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.     

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.     

HLA-B27 in the Greek Cypriot population: distribution of subtypes in patients with ankylosing spondylitis and other HLA-B27-related diseases. The possible protective role of B*2707

The major purpose of the present study was to investigate the frequency of human leukocyte antigen (HLA)-B27 alleles in healthy controls and in patients with ankylosing spondylitis (AS) and other HLA-B27–related diseases in the Greek Cypriot population. We selected 102 HLA-B27–positive individuals (60 controls and 42 patients). Typing of the HLA-B27 alleles was performed by polymerase chain reaction amplification with sequence-specific primers. Only two alleles were detected in the patient group: B*2702 (n = 31, 73.8%) and B*2705 (n = 11, 26.2%). The HLA-B*2707 allele was detected (n = 10, 16.7%) only in the healthy controls in addition to the B*2702 (n = 31, 51.7%) and B*2705 (n = 19, 31.7%) alleles. Our results show a restricted number of HLA-B27 subtypes associated with AS and other B27-related diseases and an elevated frequency of the B*2702 allele in the AS patients. The allele B*2707 seems to have a protective role in the population studied because it was found only in the healthy controls.     

Predominant association of HLA-B*2704 with ankylosing spondylitis in Chinese Han patients

Abstract
The HLA-B27 subtypes have a varied racial and ethnic prevalence throughout the world. However, the association of B27-subtypes with ankylosing spondylitis (AS) in the mainland China is unknown. To determine the association of B27-subtypes with AS in the Mainland Chinese Han population, a total of unrelated 153 patients with AS were enrolled in a large case-control association study, and 1545 unrelated, healthy, ethnically matched blood donors were included as controls. The genotyping of B27 and its subtypes was performed using the polymerase chain reaction with sequence specific primers (PCR-SSP). A total of 130 (84.97%) AS patients and 61 (3.95%) healthy controls were B27 positive. Three B27-subtypes, B*2704, B*2705 and B*2710, were further identified, of which both B*2704 and B*2705 were strongly AS associated. B*2710 was only detected in one AS patient and two other healthy controls. Considering only B27-positive cases and controls, a statistically different frequency of B27-subtypes was observed, with an over-representation of B*2704 ( P = 0.018). B*2704 was clearly more strongly associated than B*2705 with AS [odds ratio (OR ) = 2.4, P = 0.011]. Furthermore, a combined analysis including three previous studies of B27-subtype distributions in Chinese AS cases confirmed the stronger association of B*2704 with AS than B*2705 (OR = 2.5, P = 0.00094).     

HLA-B27 polymorphism in patients with juvenile and adult-onset ankylosing spondylitis in Southern China

Abstract
Distribution of B27 subtypes in juvenile and adult-onset ankylosing spondylitis (JAS and AAS) in Southern China was studied. A total of 505 patients belonged to Han population were included (145 JAS and 360 AAS patients), and 1368 healthy individuals were included as controls. Human leukocyte antigen (HLA)-B27 typing was performed by Luminex liquid array combining polymerase chain reaction-sequence specific oligonucleotide probe (PCR-SSOP) and/or serological method. HLA-B27 subtyping was performed by polymerase chain reaction-sequence specific primer (PCR-SSP). The sequence-based typing was performed for the B*2715 samples to verify the PCR-SSP results. HLA-B27 was presented in 453 of 505 patients (89.7%), compared with 74 of 1368 controls (5.41%). B*2704 subtype in AS group was significantly higher than controls and B*2705 subtype significantly lower. B*2715 and B*2702 were found in 1.32% and 0.66% of the B27-positive patients but none in controls, and there was no significant difference between either of them and controls. B27-positive patients were 134 (92.4%) in JAS group and 319 (88.6%) in AAS group. There was no significant difference for B27 subtypes distribution between JAS (B*2704, 05, 15) and AAS (B*2704, 05, 15, 02) groups. The frequency of B*2715 in two groups was 3 (2.24%) and 3 (0.94%), respectively. The onset age of three JAS patients carrying B*2715 was 5, 9 and 13 years old, respectively. Our results suggested that B*2704 was the predominant subtype in AS patients in Southern China. B*2715 was observed in AS group only and slightly more in JAS than in AAS, and the patients carrying this allele tended to have early onset, B*2715 may be disease-association subtype.     

