The HLA-DRB1 shared epitope alleles differ in the interaction with smoking and predisposition to antibodies to cyclic citrullinated peptide

OBJECTIVE: The HLA shared epitope (SE) alleles are primarily a risk factor for the presence of antibodies to cyclic citrullinated peptide (anti-CCP antibodies) rather than for the development of rheumatoid arthritis (RA). The SE alleles interact with the environmental risk factor tobacco exposure (TE) for predisposition to anti-CCP+ RA. The objectives of this study were to determine 1) whether different SE subtypes contribute differently to the presence of anti-CCP antibodies, 2) whether different SE subtypes all interact with TE for the development of anti-CCP antibodies, and 3) the effect of TE in relation to the SE alleles and anti-CCP antibodies on the risk of progression from undifferentiated arthritis (UA) to RA. METHODS: We assessed the effect of SE subtypes and TE on the presence and level of anti-CCP antibodies and on the risk of progression from UA to RA in 977 patients with early arthritis who were included in the Leiden Early Arthritis Clinic. RESULTS: The HLA-DRB1*0401, *0404, *0405, or *0408 SE alleles conferred the highest risk of developing anti-CCP antibodies (odds ratio [OR] 5.0, compared with an OR of 2.0 for the HLA-DRB1*0101 or *0102 SE alleles and an OR of 1.7 for the HLA-DRB1*1001 SE allele). Conversely, the TE-SE allele interaction was the strongest for the HLA-DRB1*0101 or *0102 SE alleles and the HLA-DRB1*1001 SE allele. TE in SE+, anti-CCP+ patients correlated with higher levels of anti-CCP antibodies and with progression from UA to RA. In logistic regression analysis, only the presence and level of anti-CCP antibodies were associated independently with RA development. CONCLUSION: The HLA-DRB1 SE subtypes differ in their interaction with smoking and in their predisposition to anti-CCP antibodies. TE contributes to the development of RA in SE+, anti-CCP+ patients, which is explained by its effect on the level of anti-CCP antibodies.     

Association of smoking with the constitution of the anti–cyclic citrullinated peptide response in the absence of HLA–DRB1 shared epitope alleles

Objective

Smoking is a risk factor for anti–cyclic citrullinated peptide (anti‐CCP) antibody–positive rheumatoid arthritis (RA) in patients with HLA–DRB1 shared epitope (SE) alleles. It is unknown whether smoking influences not only the presence of these antibodies, but also other characteristics of the anti‐CCP response, such as isotype usage. The aim of this study was to determine the influence of smoking on anti‐CCP isotypes in RA patients, and to determine whether this influence is observed in the presence and/or absence of SE alleles.

Methods

IgA, IgM, and IgG subclasses of anti‐CCP antibodies were measured by enzyme‐linked immunosorbent assay in serum obtained at the first visit to the Leiden Early Arthritis Clinic from 216 patients with anti‐CCP–positive RA whose smoking habits were also assessed. HLA genotyping data were available for 202 of these patients.

Results

IgA and IgM anti‐CCP were more frequent in RA patients who were smokers than in those who were nonsmokers (odds ratio 2.8 and 1.8, respectively). In addition, levels of all isotypes of anti‐CCP, except IgG3, were significantly higher (P < 0.05) in smokers. The number of anti‐CCP isotypes was higher in smokers compared with nonsmokers, both in SE‐negative RA (P = 0.04) and in SE‐positive RA (P = 0.07).

Conclusion

Patients with anti‐CCP–positive RA who have a current or former tobacco exposure display a more extensive anti‐CCP isotype usage in general, and IgA and IgM in particular, compared with patients with anti‐CCP–positive RA who have never smoked. In contrast to its influence on the incidence of anti‐CCP positivity, the influence of tobacco exposure on the constitution of the anti‐CCP response is significant in SE‐negative RA. These findings suggest a differential effect of tobacco exposure on the induction as compared with the propagation of the anti‐CCP antibody response.

