HLA B27 polymorphism in Western India

We have characterized HLA B27 alleles in a sample population of Maharastra, Western Indians (n = 51), with the aim to investigate the different subtypes present among this population. The study was carried out using polymerase chain reaction with sequence-specific primers (PCR-SSP) and reverse line strip (RLS) techniques. Significant new findings have arisen from this study: B*2704, B*2705, B*2707, B*2708 and B*2714 alleles were found to be present, and two novel B27 alleles, B*2708 and B*2714, were found in this Indian population. In addition, B*2714 was observed in a patient with ankylosing spondylitis. This association has not been previously reported in ethnic groups from India.     

Susceptibility to ankylosing spondylitis is independent of the Bw4 and Bw6 epitopes of HLA-B27 alleles

We have characterized HLA-B27 alleles in a sample of the population from the Azores (n=46) with the aim of investigating the contribution of different subtypes to ankylosing spondylitis (AS). The study was carried out using PCR-SSOP and in some samples genomic sequencing was conducted. Some significant new finding have arisen from this study. First, B*2705,B*2702,B*2703,B*2707 and B*2708 alleles were found to be represented in this population. The polymorphism of B27 alleles found in a sample of the population from the Azores is higher than the Caucasian groups described. B*2703 and B*2707 have not previously been described to be represented in Caucasians and this could indicate admixtures with different populations of the world. In addition, the B*2708 allele was found to be associated with AS in a large family from the Azores. This association has not been previously reported in either ethnic group and needs to be confirmed in other population studies. This is of considerable interest since has only been described as a rare subtype underrepresented in the British population and has not been previously found to be associated with AS. B*2708 carries the sequence specifying the Bw6 epitope in contrast to most B27 alleles which carry a Bw4 sequence. Differences in this region (residues 77-83) can alter the F-pocket and affect T-cell recognition. The importance that these molecular changes can play in the pathogenesis of AS is discussed.     

High variability of HLA-B27 alleles in ankylosing spondylitis and related spondyloarthropathies in the population of northern Spain

The distribution of B27 alleles (B*2701-23) was characterized by PCR-SSP in ankylosing spondylitis and related spondyloarthropathies (SpA) in a sample of B27 positive patients from northern Spain. Six B27 alleles were identified: B*2705,02,03,07,08 and B*2713. B*2705 and 02 were the most common alleles in the SpA studied: ankylosing spondylitis (AS) (n = 89), reactive arthritis (ReA) (n = 11), psoriatic arthritis (PsA) (n = 29), and inflammatory bowel disease (IBD) (n = 21). B*2707 and B*2708 were found in PsA patients and B*2703 in one patient with IBD. B*2713 was identified in a healthy control family. B*2713 has not been reported to be represented in either ethnic group. Thus, this population shows higher levels of B27 diversity than other Caucasian groups.     

HLA-B27 polymorphism and worldwide susceptibility to ankylosing spondylitis

HLA-B27 is strongly associated to ankylosing spondylitis (AS) and represents a family of eleven B27 alleles (B*2701–11). Our aim was to analyze the distribution of B27 subtypes by PCR/SSOP and genomic sequencing in a large group of populations ( n =17). 711 B27-positive samples from Caucasoid, Asian, African, Amerindian and Polynesian populations were selected to ascertain transracial gene mapping of the B27 subtypes. 476 of these were AS patients, chosen to investigate the contribution of B27 alleles to AS susceptibility. Some significant new findings have arisen from this study: 1) B*2705 was the predominant subtype in circumpolar and subarctic areas. B*2702 was found to be practically restricted to Caucasian populations, showing a higher frequency in Middle-East (Jews) and North Africa (Arabs/Berbers) groups. 2) B*2703 appears associated with AS in Western Africans. This is of remarkable interest since it was suggested that B*2703 would be negatively disease-associated. 3) Although B*2706 appears negatively associated with AS in Thais, we identified two patients from northern China carrying it. This may be a reflection of a disease heterogeneity and could indicate that more than one pathogenic agent can be involved in AS. B*2709 has been recently described as negatively associated with AS in Sardinians. The molecular changes His 114Asp (B*2706) and Asp 116His (B*2709) could modify the genetic susceptibility to AS.     

