We evaluated the association between anti-cyclic citrullinated peptide antibodies (anti-CCP) and anti-mutated citrullinated vimentin antibodies (anti-MCV) with the presence of extra-articular (ExRA) manifestations in 225 patients with rheumatoid arthritis (RA). Ninety-five patients had ExRA and 130 had no ExRA. There was no association of anti-CCP and anti-MCV levels with the presence of ExRA as total group (
Extra-articular manifestations (ExRA) in patients with rheumatoid arthritis (RA) have been observed with at a frequency of 17.8 to 40.9% [
Besides of RF and ANA, other autoantibodies have been tested as factors associated with ExRA. Anti-cyclic citrullinated peptide (anti-CCP) antibodies are commonly observed in the serum of RA patients, where the frequency varies between 55% and 69% [
Antibodies against mutated citrullinated vimentin (anti-MCV) have been tested most recently for the diagnosis of RA, showing high sensitivity and specificity for the diagnosis of the disease [
Therefore, because the information about the possible relationship between anti-MCV and ExRA is still insufficient, we designed a study to evaluate whether there is an association of anti-MCV or anti-CCP with ExRA.
We evaluated consecutive patients with RA from an outpatient secondary-care center in Guadalajara, Mexico (Department of Internal Medicine-Rheumatology, Hospital General Regional 110, IMSS). Study participants were included if they were Mexicans mestizo (defined as having at least two generations of ancestors born in Western Mexico) and only one person per family was recruited. To be included, patients with RA had to meet the ACR 1987 criteria for RA to have an established diagnosis and be 18 years old or older. Pregnant or nursing patients, those with other autoimmune disorders such as myasthenia gravis, Hashimoto tiroiditis, or any overlapping syndrome were excluded. Patients with diagnosis of chronic infections including B or C hepatitis, human immunodeficiency virus, tuberculosis, or other chronic infections were also excluded (the assessment for these exclusion criteria was based on the information obtained in a chart review of each patient).
Patients were invited to participate and after signing an informed consent they were assessed by one researcher through a structured interview about epidemiological characteristics (such as age at the time of the study) and disease antecedents; disease duration was defined as the time from the onset of first symptoms of RA until the inclusion in the present study and was assessed through DAS-28 for disease activity, HAQ-DI for functioning, and other clinical measures.
All the included patients were systematically assessed during the evolution of the disease and at the time of the study for presence of eExRA (Table
Specific extra-articular manifestations evaluated in patients with RA.
Extra-articular manifestation | Criteria |
---|---|
(1) Pericarditis | Clinical judgment and confirmed by echocardiography |
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(2) Pleuritis | Clinical judgment and thorax radiographs |
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(3) Felty’s syndrome | Clinical evidence confirmed by ultrasound and neutropenia <1.8 |
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(4) Major cutaneous vasculitis | Clinical judgment confirmed by biopsy |
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|
(5) Neuropathy | Clinical judgment and positive results poly/mononeuropathy at electromyography |
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|
(6) Scleritis, epiescleritis, uveitis, or retinal vasculitis | Identified by an specialist and if required a biopsy was performed |
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(7) Glomerulonephritis | Corroborated by nephrologist and if required a renal biopsy was performed |
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(8) Vasculitis involving other organs | Identified by an specialist and if required a biopsy was performed |
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(9) Amyloidosis | Clinical judgment and positive biopsy if required |
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(10) Keratoncunjunctivitis sicca | Clinical judgment: (a) positive Rose-Bengal staining and (b) positive Schirmer’s test <5 mm/5 mn |
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(11) Xerostomia | Clinical judgment and abnormal sialometry and if suspected minor salivary gland biopsy with lymphocytic infiltrate |
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(12) Secondary Sjögren’s syndrome | Diagnosed by two of the following criteria (a) keratonconjunctivitis sicca, (b) xerostomia, and (c) positivity for anti-Ro or anti-La antibodies |
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(13) Bronchiolitis obliterans | Clinical judgment by pulmonologist |
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(14) Organizing Pneumonia | Clinical judgment by pulmonologist |
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(15) Pulmonary fibrosis | Clinical judgment by pulmonologist plus restrictive pattern in lung function test and confirmed by positive findings in high-resolution computed tomography of the lung |
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(16) Cervical myelopathy | Clinical judgment and radiograph showing increased in atlantoaxial distance |
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(17) Subcutaneous rheumatoid nodules | Clinical judgment and biopsy if required |
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(18) Chronic anemia | Diagnosed if a hemoglobin <11 g/dL was observed in the last 6 months before the evaluation and other causes of anemia were excluded |
At the same day of the clinical evaluation, a venous blood sample was obtained to quantify the titers of anti-CCP and anti-MCV antibodies. This sample was centrifuged and the serum was stored to
RF and C-reactive protein (CRP) were quantified in serum by nephelometry using a venous blood sample taken at the same day of the evaluation; the RF was measured on IU/mL and assessed using standard nephelometric assay according to the manufacturer’s specifications (Dade Behring, DE). A positive result was defined as a level of >20 UI/mL. CRP was measured on mg/L and assessed using VITROS Chemistry systems CRP Slides, according to the manufacture’s specifications (Ortho Clinical Diagnostics, INC.100.Indigo Creek Drive; Rochester NY 14626-5101).