少儿强直性脊柱炎与幼儿类风湿性关节炎HLA-B27亚型的鉴别诊断

目的：探讨HLA－B27等位基因亚型与少年强直性脊柱炎和幼年类风湿性关节炎的关联。方法：用PCR－SSP方法对74人HLA－B27等位基因亚型进行研究，其中少年强直性脊柱炎32例，幼年类风湿性关节炎28例，5个家系中患者的父亲或母亲5例，正常对照组9例，并进行关联分析。结果：本组人群的HLA－B27等位基因由HLA－B*2704、*2705、*2702、*2707 4种亚型组成，其中少年强直性脊柱炎患者HLA－B27等位基因亚型频率为B*2704 56.25%、B*2705 40.63%、B*2702 3.13%；幼年类风湿性关节炎HLA－B27等位基因亚型频率为B*2705 60．7％、B*2704 28．57％、B*2702 3．57％及B*2707为7．14％；少年强直性脊柱炎与幼年类风湿性关节炎结果比较，HLA－B*2704基因频率在少年强直性脊柱炎组高于幼年类风湿性关节炎组（RR＝3．21，P＜0．05）。结论：少年强直性脊柱炎与HLA－B*2704等位基因亚型关联。对HLA－B27等位基因亚型的检测可成为少年强直性脊柱炎和幼年类风湿性关节炎鉴别诊断中一个有价值的实验指标。     

Possible protective role of HLA-B*2706 for ankylosing spondylitis

HLA-B27 is strongly associated with ankylosing spondylitis (AS) but the role of the HLA molecule itself is still unclear. In this study on Singapore Chinese, we have subtyped 50 B27 positive AS patients and 45 B27 positive normals and found that the B*2706 allele has a significant negative association with disease (p=0.047). Together with recent data indicating the existence of AS "protective" B27 alleles, our data shows that the HLA molecule itself plays a crucial role in disease development.     

Susceptibility to ankylosing spondylitis correlates with the C-terminal residue of peptides presented by various HLA-B27 subtypes

Susceptibility to spondyloarthropaties is strongly associated with some HLA-B27 alleles. Evidence suggests a direct pathogenic role for the B27 molecules which possibly present an arthritogenic peptide to the T cells. If this hypothesis is true, B27 subtypes that differ structurally but are disease-associated ought to be capable of presenting such peptide(s), while non-disease-associated ones would not. We have recently described a B27 subtype, B*2709, and shown its absence in ankylosing spondylitis (AS) patients. Here, we show the elution and sequence of peptides from HLA-B*2709 molecules. Similar to other B27 subtypes, these peptides are mainly nonamers with an Arg at position P2. Comparison of the C-terminal anchors of peptides eluted from B*2702 and B*2705 with those eluted from B*2709 reveals that, while B*2702 and B*2705 have a broader specificity, B*2709 molecules appear to only accept C-terminal hydrophobic residues. A common feature shared by the two caucasoid AS-associated subtypes (B*2702 and B*2705) but different from B*2709, is the presence of a Tyr as peptide C-terminal anchor. The substitution of Val for Tyr at the C terminus in one of the eluted peptides greatly reduces the binding to B*2709 molecules. This finding suggests Tyr as a discriminative amino acid allowed at the C terminus of peptides bound to the AS-associated B27 subtypes, but not to those which are not associated with AS.     

HLA-B27亚型及其与强直性脊柱炎关系的研究进展

强直性脊柱炎(AS)是与mA关联最强的疾病.HLA-B27由22个以上同种异型基因型(亚型B*2701～B*22)组成,不同亚型核苷酸序列之间只存在个别位点的差异,其亚型具有分布不同的种族和人种流行情况,以B*2705分布最广.近年来建立了大量的AS动物模型,人类B27转基因鼠实验证实B27分子是AS的原发关联成分,这种带有B27等位基因的实验动物可发生类似人类AS疾病.目前倾向于以关节源性肽假说来解释HLA-B27在AS发病中的作用.     

Relevance of residue 116 of HLA-B27 in determining susceptibility to ankylosing spondylitis

Ankylosing spondylitis (AS) is an autoimmune disorder strongly associated with HLA-B27. A direct role of B27 molecules in the disease pathogenesis has been postulated, possibly by presenting to T cells an as-yet unidentified arthritogenic peptide that triggers the autoimmune response. There are nine HLA-B27 alleles differing from each other at one or more amino acid positions. It is important, for the identification of the arthritogenic peptide, to define which alleles, and therefore which polymorphic positions, predispose to the disease. Here, we report that HLA-B^★2709 is not associated with AS, as it was not found in patients. HLA-B^★2709 differs from the most frequent and disease-associated HLA-B^★2705 allele for a single substitution (His vs. Asp) at position 116. Amino acid 116 is located at the bottom of the groove where the antigenic peptide sits, and it has been proven to influence the peptide-binding specificity of HLA class I molecules. The most likely interpretation of these data is that the differences in charge and size that accompany the His-to-Asp substitution exclude the acceptance of the arthritogenic peptide.     

