CD226 (DNAM-1) is associated with susceptibility to juvenile idiopathic arthritis.

OBJECTIVES: Juvenile idiopathic arthritis (JIA) is considered a complex genetic autoimmune disease. We investigated the association of genetic variants previously implicated in JIA, autoimmunity and/or immunoregulation, with susceptibility to JIA. METHODS: A genetic association study was performed in 639 JIA patients and 1613 healthy controls of northwest European descent. Ninety-three single nucleotide polymorphisms (SNP) were genotyped in a candidate gene approach. Results of the entire JIA patient group (all subtypes) were compared with results obtained, alternatively, with a clinically homogeneous patient group including only oligoarticular and rheumatoid factor (RF) negative polyarticular JIA patients (n=493). Meta-analyses were performed for all SNPs that have been typed in other Caucasian JIA cohorts before. RESULTS: SNPs in or near PTPN22, VTCN1, the IL2-IL21 region, ANKRD55 and TNFA were confirmed to be associated with JIA (p<0.05), strengthening the evidence for involvement of these genes in JIA. In the majority of these replicated SNPs, effect sizes were larger when analysing a homogeneous patient cohort than when analysing all subtypes. We identified two novel associations with oligoarticular and RF-negative polyarticular JIA: CD226 rs763361 (OR 1.30, 95% CI 1.12 to 1.51, p=0.0006) and CD28 rs1980422 (OR 1.29, 95% CI 1.07 to 1.55, p=0.008). Meta-analyses including reported studies confirmed the association of both SNPs with susceptibility to JIA (OR 1.16, p=0.001 and OR 1.18, p=0.001, for rs763361 and rs1980422, respectively). CONCLUSIONS: The CD226 gene has been identified as novel association with JIA, and a SNP near CD28 as a suggestive association. Both genes are probable candidate risk factors, since they are involved in costimulation of T cells.

Genetic variation in VTCN1 (B7-H4) is associated with course of disease in juvenile idiopathic arthritis.

OBJECTIVE: The course of disease in juvenile idiopathic arthritis (JIA) is unpredictable with episodes of activity and remission. In order to identify predictive factors, 93 SNPs, JIA subtype, age at onset and ANA status were studied in relation to disease course. METHODS: Genetic and clinical parameters were analysed in a cohort of 272 Caucasian patients with persistent oligoarthritis (n=129), extended oligoarthritis (n=57) and rheumatoid factor negative polyarthritis (n=86). Categories of disease course (remitting (n=65), intermediate (n=96) and unremitting (n=111)) were designed based on the cumulative time spent in active disease in the first 2 years. RESULTS: Univariate analysis revealed association of the course of disease with JIA subtype (p=5.7*10(-5)) and three SNPs; VTCN1 rs10 923 223 (p=4.4*10(-5)), VTCN1 rs12 046 117 (p=0.017) and CDK6 rs42 041 (p=0.038). In a subsequent multivariate ordinal logistic regression analysis, VTCN1 rs10 923 223 (OR 0.41, 95%-CI 0.26 to 0.63) and JIA subtype (OR 3.8, 95%-CI 2.0 to 7.2; OR 2.5, 95%-CI 1.4 to 4.2, for extended oligoarthritis and RF-negative polyarthritis vs persistent oligoarthritis, respectively) were the strongest independent factors for course of disease. CONCLUSIONS: This study provides evidence that VTCN1, encoding B7-H4, is associated with course of disease in selected subtypes of JIA. VTCN1 might be useful in predicting the course of disease.

Genetic variants in the region of the C1q genes are associated with rheumatoid arthritis.