Erythrocyte sedimentation rate (ESR) was determined using Wintrobe technique.
Quantitative variables were expressed as means and standard deviations and qualitative variables in frequency and percentages. Comparisons in proportions between groups were performed with Chi-square (or Fisher exact test when required) and comparisons in means between groups were performed by Student’s
All analyses were performed two-tailed, and statistical significance was considered when
The study was approved by the Research and Ethics Committee of the Hospital in Guadalajara, Mexico. The study was approved by the Research and Ethics Committee of the Hospital Number of approval R-2010-1303-29. All participants signed a letter of voluntary informed consent. The study protocol followed the guidelines of the Helsinki declaration.
Figure
Study flow chart. RA: rheumatoid arthritis; ExRA: extra-articular manifestations; Anti-CCP: Anti-cyclic citrullinated peptide; anti-MCV: antimutated citrullinated vimentin.
Table
Selected characteristics in patients with rheumatoid arthritis.
Characteristic | RA |
---|---|
Female, |
207 (92.0) |
Age, years |
|
Alcohol consumption*, |
28 (12.4) |
Smoke exposure, |
64 (28.4) |
RA characteristics | |
Disease duration (years), mean |
|
DAS28, mean |
|
HAQ-Di score (units) |
|
Global functional status III-IV, |
43 (19.1) |
Steinbrocker stage in hands III or IV |
94 (41.8) |
ExRA, |
95 (42.2) |
Extra-articular manifestations | |
Sjögren’s Syndrome, |
58 (25.8) |
Chronic anemia, |
33 (14.7) |
Rheumatoid nodules, |
23 (10.2) |
Peripheral neuropathy, |
12 (5.3) |
Pulmonary fibrosis, |
6 (2.7) |
Raynaud's phenomenon, |
3 (1.3) |
Felty’s Syndrome, |
2 (0.9) |
Thrombosis, |
2 (0.9) |
Scleritis, |
1 (0.4) |
Intestinal vasculitis, |
1 (0.4) |
Laboratory findings | |
ESR, mm/h |
|
CRP, mg/L |
|
Positive RF**, |
135 (66.2) |
RF titres, IU/mL |
|
Positive anti-CCP | 155 (68.9) |
Anti-CCP titres, IU/mL |
|
Positive anti-MCV | 157 (69.7) |
Anti-MCV titres, IU/mL |
|
(+) anti-CCP and (+) anti-MCV, |
140 (62.2) |
(+) anti-CCP, (+) anti-MCV and (+) RF, |
110 (53.9) |
Treatment | |
Synthetic DMARDs, |
206 (92.6) |
MTX users, |
180 (80.0) |
Anti-TNF |
19 (8.4) |
Corticosteroids, |
214 (95.1) |
Prednisone (current doses), mg/day |
|
RA: rheumatoid arthritis; *Alcohol consumption: defined as consume of at least one alcoholic beverage on a daily basis in the last year. DAS28: disease activity score; HAQ-Di: Health Assessment Questionnaire-Disability Index; ExRA: rheumatoid arthritis with extra-articular manifestations; ESR: erythrocyte sedimentation rate; **RF: rheumatoid factor (only assessed in two thousand four patients); CRP: C-Reactive Protein; anti-CCP: anti-cyclic citrullinated peptide antibodies; anti-MCV: anti-mutated citrullinated vimentin, DMARDs: disease-modifying antirheumatic drugs.
Qualitative variables are expressed as frequencies (%); quantitative variables are expressed as means and standard deviation.
Table
Comparisons of characteristics between patients with ExRA and without extra-articular manifestations (RA w/out).