67例强直性脊柱炎患者临床表现及免疫功能与HLA-B_(27)的关系

我们研究了67例强直性脊柱炎(AS)患者的临床特点及免疫学变化与HLA-B_(27)的关系。结果表明,HLA-B_(27)阳性AS患者的发病年龄较HLA-B_(27)阴性者平均早5年(P<0.05);HLA-B_(27)阳性的AS患者中17.2%并发虹膜睫状体炎或急性结膜炎;HLA-B_(27)阳性患者血清IgG、IgA、IgM及γ-球蛋白水平明显增高。提示AS患者的发病、眼部损害及体液免疫反应增强均与HLA-B_2密切相关。     

强直性脊柱炎患者HLA-B27位点等位基因相关抗原的表达

目的：探讨强直性脊柱炎（AS）患者HLA-B27位点等位基因相关抗原的表达。方法：采用补体依赖性微量淋巴细胞毒法检测418例脊柱关节炎（SpAS）患者和30例正常对照HLA-B27相关抗原。结果：418例SpAS患者73例被确诊为AS,Bw6、B27（47）、B27和B7/B27/B73/B81抗原阳性率分别为31%、28%、25%和22%。在AS中,B27阳性组与B27阴性组间有不同分布,两组B27（47）、B27、B7三种等位基因有统计学意义（P〈0.01）。在66例B27阳性组中,除B13与Bw6成负相关外,Bw4与B27（47）、B7与B27之间等均成明显的正相关。结论：在AS患者中,B13与Bw6、B60/61/47/48/81（13）,B7与B27,Bw4与B27（47）,Cw1与B42B45表达联锁失衡,B27并不是AS易感的唯一因素,其他基因位点可能起增强（如Cw1及Cw2）或抑制（如B13）AS发生的作用。     

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 subtyping by PCR-RFLP in Spanish patients with ankylosing spondyiitis

Abstract: We describe the use of restriction analysis on PCR-amplified DNA for detecting all B*27 subtypes except B*2710 and B*2711 (i.e. from B*2701 to B*2709). After detecting B*27 by Sty I, double digestions consisting of Sty I plus another informative enzyme led to subtype assignment. We used mismatched primers to create restriction sites when necessary. The method avoids group-specific amplifications and other laborious optimization procedures. It was successfully tested on a panel of well characterized cell lines covering different B*27 subtypes. Then, we studied a group of 57 ankylosing spondyiitis patients and 746 controls from the south of Spain. B*27 showed a very strong association with the disease (OR=211.27, P=\0˜7). B*2702 and B*2705 distribution in controls (20% and 77.1%, respectively) differed from previously reported data in the Spanish population. We unexpectedly found the B*2707 allele in our population (one control).     

强直性脊柱炎患者血液HLA-B27的临床参考值的预测

目的 以实时荧光定量PCR（FQ-PCR）检测可疑患者HLA-B27的定量水平,研究强直性脊柱炎（AS）与HLA-B27的相关性,明确HLA-B27的检测值范围并用于AS的确诊诊断.方法 针对本院2005年~2010年收集的168例骶髂关节及下腰部疼痛等症状疑似AS病例进行回顾性分析,获得所有病例外周静脉血标本,然后通过实时荧光定量PCR进行HLA-B27的定量检测,疑似病例分为AS患者、HLA-B27阳性的非AS患者、HLA-B27阴性的非AS患者三组,根据临床体征、影像学进行确诊病情,同时选取健康体检者52例外周静脉血标本的HLA B-27测定结果进行对照,统计分析各变量与AS存在之间的关系.结果 AS患者、HLA-B27阳性的非AS患者、HLA-B27阴性的非AS患者、健康体检者的HLA-B27循环阈值（ct）平均值分别为：26.3 copies/ml、17.5 copies/ml、6.2 copies/ml、4.9 copies/ml,单变量分析表明,HLA-B27与化脓性关节炎明显相关（P＜0.05）.结论 AS患者HLA-B27定量分析循环阈值（ct）的95%参考值范围为（17.3~35.3）copies/ml,HLA-B27定量分析可作为疾病发病的参照影响因素及预测因素.     