Rodent models for arthritis implicate a role for complement in disease development and progression. In humans, complement deposition has been observed in inflamed synovia of rheumatoid arthritis (RA) patients. In this study we analysed whether genetic variants of complement component C1q predispose to RA. We genotyped single nucleotide polymorphisms (SNPs) in and around the C1q genes, C1qA, C1qB and C1qC, in a Dutch set of 845 RA cases and 1046 controls. Replication was sought in a sample set from North America (868 cases/1193 controls), and a meta-analysis was performed in a combined samples set of 8000 cases and 23 262 controls of European descent. We determined C1q serum levels in relation to C1q genotypes. In the discovery phase, five of the 13 SNPs tested in the C1q genes showed a significant association with RA. Additional analysis of the genomic area around the C1q genes revealed that the strongest associating SNPs were confined to the C1q locus. Within the C1q locus we observed no additional signal independent of the strongest associating SNP, rs292001 [odds ratio (OR) = 0·72 (0·58-0·88), P = 0·0006]. The variants of this SNP were associated with different C1q serum levels in healthy controls (P = 0·006). Interestingly, this SNP was also associated significantly in genome-wide association studies (GWAS) from the North American Rheumatoid Arthritis Consortium study, confirming the association with RA [OR = 0·83 (0·69-1·00), P = 0·043]. Combined analysis, including integrated data from six GWAS studies, provides support for the genetic association. Genetic variants in C1q are correlated with C1q levels and may be a risk for the development of RA.

Association of the autoimmunity locus 4q27 with juvenile idiopathic arthritis.

OBJECTIVE: Juvenile idiopathic arthritis (JIA) is characterized by chronic arthritis and an autoimmune etiology. In several autoimmune diseases, including rheumatoid arthritis (RA), an association with the 4q27 locus has been reported. We undertook this study to investigate the possible role of the 4q27 locus in JIA. METHODS: A case-control association study was conducted, with a total of 655 Caucasian JIA patients and 791 healthy controls divided into 2 independent sample sets. The rs6822844 marker in the 4q27 locus was genotyped. RESULTS: In the first and larger sample set, a 5% decrease in T allele frequency was observed in patients compared with controls (allelic odds ratio [OR] 0.72 [95% confidence interval 0.55-0.95], P = 0.019), and in the second set, a 3% decrease was observed (allelic OR 0.81 [95% confidence interval 0.61-1.09], P = 0.169). The combined data set generated an OR of 0.76 (95% confidence interval 0.62-0.93, P = 7.08 x 10(-3)). When the different JIA subtypes were analyzed individually, significant decreases were seen in the subtypes with a polyarticular course of disease (extended oligoarthritis [P = 0.019] and rheumatoid factor-negative polyarthritis [P = 0.038]). CONCLUSION: Our findings suggest that the 4q27 locus, previously reported to be associated with RA, type 1 diabetes mellitus, celiac disease, and psoriatic arthritis, is also associated with susceptibility to JIA.

The TRAF1/C5 region is a risk factor for polyarthritis in juvenile idiopathic arthritis.

OBJECTIVE: Juvenile idiopathic arthritis (JIA) is a chronic disorder in which both genetic and environmental factors are involved. Recently, we identified the TRAF1/C5 region (located on chromosome 9q33-34) as a risk factor for rheumatoid arthritis (RA) (p(combined) = 1.4 x 10(-8)). In the present study the association of the TRAF1/C5 region with the susceptibility to JIA was investigated. METHODS: A case-control association study was performed in 338 Caucasian patients with JIA and 511 healthy individuals. We genotyped the single nucleotide polymorphism rs10818488 as a marker for the TRAF1/C5 region. RESULTS: The A allele was associated with the susceptibility to rheumatoid factor-negative polyarthritis with an 11% increase in allele frequency (OR 1.54, 95% CI 1.09 to 2.18; p = 0.012). This association was stronger when combining subtypes with a polyarticular phenotype (OR 1.46, 95% CI 1.12 to 1.90; p = 0.004). In addition, we observed a trend towards an increase in A allele frequency in patients with extended oligoarthritis versus persistent oligoarthritis (49%, 38% respectively); p = 0.055. CONCLUSIONS: Apart from being a well replicated risk factor for RA, TRAF1/C5 also appears to be a risk factor for the rheumatoid factor-negative polyarthritis subtype of JIA and, more generally, seems to be associated with subtypes of JIA characterised by a polyarticular course.

The PIP5K2A and RGS4 genes are differentially associated with deficit and non-deficit schizophrenia.

