the world. Studies showed that the associated HLA haplotype in Jewish pemphigus vulgaris (PV) patients is HLA-B38, DRB1*0402, and DQB1*0302; or HLA-B35, DRB1*0402, and DQB1*0302. Similar associations with class II genes were found in Spanish non-Jewish PV patients. As Jews lived in Spain for hundreds of years and many converted to Christianity, the presence of the same HLA haplotype in the Jewish and Spanish PV suggests that they may share the same founder. Microsatellite markers which span the entire major histocompatibility complex (MHC) locus were used as genetic probes. They were utilized to dissect the MHC region in the search for possible common haplotypes, besides HLA, which may provide an answer to this question. It was found that in both cohorts, in addition to HLA class II genes, there are probably genes in the class I region which are associated with PV. Alleles belonging to the associated markers were used to construct haplotypes and to estimate genetic distances. The distance between the two PV cohorts is relatively short, but the distance between the Jewish patients and the Jewish controls is greater compared to the distance between Spanish patients and Spanish controls. In both PV populations, the same microsatellite haplotypes in addition to a common class II haplotype were found, suggesting that both patient populations originated from the same genetic stock and, therefore, share the same ancestral disease gene.     

寻常型银屑病血清瘦素水平与HLA等位基因相关性研究

目的:研究中国北方汉族人寻常型银屑病(PV)患者血清瘦素(leptin)水平与HLA等位基因的关联性。方法:采用序列特异性引物-聚合酶链反应(PCR-SSP)方法对91例PV患者和102例健康人进行HLA-A、B、Cw、DRB1及DQB1分型,放射免疫分析(RIA)技术测定其血清瘦素水平。结果:(1)PV患者血清瘦素水平(7.63±4.44 ng/ml)较正常对照(5.03±2.72 ng/ml)显著增高(P<0.001);男性及女性PV患者血清瘦素水平分别较男性及女性正常对照显著增高(P=0.004,P<0.001)。(2)北方汉族人PV与HLA-A*010x(Pc=0.039 4)、A*300x(Pc=0.012 9)、B*570x(Pc=0.013 7)、Cw*0602(Pc=0.000 9)、Cw*060x(除外Cw*0602)(Pc<1×10-4)、DRB1*0701/0702(Pc=0.001 2)及DQB1*0201(Pc=0.005 6)等位基因呈正相关,与Cw*040x(Pc=0.018 8)呈负相关。(3)携带HLA-DRB1*15及DRB1*0701/0702等位基因的男性PV患者和携带HLA-A*010x、A*24、Cw*040x、DRB1*15、DQB1*0301等位基因的女性患者的血清瘦素水平均显著增高(P<0.05)。结论:(1)HLA-A*010x、A*300x、B*570x、Cw*0602、Cw*060x(除外Cw*0602)、DRB1*0701/0702及DQB1*0201等位基因可能是北方汉族人PV的易感基因或与易感基因相连锁,HLA-Cw*040x可能是预防PV发病的“保护因子”。(2)PV患者血清瘦素水平显著增高,并与HLA等位基因相关联,提示HLA等位基因和瘦素参与PV发病过程。     

Pathogenic activity of circulating anti-desmoglein-3 autoantibodies isolated from pemphigus vulgaris patients

Introduction

There are scarce data on immunochemical properties of pemphigus antibodies detected in clinical remission in pemphigus vulgaris (PV) patients. The aim of the study was to compare biological activity of anti-Dsg3 autoantibodies purified from the sera of PV patients in active stage and in clinical remission.

Material and methods

The effect of purified antibodies on expression of procaspase-3, Bax, Bcl-2, uPAR, IL-1β, IL-6, and TNF-α mRNAs in the HaCaT keratinocytes was evaluated by Western blot and RT-PCR method.

Results

Incubation of HaCaT cells with anti-Dsg-3 autoantibodies caused their binding to cell membranes surfaces. Anti-Dsg3 autoantibodies isolated from the patients in active stage and clinical remission showed proapoptotic effect, caused enhanced expression of analyzed proinflammatory cytokines’ mRNAs and uPAR mRNA.

