Year : 2010 | Volume
: 53 | Issue : 3 | Page : 433--438
Evaluation of the rheumatoid factors of the IgG, IgM and IgA isotypes as prognostic parameters for rheumatoid arthritis among Iraqi patients
Mohanad M Ahmed, Khalid A Obaid Al-Ruhaimi, Suhad Hadi Mohammed
Department of Rheumatology and Rehabilitation, Al Hussain Hospital, Kerbala Health Directory, College of Medicine, Kerbala University, Kerbala, Iraq
Mohanad M Ahmed
Department of Microbiology, College of Medicine, Kerbala University, Kerbala
Context: Rheumatoid arthritis (RA) has a heterogeneous course, spanning from mild forms tending to remission and reacting well to treatment, to aggressive forms resistant to classical therapeutic measures. Reliable predictive parameters of the disease course in RA are needed. Raised levels of Rheumatoid Factors (RFs) are associated with RA and that this RF is found in IgM, IgA and IgG classes (isotypes). Aims: To figure out the value of RF isotypes titers as predictors for RA processes and outcomes. Materials and Methods: Fifty three RA patients were enrolled in this study. The patients were diagnosed based on ACR criteria. Blood sample was taken from each patient at time of attending; sera were separated immediately and kept frozen at -70oC until used. Disease Activity Score (DAS) was calculated using DAS28-3 formula. Radiographs were read by expert radiologists. Sandwich Enzyme-linked Immunosorbent Assay (ELISA) was used for the separate quantitative detection of RF of the IgG, IgM and IgA classes in serum. Statistical Analysis Used: Chi-square, Pearson«SQ»s correlation coefficient and ROC statistical analyses was performed using SPSS version 15.0. Results: Among the 53 RA patients who were enrolled in this study, there were statistically significant positive correlations between the presence of radiological joint changes with serum levels of IgG-RF, IgM-RF and IgA-RF as measured by calculation of area under curve (0.772, 0.703 and 0.769, respectively). However, no correlation could be found between those RF isotypes with any of other disease processes and outcomes. Conclusions: These results may indicate the importance of the titers of those isotypes as good predictors of erosive RA and may reflect a causal relationship between their titers and joint damage during the course of RA.
|How to cite this article:|
Ahmed MM, Obaid Al-Ruhaimi KA, Mohammed SH. Evaluation of the rheumatoid factors of the IgG, IgM and IgA isotypes as prognostic parameters for rheumatoid arthritis among Iraqi patients.Indian J Pathol Microbiol 2010;53:433-438
|How to cite this URL:|
Ahmed MM, Obaid Al-Ruhaimi KA, Mohammed SH. Evaluation of the rheumatoid factors of the IgG, IgM and IgA isotypes as prognostic parameters for rheumatoid arthritis among Iraqi patients. Indian J Pathol Microbiol [serial online] 2010 [cited 2021 Nov 27 ];53:433-438
Available from: https://www.ijpmonline.org/text.asp?2010/53/3/433/68265
Rheumatoid arthritis (RA) is a chronic inflammatory autoimmune disease characterized by synovial cell over-proliferation, mononuclear cell infiltration and pannus formation, which results finally in the destruction of cartilage and bone. It has a heterogeneous course, spanning from mild forms tending to remission and reacting well to treatment, to aggressive forms resistant to classical therapeutic measures. Extra-articular manifestations (vasculitis, rheumatoid nodules) occur, especially in patients with long-standing RA. The precise etiology and pathogenesis of RA remain elusive. Genetic background and environmental triggers are both important in the disease determination of RA. ,
One of the characteristics of RA is the presence of certain autoantibodies. Rheumatoid factor (RF) is a family of autoantibodies that recognizes the 'fraction crystallizable' (Fc) part of IgG molecules and exists as IgA-, IgG- and IgM-isotypes. Rheumatoid factor is detected in majority of patients with established disease, and constitutes one of the American College of Rheumatology (ACR) classification criteria.  Rheumatoid factor has been the subject of intensive study, but definite conclusions on its role in RA have not been drawn. Moreover, several recent studies have generated interest in the value of positive titers of autoantibodies as markers of rheumatic diseases. The levels of RF give some indication of prognosis, albeit a rather poor one in this highly variable disease.  In this context, various prospective studies showed a clear association between RF at baseline and the later development of cartilage and bone erosions. ,
The pathogenic implication of RF in RA may involve formation of immune complexes able to mediate tissue damage by complement activation or Fc receptor ligation.  Fc receptors (FcγRI, FcγRII and FcγRIII) are cell surface receptors expressed on various leukocytes specifically binding IgG. IgG immune complexes (ICs) crosslink these FcγRs and activate leukocytes effector functions, such as respiratory cellular burst, cytokine secretion, antibody-dependent cellular cytotoxicity and phagocytosis. ,, These FcγRs have a well-defined role in antibody-induced inflammation and clearance of antigen-antibody complexes.  Experimental studies using mice deficient in various FcγRs have demonstrated an important role of FcγRs in the pathology of RA. , The interaction of IC with particular FcγRs, in concert with other factors, influences aspects of the disease including joint inflammation, matrix and cartilage destruction, metalloprotease activation and proinflammatory cytokine secretion. ,, In human RA studies, expression levels of FcγRII and FcγRIII upregulated in monocytes and macrophages resulted in proinflammatory cytokine production and arthritis inflammation. 
