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.     

Molecular genetic analysis of HLA class II alleles in Japanese patients with melanoma

Distribution of HLA-DQA, -DQB and -DPB alleles in ninety-six Japanese patients with melanoma was analyzed using the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method, and the association between clinical parameters and the presence of certain HLA class II alleles investigated. The frequency of HLA-DQB 1*0302 was increased, while those of DQA1*0101(04), -DQA1*0401 and DRB1*0802 were decreased in melanoma patients compared with controls. Moreover, the frequency of HLA-DQA 1*0103 in patients with acral lentiginous melanoma was increased compared with controls. However, none of these HLA class II alleles showed significant positive or negative associations after correction of the P value. In addition, there was no correlation between these antigens and clinical parameters. These results indicate that HLA class II alleles may not contribute to a strong susceptibility to melanoma in the Japanese     

HLA class I and II polymorphisms in Azores show different settlements in Oriental and Central islands

Human leucocyte antigen-A, -B, -Cw, -DRB1, -DQA1 and -DQB1 polymorphisms were examined in the Azorean population. The data were obtained at high-resolution level, using polymerase chain reaction (PCR) with sequence-specific primer, PCR-sequence-specific oligonucleotides and sequence-based typing. The most frequent allele in each locus was: A*0201 (24.5%), B*510101 (9.8%), Cw*0401 (14.8%), DRB1*070101 (18.3%), DQA1*0201 (17.4%) and DQB1*0301 (19.4%). The predominant extended haplotype was A*0202-B*1503-Cw*0202-DRB1*090102-DQA1*0303- DQB1*0202 (1.9%), which was found to be absent in the Portuguese mainland. The present study corroborates historical sources that say the Azores were populated not only by Portuguese but also by other Europeans, mostly Flemish people. Despite dendrogram analysis showing some remote Asian genetic affinities, the lack of specific alleles and haplotypes from those populations does not allow us to conclude for direct influence. Haplotype and allele frequencies in Azores show no homogeneous distribution between Oriental and Central islands of this archipelago. The Oriental islands harbour several haplotypes already found in mainland Portugal and identified as Mediterranean and European. The Central group of islands on the contrary clearly shows an influence of north Europeans (most probably derived from a well-documented Flemish settlement), with much less affinity to mainland Portugal.     

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。     

A sensitive fluorometric assay for quantitatively measuring specific peptide binding to HLA class I and class II molecules

A sensitive, highly reproducible assay was developed for measuring binding of peptides to various HLA class I and II alleles. The assay is based on competition for binding to HLA between a peptide of interest and a fluorescent labelled standard peptide. This mixture is incubated with HLA to obtain equilibrium binding, and subsequently separated on an HPLC size-exclusion column in (i) a protein fraction containing HLA and bound peptide and (ii) a free peptide fraction. Each assay uses only 100 fmol labelled peptide and approximately 10 pmol of HLA. The analytical system contains an autosampler that samples from 96-well microtiter plates. Injections and data recording/evaluation is fully automated. Typical analysis time is 10–12 min per sample. The fluorescence in the HLA-bound peptide and free peptide containing fractions is measured on-line. The ratios of fluorescence signal in protein and peptide fractions at various concentrations of the peptide of interest are determined. IC₅₀ values are calculated from the binding curve as obtained by curve fitting of the data. Here we show results for peptide binding to HLA-DR1 and -DR17 molecules purified from detergent solubilized cell lysates, and for recombinant HLA-A^* 0201 and HLA-A^* 0301 expressed in E. coli.

The assay reported is sensitive and reproducible. It is non-radioactive and is non-labor intensive due to the high degree of automation.     

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.     

MHC class II alleles and haplotypes in patients with pemphigus vulgaris from India