Several putative schizophrenia susceptibility genes have recently been reported, but it is not clear whether these genes are associated with schizophrenia in general or with specific disease subtypes. In a previous study, we found an association of the neuregulin 1 (NRG1) gene with non-deficit schizophrenia only. We now report an association study of four schizophrenia candidate genes in patients with and without deficit schizophrenia, which is characterized by severe and enduring negative symptoms. Single-nucleotide polymorphisms (SNPs) were genotyped in the DTNBP1 (dysbindin), G72/G30 and RGS4 genes, and the relatively unknown PIP5K2A gene, which is located in a region of linkage with both schizophrenia and bipolar disorder. The sample consisted of 273 Dutch schizophrenia patients, 146 of whom were diagnosed with deficit schizophrenia and 580 controls. The strongest evidence for association was found for the A-allele of SNP rs10828317 in the PIP5K2A gene, which was associated with both clinical subtypes (P = 0.0004 in the entire group; non-deficit P = 0.016, deficit P = 0.002). Interestingly, this SNP leads to a change in protein composition. In RGS4, the G-allele of the previously reported SNP RGS4-1 (single and as part of haplotypes with SNP RGS4-18) was associated with non-deficit schizophrenia (P = 0.03) but not with deficit schizophrenia (P = 0.79). SNPs in the DTNBP1 and G72/G30 genes were not significantly associated in any group. In conclusion, our data provide further evidence that specific genes may be involved in different schizophrenia subtypes and suggest that the PIP5K2A gene deserves further study as a general susceptibility gene for schizophrenia.

Strong association of MuSK antibody-positive myasthenia gravis and HLA-DR14-DQ5.

The authors studied the HLA profile of 23 white Dutch patients with muscle-specific kinase antibody-positive myasthenia gravis (MuSK Ab+ MG) and found an association with HLA-DR14-DQ5 (odds ratio 8.5; 95% CI 3.9 to 18.7; p = 4.9 x 10(-5)). Fifty-two percent of the patients carried the DR14 allele compared with 5% of the controls (p = 1.0 x 10(-8)). This association between MuSK Ab+ MG and a relatively rare HLA haplotype differs from the previously described association of early-onset acetylcholine receptor Ab+ MG with HLA-B8-DR3.

Five newly identified HLA alleles: A*0310, A*2907, B*4435, Cw*0206, Cw*0506, and confirmation of A*3106, B*3924, Cw*0314, DRB1*0322, and DRB1*1433 alleles.

A number of HLA alleles have been newly identified. This concerns HLA-A*0310, A*2907, B*4435, Cw*0206, Cw*0506, of which Cw*0206 was found in three unrelated individuals, all B*4002 positive. Some other alleles are also presented but confirm earlier detected sequences: A*3106, Cw*0314, DRB1*0322, and DRB1*1433. Moreover, we identified B*3924 in a bone marrow transplant recipient and in five of six unrelated stem cell donors, selected for this patient. In all cases, B*3924 was found on a haplotype combining A*0201, B*3924, Cw*0701, and DRB1*1303. The observation of this extended haplotype is of importance for the selection for stem cell transplantation. Cells expressing B*3924 and B*4435 were typed by serology as B39 and B44, respectively. Cells expressing HLA-A*0310 do not express A3 but type as A-Blank.

Three newly identified HLA-B alleles: B*5124, B*5306, B*5307 and confirmation of B*0809 and B*5606.

Five HLA-B sequences are described which have been detected as irregular patterns during routine molecular typing. Sequencing of HLA-B exon 2 and 3, both heterozygous and after group specific amplification, revealed three new HLA-B alleles: B*5124, B*5306 and B*5307, whereas the sequences of B*0809 and B*5606 were confirmed. Serological typing showed that B*0809 is expressed as a regular B8, B*5124 as a regular B51, B*5306 as a B51/B53-like variant and B*5606 as a B"blank"-Bw6.

HLA class II associations with Type 1 diabetes mellitus: a multivariate approach.