TAP polymorphism is not associated with ankylosing spondylitis and complications with acute anterior uveitis in HLA-B27-positive Japanese

Abstract: HLA-B27 is associated with the etiology of ankylosing spondylitis (AS) and acute anterior uveitis (AAU). Transporter associated with antigen processing (TAP) 1 and TAP2 polymorphism influences the range of peptide presented by HLA class I molecules. In this report, contribution of TAP polymorphism to the susceptibility to AS and AAU was studied in HLA-B27-positive Japanese individuals. Patients were classified into three groups: 16 AS patients, 14 AAU patients and 22 patients with both AS and AAU. Twelve HLA-B27-positive healthy individuals were included as a control. TAP polymorphism was detected by PCR-RFLP methods. Significant differences in frequencies of TAP1 alleles were not found between patient groups. None of the TAP2 frequencies showed increased or decreased frequencies compared with HLA-B27-positive healthy controls. In comparison with a random Japanese control, TAP2D allele frequency was significantly increased in the AAU group, but failed to reach a significant level in a group consisting of the AAU-only patients and the patients with both AS and AAU. All of the patient groups were noted to have a significantly increased prevalence of the TAP2H allele as compared to random controls; however, the higher frequency of this allele was detected in HLA-B27 healthy controls as well. These observations suggest a linkage disequilibrium between TAP2D, TAP2H and HLA-B27 in Japanese.     

Antibodies to the peptide from the plasmid-coded Yersinia outer membrane protein (YOP1) in patients with ankylosing spondylitis

Seventy-five Norwegian patients with ankylosing spondylitis (AS) were studied for class-specific antibody response against synthetic peptide, P81, representing the sequence of plasmid-coded outer membrane protein of Yersinia (YOP1) containing four amino acid homology (TDRE) with HLA-B27 sequence. Ten (16.7%), five (8.3%) and seven (11.2%) of 60 male AS patients showed elevated anti-YOP1 P81 antibody of IgA, IgG, and IgM class, respectively, whereas for each isotype only one (4%) of 25 healthy male controls was positive. Differences were not observed between female patients and controls. In all isotypes, antibody-positive patients were more frequently found in patients with active disease. The anti-YOP1 P81 antibody levels of the patients were generally not correlated with the antibody levels against the peptide representing the hypervariable region of HLA-B27 (B27 peptide). However, in one patient the antibody was shown to react with both peptides by cross-inhibition analysis. Overall, it appears that any causal relationship between YOP1 and pathogenesis of AS is not strong. Immunogenicity and cross-reactivity of the YOP1 region encompassing the TDRE sequence particularly at the T cell level require further study.     

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.     

Activating killer immunoglobulin-like receptors genes are associated with increased susceptibility to ankylosing spondylitis

The aim of this study was to analyse the association of specific killer cell immunoglobulin-like receptors (KIR) genes and haplotypes with susceptibility to ankylosing spondylitis (AS) and its different clinical manifestations in a Spanish population. The presence or absence of all KIR genes was studied for their association with AS. A total of 176 patients with AS and 435 healthy control subjects were selected for this study based on clinical criteria. The commercial KIR-sequence-specific oligonucleotides (SSO) typing kit was used to investigate KIR typing. Frequencies of KIR2DS1 and KIR3DS1 genes were increased significantly in patients compared with healthy controls [52·8 versus 38·2%, P_Bonf < 0·01, odds ratio (OR) = 1·81 (1·28–2·59); 51·7 versus 37·5%, P_Bonf < 0·01, OR = 1·79 (1·25–2·54)]. Moreover, the frequency of activating genotypes in the AS patient group was significantly higher than in the healthy control group (P < 0·05). KIR2DS1 and KIR3DS1, in addition to human leucocyte antigen (HLA)-B27, may play an important role in the pathogenesis of AS. However, we show that the contribution of the KIR genes to AS susceptibility extends beyond the association with individual KIRs, with an imbalance between activating and inhibitory KIR genes seeming to influence the susceptibility to AS.     

HLA-B27 and the pathogenesis of spondyloarthritis

The association of HLA-B27 with ankylosing spondylitis and other spondyloarthropathies ranks among the strongest between any HLA antigen and a human disease. Yet, in spite of intense research and advanced knowledge of the biochemistry and biology of major histocompatibility complex molecules, the mechanism of this association remains unknown. This review attempts a critical assessment of current pathogenetic hypotheses from evidence concerning the epidemiology of HLA-B27 association with disease, its peptide-binding specificity, and other aspects of the molecular biology and immunology of this molecule.