     

Regulation of anti–cyclic citrullinated peptide antibodies in rheumatoid arthritis: Contrasting effects of HLA–DR3 and the shared epitope alleles

Objective

To examine the association between HLA–DRB1 alleles and the production of anti–cyclic citrullinated peptide (anti‐CCP) and rheumatoid factor (RF) autoantibodies in patients with rheumatoid arthritis (RA).

Methods

We studied 1,723 Caucasian RA patients enrolled in the North American Rheumatoid Arthritis Consortium (NARAC) family cohort and the Study of New Onset Rheumatoid Arthritis (SONORA) cohort. All patients were tested for anti‐CCP antibodies (by enzyme‐linked immunosorbent assay), RF (by nephelometry), and HLA–DR genotype (by polymerase chain reaction and sequence‐specific oligonucleotide hybridization).

Results

When controlled for the presence of RF, anti‐CCP positivity was strongly associated with the HLA–DRB1 shared epitope (SE). In RF+ patients, the presence of the SE was very significantly associated with anti‐CCP positivity, with an odds ratio (OR) of 5.8 and a 95% confidence interval (95% CI) of 4.1–8.3. This relationship was also seen in RF– patients (OR 3.1 [95% CI 1.8–5.3]). In contrast, RF positivity was not significantly associated with presence of the SE independently of anti‐CCP antibodies. Strikingly, HLA–DRB1*03 was strongly associated with reduced anti‐CCP titers, even after controlling for the presence of the SE and restricting the analysis to anti‐CCP+ patients. HLA–DR3 was also associated with anti‐CCP– RA in our population.

Conclusion

The HLA–DRB1 SE is strongly associated with the production of anti‐CCP antibodies, but not RF. In contrast, HLA–DR3 alleles are associated with anti‐CCP– disease and with lower levels of anti‐CCP antibodies, even when controlling for the SE. These data emphasize the complexity of the genetic effects of the major histocompatibility complex on the RA phenotype.

     

The HLA–DRB1 shared epitope alleles are primarily a risk factor for anti–cyclic citrullinated peptide antibodies and are not an independent risk factor for development of rheumatoid arthritis

Objective

The shared epitope (SE)–containing HLA–DRB1 alleles represent the most significant genetic risk factor for rheumatoid arthritis (RA). Recent studies indicate that the SE alleles are associated with only RA that is characterized by the presence of anti–cyclic citrullinated peptide (anti‐CCP) antibodies, and not with anti‐CCP–negative disease. In this study we investigated whether the SE alleles contribute to the development of anti‐CCP–positive RA, or whether they are associated solely with the presence of anti‐CCP antibodies. We therefore determined the influence of the SE alleles and anti‐CCP antibodies on the progression from recent‐onset undifferentiated arthritis (UA) to RA.

Methods

Patients with recent‐onset UA at the 2‐week visit (n = 570) were selected from the Leiden Early Arthritis Cohort. SE alleles, rheumatoid factor (RF) status, and anti‐CCP antibody levels were determined. Progression to RA or other diagnoses was monitored.

Results

One hundred seventy‐seven patients with UA developed RA during the 1‐year followup, whereas the disease in 393 patients remained unclassified or was given other diagnoses. The SE alleles correlated with the presence of anti‐CCP antibodies, but not with the presence of RF. Both in SE‐positive and in SE‐negative patients with UA, the presence of anti‐CCP antibodies was significantly associated with the development of RA. More intriguingly, however, no apparent contribution of the SE alleles to the progression to RA was found when analyses were stratified according to the presence of anti‐CCP antibodies. In patients with anti‐CCP–positive disease, the presence of SE alleles was associated with significantly higher levels of anti‐CCP antibodies, suggesting that the SE alleles act as classic immune response genes.

Conclusion

The SE alleles do not independently contribute to the progression to RA from UA, but rather contribute to the development of anti‐CCP antibodies.