Clinical characteristics | RA w/out |
ExRA |
|
---|---|---|---|
Female, |
116 (89.2) | 91 (95.8) | 0.73 |
Age, years |
|
|
0.006 |
Alcohol consumption, |
21 (16.2) | 7 (7.4) | 0.49 |
Smoke exposure, |
30 (23.1) | 34 (35.8) | 0.04 |
Disease duration, years |
|
|
<0.001 |
DAS28 |
|
|
0.91 |
Tender joint count |
|
|
0.004 |
Swollen joint count |
|
|
0.43 |
VAS global |
|
|
0.16 |
HAQ-Di score (units) |
|
|
0.18 |
Global functioning status III-IV, |
23 (17.7) | 20 (21) | 0.53 |
Radiological Stage III or IV in hands, |
45 (34.7) | 49 (51.6) | 0.01 |
ESR, mm/h |
|
|
0.79 |
CRP, mg/L |
|
|
0.01 |
RF, IU/mL |
|
|
0.99 |
Anti-CCP, IU/mL |
|
|
0.40 |
Anti-MCV, IU/mL |
|
|
0.91 |
MTX users, |
112 (86.2) | 68 (71.6) | 0.007 |
MTX dose, mean |
|
|
0.12 |
Anti-TNF agents users, |
9 (6.9) | 10 (10.5) | 0.34 |
RA w/out: rheumatoid arthritis (RA) without extra-articular manifestations; ExRA: RA with extra-articular manifestations; DAS28: disease activity score; HAQ-Di: Health Assessment Questionnaire-Disability Index; ESR: erythrocyte sedimentation rate; RF: rheumatoid factor; CRP, C-Reactive Protein; anti-CCP: anti-cyclic citrullinated peptide antibodies; anti-MCV: anti-mutated citrullinated vimentin, DMARDs: disease-modifying antirheumatic drugs. Qualitative variables are expressed as frequencies (%); quantitative variables are expressed as mean and standard deviation. Comparisons between proportions were compared with Chi-square or Fisher exact test (when required). Comparisons between means were evaluated with Student’s
In Table
Comparisons of frequencies of ExRA and specific extra-articular manifestations according to the findings of positive or negative anti-CCP or anti-MCV.
Characteristics | (−) RF |
(+) RF |
|
(−) anti-CCP |
(+) anti-CCP |
|
(−) anti-MCV |
(+) anti-MCV |
|
---|---|---|---|---|---|---|---|---|---|
Presence of ExRA, |
30 (43.5) | 57 (42.2) | 0.86 | 27 (38.6) | 68 (43.9) | 0.46 | 26 (38.2) | 69 (43.9) | 0.43 |
Specific manifestations | |||||||||
Sjögren syndrome, |
23 (33.3) | 29 (21.5) | 0.07 | 18 (25.7) | 40 (25.8) | 1.00 | 16 (23.5) | 42 (26.8) | 0.61 |
Chronic anemia, |
9 (13.0) | 23 (17.0) | 0.46 | 7 (10.0) | 26 (16.8) | 0.18 | 9 (13.2) | 24 (15.3) | 0.69 |
Rheumatoid nodules, |
2 (2.9) | 19 (14.1) | 0.01 | 3 (4.3) | 20 (12.9) | 0.048 | 2 (2.9) | 21 (13.4) | 0.02 |
P. neuropathy, |
6 (8.7) | 5 (3.7) | 0.19 | 5 (7.1) | 7 (4.5) | 0.52 | 4 (5.9) | 8 (5.1) | 0.76 |
The cut-off value to be considered anti-CCP as positive was
In data not shown in tables, no differences were observed in anti-CCP levels, in patients with SS versus patients without these manifestations (69.92 versus 80.15 IU/mL resp.,
Levels of anti-CCP correlated with the score of the Health Assessment Questionnaire-Disability Index (HAQ-Di) (
Table
Comparison in characteristics between specific ExRA manifestations and w/out these manifestations.
