Interleukin-18-promoter polymorphisms are not relevant in rheumatoid arthritis.

Interleukin-18 (IL-18), a member of the IL-1 family, is known to play a relevant role in rheumatoid arthritis (RA) physiopathology mainly by promoting the inflammatory response. The aim of this work was to investigate the possible implication of two single-nucleotide polymorphisms (SNPs) [-607 A/C (rs1946518) and -137 G/C (rs187238)] within the IL-18-promoter region in RA predisposition and clinical course. A total of 362 unrelated RA patients and 339 healthy controls were genotyped using a real-time polymerase chain reaction (PCR) method for the -607 A/C SNP and a sequence-specific PCR method (PCR-SSP) for the -137 G/C polymorphism. No statistically significant differences were observed for both -607 and -137 IL-18-promoter polymorphisms between RA patients and controls, considering either allelic or genotypic frequencies. In addition, no association was found with the haplotypes inferred by the two polymorphisms and RA susceptibility. This was also the case when RA patients were stratified according to sex, age at the onset of the disease, rheumatoid factor status, and extraarticular manifestations. Our data suggest that -607 A/C (rs1946518) and -137 G/C (rs187238) polymorphisms within the IL-18-promoter region do not play a major role in RA predisposition.

HLA haplotypes and susceptibility to rheumatoid arthritis. More than class II genes.

Our aim was to examine, using microsatellite (ms) markers, the contribution of the telomeric part of the HLA region to rheumatoid arthritis (RA) predisposition in the Spanish population. We have looked at the distribution of DQB1, DRBI and five ms loci (D6S1014, D6S273, D6STNFa, MIB and C1-2-5) within the HLA region in 147 Spanish RA patients and 202 control subjects. A total of 19 conserved ms configurations were observed, twelve of them in linkage disequilibrium with particular DQB1-DRB1 haplotypes. Interestingly, haplotype c1 (DQB1*0201-DRB1*0301-D6S1014*143-D6S273*139-D6STNFa*99-MIB*350-C1-2-5*196) was significantly associated with RA predisposition. As part of this haplotype, the MIB*350 allele was found to be a risk factor independently of the RA-predisposing haplotypes. The present results along with data from others prove the existence of a second predisposing locus located inside the MHC region, and suggest that might be located within the TNFa-HLA-B region.

Rheumatoid arthritis in southern Spain: toward elucidation of a unifying role of the HLA class II region in disease predisposition.

OBJECTIVE: To evaluate the contributions of HLA-DQ and -DR polymorphisms to susceptibility to rheumatoid arthritis (RA) in a population in southern Spain, and to compare the value of the shared epitope (SE) and RA protection (RAP) models in accounting for the HLA class II region's contribution to RA predisposition. METHODS: One hundred sixty RA patients and 153 healthy controls were typed for HLA-DRB1 and -DQB1 using high-resolution DNA techniques. Distributions of predisposing DRB1 alleles in patients and control subjects according to the SE model were compared with distributions of predisposing DQ and protective DERAA-positive DRBI alleles according to the RAP model. RESULTS: DQ3 (DQBI*03 and *04 combined with DQA1*03) and DQ5 (DQB1*0501/DQA1*0101) alleles predisposed individuals to RA independently of SE-positive DRB1 alleles. DQ3/3-homozygous individuals had the strongest risk of developing RA. DQ3 molecules predisposed to RA more than did DQ5 molecules. The weaker predisposition mediated by DQ5 included the DRB1*1001-carrying haplotype; no DRB1*1001-homozygous patients were observed. DRBI*0401 played a unique role in the contribution of DQ3-DR4 haplotypes to RA, in spite of its low frequency in southern Spain. CONCLUSION: The low prevalences of RA and of mild disease observed in Spain, and in southern Europe in general, can be explained in great part by the low frequency of DQ3-DR4 haplotypes, especially those carrying DRB1*0401. However, the overall distribution of HLA-DQ and -DR alleles in RA patients compared with control subjects is similar to that in other European and North American populations. A model involving both DQ and DR can best account for the contribution of HLA to RA.

TAP1 and TAP2 polymorphism in Spanish patients with ankylosing spondylitis.