     

Refining the complex rheumatoid arthritis phenotype based on specificity of the HLA–DRB1 shared epitope for antibodies to citrullinated proteins

Objective

The main genetic risk factor for rheumatoid arthritis (RA), the HLA region, has been known for 25 years. Previous research has demonstrated, within the RA population, an association between HLA–DRB1 alleles carrying the shared epitope (SE) and antibodies directed against cyclic citrullinated peptides (anti‐CCP antibodies). We undertook this study to make the first comparison of SE allele frequencies in the healthy population with those in RA patients who do or do not harbor anti‐CCP antibodies.

Methods

HLA–DRB1 typing was performed in 408 RA patients from the Leiden Early Arthritis Clinic (the Leiden EAC; a Dutch population‐based inception cohort in which disease course was followed up over time), in 423 healthy Dutch controls, and in 720 affected members of 341 US multiplex (sibpair) families of Caucasian origin from the North American RA Consortium (NARAC) with well‐established disease and fulfilling the American College of Rheumatology classification criteria for RA. The presence of anti‐CCP antibodies was determined by enzyme‐linked immunosorbent assay.

Results

For the Leiden EAC, the odds ratio (OR) describing the association of 2 copies of the SE allele with anti‐CCP positivity (using no copies of the SE allele in the healthy control group as the referent) was 11.79 (P < 0.0001), while the OR for 1 SE allele was 4.37 (P < 0.0001). No association with the SE was observed in the Dutch anti‐CCP–negative RA patients. For the NARAC families, linkage and association analysis revealed the SE to be associated only with anti‐CCP–positive disease and not with anti‐CCP–negative disease. Stratified analyses indicated that anti‐CCP antibodies primarily mediated association of the SE with joint damage or disease persistence.

Conclusion

HLA–DRB1 alleles encoding the SE are specific for disease characterized by antibodies to citrullinated peptides, indicating that these alleles do not associate with RA as such, but rather with a particular phenotype.

     

Smoking increases rheumatoid arthritis susceptibility in individuals carrying the HLA–DRB1 shared epitope,regardless of rheumatoid factor or anti–cyclic citrullinated peptide antibody status

Objective

Smoking is associated with rheumatoid arthritis (RA) in individuals with the HLA–DRB1 shared epitope (SE). SE alleles have been shown to be predominantly associated with anti–cyclic citrullinated peptide (anti‐CCP)–positive RA. These risk factors have not been identified for anti‐CCP–negative RA. The aim of this study was to investigate whether SE‐containing HLA–DRB1 alleles, smoking, or the combination of these factors contributes to the development of RA, depending on the presence or absence of serologic markers, in a Korean population.

Methods

All of the patients with RA (n =1,482) and all of the control subjects (n = 1,119) were Korean. Four‐digit HLA–DRB1 typing was performed by a conventional polymerase chain reaction–sequence‐based typing method. Information about smoking history was obtained through a questionnaire. The patients with RA were tested for anti‐CCP antibodies and rheumatoid factor (RF).

Results

The SE alleles had significant effects on anti‐CCP antibody and RF formation. The DRB1*0901 allele was associated with the presence of anti‐CCP antibodies (odds ratio [OR] 2.49) and RF (OR 2.09). SE alleles and smoking were associated with both anti‐CCP–positive and anti‐CCP–negative RA. The combination of smoking and double copies of the SE allele increased the risk of anti‐CCP–positive RA 36.11‐fold and increased the risk of anti‐CCP–negative RA 12.29‐fold, compared with the risk among nonsmokers not carrying SE alleles. Interactions between SE alleles and smoking were observed for both anti‐CCP–positive and RF‐positive RA, although the associations of RF‐positive RA could be consequences of the underlying anti‐CCP antibody status.

