Abstract
Background: Rheumatoid arthritis (RA) is a chronic inflammatory joint disease hallmarked by irreversible damage of cartilage and bone. Matrix metalloproteinases (MMPs) involved in connective tissue remodeling play an important role in this process. Numerous MMPs have been examined in humans and animals, but their functions are still not fully understood. Therefore, we investigated the role of MMPs in the K/BxN serum-transfer model of RA with the broad-spectrum MMP inhibitor subantimicrobial dose doxycycline (SDD) using complex in vivo and in vitro methodolgy.Methods: Chronic arthritis was induced by repetitive i.p. injections of K/BxN serum in C57BL/6J mice. SDD was administered daily in acidified drinking water (0.5 mg/mL, 80 mg/kg) during the 30 days experimental period. Mechanonociceptive threshold of the paw was evaluated by aesthesiometry, grasping ability by grid test, arthritis severity by scoring, neutrophil myeloperoxidase activity by luminescence, vascular hyperpermeability and MMP activity by fluorescence in vivo imaging and the latter also by gelatin zymography, bone structure by micro-computed tomography (micro-CT). Plasma concentrations of doxycycline were determined by liquid chromatography-mass spectrometry analysis.Results: K/BxN serum induced significant inflammatory signs, mechanical hyperalgesia, joint function impairment, increased myeloperoxidase activity and vascular hyperpermeability. Significant increase of MMP activity was also observed both in vivo and ex vivo with elevation of the 57–60, 75, and 92 kDa gelatinolytic isoforms in the arthritic ankle joints, but neither MMP activity nor any above described functional parameters were influenced by SDD. Most importantly, SDD significantly reduced bone mineral density in the distal tibia and enhanced the Euler number in the ankle. Arthritis-induced microarchitectural alterations demonstrating increased irregularity and cancellous bone remodeling, such as increased Euler number was significantly elevated by SDD in both regions.Conclusion: We showed increase of various MMP activities in the joints by in vivo fluorescence imaging together with ex vivo zymography, and investigated their functional significance using the broad-spectrum MMP inhibitor SDD in the translational RA model. This is the first demonstration that SDD worsens arthritis-induced bone microarchitectural alterations, but it appears to be independent of MMP inhibition.
Highlights
Rheumatoid arthritis (RA) is a progressive, chronic inflammatory joint disease leading to irreversible articular cartilage and bone destruction
In the present study we showed the increase of various matrix metalloproteinase (MMP) activities in the joints by in vivo fluorescence imaging together with ex vivo zymography, as well as demonstrated for the first time that subantimicrobial dose doxycycline (SDD) worsens arthritis-induced bone microarchitectural alterations most probably independently of MMP inhibition
Arthritis induced a significant increase of fluorescent signal indicating in vivo MMP activity in the ankle joints on day 4, but significant difference did not occur between the tap waterand SDD-treated arthritic groups (Figures 2A,B)
Summary
Rheumatoid arthritis (RA) is a progressive, chronic inflammatory joint disease leading to irreversible articular cartilage and bone destruction. It is one of the most common musculoskeletal disorder causing physical disability with a worldwide prevalence of approximately 1% (Gibofsky, 2012). Further research is needed to precisely explore its pathophysiological mechanisms, identify crucial mediators, and find new potential drug targets. These may include matrix metalloproteinases (MMPs), which are important players of joint damage in arthritic conditions, most importantly in RA (Rose and Kooyman, 2016). We investigated the role of MMPs in the K/BxN serum-transfer model of RA with the broad-spectrum MMP inhibitor subantimicrobial dose doxycycline (SDD) using complex in vivo and in vitro methodolgy
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