HLA-B*27 subtypes in Northern and Northeastern Thais, Karens, and Bamars determined by a high-resolution PCR-SSP technique

Human leukocyte antigens (HLA), class I, are a group of antigens expressed on most nucleated cell surfaces. They transport endogenous peptides to the cell surface for recognition by T-cell receptors. Their functions are involved in immune responses. Many diseases are associated with HLA alleles, especially HLA-B*27 that is strongly associated with ankylosing spondylitis (AS). HLA-B*27 consists of 42 subtypes. Different subtypes of HLA-B*27 were reported in different ethnic groups of AS patients. In this study, a high-resolution polymerase chain reaction–sequence-specific primer technique has been developed to define all the HLA-B*27 subtypes with a total of 29 primer mixtures. Two of the primer mixes were used to detect the HLA-B*27 -specific group, and 27 primer mixes were used to identify 42 subtypes ( B*2701–B*2721 and B*2723–B*27 43). The HLA-B*27 -group-specific primers have been tested in unrelated healthy subjects; 846 Northeastern Thais (NET), 334 Northern Thais (NT), 264 Karens, and 310 Bamars. Sixty-three NET (phenotype frequency, PF = 7.4%), 24 NT (PF = 7.1%), 5 Karens (PF = 1.8%), and 12 Bamars (PF = 3.9%) were positive for HLA-B*27 . Only B*2704 was found in Karens, whereas B*2704 , B*2705/37/39 , B*2706 , and B*2707 were found in NET and NT. In Bamars, B*2704 , B*2705/37/39 , B*2706 , and B*2725 were found. The distribution of HLA-B*27 subtypes was compared with other studies in Asian and Caucasian populations. Significant differences of the distribution of HLA-B*27 subtypes were found in most of the populations. This study established a simple technology for HLA-B*27 subtyping and provided basic information for anthropology and further studies in disease associations.     

Diversity of human leukocyte antigen-B27 alleles in Han population of Hunan province, southern China

This study was to investigate the frequency of HLA-B27 and its subtypes in the Han population of Hunan province, southern China. One hundred and sixty-nine healthy unrelated donors were tested for HLA-B27 by polymerase chain reaction-sequence-specific primer (PCR-SSP). One hundred and twenty-eight B27-positive spondyloarthropathy patients and 18 B27-positive healthy controls were subtyped using the high-resolution PCR-SSP. The phenotype frequency of human leukocyte antigen (HLA)-B27 was found to be 2.36% in healthy population. Five B27 alleles were identified: B*2704, B*2705, B*2706, B*2707, and B*2724. No significant difference was found in the distribution of HLA-B27 subtypes between the patients and controls studied. Notably, B*2724 was observed in a juvenile patient with ankylosing spondylitis. This subtype has not been previously reported in Chinese ankylosing spondylitis (AS) patients and other ethnic groups.     

HLA-B27 subtypes in Asian patients with ankylosing spondylitis Evidence for new associations

Abstract: The aim of this study was to investigate the contribution of the different B27 subtypes to ankylosing spondylitis (AS) susceptibility. The polymerase chain reaction (PCR) in combination with the sequence-specific oligonucloetide probes (SSOs) was used to analyse the polymorphism in exon 2 and 3 of HLA-B27 in two Asian groups with different genetic HLA structures: Indian (I) and Thai (T) populations. The same number of AS patients (45) and healthy B27 positive donors (n=17) from both populations were analysed in order to ascertain the B27 subtypes. Three different findings can be concluded from this study: 1) B*2707 has been found to be associated with AS in both populations. This association has not been previously reported in either ethnic group. 2) B*2704 is strongly associated with AS in the Thai patients (91% in AS vs. 47% in C; RR=11.5; EF=0.83). In contrast, B*2704 was found with similar frequency in Asian Indians AS patients and controls (41% in AS vs. 41% in G). 3) B*2706 was found overrepresented in control populations and absent in AS patients (0% in AS vs. 47% in C; p_c<10^-6) showing the maximum value of protective fraction (PF=1). The B*2706 negative association with AS has not been previously described in other ethnic groups and could indicate a protective effect of this subtype on AS susceptibility. The B*2706 allele has two changes relative to B*2704 at residue 114 (His to Asp) and 116 (Asp to Tyr) in the pockets D/E. The importance that these differences can play in the pathogenesis of AS are discussed.     