Conclusion

We demonstrated that the combination of SE alleles and smoking is associated with RA susceptibility regardless of anti‐CCP antibody or RF status, but that the combination shows stronger effects in anti‐CCP–positive/RF‐positive patients with RA than in anti‐CCP–negative/RF‐negative patients with RA. The SE–smoking interactions were present in anti‐CCP–positive and RF‐positive RA.

     

New classification of HLA–DRB1 alleles supports the shared epitope hypothesis of rheumatoid arthritis susceptibility

Objective

The shared epitope hypothesis was formulated to explain the involvement of HLA–DRB1 in rheumatoid arthritis (RA). However, several studies, which considered only the HLA–DRB1 alleles shown to be associated with RA risk, rejected this hypothesis. In this report, we propose that a different classification of HLA–DRB1 alleles be considered, based on the amino acid sequence at position 70–74.

Methods

The fit of both HLA–DRB1 classifications was tested in 2 groups of RA patients. All subjects were recruited through the European Consortium on Rheumatoid Arthritis Families, and included 100 patients with isolated RA and 132 patients with at least 1 affected sibling.

Results

The new classification produced risk estimates that fit all of the observed data, i.e., the distribution of the HLA–DRB1 genotype in the 2 patient groups, and the distribution of parental alleles shared by affected sibpairs. The risk of developing RA under this new classification depends on whether the RAA sequence occupies position 72–74 but is modulated by the amino acid at position 71 (K confers the highest risk, R an intermediate risk, A and E a lower risk) and by the amino acid at position 70 (Q or R confers a higher risk than D).

Conclusion

A new classification based on amino acid sequence allows us to show that the shared epitope RAA sequence at position 72–74 explains the data, with the risk of developing RA modulated by the amino acids at positions 70 and 71.

     

Gene–environment interaction between the DRB1 shared epitope and smoking in the risk of anti–citrullinated protein antibody–positive rheumatoid arthritis: All alleles are important

Objective

An interaction effect for developing rheumatoid arthritis (RA) was previously observed between HLA–DRB1 shared epitope (SE) alleles and smoking. We aimed to further investigate this interaction between distinct SE alleles and smoking regarding the risk of developing RA with and without anti–citrullinated protein antibodies (ACPAs).

Methods

We used data regarding smoking habits and HLA–DRB1 genotypes from 1,319 patients and 943 controls from the Epidemiological Investigation of Rheumatoid Arthritis, in which 972 patients and 488 controls were SE positive. Subsequently, 759 patients and 328 controls were subtyped for specific alleles within the DRB1*04 group. Odds ratios with 95% confidence intervals (95% CIs) were calculated by means of logistic regression. Interaction was evaluated by calculating attributable proportion due to interaction, with 95% CIs.

Results

A strong interaction between smoking and SE alleles in the development of ACPA‐positive RA was observed for all DRB1*04 SE alleles taken as a group (relative risk [RR] 8.7 [95% CI 5.7–13.1]) and for the *0401 and *0404 alleles (RR 8.9 [95% CI 5.8–13.5]) and the *01 and *10 alleles (RR 4.9 [95% CI 3.0–7.8]) as specific, separate groups, with similar strength of interaction for the different groups (attributable proportion due to interaction 0.4 [95% CI 0.2–0.6], 0.5 [95% CI 0.3–0.7], and 0.6 [95% CI 0.4–0.8], respectively).

Conclusion

There is a statistically significant interaction between distinct DRB1 SE alleles and smoking in the development of ACPA‐positive RA. Interaction occurs with the *04 group as well as the *01/*10 group, demonstrating that regardless of fine specificity, all SE alleles strongly interact with smoking in conferring an increased risk of ACPA‐positive RA.

     

Relationship among the HLA–DRB1 shared epitope,smoking, and rheumatoid factor production in rheumatoid arthritis

Objective

Rheumatoid factor (RF) production in rheumatoid arthritis (RA) is generally associated with more severe disease. In some studies, RF production has been associated with carriage of HLA–DRB1 alleles encoding the RA‐associated shared epitope (SE). Patients who smoke are also more likely to be RF positive. In this study, we investigated whether the association between RF production and smoking was influenced by carriage of the SE.