Clinical characteristics | Sjögren’s syndrome |
|
Chronic anemia |
|
Rheumatoid nodules |
|
Peripheral neuropathy |
| ||||
---|---|---|---|---|---|---|---|---|---|---|---|---|
Yes |
No |
Yes |
No |
Yes |
No |
Yes |
No |
|||||
Female, |
56 (96.6) | 151 (90.4) | 0.17 | 32 (97.0) | 175 (91.1) | 0.48 | 22 (95.7) | 185 (91.6) | 0.70 | 12 (100) | 195 (91.5) | 0.29 |
Age, years |
|
|
0.001 |
|
|
0.17 |
|
|
0.22 |
|
|
0.07 |
Disease duration, years |
|
|
0.005 |
|
|
0.74 |
|
|
0.001 |
|
|
0.19 |
Year of diagnosis before 2000 | 20 (34.5) | 30 (18.0) | 0.009 | 6 (18.2) | 44 (22.9) | 0.55 | 10 (43.5) | 40 (19.8) | 0.010 | 4 (33.3) | 46 (21.6) | 0.31 |
DAS28 |
|
|
0.29 |
|
|
0.005 |
|
|
0.79 |
|
|
0.21 |
HAQ-Di score (units) |
|
|
0.11 |
|
|
0.81 |
|
|
0.29 |
|
|
0.49 |
Radiological III-IV hands, |
28 (48.3) | 66 (39.5) | 0.24 | 17 (51.5) | 77 (40.1) | 0.22 | 16 (69.6) | 78 (38.6) | 0.004 | 7 (58.3) | 87 (40.8) | 0.23 |
ESR, mm/h |
|
|
0.08 |
|
|
0.33 |
|
|
0.51 |
|
|
0.09 |
CRP, mg/L |
|
|
0.62 |
|
|
0.015 |
|
|
0.26 |
|
|
0.19 |
(+) RF*, |
29 (55.8) | 106 (69.7) | 0.07 | 23 (71.9) | 112 (65.1) | 0.46 | 19 (90.5) | 116 (63.4) | 0.01 | 5 (45.5) | 130 (67.4) | 0.19 |
(+) Anti-CCP, |
40 (69.0) | 115 (68.9) | 0.99 | 26 (78.8) | 129 (67.2) | 0.18 | 20 (87.0) | 135 (66.8) | 0.048 | 7 (58.3) | 148 (69.5) | 0.52 |
(+) Anti-MCV, |
42 (72.4) | 115 (68.9) | 0.61 | 24 (72.7) | 133 (69.3) | 0.69 | 21 (91.3) | 136 (67.3) | 0.02 | 8 (66.7) | 149 (70.0) | 0.76 |
Chloroquine users, |
14 (24.1) | 17 (10.2) | 0.008 | 4 (12.1) | 27 (14.1) | 1.00 | 4 (17.4) | 27 (13.4) | 0.53 | 1 (8.3) | 30 (14.1) | 1.00 |
MTX users, |
41 (70.7) | 139 (83.2) | 0.04 | 26 (78.8) | 154 (80.2) | 0.85 | 12 (52.2) | 168 (83.2) | 0.001 | 8 (66.7) | 172 (80.8) | 0.26 |
MTX doses, mean |
|
|
0.15 |
|
|
0.46 |
|
|
0.32 |
|
|
0.10 |
Azathioprine users, |
6 (10.3) | 13 (7.8) | 0.55 | 5 (15.2) | 14 (7.3) | 0.17 | 9 (39.1) | 10 (5.0) |
|
2 (16.7) | 17 (8.0) | 0.27 |
Anti-TNF agents, users, |
6 (10.3) | 13 (7.8) | 0.59 | 2 (6.1) | 17 (8.9) | 1.00 | 4 (17.4) | 15 (7.4) | 0.11 | 0 (0.0) | 19 (8.9) | 0.61 |
RA: rheumatoid arthritis; DAS28: disease activity score; HAQ-Di: Health Assessment Questionnaire-Disability Index; ExRA: rheumatoid arthritis with extra-articular manifestations; ESR: erythrocyte sedimentation rate; CRP: C-Reactive Protein; *RF: rheumatoid factor (only assessed in two thousand four patients); anti-CCP: anti-cyclic citrullinated peptide antibodies; anti-MCV: anti-mutated citrullinated vimentin, DMARDs: disease-modifying antirheumatic drugs. Qualitative variables are expressed as frequencies (%); quantitative variables are expressed as means and standard deviation. Comparisons between proportions were made with Chi-square (or Fisher exact test if required). Comparisons between means were made using Student’s
Table
Multivariate logistic regression testing for variables associated with ExRA.