A new HLA-B*27 allele (B*2719) identified in a Lebanese patient affected with ankylosing spondylitis

Eighteen different HLA-B*27 alleles (B*2701-B2718) have so far been recognized by the WHO Nomenclature Committee for Factors of the HLA System. Frequency and disease association of these alleles with spondyloarthropathies differ among ethnic groups. We describe here a novel HLA-B*27 subtype identified in a Lebanese patient suffering from ankylosing spondylitis (AS). This new variant differs from the common HLA-B*2705 DNA sequence at five different nucleotide positions. These nucleotide changes lead to three amino acid differences in the alpha2 domain; Thr to Ile at position 94, Leu to Ile at position 95 and Asn to Arg at position 97. Since this novel allele is encountered in an AS patient, the associated sequence changes are not expected to affect significantly neither the presentation of a putative arthritogenic peptide nor the conformation-dependent recognition by effector cells.     

HLA-B27 alone rather than B27-related class I haplotypes contributes to ankylosing spondylitis susceptibility

Characterization of non-B27 susceptibility genes will be required to know the pathogenesis of AS. The aim of this study was to examine whether ankylosing spondylitis (AS) susceptibility is controlled by B27 alone rather than B27 haplotypes and, whether other closely related class I loci, such as MICA and TNFA genes might play a role in AS. Three hundred eleven B27-positive samples from Caucasoid, Asian, and African populations were selected and genotypes were carried out by PCR/SSOP (HLA-B27 and HLA-C), PCR/SSP (MICA-TM polymorphism in the transmembrane region), PCR/SSCP (MICA alleles), and PCR-RFLP (TNF-alpha). Of these, 161 were AS patients, chosen in order to investigate the contribution of TNFA and MICA loci to AS in HLA-B27 positive individuals. Some findings can be concluded from the study: (a) No significant differences of TNF-alpha promoter alleles at position -308 and -238 (A/G) were found between AS patients and B27 matched alleles from healthy controls; (b) strong linkage disequilibrium was found between the B27 and the MICA alleles. The MICA-A4 was found to be in association with B*2705,02,03 and 08; MICA-A5 with B*2704 and B*2707 and MICA-A.5.1 with B*2706; (c) no significant differences of MICA alleles were found between AS and controls carrying the B27-associated alleles, and therefore no evidence is provided for an additional role of MICA gene in AS susceptibility; (d) there are a striking correlations between the structure of B27 extended haplotypes (from MICA region to HLA-C) and the ethnic distribution of these subtypes. The results of differential linkage disequilibrium with HLA-B27 subtypes suggest that B27 itself remains the primary gene for AS susceptibility, and TNFA and MICA are not involved in the pathogenesis of the disease.     

HLA-B27 subtypes determination in patients with ankylosing spondylitis from Zulia,Venezuela

To perform an investigation regarding the distribution of the human leukocyte antigen (HLA)-B27 subtypes in the Zulian population with ankylosing spondylitis (AS), 48 unrelated Mestizos, HLA-B27 positive by serology, were studied using the polymerase chain reaction-specific sequence oligonucleotides probe (PCR-SSOP) and specific sequence primers (SSP) to analyze the polymorphism in exons 2 and 3 of the HLA-B27 gene. Only two of eight HLA-B27 subtypes studied (B*2701-B*2708) were found. The distribution of these alleles in the population of patients was: B*2705, 68.8%, and B*2702, 31.2%. B*2705 subtype showed significant association with patients being male. In the healthy controls, the most common subtype was B*2708. These results were compared with frequencies reported in other Mestizo and Spanish populations and showed significant differences, such as a high frequency of B*2702. Such results show that HLA*B2705 and HLA*B2702 are the subtypes most frequently associated with AS in our Mestizo population and suggest a possible protector role for HLA*B2708, which was found only in the healthy population.     