Methods

The smoking histories of 371 RA patients attending a hospital clinic were recorded. RF levels and SE status were determined for every patient, and the associations between the SE, smoking, and RF production were examined. HLA–DRB1 typing was performed using polymerase chain reaction. Results were analyzed using chi‐square tests and logistic regression analysis.

Results

Patients who had ever smoked were significantly more likely to be RF positive than nonsmokers (odds ratio 2.2, P < 0.0001). This remained significant (P = 0.003) after correction for age, sex, and disease duration in a logistic regression model. An association was also found between RF positivity and carriage of the SE (P = 0.03, after correction for age, sex, and disease duration), but significance was reduced or lost after correction for previous or current smoking (P = 0.05 and 0.09, respectively). Examination of the major SE phenotypes in this RA population by multivariate logistic regression analysis revealed that only DRB1*0401 was associated with RF positivity, and that this was independent of the influence of smoking.

Conclusion

Our data confirm that RF production in RA patients is associated with smoking. This does not appear to depend on an HLA–DR‐restricted immune response. The association of the SE with RF positivity is primarily due to HLA–DRB1*0401. This appears to be independent of the association with smoking, although smoking further increases the likelihood of RF production in DRB1*0401 patients.

     

Particular HLA–DRB1 shared epitope genotypes are strongly associated with rheumatoid vasculitis

Objective

To examine the relationship of the HLA–DRB1 shared epitope (SE) to rheumatoid vasculitis, using individual patient data (IPD) meta‐analytic methods.

Methods

Published studies that enrolled adult patients with rheumatoid arthritis (RA) were identified by searches of Medline and Embase, and by manual searches of medical journals. All authors were contacted for IPD. Meta‐analyses were performed to assess the association of SE presence, dose, and genotype with rheumatoid vasculitis.

Results

A total of 14 studies and 1,568 patients (129 with vasculitis) were included in the analysis. RA patients with vasculitis were significantly more likely to have rheumatoid nodules (odds ratio [OR] 2.5, 95% confidence interval [95% CI] 1.5–3.9], but there was no significant association with male sex, rheumatoid factor positivity, or erosive disease. No significant association was observed between the presence of the SE (i.e., 1 or 2 alleles versus 0 alleles) and rheumatoid vasculitis (summary OR 1.4, 95% CI 0.7–2.7). Analysis by SE genotype, however, demonstrated a striking relationship of vasculitis to 3 genotypes containing a double dose of the SE, specifically HLA–DRB1*0401/*0401 (OR 6.2, 95% CI 1.01–37.9), *0401/*0404 (OR 4.1, 95% CI 1.1–16.2), and *0101/*0401 (OR 4.0, 95% CI 1.4–11.6).

Conclusion

The HLA–DRB1 SE genotypes *0401/*0401, *0401/*0404, and *0101/*0401 may be of particular importance to rheumatoid vasculitis. It is hoped that additional investigation of these and other SE genotypes will lead to improved insight into the mechanisms influencing the clinical expression of RA.

     

Antibodies against cyclic citrullinated peptide are associated with the DRB1 shared epitope and predict joint erosion in rheumatoid arthritis