OBJECTIVE: HLA-B27 is strongly associated with ankylosing spondylitis (AS); however, the association is not absolute and additional susceptibility factors in the MHC region could play a role. We studied the influence of polymorphism in the transporter associated with antigen processing (TAP) genes, including point mutations not previously analyzed. METHODS: HLA-B*27 typing and subtyping as well as TAP1 and TAP2 typing were performed by PCR-RFLP. Forty-four AS individuals were compared to 61 ethnically matched random individuals and 35 B*27-positive healthy unrelated individuals as controls. RESULTS: The frequency of the TAP1B allele was significantly greater in the patient group compared with the random controls (corrected p value (p(c)) = 0.035; odds ratio = 15.8, p = 0.01). A greater frequency was also evident when B*27-positive patients and B*27- positive healthy controls were compared, although it did not reach statistical significance. No differences were observed in TAP2 alleles between the groups studied. DISCUSSION: We did not find a primary association between TAP2 polymorphism and AS susceptibility. Formal confirmation of a linkage between the TAP and HLA-B loci would probably require family studies.

Interleukin 6 gene promoter polymorphism is not associated with ankylosing spondylitis.

OBJECTIVE: To investigate the possible association between the recently described interleukin 6 (IL-6) promoter polymorphisms at position -174, and susceptibility to ankylosing spondylitis (AS). METHODS: Ninety-two patients with AS, 157 healthy controls, and an additional group of 52 HLA-B27 positive unrelated individuals were included in this study. The -174 polymorphic site in the promoter region of IL-6 gene was typed by polymerase chain reaction-restriction fragment length polymorphism. RESULTS: No statistically significant differences were observed when IL-6 promoter genotype and allele distribution between patients with AS and healthy controls were compared. CONCLUSION: Our results suggest the -174 IL-6 polymorphism does not play an important role in susceptibility to AS. Larger studies are needed to provide more conclusive evidence on the role of -174 IL-6 polymorphism in AS.

Involvement of Fcgamma receptor IIIA genotypes in susceptibility to rheumatoid arthritis.

OBJECTIVE: To investigate a possible association of Fcgamma receptor IIIA (FcgammaRIIIA) gene polymorphism at position 158 with susceptibility to, and the outcome of, rheumatoid arthritis (RA). METHODS: One hundred seventeen RA patients and 142 unrelated healthy control subjects from the same geographic area were studied. Genotyping for FcgammaRIIIA-158V/F was performed by a method based on polymerase chain reaction (PCR) and restriction fragment length polymorphism analysis using amplification-created restriction sites. HLA-DRB1 typing by PCR-sequence-specific oligonucleotide hybridization (reverse hybridization) was also performed. RESULTS: Allele and genotype distributions in healthy controls were similar to those reported in other populations. The overall distribution of genotypes in the patients was significantly different from that in the controls (P = 0.023, by chi-square test from 3 x 2 contingency table). An overrepresentation of the FcgammaRIIIA-158FF genotype in the patients was observed (for 158FF versus non-158FF P = 0.01, odds ratio [OR] 1.98, 95% confidence interval [95% CI] 1.16-3.4). However, the FcgammaRIIIA-158VF genotype was increased in controls (for 158VF versus non-158VF P = 0.021, OR 0.54, 95% CI 0.32-0.92). No associations were found with any of a series of clinical parameters. Analysis of FcgammaRIIIA-158FF along with shared epitope showed that the presence of both factors increased the susceptibility to RA (P = 0.0009, OR 3.6, 95% CI 1.63-8.01); however, they probably do not interact to produce this effect. CONCLUSION: These results indicate that the FcgammaRIIIA-158 genotypes confer differential susceptibility to RA in our study population. Further studies to elucidate the role of this polymorphism in the pathogenesis of RA and other autoimmune diseases are warranted.

IL-6 promoter polymorphisms in rheumatoid arthritis.