B*27 in molecular diagnostics: impact of new alleles and polymorphism outside exons 2 and 3

HLA-B*27 is known to be associated with ankylosing spondylitis and several methods have been applied to determine its presence or absence. In this report two molecular methods were used for detection of B*27. The polymerase chain reaction sequence-specific primer (PCR-SSP) method was performed to detect the presence or absence of B*27, whereas the sequence-based typing method (SBT) was used to identify the B*27 subtype. The PCR-SSP method used to detect B*27 was updated to enable the detection of all B*27 alleles. The typing results obtained by this method were compared with the serological typings of 262 individuals. Fifty of them were found to be B*27 positive by PCR-SSP and 46 also showed positive serological reactions with B27-specific sera. The four discrepancies were the result of the presence of B*2712 in three individuals and B*2715 in one individual; both alleles showed no serological reactions with B27-specific antisera. With SBT the sequences of exons 1 through 4 were determined to unequivocally assign the B*27 alleles. Eleven different subtypes were detected in 78 individuals, including three new B*27 alleles: B*27054, B*2715 and B*2717. The allele B*27054 showed an allelic drop out when exon 3 was amplified. Three differences with B*27052 were demonstrated; one in exon 1, one in intron 1 and one in intron 2, the latter being responsible for the allelic drop out. The B*2715 allele was serologically not detectable with several B27-specific sera, but showed Bw4-positive reactions. The sequence of B*2715 showed two mismatches with B*2704. The sequence of B*2717 showed one mismatch with B*27052 at position 248 (A-->T), which was considered to be a conserved position in all B alleles.     

抗原处理相关转运体等位基因与HLA-B27检测呈阳性的强直性脊柱炎的相关研究

目的探讨汉族人群抗原处理相关转运体(transporter associated with antigen processing, TAP)等位基因与HLA-B27及强直性脊柱炎(ankylosing spondylitis, AS)的相关性。方法用聚合酶链反应-序列特异性寡核苷酸探针杂交技术,对48例AS患者(B27+)及123名正常对照人群(B27+27名、B27-96名)进行TAP1、TAP2等位基因分型及变异位点氨基酸表型频率分析。结果汉族人群TAP1表现型主要为Ile/Ile和Asp/Asp,而TAP2则以Val/Val、Ala/Thr和Stop/Stop占优势。TAP1和TAP2至少各有4种等位基因型,分别为TAP1*0101、TAP1*0201、TAP1*0301、TAP1*0401和TAP2*0101、TAP2*0102、TAP2*0201、TAP2*0202。研究对象中有9.9%(17/171)TAP1探针无法定型,15.8%(27/171)TAP2无法定型,呈杂交空白。病例组与对照组间TAP等位基因型分布无差异(P＞0.05)。AS(B27     

Diversity of HLA-B61 alleles and haplotypes in East Asians and Spanish Gypsies

Abstract: The distribution of HLA-B61 alleles and their association with HLA-C and DRB1 alleles were investigated in six East Asian populations (South Korean, Chinese Korean, Man (Manchu), Northern Han, Mongolian and Buryat) and Spanish Gypsies and compared to our previous report on the Japanese population. The alleles were identified using a group-specific polymerase chain reaction (PCR) and genomic DNA followed by hybridization with sequence-specific oligonucleotide probes (SSOP)- Both HLA-B*4002 and B*4006 were commonly detected in the South Korean, Chinese Korean, Man, Northern Han and Japanese populations, while HLA-B*4002 was predominant in the Mongolian and Buryat populations. Strong associations of B*4002 with Cw*0304 and of B*4006 with Cw*0801 were commonly observed in these East Asian populations. In contrast, in Spanish Gypsies, only HLA-B*4006 was found and the allele exhibited a strong association with Cw*1502. HLA-B*4003 was also identified in the South Korean, Chinese Korean, Northern Han, Mongolian and Japanese populations at relatively low frequencies, and exhibited an association with Cw*0304. Moreover, the association of these B61 alleles with the DRB1 alleles revealed considerable diversity among the different populations. HLA-B*4004 and B*4009 were not observed in these populations. Consequently, the frequencies of the B61 alleles varied among the different East Asian populations, but the individual B61 alleles were carried by specific haplotypes often regardless of the ethnic differences.     