Conclusions

Our data revealed similar pathogenic activity of anti Dsg-3 autoantibodies isolated from active and clinical remission PV patients.     

Human leukocyte antigen class II alleles in Caucasian women with primary biliary cirrhosis

In the current study, we investigated human leukocyte antigen (HLA) class II alleles in Caucasian women with primary biliary cirrhosis (PBC), a disease that preferentially affects women. Alleles of DRB1, DQA1, and DQB1 were determined by DNA-based HLA typing for women with PBC (n = 72) and healthy women (n = 381). All study subjects were Caucasian. HLA DRB1*08 was significantly increased in women with PBC compared to healthy women. The increase was primarily due to the DRB1*0801 allele, also the most common DRB1*08 allele among controls. DQB1*04 and DQA1*0401 were significantly increased. DRB1*1501, DQA1*0102, and DQB1*0602 were associated with decreased risk. Analyses conducted comparing parous women with PBC to parous healthy women (n = 68 and n = 282, respectively) yielded similar significant results. Although the DRB1*08-DQA1*0401-DQB1*04 haplotype was significantly associated with PBC, consistent with other studies, this haplotype nevertheless represented only 19% (14/72) of all PBC patients and can account for only a minority of the risk of PBC.     

Role of HLA-B and C alleles in development of psoriasis in patients from North India

Abstract: The association of HLA antigens with susceptibility for development of psoriasis vulgaris has been studied mostly using serologic methods. A number of different HLA class I antigens have been reported to be associated with predisposition to develop this disease. While Cw6 is the allele most commonly thought to confer risk for psoriasis, a number of HLA-B locus alleles have also been thought to be involved and, recently, in at least three studies, a specific amino acid in the HLA-C heavy chain has been implicated. With the recent development of molecular methods for typing for HLA class I alleles, it has become possible to re-examine this association by performing high resolution typing. In the present study we have investigated 38 psoriasis patients and 84 ethnically and geographically matched controls from North India. They were typed for HLA-B and HLA-C. The results showed the Cw*0602 was the main allele that was increased in this group of patients. The previously reported association with alanine-73 was not found to be significant. Cw*0602 was found in 71% of the patients. B*5701 and B*3701 were also increased but appeared to be secondary to linkage disequilibrium with Cw*0602.     

Role of direct immunofluorescence on Tzanck smears in pemphigus vulgaris

The Tzanck smear is a simple, sensitive, and rapid test to diagnose pemphigus vulgaris (PV), a life threatening autoimmune blistering disorder. The presence of acantholytic cells in cytology is indicative of but not specific for PV. Hence, a direct Immunofluorescence (DIF) test to demonstrate immunoglobulin deposits on the acantholytic cells would make the Tzanck test more specific, in addition to being a rapid test. Twenty untreated patients with PV confirmed histopathologically were enrolled to evaluate the efficacy of using DIF technique using IgG on Tzanck smear samples. The DIF smears were compared with DIF on skin biopsies in the same patient. This prospective pilot study approved by the institutional ethics committee was carried out in a tertiary health care hospital in a developing country. Of the 15 patients presenting within 3 mo of onset of the illness, 40% (n = 6) showed DIF positivity on Tzanck smear, when compared with 46.67% (n = 7) on skin biopsy. On the other hand, of the five patients presenting beyond 3 mo of their illness, only 20% (n = 2) showed positivity on Tzanck, when compared with all 100% (n = 5) on skin biopsy. The study, thus, suggests that DIF on skin biopsy is comparable to biopsy in diagnosing early PV. This preliminary study proposes that the use of DIF on Tzanck smear is a simple, rapid, painless, and user-friendly out-patient procedure for the diagnosis of early PV, even for relatively inaccessible lesions in the oral cavity and flexural regions. This methodology would be of great help in outlying and rural facilities lacking proper histological equipment, thus avoiding the need for a surgical or punch biopsy or heavy investment in laboratory equipment and expertise. Probable reasons for DIF negativity on Tzanck smears are also discussed.     

