SEROPREVALENCE OF ANTI-MANNOSE BINDING LECTIN AUTOANTIBODIES IN PATIENTS WITH RHEUMATOID ARTHRITIS IN SANA'A CITY- YEMEN

Rheumatoid arthritis (RA) characterized by synovial inflammation and destruction of cartilage and bone. Until now there is no single test that diagnoses RA, however, several blood tests may suggest the presence of this disease. RA is associated with the presence of a number of autoantibodies as such as rheumatoid factor (RF), anti-cyclic citrullinated peptide (ACPA) and antimannose binding lectin (anti-MBL). This study aimed firstly to investigate the presence of anti-MBL autoantibodies in the sera of RA patients and healthy controls and secondly to determine the diagnostic value of this marker in comparison with the classical RF, Creactive protein (CRP) and ACPA among RA cases. This case-control study was conducted at four health establishments; two public (Al-Thawra Modern General Hospital and National Center of Central Public Health Laboratories) and two private (University of Science and Technology Hospital and Aulqi Specialized Medical Laboratories) in Sana'a city. Ninety-four individuals were enrolled in this study. Forty-seven persons were clinically diagnosed to have RA by a rheumatologist and 47 healthy subjects without RA were used as controls. Sera were separated and tested for presence of serum anti-MBL autoantibodies, ACPA, RF and CRP by a commercially available enzyme linked immunosorbent assay (ELISA) and latex agglutination technique. Study results showed that the meanSD for the levels of serum anti-MBL autoantibodies among RA cases were 394243 ng/ml which were significantly higher than that recorded among healthy controls (217173 ng/ml). The levels of serum anti-MBL autoantibodies were associated with positive RF and CRP tests (p=.02 and .007 respectively), but not with positive ACPA test (p=.42). The result of this study showed higher levels of serum anti-MBL autoantibodies among RA cases comparing with the healthy controls and reveal an association with positive results for RF and CRP, but not with ACPA. Therefore, the anti-MBL antibody levels may associated with systemic autoimmune diseases and might not exclusive to RA.


INTRODUCTION
Rheumatoid arthritis (RA) is a chronic autoimmune disease that affects 0.5%-1% of the worldwide population. It is characterized by chronic and erosive polyarthritis which in the majority of patients leads to partial disability or to permanent handicap. The exact etiology of RA is unknown but is believed to be influenced by genetic, environmental, hormonal, immunologic, and infectious factors 1,2 . The diagnosis of RA based on physical exam, radiographic and laboratory tests. Unfortunately, there is no single test can confirm the diagnosis of RA. The blood tests, erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP) are only markers of inflammation. RA is characteristically associated with the presence of many different autoantibodies directed against multiple autoantigens. The first discovered autoantibody in RA was rheumatoid factor (RF). It was found to react with the Fc portion of IgG antibodies. Typically, RF is of IgM isotype, but IgG and IgA may also occur. RF is found in 60-80% of RA patients and thus is fairly sensitive in the RA diagnosis. However, the specificity of RF as a diagnostic marker for RA is low since it is frequently detected in many other conditions such as connective tissue diseases 3,4 . Anti-citrullinated peptide antibody (ACPA) target peptides that are posttranslationally modified by conversion of arginine to citrulline, occurs primarily in patients of RA and demonstrate a much higher specificity of 95.9% for RA. In addition, ACPA seem to be a better marker of poor prognostic features of progressive joint destruction 5,6 . Mannose-binding lectin (MBL) is a liver-derived acute phase protein. This C type lectin has specific binding affinity to mannose and N-acetyl glucoseamine which structurally homologus to C1q, the component of classical complement pathway. When MBL binds to the specific carbohydrate, its associated serine proteases (MBL associated serine proteases "MSAPs") become activated, leading to activation of lectin pathway of the complement system 7-10 . MBL has an important role in provoking an inflammatory response and consequently has been well associated with the pathogenesis of different infectious and autoimmune diseases 11, 13 . It has been shown that MBL binds both dimeric and polymeric IgA and activates complement 14 . Moreover, previous studies demonstrated that MBL can bind agalactosyl IgG and IgM including IgM RF complexes in RA patients, leading to generation of an inflammatory response 13, 15 .The presence of autoantibodies against MBL in serum as well as in synovial fluid from several RA patients has been reported by many studies [16][17][18][19] , and they demonstrated that the anti-MBL decrease the functional activity of MBL in patients in SLE. This study aimed firstly to investigate the presence of anti-MBL autoantibodies in the sera of patients with RA and healthy controls and secondly to determine the diagnostic value of this marker in comparison with the classical RF, CRP and ACPA among RA cases.

