Rheumatoid Factor and Anti-immunoglobulin Antibodies
The serological hallmark of rheumatoid arthritis is the detection of rheumatoid factor (RF) and other anti-immunoglobulin (Ig) antibodies. By definition, classical RF is an IgM anti-body to autologous IgG. The more encompassing designation of anti-immunoglobulin anti-bodies is applicable to anti-IgG antibodies of IgG or IgA isotypes. As a rule, the affinity of IgM rheumatoid factor for the IgG molecule is relatively low and does not reach the mean affinity of other IgM antibodies generated during an induced primary immune response.
Rheumatoid factors from different individuals show different antibody specificity, reacting with different determinants of the IgG molecule. In most cases, the antigenic de-terminants recognized by the antigen-binding sites of these IgM antibodies are located in the Cγ 2 and Cγ 3 domains of IgG; some of these determinants are allotype-related. Circu-lating RF reacts mostly with IgG1, IgG2, and IgG4; in contrast, RF detected in synovial fluid react more frequently with IgG3 than with any other IgG subclasses. The significance of these differences is unknown, but they suggest that different B-cell clones may produce circulating RF and synovial RF. Other RF react with determinants that are shared between species, a fact that explains the reactivity of the human RF with rabbit IgG as well as with IgG from other mammalians.
The frequent finding of RF reactive with several IgG subclasses in a single patient suggests that the autoimmune response leading to the production of the RF is polyclonal. This is supported by the fact the idiotypes of RF are heterogeneous, being obviously the product of several different V-region genes.
Rheumatoid factor and anti-Ig antibodies can be detected in the serum of affected patients by a variety of techniques.
The Rose-Waaler test is a passive hemagglutination test which uses sheep or human erythrocytes coated with anti-erythrocyte antibodies as indicators. The agglutination of the IgG-coated red cells to titers greater than 16 or 20 is considered indicative of the presence of RF. These tests detect mostly the classic IgM rheumatoid factor specific for IgG.
The latex agglutination test employs IgG-coated polystyrene particles mixed with serum suspected of containing RF or anti-Ig antibodies . The agglutination of latex particles by serum dilutions greater than 1:20 is considered a positive result. This test detects anti-immunoglobulin antibodies of all isotypes.
As with many other autoantibodies, the titers of RF are a continuous variable within the population studied. Thus, any level intending to separate the seropositive from the seroneg-ative is arbitrarily chosen to include as many patients with clinically defined RA in the seropositive group, while excluding from it as many nonrheumatoid subjects as possible.
Even with these caveats, RF is neither specific nor diagnostic of RA. First, it is found in only 70–85% of RA cases, while it can be detected in many other conditions, particularly in patients suffering from Sjögren’s syndrome. Also, RF screening tests can be positive in as many as 5% of apparently normal individuals, sharing the same V-region idiotypes (and by implication, the same V-region genes) as the antibodies detected in RA patients.
The finding of RF in normal individuals indicates that RF may have a normal, physiologi-cal role, such as to ensure the rapid removal of infectious antigen-antibody complexes from circulation. The synthesis of anti-Ig antibodies in normal individuals follows some inter-esting rules:
1. Anti-Ig antibodies are detected transiently during anamnestic responses to common vaccines and, in these cases, are usually reactive with the dominant immunoglobu-lin isotype of the antibodies produced in response to antigenic stimulation.
2. Anti-Ig antibodies are also found in relatively high titers in diseases associated with persistent formation of antigen-antibody complexes such as subacute bac-terial endocarditis, tuberculosis, leprosy, and many parasitic diseases.
3. The titers of vaccination-associated RF follow very closely the variations in titer of the specific antibodies induced by the vaccine; similarly, the levels of RF de-tected in patients with infections associated with persistently elevated levels of circulating immune complexes decline once the infection has been successfully treated. In contrast, the anti-immunoglobulin antibodies detected in patients with rheumatoid arthritis persist indefinitely, reflecting their origin as part of an autoimmune response.