We investigated the possible association between the IL-6 promoter polymorphisms, at positions -622 and -174, and susceptiblity to, and/or outcome of, rheumatoid arthritis (RA). A total of 163 patients with RA and 157 healthy controls were genotyped for IL-6 using a PCR-RFLP method. The -622 and -174 alleles were in complete linkage disequilibrium. No difference was observed in the distribution of IL-6 promoter genotype or allele frequencies between RA patients and controls. However, a significant difference in the mean age at disease onset between IL-6 genotypes was observed. The present data appear to rule out an important role of IL-6 promoter polymorphisms in the susceptibility to RA. However, IL-6 genotypes may contribute to the pathogenesis of the disease by influencing the age at disease onset.

HLA-B*27 subtyping by PCR-RFLP in Spanish patients with ankylosing spondylitis.

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 spondylitis patients and 746 controls from the south of Spain. B*27 showed a very strong association with the disease (OR=211.27, P=10(-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).

HSP70 gene polymorphisms in ankylosing spondylitis.

Despite the strength of the association of ankylosing spondylitis (AS) with HLA-B27, other genetic elements could play a possible role in the pathophysiology of AS. Because of the localization, in the proximity of the HLA-B locus, and the involvement of heat shock proteins (HSP) in the immune response, we analyzed the influence of HSP70 gene polymorphism on the susceptibility to AS. HSP70-1, HSP70-2 and HSP70-hom genotypes were analyzed by PCR-RFLP in patients with AS and in healthy controls. The results obtained in the present study showed that there are not significant differences in the distribution of HSP70-hom genotypes, whereas significant differences in HSP70-1 and HSP70-2 genotypes between AS patients and random controls were found. However, when the distribution of these genotypes were compared in B*27-matched AS patients and controls, the differences disappeared. These data suggest that the polymorphism of HSP70 genes was not independently associated with AS, and that the differences in HSP70-1 and HSP70-2 genotypes among AS patients and controls appears to be due to the linkage disequilibrium between HSP70 alleles and HLA-B*27.

Tumor necrosis factor gene polymorphisms in ankylosing spondylitis.

Despite the strength of the association of ankylosing spondylitis (AS) with HLA-B27, other genetic elements could play a possible role in the pathophysiology of AS. In view of its gene location, in the proximity of the HLA-B locus, and biological effects, tumor necrosis factor (TNF) genes are potential candidates for additive susceptibility factors to AS. TNFalpha and TNFbeta genotypes were analyzed by PCR-RFLP in 57 patients with AS, 102 random controls and 30 HLA-B*27-positive controls. No significant differences of TNFalpha promoter variations at position -308 and -238 were found in AS patients in comparison with controls. The -244 polymorphism was not detected in our population. The TNFbeta genotype frequency was significantly different between AS patients and random controls. However, when the distribution of the TNFbeta genotype was compared in B*27-positive AS patients and controls, these differences disappeared. In addition, we demonstrated that the TNFbeta*1 was in strong linkage disequilibrium with the B*27 allele, which may explain the differences observed for the TNFbeta genotype among AS patients and random controls. Our data suggest that the polymorphisms of TNFalpha and TNFbeta genes do not have an independent effect on AS susceptibility.

Analysis of LMP and TAP polymorphisms by polymerase chain reaction-restriction fragment length polymorphism in rheumatoid arthritis.

OBJECTIVE: The aim of this study was to investigate the relation between the polymorphism of large molecular weight proteasome (LMP) (LMP2-LMP7) and transporter associated with antigen processing (TAP) (TAP1-TAP2) genes and rheumatoid arthritis (RA). METHODS: Sixty RA patients and 102 ethnically matched unrelated healthy subjects were typed for LMP, TAP, and disease associated HLA-DRB1 alleles by using a new strategy based on polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) with amplification created restriction sites. RESULTS: The polymorphism of LMP (LMP2-LMP7) and TAP (TAP1-TAP2) genes was examined in shared epitope positive and negative RA patients and controls. No significant differences in the LMP or TAP allele frequencies were observed between the total patient and control groups or the patients and controls groups or the patients and controls positive or negative for the shared epitope. CONCLUSION: The data suggest that the polymorphisms of LMP and TAP genes do not have an important influence in the pathogenesis of RA, although larger studies will be needed to provide more conclusive evidence on the role of these genes in RA. A new, highly reliable strategy for typing LMP alleles is also described.