B*2707 differs in peptide specificity from B*2705 and B*2704 as much as from HLA-B27 subtypes not associated to spondyloarthritis

HLA-B*2707 is associated with ankylosing spondylitis in most populations. Like the non-associated allotypes B*2706 and B*2709, it lacks Asp116 and shows preference for peptides with nonpolar C-terminal residues. The relationships between the peptide specificity of B*2707 and those of the disease-associated B*2705 and the non-associated subtypes were analyzed by determining the overlap between the corresponding peptide repertoires, the sequence of shared and differential ligands, and by comparing allospecific T cell epitopes with peptide sharing. The B*2707-bound repertoire was as different from that of B*2705 as from those of B*2706, B*2709, or the two latter subtypes from each other. Differences between B*2707 and B*2705 were based on their C-terminal residue specificity and a subtle modulation at other positions. Differential usage of secondary anchor residues explained the disparity between the B*2707-, B*2706-, and B*2709-bound repertoires. Similar differences in residue usage were found between B*2707 and both B*2704 and B*2706, as expected from the high peptide overlap between the two latter subtypes. T cell cross-reaction paralleled peptide sharing, suggesting that many shared ligands conserve their alloantigenic features on distinct subtypes. Our results indicate that association of HLA-B27 subtypes with ankylosing spondylitis does not correlate with higher peptide sharing among disease-associated subtypes or with obvious peptide motifs.     

Lack of carboxyl–terminal tyrosine distinguishes the B2706–bound peptide repertoire from those of B2704 and other HLA–B27 subtypes associated with ankylosing spondylitis

B*2704 and B*2706 are two closely related HLA-B27 subtypes, which differ from the common B*2705 by the Asp>Ser77, Val>Glu152, and Ala>Gly211 amino acid changes. In addition, B*2706 differs from B*2704 by the His>Asp114 and Asp>Tyrl 16 changes. In spite of their similarity B*2704, but not B*2706, was associated to ankylosing spondylitis in a same population. We have carried out pool sequence analyses of the peptides naturally bound to each of these subtypes, and of several individual peptide ligands. B*2704 and B*2706 shared with B*2705, among other features, their selectivity for Arg² and their allowance for some aliphatic and aromatic C-terminal residues in their bound peptides. The main features that distinguished both subtypes from B*2705 were: 1) their failure to present peptides with C-terminal basic residues, and 2) their allowance for both polar and nonpolar residues at peptide position 3. A major difference between B*2704 and B*2706 was that C-terminal Tyr was prominent among the peptides bound to B*2704, but was not detected among those from B*2706. The use of Tyr as a C-terminal anchor motif is the only functional feature shared by the disease-associated B*2705, B*2702, and B*2704 subtypes that is absent in B*2706. This suggests that the ability of HLA-B27 to present peptides with C-terminal Tyr might be critical for its association to spondyloarthropathy,     

HLA-B27 subtypes in chinese patients with ankylosing spondylitis

The HLA-B27 allele has been extensively studied due to its strong association with ankylosing spondylitis (AS). In order to identify B27 alleles in Chinese patients with AS from the Shanghai area, we joined the AHS#5 of the 12 IHW and total of 68 B27 positive patients and 7 B27 positive normal persons have been investigated using polymerase chain reaction and Dig-ddTUP labeled oligonucleotides. Three primer pairs, E403 and E90as, E91As and E136as, E91Bs and E18as, were used to amplify codons 40-90 of HLA-B related alleles, codon 91 to 136 of HLA-B^*2701-B^*2706 and B^*2708 and codons 91-180 of B^*2707. A total of 11 probes were used to distinguish 8 B^*27 alleles from B^*2701 to B^*2708. 68 AS patients contain 69 B27 alleles because one patient is heterozygous B^*2704/B^*2705. A total of 4 alleles of B^*27 were detected in the AS-patient group. B^*2704 is the most common B^*27 allele in both AS patients and controls with similar frequency, 76.8% and 71.4%, respectively. We found a high proportion of B^*2705 in both AS patient (20.3%) and control (28.6%) groups. Although the control group is quite small we are still able to deduce that B^*2704 and probably also B^*2705 seem to be associated with AS in Shanghai area patients We also found one AS allele typed as B^*2707. Interestingly, for the first time we detected B^*2706 in an AS patient, which would argue against a protective effect of B^*2706 on AS susceptibility in Shanghai Chinese. The conclusion from this study is that the distribution of B^*27 alleles is not significantly different between AS patients and controls. Expanded numbers of AS patients and especially of healthy controls in different ethnic groups will be necessary to assess the contribution of different B27 subtypes to AS susceptibility.     