Pemphigus vulgaris (PV) is a blistering disease of the skin and mucous membranes characterized by an autoantibody response against a keratinocyte adhesion molecule, desmoglein 3, causing acantholysis and blister formation. We compared high resolution MHC class II alleles and haplotype frequencies (HLA-DRB, DQA1 and DQB1) in 37 patients with PV to 89 haplotypes of normal relatives from New Delhi and Ahmedabad. We found that PV patients had significantly increased frequencies of DRB1*1404 (P<0.0001), DQA1*0101 (P=0.001), and DQB1*0503 (P<0.0001). These associations were due to the increased frequencies of the haplotype HLA-DRB 1 * 1404, DRB3*0202, DQA1*0101, DQB1*0503 in patients compared to control haplotypes (p<0.0001). Also, patients from Ahmedabad had a significant increase in HLA-DQB 1*0302 (p=0.03). An identical amino acid sequence (Leu-Leu-Glu-Arg-Arg-Arg-Ala-Glu), in positions 67–74 of the P domain of DRB alleles is restricted to some DR14 alleles. Therefore, there are three possible explanations for class II allele involvement in autoantibody in PV patients with class II haplotypes marked by HLA-DR14. First, the class II alleles could be markers for an unidentified susceptibility gene in linkage disequilibrium with them. Second, the primary association could be with DQB 1*0503 and the association with HLA-DR14 alleles would be the result of linkage disequilibrium. Third, the HLA-DRB 1 locus susceptibility could involve a specific amino acid sequence in the third hypervariable region shared by several HLA-DR14 alleles.     

Possible impact of HLA class I and class II on malignancies driven by a single germ-line BRCA1 mutation

This study provides the first immunogenetic preliminary evidence that specific human leucocyte antigen (HLA) class I and class II alleles and haplotypes may be relevant for BRCA1 c.5263_5264insC driven oncogenesis. Observed HLA associations might have practical implications for establishment of predictive markers for the response to immunotherapies in malignancies driven by this germ-line mutation.     

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.     

A one-step monoclonal antibody typing procedure that simplifies HLA class I and class II typing

Abstract: We have developed monoclonal antibodies to most HLA specificities, making it possible for us to devise a simple, rapid, one-step micro-cytotoxicity test. The test is performed by adding 1 μl of cells to 1 μl of antibody-complement mixture predotted on the microtest tray. The reactions are read following a 1-hour incubation period (30 minutes in some instances). The analysis of reactions seen on testing 105 class I antibodies and 50 class II antibodies is shown. A comparison of typing by the standard NIH method and the new one-step procedure showed a > 96% concordance in the 500 T cells and 200 B cells we examined. Class I and class II typing could be performed using B cells, thus obviating the need to isolate both T and B cells for HLA typing.     

Distributions of HLA class I alleles and haplotypes in Northern Han Chinese

Human leukocyte antigen (HLA) class I allelic genotypes were determined in 105 unrelated Han ethnic individuals inhabiting the northern China area. A total of 19 HLA-A alleles, 49 HLA-B alleles and 24 HLA-Cw alleles were detected. Through the analyses of two and three loci haplotypes of HLA-A, HLA-B, and HLA-C loci, 11 HLA-A-B-Cw haplotypes, 19 HLA-A-B haplotypes, 18 HLA-A-Cw haplotypes, and 24 HLA-B-Cw haplotypes with the frequencies of higher than 0.01 were revealed. The Nei's genetic distance (GD) was estimated, and the NJ dendrogram showed that Northern Han had a higher GD to Southern Han (0.233) than those to the Korean (0.138) or other Northern ethnic groups, suggesting that Northern Han had more mixed blood with the ethnic groups originally in Northeast Asia. Our results provide useful information on the further study of evolution and relationships of Chinese ethnic groups and disease association in terms of HLA class I genes.     

HLA class I and class II genotyping in patients with Behcet's disease: a regional study of eastern part of Turkey

Class I human leucocyte antigen (HLA)-B51 is well known to be associated with Behcet's disease in many ethnic groups. However, there has been no published paper with respect to its association with HLA class I and class II among the Turkish people who live in the eastern region of Turkey. Moreover, as it is known that B51 antigen is encoded by 21 alleles, B*5101-5121, HLA-B51 allele typing was performed, as well as HLA class I and class II genotyping of 75 patients with the disease and the 54 individuals in the matched control group. The result shows that the HLA-B51 frequency was significantly higher (58.66%) in the patient group, compared to that in the control group (18.51%) (OR = 6.245). In the subtyping of B51 alleles, 44 B51-positive patients possessed B*5101 (45.5%), B*5108 (25%), B*5105 (9.1%) and B*5104 (4.5%). There was no significant difference in the HLA-B51 allelic distribution between the patient group and the control group. However, homozygous carriers of HLA-B51 showed considerably high risk (OR = 2.647) in the patient group, compared to that in the control group. In the genotyping of class II HLA alleles, while HLA-DRB1*04 (45.3%) and HLA-DRB1*07 (24%) were the predominant alleles in the patient group, DRB1*11 (50%) appeared to be more common in the control group.     