Analysis of the reactivity of indirect immunofluorescence in patients with pemphigus foliaceus and pemphigus vulgaris using rat bladder epithelium as a substrate

OBJECTIVES:

To evaluate the reactivity of indirect immunofluorescence using rat bladder epithelium as a substrate in patients with pemphigus foliaceus and pemphigus vulgaris from the Department of Dermatology, University of São Paulo Medical School, Brazil.

METHODS:

Thirty-two patients (8 male and 24 female) from the Department of Dermatology, University of São Paulo Medical School, were selected. Three had mucosal pemphigus vulgaris, 20 had mucocutaneous pemphigus vulgaris, and 9 had pemphigus foliaceus. Patients'' sera were tested by indirect immunofluorescence performed on human foreskin and rat bladder epithelium and by ELISA assays utilizing baculovirus-expressed recombinant desmoglein 3 and desmoglein 1.

RESULTS:

No patients with mucosal pemphigus vulgaris, 5 of 20 patients with mucocutaneous pemphigus vulgaris (25%) and 4 of 9 patients with pemphigus foliaceus (44%) had positive indirect immunofluorescence using rat bladder epithelium as a substrate.

CONCLUSION:

Indirect immunofluorescence using rat bladder epithelium as a substrate is recommended whenever a diagnosis of paraneoplastic pemphigus is considered. The identification of a subset of pemphigus foliaceus and pemphigus vulgaris patients that recognizes desmoplakins by this laboratory tool is critical to avoid the misdiagnosis of paraneoplastic pemphigus.     

HLA-DQA1, -DQB1 and -DRB1 genotyping in Japanese pemphigus vulgaris patients by the PCR-RFLP method

Abstract: We performed HLA-DQA1, -DQB1 and -DRB1 genotyping using the PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism) method for 32 Japanese pemphigus vulgaris (PV) patients. There was a significant association of either DQB1*0503 or DRB1*1405 with PV, and a negative association of either DQA1*0103 or DQB1*0601 with PV was found. Since the DQB1*0503+ patients had various DR14-related alleles, we concluded that the association with DQB1 is primary and that the association with DRB1 is simply due to linkage disequilibrium between the DQ and DR genes. These results may indicate that specific HLA class II antigens confer the susceptibility to PV among Japanese.     

Association between HLA-DQB1 polymorphisms and pemphigus vulgaris: A meta-analysis

Objective: This study was performed to systematically summarize the results on the association of HLA-DQB1 polymorphisms with pemphigus vulgaris (PV) and other related factors.

Methods: A comprehensive literature search of PubMed, The Cochrane Library, Embase, and Google Scholar database was conducted to identify relevant articles in English, with the last report up to November 1, 2016. Heterogeneity test was performed, and publication bias was evaluated. Stata software 12.0 was used to perform the meta-analysis. Odds ratios (ORs) and 95% confidence intervals (CI) were used to describe the correlation by random-effects model.

Results: 18 studies were obtained after searching databases: 10 studies were about Caucasian, and 8 articles were about non-Caucasian. Meta-analysis revealed that the allele and phenotype frequencies of DQB1*05 were markedly higher in PV patients than in controls [P < 0.001, OR: 2.640, 95%CI: 1.570–4.441; P = 0.030, OR 3.688, 95%CI: 1.138–11.946]. In addition, DQB1*03 was significantly increased at the allele level [P < 0.001, OR: 2.080, 95%CI: 1.507–2.869], and DQB1*02 was significantly decreased in PV at the allele and phenotype levels [P = 0.002, OR: 0.450, 95%CI: 0.289–0.702; P = 0.001, OR: 0.293, 95%CI: 0.146–0.587]. When based on each subtype of HLA-DQB1, DQB1*05:03 and DQB1*03:02 may play susceptibility roles in PV, and DQB1*03:03, DQB1*05:01 and DQB1*06:01 are negatively associated with PV.

Conclusion: In summary, our study suggests that alleles from the groups DQB1*05 and DQB1*03, concretely DQB1*05:03 and DQB1*03:02, respectively, may be the susceptibility factors for PV at allele and phenotype levels, whereas DQB1*05:01, DQB1*02, DQB1*06:01, and DQB1*03:03 are negatively associated with PV.