OBJECTIVE: To evaluate antibodies against cyclic citrullinated peptide (anti-CCP antibodies) for their predictive value for severe joint destruction in rheumatoid arthritis (RA) and to examine their relationship to shared epitope (SE)-positive DRB1 alleles. METHODS: Concentrations of anti-CCP antibodies were determined in sera from 126 patients with recent onset RA who had been followed prospectively for 6 yr. Progression of joint destruction was evaluated according to Larsen by scoring radiographs from the hand and feet taken at baseline and after 1, 2, 4 and 6 yr of observation. In addition to clinical parameters, the presence of SE-positive DRB1 alleles and of rheumatoid factor IgM and IgA was determined. RESULTS: Anti-CCP antibodies were found more frequently and in higher concentrations in both DRB1*01-positive and in DRB1*04-positive SE-positive patients compared with SE-negative patients. Severe joint destruction as defined by a Larsen score in the upper third of the study population was predicted by positivity for anti-CCP antibodies, by the presence of SE-positive DRB1*04 alleles and by the presence of erosive disease at initial presentation. Multiple logistic regression analysis revealed that SE-positive DRB1*04 alleles and anti-CCP antibodies exerted a significant influence on the progression of joint destruction. CONCLUSION: The association of anti-CCP antibodies with DRB1*01 and with SE-positive DRB1*04 alleles implies a functional role for the SE sequence motif. The determination of SE-positive DRB1*04 alleles and of anti-CCP antibody positivity facilitates the prediction of disease course and prognosis at the time of initial presentation.     

中国汉族类风湿关节炎患者HLA-DR4基因与抗环瓜氨酸肽抗体的相关性研究

目的：探讨中国汉族类风湿关节炎（RA）患者中抗环瓜氨酸肽抗体（ACCP）与人类白细胞抗原（HLA）-DR4基因的相关性。方法：入选RA104例、正常对照122名。ACCP检测采用酶联免疫吸附法（ELISA）,类风湿因子检测采用散色比浊法,HLA-DR4基因采用序列特异性引物-聚合酶链方法（PCR-SSP）检测。结果：RA患者中HLA-DR4基因携带率为34.6%,主要亚型为HLA-DRB1＊0405,正常对照组为17.2%,差异有统计学意义（P=0.01）。RA患者的共同表位（SE）携带率为30.9%,与国内相关研究结果（33.2%,36.8%）相似,但明显低于国外相关研究（78.5%,65.4%,85%,67%）,差异有统计学意义（P〈0.01）。RA患者中ACCP阳性率为76.5%,与国内外报道相符,正常对照组为0,两者差异有统计学意义。SE（＋）患者的ACCP阳性率为84%,SE（-）患者的ACCP阳性率73.2%,两者差异无统计学意义。RA患者中ACCP的滴度与X线分期相关（r=0.233,P〈0.05）。结论：我国汉族RA患者中ACCP与HLA-DR4或SE无明显的相关性。ACCP可能与关节破坏的严重程度相关。     

Influence of shared epitope–negative HLA–DRB1 alleles on genetic susceptibility to rheumatoid arthritis

Objective

Most patients with rheumatoid arthritis (RA) express the shared epitope (SE). It is not known whether SE‐negative HLA–DRB1 alleles influence the development of RA. This study examined the influence of SE‐negative HLA–DR alleles (DRB1*X) on the development of RA in 3 different French populations.

Methods

HLA–DRB1 alleles were defined by polymerase chain reaction with sequence‐specific oligonucleotide hybridization or sequence‐specific primers. SE‐negative alleles were classified according to the electric charge of their P4 pocket. HLA–DRB1 alleles *0103, *0402, *07, *08, *11 (except *1107), *12, and *13 have a neutral or negative P4 charge and are called DRB1*XP4n. HLA–DRB1*03, *0403, *0406, *0407, *0901, *1107, *14, *15, and *16 have a positive P4 charge and are called DRB1*XP4p.

Results

Among the SE‐negative subjects, DRB1 genotypes with 1 or 2 DRB1*XP4n alleles were significantly overrepresented in the control subjects compared with the RA patients, whereas DRB1*XP4p/XP4p genotypes were equally represented in the patients and controls. In single‐dose SE–positive subjects, SE/XP4n genotypes were equally represented in the patients and controls. However, SE/XP4p genotypes were significantly overrepresented in the RA patients.