Expression of HLA class I/II antigens and T cell immune response in human neuroendocrine tumors of the pancreas

Peptide presentation by HLA class I and II antigens regulates specific antigen recognition by T cells. The present study aimed to investigate T cell infiltration and its relation to HLA antigen expression in pancreatic neuroendocrine tumors. Fresh tissue samples were collected from five insulinomas and six other neuroendocrine tumors (one gastrinoma, one glucagonoma, two carcinoid, and two neuroendocrine carcinomas). Normal pancreatic and splenic tissue samples were used as controls. Investigation of infiltrating lymphocyte populations, as well as staining of HLA class I and II antigens, were performed by standard immunohistochemistry. The majority of investigated tumors demonstrated an intratumoral infiltration by CD3+, CD4+ and CD8+ T cells that was significantly higher than in normal pancreatic islets. Only a minority of tumor-infiltrating T cells showed the CD45RO+ phenotype. The expression of HLA class I antigen was altered in 10 of 11 tumors. A loss of beta-2microglobulin represented the most frequent type of alteration to HLA class I expression, although the total loss of HLA class I was found in only one case of neuroendocrine carcinoma. HLA class II molecules were expressed by endothelial and lymphoid cells and not by tumor cells. In conclusion most neuroendocrine pancreatic tumors induce a T cell mediated immune response resulting in an intratumoral infiltration with CD3+, CD4+ and CD8+ T cells. Loss of beta-2microglobulin is a frequent alteration in these tumors, which may influence the normal function of the HLA class I antigen complex. In contrast to malignant tumors of the exocrine pancreas, expression of HLA class II was absent in neuendocrine pancreatic tumor cells.     

Soluble HLA class I and HLA-DR plasma levels in patients with anterior uveitis

Anterior uveitis (AU) is an autoimmune disease frequently associated with HLA-B27 antigen. Because of the immune regulatory properties of soluble human leukocyte antigen (sHLA) molecules, we quantified sHLA class I (sHLA-I) and sHLA-DR plasma levels in HLA-typed AU patients (n = 60). Randomly selected healthy individuals (n = 128) and HLA-B27 antigen-positive individuals (n = 24) with HLA phenotype frequencies similar to the HLA-B27 antigen-positive AU patients served as control panels. As expected, HLA-B27 phenotype was significantly increased in AU patients (n = 60), compared to healthy controls. Mean sHLA-I levels in AU patients were slightly higher than in randomly selected healthy controls. Regarding AU subgroups, elevated sHLA-I levels were only found in HLA-B27 antigen-negative patients. Compared to controls, sHLA-DR levels were significantly increased in AU patients and the subgroups of HLA-B27 antigen-negative and -positive patients but not Fuchs' heterochromic cyclitis (FHC). AU patients negative for HLA-B27 antigen with a chronic course had higher sHLA-DR levels than those with an acute course. The presence of associated systemic diseases in AU patients was related to elevated sHLA-DR levels. Secretion of sHLA-DR in blood differs among the various forms of AU. Systemic immune activation was present in AU but not in FHC.     

HLA class II antigens in South African Blacks with type I diabetes

Type 1 diabetes mellitus is poorly characterised in many African communities, including South Africa, where little is known of the disease epidemiology. This study aimed to identify the HLA class II alleles associated with type 1 diabetes in a group of Zulu subjects in Durban, KwaZulu-Natal by PCR-SSP. The HLA alleles associated with type 1 diabetes included HLA-DQB*0302 (P<0.0001), DRB1*O9 (P<0.0001), DRB1*04 (P=0.002), DRB1*0301 (P=0.003), DQB*02 (P=0.004) and DQA*03 (P=0.035). Estimated haplotypes positively associated with type 1 diabetes included HLA-DRB1 *0301-DQA*0501, DRB1*04-DQA*03, DRB1*04-DQB*0302, DRB1*0301-DQB*0201, DQA*0501-DQB*0201 and DQA*03-DQB*0302. These findings are similar to those reported from Zimbabwe and other populations with type 1 diabetes.