Heat shock protein 70 gene polymorphisms in rheumatoid arthritis.

A role for heat shock proteins (hsp) in rheumatoid arthritis has been suggested. In addition, the specific binding of human HSP70 protein to QKRAA and RRRAA motifs within the HV3 region of disease-associated DRB1*0401 and DRB1*1001 molecules, respectively, has been proposed as being relevant to rheumatoid arthritis. The purpose of this work was to analyze the influence of HSP70 gene polymorphism on the susceptibility to or severity of rheumatoid arthritis and to investigate the possible contribution of these HSP70 polymorphisms in determining HLA-DRB1*0401/*1001 disease association. The frequencies of the HSP70-1, HSP70-2 and HSP70-hom genotypes were analyzed by PCR-RFLP using BsrBI, PstI and NcoI enzymes, respectively, in patients with rheumatoid arthritis and in healthy controls. No significant differences were observed when HSP70 allele distribution between the groups under study were compared. Moreover, we did not observe any significant difference in HSP70 allele frequencies between patients positive for HLA-DRB1*0401/*1001 alleles and matched controls. Our data indicate that HSP70 gene polymorphisms do not appear to be relevant in the susceptibility to or severity of rheumatoid arthritis.

Polymorphism at the TNF loci in rheumatoid arthritis.

The purpose of this work was to analyze the possible influence of TNF loci polymorphism on the susceptibility and/or the disease profile of rheumatoid arthritis (RA). Tumor necrosis factor (alpha and beta) genotypes were determined in 60 patients with RA and 102 healthy subjects by a method based on PCR-RFLP with amplification-created restriction sites. The results obtained in the present study showed that there is not a significant association of either TNF alpha promoter variation (at positions -308 and -238) or TNF beta polymorphism with susceptibility to RA. However, a significant difference in the mean age at disease onset was found between -238 TNF alpha genotypes. In addition, a difference in the presence of nodular disease was observed between -308 TNF alpha genotype. The results of this study suggests that the TNF alpha gene may play a role in the disease profile of rheumatoid arthritis.

Association of HLA-DR5 with mucocutaneous lesions in patients with rheumatoid arthritis receiving gold sodium thiomalate.

OBJECTIVE: To investigate the possible association between HLA antigens and adverse reactions to gold sodium thiomalate therapy (GSTM). METHODS: Ninety consecutive patients with rheumatoid arthritis (RA) were studied for possible association between HLA antigens and adverse reactions to GSTM therapy. RESULTS: HLA-DR5 was significantly increased in patients who developed gold induced mucocutaneous lesions. On the other hand, patients with RA carrying B8 and DR3 antigens are of a high risk of developing proteinuria after gold therapy. A very interesting finding was the low incidence of DR7 antigen in patients who developed adverse reactions to GSTM. We also report the relationship between B27 antigen and chrysiasis due to gold therapy. CONCLUSION: Our results support suggestions that the DR7 antigen provides a protective effect against gold toxicity. We also found a strong association between DR5 and mucocutaneous lesions in patients with RA treated with GSTM.

[Comments with respect to 2 patients with anti-Jo 1 antibodies]. / Comentarios a dos enfermos con anticuerpos anti-Jo 1.

Anti-Jo 1 antibodies define a subgroup of patients with polymyositis, which are characterized by the presence of extra-muscular involvement as pulmonary fibrosis, although Raynaud's phenomena, arthritis, tenosynovitis, pleuritis and pericarditis may also be present. Given the fact that extra-muscular signs may precede the clinical diagnosis of polymyositis, as in the two cases that we present here, these antibodies may help us to diagnose the disease before the onset of myositis.

DNA-RFLP analysis and genotyping of HLA-DR alleles in the south of Spain.

42 healthy individuals previously HLA-DR typed by serology, were HLA class II typed using RFLPs. The pattern of hybridization revealed allele specific DNA fragments for some DR specificities. Three fragments highly associated with HLA-DR antigens: a 7 kb RFLP with DR1; a 10 kb RFLP with DR-3/DR-5 and a 13 kb band with DR4/DR7.