The association of HLA class II phenotype with the development of insulin-dependent (Type 1) diabetes mellitus (IDDM) is well established but the contribution of the various HLA-DR and -DQ alleles and haplotypes to disease predisposition is not fully understood. We have determined haplotype and genotype odds ratios, and further employed multivariate tree analysis to explore the contribution of individual HLA-DRDQ haplotypes to the genetic risk for developing IDDM in the Dutch population. Next to haplotype and genotype odds ratios, multivariate tree analysis techniques provide overall risk calculations for each modeled parameter, and offer insight in the interaction of the model parameters via tree-shaped reports, in which subsequent stratifications on the data can easily be followed. We compared 206 Dutch IDDM patients with 840 serologically typed random healthy unrelated Dutch Caucasoid controls. The multivariate tree analysis showed that the HLA-DR7DQ9 and DR15DQ6 haplotype were strongly associated with disease protection (OR = 0.04, P = 0.0003, and OR = 0.07, P < or = 0.0001, respectively). The highest ORs were found for the DR4DQ8/DR8DQ4 genotype (OR = 21.04, P = 0.001), followed by DR4DQ8/DR17DQ2 (OR = 12.45, P < 0.0001) and DR9DQ9/DR17DQ2 (OR = 10.87, P = 0.02). DR4DQ8 homozygous and DR17DQ2 homozygous individuals have a disease OR of 9.0 and 3.0 (P = 0.01 and 0.03), respectively. In conclusion, the results from haplotype, genotype, and tree analyses provide insight into the disease associations for combinations of HLA-DRDQ haplotypes. We confirm that the DR9DQ9/DR17DQ2 genotype is associated with susceptibility in the Dutch population, which has previously been noticed as a HLA risk genotypes in Asian populations only.

Identification of two new alleles HLA-DRB1*0312, DRB1*0432 and of a DRB3-negative DRB1*1313-positive haplotype.

Two new HLA-DRB1 alleles were identified in the course of routine class II molecular typing in Dutch Caucasoid. HLA-DRB1*0312 is similar to *03011 except for codon 57 (GAT-->AGC). DRB1*0432 is similar to *0413 but with a mutation at position 215, changing codon 72 (CGG-->CAG; Arg-->Gln). This sequence has never before been identified at this position. A DRB3-negative DRB1*1313 haplotype was identified in an individual from Indonesia. Monoclonal antibodies against DR52 were nonreactive with lymphocytes of this individual. The DRB1*1313-DRB3-negative haplotype probably represents a recombination of DRB1*13 and *08 haplotypes where the sequences telomeric of HV1 are derived from the DRB3-negative DRB1*0803 haplotype.

HLA-DRB and -DQB1 polymorphism in the Macedonian population.

HLA-DRB1, DRB3/4/5 and DQB1 polymorphism has been studied in a population of 80 unrelated healthy Macedonians using molecular methods. Twenty-five different DRB1 alleles were identified of which DRB1*1104, *1501, *1601, and *1101 were found most frequently. Among the 15 identified DQB1 alleles, two were predominant: DQB1*0301 and *0502. The most frequent three-locus haplotypes were DRB1*1104-DRB3*02-DQB1*0301 (18%/), DRB1*1101-DRB3*02-DQB1*0301 (9%) and DRB1*1601-DRB5*02-DQB1*0502 (10%). Polymorphism for DRB1*04, *13 and *15 haplotypes was extensive. Eleven different DR2-related haplotypes were found, some of which were unusual for European populations: DRB1*1501-DRB5*0102-DQB1*0502, DRB1*1501-DRB5*02-DQB1*0502, DRB1*1501-DRB5*0102-DQB1*0601.

HLA and elephantiasis revisited.

A recent case-control study in Indonesia suggested that the course of Brugian filariasis and in particular resistance to the development of elephantiasis was associated with certain HLA class II alleles. In order to see whether these data could be confirmed we conducted a similar study in another Indonesian population from South Sulawesi. We could not confirm our earlier results and therefore concluded that HLA-DR and -DQ alleles are at least not strongly associated with progression to elephantiasis in Brugian filariasis. The complete data are presented also for anthropological reference purposes.

Polymorphism of HLA-DRB, -DQA1, and -DQB1 in rheumatoid arthritis in Asian Indians: association with DRB1*0405 and DRB1*1001.