Criterion predictor | Method Enter | Method forward stepwise | ||||
---|---|---|---|---|---|---|
OR | 95% CI |
|
OR | 95% CI |
| |
Age, years | 1.08 | 1.02–1.13 | 0.002 | 1.05 | 1.01–1.10 | 0.015 |
Disease duration, years | 1.13 | 1.04–1.23 | 0.002 | 1.09 | 1.02–1.16 | 0.007 |
Smoke exposure | 0.70 | 0.25–2.00 | 0.41 | Not in the model | — | — |
DAS-28 | 2.24 | 1.42–3.52 | <0.001 | 1.86 | 1.27–2.73 | 0.002 |
Functional Impairment (HAQ-Di) | 5.62 | 1.78–17.75 | 0.003 | 4.46 | 1.51–13.14 | 0.007 |
CRP mg/L | 1.02 | 1.00–1.05 | 0.025 | Not in the model | — | — |
ESR mm/hr | 0.95 | 0.90–0.99 | 0.02 | Not in the model | — | — |
RF IU/mL | 1.27 | 0.45–3.54 | 0.64 | Not in the model | — | — |
Anti-CCP IU/mL | 1.00 | 0.99–1.01 | 0.84 | Not in the model | — | — |
Anti-MCV IU/mL | 1.00 | 0.99–1.002 | 0.63 | Not in the model | — | — |
Anti-TNF agents | 1.63 | 0.36–7.50 | 0.53 | Not in the model | — | — |
DAS28: disease activity score 28-joints assessed; HAQ-Di: Health Assessment Questionnaire-Disability Index; MTX: methotrexate; Anti-CCP: anti-cyclic citrullinated peptide antibodies, OR: odds ratio, 95% CI, 95% confidential intervals.
Our results showed that 42% of our patients had ExRA; the variables associated with this involvement in the adjusted analysis are older age, longer disease duration, DAS-28 score, and HAQ-DI score, whereas no association was observed between anti-CCP or anti-MCV antibodies and ExRA as general group. For specific ExRA, there was a weak association between the presence of rheumatoid nodules and positivity for RF, anti-CCP, or anti-MCV antibodies but no relationship was found between these autoantibodies and SS, chronic anemia, or peripheral neuropathy.
The frequency of ExRA in RA observed in our study is similar to that described by Turesson et al. [
Because the main objective of the study was to identify the possible association of anti-CCP or anti-MCV autoantibodies with ExRA, we examined primarily all the patients that had one or more ExRA and subsequently, those patients were with a specific ExRA. We did not observe an association between the presence of ExRA as total group and positivity for anti-CCP. Similar results were reported by Sghiri et al. who did not observe association between ExRA and anti-CCP antibodies [
We did not observe an association between anti-MCV and ExRA as total group. Our data are similar to those described by Sghiri et al. [
Our study has several limitations, first because our study was designed to include consecutive patients with RA tested for presence of ExRA and although we evaluated 225 patients, we were unable to identify a significant number of severe ExRA, therefore, we did not test associations in these subgroups. Nevertheless, we identified a significant number of patients with nonsevere ExRA and found no associations except for rheumatoid nodules. Another limitation inherent to this cross-sectional design is that we only tested for anti-CCP and anti-MCV antibodies in one occasion, being necessary in further studies to test these auto-antibodies several times to identify variations in their titers. Hence, further longitudinal studies are required to test if patients with RA with higher titers for these autoantibodies will develop a higher rate of ExRA in the long-run. Another limitation of our study is the long disease duration observed in some of our patients, in this case is expected that the age of our patients at the time of the study is closely related with RA duration. Therefore we observed in the multivariate analysis that disease duration is relevant confounder in the final model.
Relevantly, in the best of our knowledge this is the first study performed in Mexican patients that test the association between anti-CCP or anti-MCV anti-bodies with ExRA, and also is one of the few studies where patients are obtained from a secondary-care center; this last characteristic constitutes an advantage compared with studies performed in tertiary-care centers where ExRA is expected be more severe and are more likely to have referral bias.
In conclusion, we identified that 42% of our patients with RA have extra-articular manifestations. Variables associated with ExRA were age, disease duration, DAS-28 score, and HAQ-DI score. This study did not observe an association between ExRA as total group and the positivity or titers of anti-CCP or anti-MCV antibodies. Although a weak association was observed between positivity for these autoantibodies and rheumatoid nodules. Further studies with more severe ExRA should be performed to identify if these autoantibodies are associated with these specific manifestations.
All authors declare that there is no conflict of interests regarding the publication of this paper.
This project was financed by a Grant from the Mexican Institute for Social Security (IMSS),