Conclusion

The DRB1*X allele polymorphism influences susceptibility to RA. Alleles that have a neutral or negative electric charge in their P4 pocket (DRB1*XP4n), such as DRB1*0103, *0402, *07, *08, *11 (except *1107), *12, and *13, protect against RA. Alleles that have a positive electric charge in their P4 pocket (DRB1*XP4p), such as DRB1*03, *0403, *0406, *0407, *0901, *1107, *14, *15, and *16, have no influence on the predisposition to RA.

     

The HLA-DRB1 shared epitope is not associated with antibodies against cyclic citrullinated peptide in Chinese patients with rheumatoid arthritis

OBJECTIVE: Rheumatoid arthritis (RA) is a destructive autoimmune polyarthritis that has been associated with a group of human leucocyte antigen (HLA)-DRB1 alleles that share a common amino-acid sequence at residues 70-74 called the shared epitope (SE). Recently, anti-cyclic citrullinated peptide (CCP) antibodies have also been reported to be associated with HLA-DR4 and have gained wide acceptance as early diagnostic markers for RA in Caucasian patients. The current study was performed to investigate whether the association between the SE (HLA-DRB1 0401/04/05/10) and anti-CCP antibodies is also present in Chinese Han patients with RA. METHODS: One hundred and four RA patients and 122 healthy controls were recruited. HLA-DR4 was detected by the sequence-specific primer polymerase chain reaction (SSP-PCR) phototyping method. Anti-CCP antibodies and immunoglobulin M rheumatoid factor (IgM-RF) were measured by enzyme-linked immunosorbent assay (ELISA) and laser nephelometry, respectively. RESULTS: Of the Chinese patients with RA, 76.5% exhibited anti-CCP antibodies compared with none of the controls (76.5% vs. 0%, p<0.001). The prevalence of the SE was significantly higher in patients with RA compared with controls [p = 0.010, odds ratio (OR) = 2.42, 95% confidence interval (CI) = 1.16-5.07]. Among the HLA-DR4 alleles, the presence of HLA-DRB1 0401 was significantly higher in RA patients than in controls (p = 0.0118, OR = 9.68, 95% CI = 1.13-448.8). In our study we found that the SE was not associated with production of anti-CCP antibodies (p = 0.2899, OR = 1.920, 95% CI = 0.52-8.89). CONCLUSIONS: The prevalence of the SE is significantly lower in Chinese RA patients, as compared with previous reports of a study using a Caucasian cohort, indicating that distinct genetic risk factors might be associated with anti-CCP antibody production. These data emphasized the complexity of the genetic effects of the major histocompatibility complex on the RA phenotype.     

Association of smoking with the constitution of the anti-cyclic citrullinated peptide response in the absence of HLA-DRB1 shared epitope alleles

OBJECTIVE: Smoking is a risk factor for anti-cyclic citrullinated peptide (anti-CCP) antibody-positive rheumatoid arthritis (RA) in patients with HLA-DRB1 shared epitope (SE) alleles. It is unknown whether smoking influences not only the presence of these antibodies, but also other characteristics of the anti-CCP response, such as isotype usage. The aim of this study was to determine the influence of smoking on anti-CCP isotypes in RA patients, and to determine whether this influence is observed in the presence and/or absence of SE alleles. METHODS: IgA, IgM, and IgG subclasses of anti-CCP antibodies were measured by enzyme-linked immunosorbent assay in serum obtained at the first visit to the Leiden Early Arthritis Clinic from 216 patients with anti-CCP-positive RA whose smoking habits were also assessed. HLA genotyping data were available for 202 of these patients. RESULTS: IgA and IgM anti-CCP were more frequent in RA patients who were smokers than in those who were nonsmokers (odds ratio 2.8 and 1.8, respectively). In addition, levels of all isotypes of anti-CCP, except IgG3, were significantly higher (P < 0.05) in smokers. The number of anti-CCP isotypes was higher in smokers compared with nonsmokers, both in SE-negative RA (P = 0.04) and in SE-positive RA (P = 0.07). CONCLUSION: Patients with anti-CCP-positive RA who have a current or former tobacco exposure display a more extensive anti-CCP isotype usage in general, and IgA and IgM in particular, compared with patients with anti-CCP-positive RA who have never smoked. In contrast to its influence on the incidence of anti-CCP positivity, the influence of tobacco exposure on the constitution of the anti-CCP response is significant in SE-negative RA. These findings suggest a differential effect of tobacco exposure on the induction as compared with the propagation of the anti-CCP antibody response.     