4. Infection-associated RF binds to IgG molecules whose configuration has been altered as a consequence of binding to exogenous antigens. The resulting RF-IgG-Ag complexes are large and quickly cleared from circulation. The adsorp-tion of IgG to latex particles seems to induce a similar conformational alteration of the IgG molecule as antigen-binding, and as a result IgG-coated latex particles can also be used to detect this type of RF.
The transient nature of anti-Ig antibodies in normal individuals suggests that the au-toreactive clones responsible for the production of autoantibodies to human immunoglob-ulins are not deleted during embryonic differentiation. The persistence in adult life of such autoreactive clones is supported by the observation that the bone marrow contains precur-sors of RF-producing B cells. Their frequency is surprisingly high in mice, where it is rel-atively easy to induce the production of RF in high titers after polyclonal B-cell stimula-tion. Human bone marrow B lymphocytes can also be stimulated to differentiate into RF-producing plasmablasts by mitogenic stimulation with PWM or by infection with Ep-stein-Barr virus. In addition, tolerance to self-IgG must be ensured by a strong negative feedback mechanism(s), since tolerance is broken only temporarily.
In mice and in humans, B lymphocytes capable of differentiating into RF-producing plas-mablasts express CD5 in addition to the classical B-cell markers, such as membrane IgM and IgD, CR2, CD19, and CD20. CD5, expressed by less than 2% of the B lymphocytes of a normal individual, was first detected in patients suffering from very active rheumatoid arthritis. It is considered a marker characteristic of autoimmune situations.
RF titers are highly variable, even in patients with full-blown RA, and do not seem to cor-relate very closely with the activity of the disease. However, high titers of RF tend to be as-sociated with a more rapid progression of the articular component and with systemic man-ifestations, such as subcutaneous nodules, vasculitis, intractable skin ulcers, neuropathy, and Felty’s syndrome. Thus, the detection of RF in high titers in a patient with symptomatic RA is associated with a poor prognosis.
The pathogenic properties of RF are likely to be derived from the biological charac-teristics of the antibodies involved. Classical IgM RF activates complement via the classi-cal pathway, and the ability of RF to fix complement is of pathogenic significance, because it may be responsible, at least in part, for the development of rheumatoid synovitis.
The source of the anti-Ig antibodies that are likely to play an important role in caus-ing arthritic lesions is predominantly the synovium of the affected joints. The joints are the principal sites of RF production in RA patients, and it should also be noted that in some in-dividuals the locally produced anti-Ig antibodies are of the IgG isotype. When this is the case, the joint disease is usually more severe, because anti-IgG antibodies of the IgG iso-type have a higher affinity for IgG than their IgM counterparts; consequently, they form stable immune complexes, which activate complement very efficiently.
Some patients with RA may have negative results on the screening tests for RA. True seronegative RA cases exist, particularly among agammaglobulinemic patients. In spite of their inability to synthesize antibodies, these patients develop a disease clinically indistin-guishable from RF-positive rheumatoid arthritis. This is a highly significant observation since it argues strongly against the role of the RF or other serological abnormalities as a ma-jor pathogenic insult in rheumatoid arthritis and suggests that the inflammatory response in the rheumatoid joint could be largely cell mediated. However, in many instances negative serologies in patients with RA are falsely negative. Three different mechanisms may ac-count for false-negative results in the RA test:
1. Anti-Ig antibodies of isotypes other than IgM, less efficient than IgM RF in caus-ing agglutination (particularly in tests using red cells) and therefore more likely to be overlooked, may be present.
2. The reaction between IgG RF and endogenous IgG results in the formation of soluble immune complexes that, if the affinity of the reaction is relatively high, will remain associated when the RF test is performed. Under these conditions, the RF-binding sites are blocked, unable to react with the IgG coating indicator red cells or latex particles
3. RF may be present in synovial fluid but not in peripheral blood.
In clinical practice, it is very seldom necessary to investigate these possibilities, since a pos-itive test is not necessary for the diagnosis.