Free radical damage in facsimile synovium: correlation with adhesion formation in osteoarthritic TMJs.

PURPOSE: The purpose of this study was to use the rat air pouch model of facsimile synovium to evaluate oxidative stress as a primary mechanism in the pathogenesis of degenerative temporomandibular joint (TMJ) disease. MATERIALS AND METHODS: Forty-nine Sprague-Dawley adult female rats were used to generate the standard rat air pouch model of facsimile synovium. This was accomplished by daily air injections (20 cc) subdermally through the dorsal skin. Hydrogen peroxide and ferrous iron (components of the Fenton reaction which generate free radicals) were introduced into the pouches of the 4-, 7-, and 14-day groups to generate oxidative stress. Control rats were injected with phosphate-buffered solution (PBS), pH 7.4. Either N-acetylcysteine (NAC), a powerful free radical scavenger, or ibuprofen were simultaneously injected with the Fenton reagents into the pouches of the 14-day treatment groups to modulate free radical-mediated protein damage to the synovium. Animals were euthanized at appropriate experimental intervals and biopsies obtained from specimens to analyze: (1) proteins' amino acid modification (carbonyl group formation), (2) protein hydrophobicity, (3) detection of low molecular weight protein degradation products, and (4) histological and gross anatomical observations. RESULTS: Free radicals introduced into the rat air pouch interacted with synovial tissues causing oxidation and breakdown of proteins. Clinical evidence of adhesion formation consistent with features found in osteoarthritis of the TMJ developed. The groups subjected to oxidative stress experienced statistically significant (p < 0.05) increases in carbonyl formation, carbonyls/protein, and low molecular weight protein fragments. These groups also showed significant (p < 0.05) hydrophobicity changes consistent with free radical attack. Control synovial tissues were statistically undamaged. The 14-day NAC and ibuprofen treatment groups experienced statistically significant (p < 0.05) decreases in total carbonyl formation, carbonyls/protein, and hydrophobicity. Histological and gross observations in free radical damaged synovium exhibited features consistent with known arthoscopic and arthrocentesis findings in diseased TMJs. CONCLUSIONS: This study suggests that the rat air pouch model of facsimile synovium develops clinical evidence of adhesions and biochemical signs of protein modification when subjected to free radical attack. NAC and ibuprofen prevented carbonyl formation as well as hydrophobicity changes indicative of oxidative stress damage in facsimile synovium. These findings are consistent with features of degenerative human TMJ disease. Future direction may be taken from this study to postulate new analysis techniques and treatment modalities for patients with degenerative TMJ disease.

Crosslinking of fibrinogen and fibronectin by free radicals: a possible initial step in adhesion formation in osteoarthritis of the temporomandibular joint.

PURPOSE: Adhesion formation in osteoarthritis (OA) of the temporomandibular joint (TMJ) typically results in a sustained limitation of joint movement. We propose the hypothesis that free-radical-mediated crosslinking of proteins underlies this adhesion formation in affected joints. Free radicals may cause oxidative modification of proteins, creating an opportunity for the formation of intramolecular and intermolecular crosslinks via covalent bonds. This may stabilize protein aggregates, rendering them more resistant to degradation. In this study, the free-radical-mediated crosslinking of model proteins (fibrinogen and fibronectin) was investigated to test our hypothesis that free radicals contribute to adhesion formation via this mechanism in OA of the TMJ. MATERIALS AND METHODS: Physiological clot formation of fibrinogen by thrombin and free-radical-induced crosslinking of fibrinogen and of fibronectin were analyzed using spectrophotometric turbidity measurements, light-scattering techniques, polyacrylamide gel electrophoresis (PAGE), and rotary shadowing. RESULTS: Fibrinogen was shown to aggregate after free radical treatment, as detected using turbidity measurements and light-scattering techniques. Using PAGE, fibrinogen as well as fibronectin was shown to degrade under low oxidative stress. Under high oxidative stress, however, fragments from both proteins were found to be covalently crosslinked, resulting in high-molecular-weight protein aggregates. The aggregation was shown to be at random with rotary shadowing. CONCLUSION: The study shows that high oxidative stress contributes to the formation of crosslinked proteins that may serve as an initial scaffolding for the development of adhesions frequently seen in OA of the TMJ.