Association of the HLA–DRB1 epitope LA67, 74 with rheumatoid arthritis and citrullinated vimentin binding

Objective

Although rheumatoid arthritis (RA) has long been associated with an HLA–DRB1 shared epitope, a systematic search for other epitopes has never been conducted. In addition, the relationship between these epitopes and the binding of citrullinated autoantigens has not been investigated. We developed a program that can analyze HLA data for all possible epitopes of up to 5 amino acids and used this program to assess the shared epitope hypothesis in RA.

Methods

We analyzed high‐resolution data from the International Histocompatibility Working Group, which included a group of 488 patients with RA and a group of 448 racially and ethnically balanced control subjects, for all combinations of up to 5 amino acids among polymorphic HLA–DRB1 positions 8–93. Statistical significance was determined by chi‐square and Fisher's exact tests, with a false discovery rate correction.

Results

Three residues (V¹¹, H¹³, and L⁶⁷) were found to have the highest degree of association with RA susceptibility (P < 10^–11), and D⁷⁰ was found to correlate best with RA resistance (P = 2 × 10^–11). Of >2 million epitopes examined, LA^{67, 74} exhibited the highest correlation with RA susceptibility (P = 2 × 10^–20; odds ratio 4.07 [95% confidence interval 3.07–5.39]). HLA alleles containing the LA^{67, 74} epitope exhibited significantly greater binding to citrullinated vimentin^65–77 than did alleles containing D⁷⁰. Only 1 allele (DRB1*16:02) contained both LA^{67, 74} and D⁷⁰; it bound citrullinated vimentin weakly and was not associated with RA.

Conclusion

The findings of these studies confirm the importance of HLA–DRB1 amino acids in pocket 4 for the binding of citrullinated autoantigens and susceptibility to RA.

     

Independent association of rheumatoid factor and the HLA–DRB1 shared epitope with radiographic outcome in rheumatoid arthritis

Objective

Findings of a recent study suggested that HLA–DRB1 alleles encoding the rheumatoid arthritis (RA) “shared epitope” (SE) were not predictive of erosive damage at 2 years in patients with early inflammatory arthritis who were rheumatoid factor (RF) positive, but were predictive in those who were RF negative. The present study was undertaken to determine whether RF status was also important in the association between the SE and radiographic outcome in patients with longstanding RA.

Methods

The association between radiographic outcome, HLA–DRB1, and RF status was examined in 299 RA patients with established disease (5–30 years). Radiographic outcome was measured by scoring radiographs of the hands and feet using the standard radiographs of Larsen. HLA–DRB1 typing was performed using polymerase chain reaction methodology. Results were stratified by RF status and analyzed by multiple regression.

Results

An association between radiographic severity and the SE was found in RF−, but not RF+, patients. RF− patients carrying an SE allele had higher Larsen scores than RF− patients lacking the SE, although there was no association with SE dosage. The mean Larsen score was significantly higher in RF+ patients than in RF− patients, but there were no differences between RF+ patients with 0, 1, or 2 SE alleles. Multiple regression analysis confirmed independent associations of RF and SE positivity with radiographic outcome. No significant associations were found between RF and the SE, or RF and individual SE alleles.

Conclusion

Our data indicate that RF and the SE are independently associated with radiographic outcome in RA. In RF+ patients with longstanding RA, there is no apparent association between the presence of the SE and radiographic damage. However, in RF− patients, although radiographic outcome is generally less severe, there is an association between severity and presence of the SE.