The main objective of this study is to figure out the value of RF isotypes titers in term of predictors for RA processes and outcomes. To achieve our objectives, we examine the RF serum levels of each of the three main classes, IgG, IgM and IgA and compared them with various diseases processes and outcomes such as Disease Activity Score (DAS), radiological changes, disease presentation and presence of extra-articular manifestations to see if any can be linked to a particular isotype of RFs.
Materials and Methods
Fifty three RA patients were enrolled in this study. The patients were diagnosed based on clinical and simple laboratory investigations. Baseline data about patients obtained from clinical and laboratory examinations were arranged in a questionnaire for each patient. Blood sample was taken from each patient at the time of attending; sera were separated immediately and kept frozen until used.
Disease Activity Score (DAS) was calculated from the tender joint count (TJC), swollen joint count (SJC) and erythrocyte sedimentation rate (ESR) according to the following formula: 
DAS28-3= (0.56 Χ √(TJC28) + 0.28 Χ √(SJC28) + 0.70(In) (ESR)) 1.08
where TJC28 = number of tender joints from 28 joints; SJC28 = number of swollen joints from 28 joints and ESR expressed in mm/1st hour.
Twenty-eight tender and swollen joint scores include shoulders, elbows, wrists, metacarpophalangeal joints, proximal interphalangeal joints and knees. The DAS has a continuous scale ranging from 0 to 10, and according to the EULAR criteria,) the level of disease activity can be interpreted (>5.1 = High disease activity, <5.1-> 3(.)2 = Moderate disease activity, <3.2 = Low disease activity, and <2.6 = Remission. Radiographs of hands and feet were used to assess the structural joint damage (erosions) in RA. Those radiographs were read by at least two expert radiologists.
Sandwich Enzyme-linked Immunosorbent Assay (ELISA) was used for the separate quantitative detection of RF of the IgG, IgM and IgA classes in serum. The assay is based on the use of ELISA plate coated with highly purified Fc fragment of human IgG, which is then used to 'capture' the relevant autoantibody in the test serum. The antibody-autoantibody complex is then reacted with third antibody specific to certain autoantibody isotype. This complex is then detected by measuring the activity of an appropriate enzyme that had previously been covalently attached to the third* antibody. ELISA kits were purchased from Aida GMBH, Germany. Due to the lack of international reference calibration this assay is calibrated in arbitrary units (U/ml) for IgG and IgA RFs. For IgM RFs, the assay is calibrated against the international WHO standard and results are given in IU/ml. According the manufacturer instructions the following cutoff values were used: 18 U/ml for IgG-RF, 18 IU/ml for IgM-RF and 15 U/ml for IgA-RF.
Statistical analysis was performed using SPSS statistical package for social and medical science.
Patient's characteristics of the study cohort are summarized in [Table 1]. The mean age of patients was 49.5 years with the majority less than 55 years. Most of the patients were female (88.7%) with mean disease duration of (40.4 months). The mean value of DAS28 was 4.6 (SD: 1.7). More than half of the patients were found to have radiological changes whereas extra-articular manifestations were sought in 20.8% of them.
When RF were examined qualitatively (positive versus negative) using a specified cut-off value (advised by the ELISA kit manufacturer), the highest positivity was found with the IgG isotype (83.03%), followed by IgM (62.26%) and the least positive was IgA (41.5%) as shown in [Figure 1]. Moreover, all cases were found to be positive for at least one isotype (data not shown), a result giving a sensitivity of hundred percent for the simultaneous use of combination of the three isotypes. What is important to note here is that the traditional latex RF test determine IgM RF, and IgM was found be present in lower percentage of patients, in another word, latex RF miss about 38% of the cases.