HLA-A and HLA-B in Kenya,Africa: allele frequencies and identification of HLA-B*1567 and HLA-B*4426

HLA-A and HLA-B alleles of a population from Kenya, Africa were examined by sequencing exon 2 and exon 3 DNA and typing using a Taxonomy-based Sequence-analysis (TBSA) method. Extensive diversities were observed at both HLA-A and HLA-B loci in this population. Forty-one HLA-A alleles were identified from 159 unrelated individuals. The most frequently observed alleles were A*6802 (11.64%), A*02011/09 (9.75%), A*7401/02 (9.43%), A*3001 (7.86%), A*3002 (7.23%) and A*3601 (6.6%). Forty-nine HLA-B alleles were identified in 161 unrelated individuals, including two novel alleles, B*1567 and B*4426. The most frequently observed HLA-B alleles were B*5301 (9.01%), B*5801 (8.38%), B*4201 (7.76%), B*1503 (7.14%), B*1801 (6.21%), and B*5802 (5.90%). The most frequently observed HLA-A-B haplotypes were A*3601-B*5301 (3.55%) and A*3001-B*4201 (3.19%), followed by A*7401/02-B*5801 (2.84%), A*7401/02-B*5802 (2.84%) and A*02011/09-B*1503 (2.13%). Linkage disequilibrium and chi2 analysis showed the association of these HLA-A-B haplotypes at the antigen level to be significant. The frequencies of HLA-A and HLA-B alleles from the Kenyan population were compared with that of a population from Cameroon. The difference in allele and haplotype frequency distributions partly reflected the different ethnic composition of these two African populations.     

Discrimination of HLA-B27 alleles by group-specific amplification followed by solid-phase sequencing

HLA-B27 is known to be highly associated with ankylosing spondylitis. Until now, nine B27 subtypes have been sequenced and may contribute in different fashions to ankylosing spondylitis. Additionally, the divergent subtypes may be of clinical importance in bone marrow transplantation with alternative donors. The purpose of this study was to determine the different subtypes of HLA-B27 by a direct sequencing approach. The typing strategy is based on a group-specific amplification of the second and third exon followed by automated fluorescence sequencing of the polymorphic regions. The extensive sharing of sequence motifs between the different B alleles made it impossible to specifically amplify the B27 group under the precondition of including all sequence variations necessary for a postamplification specificity step. Therefore, for setting up a direct sequencing approach of B27, co-amplified B alleles had to be taken into account. In order to get unambiguous sequencing chromatograms without any heterozygous positions, nested sequencing primers were used which selectively matched sequence motifs only present in the second and third exon of the amplified B27 alleles. This strategy allowed in all cases investigated a clear separation of the haplotypes, revealing unequivocal sequencing results. Using this method, we have investigated 93 B27-positive individuals. Sequencing identified the alleles B*2702, 2703, 2704, 2705, and 2707. B*2701, 2706, 2708, and 2709 were not represented in the population studied.     

HLA-B27 and ankylosing spondylitis: tales from China

Abstract
The two most frequent HLA-B27 subtypes worldwide are B*2704 and B*2705. In the Han population of China B*2704 and, to a lower extent, B*2705 are found with significant frequency, and both are associated to ankylosing spondylitis (AS). Two articles in this issue report that the association to AS in this ethnic group is stronger for B*2704 than for B*2705. Thus, at least among the Han, B*2704 would be the strongest known susceptibility factor for AS.     

Complete coding sequence of HLA-B*2712: a serologic B27-negative antigen associated to Bw6

Abstract: We report the complete coding sequence of a new HLA-B27 subtype, B*2712, which was found in a Caucasian Spanish family within the chromosome A2-Cw2-B*2712-DR15-DQ6. B*2712 was first detected as a segregating B blank Bw6-associated antigen. Extensive serologic analysis demonstrated that this new B27 subtype was not recognised by any of the B27-monospecific antibodies, giving positive reactions only with some monoclonal reagents against B40 or B27,40. Sequencing analysis showed a high similarity with B*2708, only differing in three clustered amino acid residues at positions 69 to 71 located in the α helix of the α1 domain. Residues 69 and 71 point towards the T-cell receptor, while amino acid 70 points to the antigen binding site. Loss of the conserved structure of pocket B as well as the differentiated pocket F configuration suggests that B*2712 does not confer ankylosing spondylitis susceptibility, Misleading serologic definition supports the usefulness of DNA-typing methods to complement HLA class I typing.