We investigated the DRB, DQA1, and DQB 1 polymorphism and haplotypes in sporadic and familial RA subjects of Asian Indian origin by PCR oligotyping using biotinylated SSOPs. Molecular subtyping of DRB 1*04 in RA patients showed strongest association with highest relative risk with DRB 1*0405, followed by DRBI*0401. A significant decreased frequency of DRBI*1502 was observed in patients compared to controls (chi 2 = 4.5). Among other alleles, DRBI*1001 was found to be significantly increased. A total of 73.3% of patients carried the shared sequence of the third HVR (67-74) of DRB1 domain compared to its presence in only 37.6% of controls. A significant number of patients carried DR4 haplotypes on DQBI*0302 (58%) as against DQBI*0301 which was present only on 10.5% of the haplotypes. When compared to controls, the difference was significant for the latter allele only. Few unique DRDQ haplotypes were observed in Asian Indians. Among DR-DQ haplotypes, DRB1*0401-DQB1*0302 gave the highest risk whereas DRB1*0403-DQB1*O301 was negatively associated. Alleles with negative charge at position 70 confer protection or are negatively associated with RA whereas among the associated alleles, glycine at position 86 resulted in higher risk than those with valine at this position. A heterogenous association of DQB1 alleles with DR4 subtypes, influencing susceptibility to RA, suggests the DQB locus is not primarily associated with RA and susceptibility lies in the sequence 67-74 of the DRB1 loci.

Analysis of HLA-DR2-associated polymorphisms by oligonucleotide hybridization in an Asian Indian population.

Among major histocompatibility complex class II antigens, HLA-DR2 appears to have a much larger degree of polymorphism than usually recognized by routine serology or restriction fragment length polymorphisms. We have utilized oligonucleotide probes to further identify the DR2 specificity and its molecular subtypes on the basis of specific DNA sequences as they occur in a select sample from the Asian Indian population. In addition, oligonucleotide typing of HLA-DQA1 and -DQB1 genes allowed us to determine specific associations of DRB1, DRB5, DQA1, and DQB1 alleles in DR2 individuals. A set of 60 oligonucleotide probes were hybridized to polymerase chain reaction (PCR)-amplified DNA from DR2 homozygous or heterozygous individuals. The most common DR2 subtypes that occurred in this selected population are: DRB1*1501 (60%), DRB1*1502 (33.8%), and DRB1*1602 (6.2%). No example of DRB1*1601 was detected. By combining these results with the allelic variations at DQA1 and DQB1, we were able to detect at least seven different haplotypes, the most common being DRB1*1502-DRB5*0102-DQA1*0103-DQB1*0601 and DRB1*1501-DRB5*0101-DQA1*0102-DQB1*0502. At least five unexpected combinations, not reported among Western Caucasians, were noticed in this sample. Thus oligonucleotide typing is a valuable tool for defining further polymorphisms in the HLA-D region as exemplified by its applications to typing DR2-positive patients with tuberculoid leprosy and pulmonary tuberculosis.

Internalization of MHC class I molecules is a prerequisite for endocytosis of endorphin by lymphocytes.

The nature of the interaction between gamma-type endorphins and the HLA class I molecules was studied by immunoelectronmicroscopy. The HLA molecules were not involved in the actual binding of endorphin to the cell. In contrast, for the endocytosis of gamma-endorphin, co-internalization of the HLA class I molecules is essential. The internalization process starts with clustering of gamma-endorphin and HLA class I molecules in coated pits. Cells that do not carry HLA class I molecules (Daudi) or do not internalize HLA class I molecules (EBV-transformed B cells) bind but do not internalize gamma-endorphin. On the basis of these observations, we suggest that the MHC class I molecules may function as transport molecules. Whether it is a general phenomenon that non-immunological ligands use the HLA class I molecules to get into the cell and immunological ligands (viral proteins) to reach the cell surface, remains to be established.

The Eurotransplant HLA-DRB oligonucleotide typing set.

Several sequence-specific oligonucleotides (SSO) were designed based upon available sequences of HLA-DRB1 genes which are associated with HLA-DR serotypes. Targets for these SSO were polymerase chain reaction (PCR) products using primers specific for the second exon of DRB genes. In view of practically, a hybridization system was developed which was independent of temperature or composition of the hybridization buffers, hence uniformly applicable to all SSO, yet allowing discrimination based upon one nucleotide mismatch only. The SSO typing results were compared with HLA serological typing and a high correlation was observed (99%).