Markers in rheumatoid arthritis

Proteinases play an important role in the pathogenesis of joint destruction, and matrix metalloproteinases (MMP) are believed to play a crucial role as they have the ability to degrade various compounds of the extra-cellular matrix of the joints, including collagen. The work described in this presentation focuses on the question of which MMPs and in what forms are they present in synovial fluid (SF) and the circulation of patients with joint pathology. Also, the applicability of the MMP measurements as markers of disease activity or joint damage progression is investigated. High levels of proMMP-3, proMMP-8 and proMMP-9 were found in SF and in the circulation of the rheumatoid arthritis (RA) patients as compared with the control population or osteoarthritis patients. Also MMP activity in MMP/α₂-macroglobulin (α₂M) complexes was shown to be higher in RA patients. Moreover, it was shown that proMMP-8 and proMMP-9 (collagenase 2 and gelatinase B) levels are related to the levels of collagen degradation products (hydroxyproline) in SF. Although there is only a trend towards correlation between proMMP-8 and hydroxyproline in SF, levels of proMMP-3 were not correlated with hydroxyproline levels. Comparison between systemic levels of inflammation (C-reactive protein) and MMP levels showed the opposite situation. Only proMMP-3 levels were correlated with C-reactive protein, and not proMMP-8 or proMMP-9 levels. These observations suggested that the aforementioned enzymes can be seen as candidate markers for the pathophysiological processes of joint inflammation and degradation. Moreover, MMP activity in MMP/α₂M complexes was considered a possible marker of the proteolytic system since increased levels of MMP activity were found in SF and in the serum of RA patients as compared with osteoarthritis patients. To investigate the predictive role of MMP levels in the circulation, we measured MMPs in patients in an Early Arthritis Clinic. The results show that proMMP-3 levels at the onset of the disease were predictive of cartilage loss at the end of the second year of the follow-up period. Moreover, proMMP-3 predicted joint damage progression independently from other known parameters such as sheared epitope or rheumatoid factor. Collagenase-2 (MMP-8) was not predictive of joint damage, and neither were high levels of proMMP-9. Based on these findings, we conclude that whereas MMP-8 and MMP-9 levels reflect the current status of the protheolytic system, MMP-3 levels at the onset of RA show the potential of the system to be destructive. It is extensively documented that protein levels of tissue inhibitor of metalloproteinase (TIMP)-1 in SF and in the circulation of RA patients are insufficient to counteract the increased production of MMPs. However, little is known about the activation of proMMPs in vivo. It was previously indicated that activated, but not TIMP-inhibited, MMPs can be neutralized by α₂M. We hypotheized that in pathological situations involving joint inflammation and destruction the high production of proMMPs leads to increased levels of activated MMPs, and because of the MMP/TIMP imbalance high levels of MMP/α₂M complexes may be found in these conditions. Thus, high levels of MMP/α₂M complexes would support the idea of a MMP/TIMP imbalance. To test this hypothesis, a method to measure MMP activity in MMP/α₂M complexes using small fluorogenic substrates, such as TNO211-F, was developed. Indeed, high levels of MMP in complexes with α₂M are found in SF and in the circulation of patients with joint pathology. The presence of increased levels of α₂M/MMP complexes shows that TIMP levels are insufficient to inhibit all activated MMPs; thus, the MMP/TIMP imbalance theory is supported by these findings. In conclusion, the results to be discussed show that high proMMP production leads to increased levels of activated MMP in SF and the systemic circulation of RA patients. Upon activation MMPs form complexes with α₂M in SF and in the systemic circulation of RA patients, which is at least partly responsible for neutralization of MMPs. MMP-1, MMP-3, MMP-8 and MMP-9 are likely to be involved the pathogenesis of RA as shown by the MMP profile in SF and the serum of RA patients. proMMP-3 levels at the onset of RA are predictive of the progression of joint damage. The MMP/TIMP imbalance is present in various joint pathologies. MMP/α₂M complexes are reduced in the systemic circulation of RA patients upon treatment with disease-modifying anti-rheumatic drugs.