Using chi-square, we tried to find out whether there is any association between the RF isotypes measured qualitatively with radiological changes, presence of extra-articular manifestations and disease presentation. Only IgG RF was found to be highly associated with presence of radiological changes as measured by Fisher exact test (P = 0.001) [Table 2].
Current disease activity can be assessed, both for clinical and research purposes, by a joint disease activity score (DAS28-3) comprising SJC, TJC and ESR. 
In the current study, DAS28-3 was assessed only in 42 patients and the majority of cases were found to have a moderate DAS (25 out of 42).
When RF were examined qualitatively (positive versus negative) using the specified cut-off value, no association was found with the DAS measures using DAS28-3 [Table 3].
We analyzed the correlation of RF isotypes as measured quantitatively (in a continuous scale) with the studied parameters using Pearson's correlation coefficient. This analysis yielded a correlation matrix that is shown in [Table 4]. Interrelation among IgG, IgM and IgA titers was highly significant, where IgG was found to have highly significant positive correlation with IgM and IgA titers (r = 0.892, r = 0.648, respectively) in addition to highly significant positive correlation between IgM and IgA (r = 0.660). As shown in [Table 4], a positive correlation between the newly diagnosed cases and DAS28-3 was clear in this study (r = 0.361, P = 0.016). As shown in [Table 5], ROC analysis was used to find the correlation of IgG-RF, IgM-RF and IgA-RF, time of presentation, atypical RA presentation and DAS28-3 score with the radiological changes and extra-articular manifestations. All of the three RF isotypes showed statistical significant correlation with radiological changes, where the calculated areas under curves were comparable (0.77, 0.70 and 0.77, respectively); however, no significant correlation was found with the extra-articular manifestations [Table 5]. Moreover, a negative correlation was found between newly diagnosed cases and erosions (AUC: 0.771).
It became clear that reliable predictive parameters of the disease course in RA are needed. The objective of this study is to evaluate the serum levels of RF isotypes as predictors for RA processes and outcomes.
Evidence shows that the radiographic progression occurs at a constant rate in the natural course of RA. ,, Since this radiological damage is considered to be largely irreversible, especially outside the context of biological treatment, it represents a cumulative process of joint destruction overtime. The availability of early, specific prognostic markers for RA is becoming increasingly important, as adequate therapy can halt the structural damage. , In our study; there were statistically significant positive correlations between the presence of radiological joint changes with serum levels of IgM, IgG and IgA as measured by ROC analysis. These results may indicate that RF isotypes titers could be used as prognostic marker for joint damage. In addition, these results confirm abroad prospective studies that showed a clear association of RF at baseline and the later development of erosions. , Moreover, in one of these studies, baseline RF also correlated with the elapsed time until first erosions. , Van Leeuwen et al. found a moderate association between time-integrated RF and the development of erosions in patients with RA.  However, in the above-cited studies, RFs were measured by qualitative or semi-quantitative methods (Waaler-Rose or latex fixation tests) or by more quantitative assays that still apply several dilutions. In the current study, we used a newly developed and highly sensitive ELISA kit that applies single dilution. This method measures RF as a continuous variable, with a wide range of measurements, in one single dilution. Consequently, a matrix effect caused by different dilutions of patient serum can be avoided and reliability is improved.
Extra-articular lesions include cutaneous vasculitis, neuropathy, Felty's syndrome, pericarditis, (interstitial) lung disease that may be life threatening. These extra-articular RA (ExRA) manifestations can have a defining impact on disease outcome, including increased premature mortality compared with RA in general. ,,, Severe ExRA occurs both in patients recently diagnosed with RA and in those with longstanding disease.  The reason why extra-articular features will develop in rare RA patients is unknown.
In this study, interrelation among IgG, IgM and IgA titers was highly significant, as in most of the abroad studies. ,, This result may be consistent with understanding that those isotypes comprise members of the same family of autoantibodies that rise against the same antigen and within the same immune response.
The positive correlation between the newly diagnosed cases and DAS28-3 may be explained in the attitude of the Iraqi patients who seek medical care only the disease become overt and non-negligible, while the significant negative correlation between the newly diagnosed cases and erosions is highly expected where erosions develop late in the disease course.
As a conclusion, RF isotypes may serve as prognostic markers for radiological changes; however, a larger cohort study with moderately long-time follow-up may further solidify this concept.
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