SUBJECTS AND METHODS Subjects
The type of study was a case-control study which was carried out during a period from May, 2015 to May, 2016 at Al-Thawra Modern General Hospital,

RESULTS
Out of the 47 cases with RA, females represented 81%, while males represented 19% with a mean age 43.3±14.8 years and 49% of ages were in group ≥50 years. In addition, out of the 47 healthy controls, females represented 83%, while males represented 17% with a mean age 43.2±14.9 years and 47% of ages were in group ≥50 years,( Table 1). The levels of serum anti-MBL autoantibodies were significantly increased among RA cases, in which the mean±SD of anti-MBL levels among RA cases was 394±243 ng/ml higher than the mean±SD of healthy controls which was 217±173 ng/ml with a statistical significance (p=0.001), ( Table  2). As regard the association of anti-MBL with other serological markers. The levels of serum anti-MBL autoantibodies were significantly higher among cases with RF and CRP positive than negative (418.4232, 418.8229 ng/ml, respectively) and the other statistical variables as median, mode and ranges were also higher for cases than controls. On the other hand, the levels of anti-MBL were higher among cases with ACPA negative 415.4208 ng/ml than positive 376.8257 ng/ml (Table 3). The results of this study showed an obvious measurable and a higher significant presence of anti-MBL autoantibodies in the sera of enrolled RA cases (meanSD=394±243 ng/ml) as compared with healthy controls (meanSD=217±173 ng/ml)(p=0.001). This result was similar to that reported by Gupta et al., 26 in India which showed a detectable significant presence of anti-MBL autoantibodies in the sera of RA patients as compared to the controls. This study demonstrated that the anti-MBL were more often in RA patient sera and suggested that it have a diagnostic value for RA. When considered the association of the levels of anti-MBL with the levels of RF and CRP in this patients, the anti-MBL autoantibody levels were significantly associated with the positive cases of these markers in which the meanSD of anti-MBL levels equal to 418.4232 and418.8229 ng/ml, respectively. Current study results were in agreeing with study by Di Muzio et al., 27 , and disagree with study by Gupta et al., which reported that the levels of anti-MBL antibodies were still positive and high in negative cases of RF and CRP 26 . As regard ACPA, there was no association between positive ACPA and the levels of anti-MBL autoantibodies, in which the meanSD of anti-MBL levels for ACPA positive cases was 376.8257 ng/ml, and for negative ACPA were 415.4208 ng/ml. To our knowledge, there is no previous study in the relation between anti-MBL levels and ACPA in RA patients that agreed or disagreed with current results. and according to the association of anti-MBL autoantibody levels with positivity of RF and CRP but not with ACPA in current results, we can have concluded that the anti-MBL antibody levels may associated with systemic autoimmune diseases and might not exclusive to RA.

CONCLUSION AND RECOMMENDATIONS
In current study, the levels of serum anti-MBL autoantibody were significantly increased in RA cases when compared with the healthy controls and a significant association was found between the levels of serum anti-MBL autoantibody, RF and CRP positive results, but no association with ACPA positive results.
Further studies should be applied to detect the diagnostic value of serum anti-MBL autoantibody levelsin RA as well as the molecular mechanism of its interaction in RA.

AUTHOR'S CONTRIBUTION
The manuscript was carried out, written, and approved in collaboration with all authors.