Iron-dependent generation of free radicals: plausible mechanisms in the progressive deterioration of the temporomandibular joint.
OBJECTIVE
The purposes of this study were 1) to determine whether iron concentrations detected in temporomandibular joint (TMJ) lavage fluid samples obtained from symptomatic patients are sufficient to catalyze the degradation of specific extracellular matrix (ECM) molecules in vitro, and 2) to provide evidence of oxidative stress in symptomatic TMJs by the detection of protein carbonyls in lavage fluids.


METHODS
Iron concentrations in TMJ lavage samples (19 joints in 14 patients) were determined colorimetrically, and the ability of the sample to produce free radicals in the presence of hydrogen peroxide was determined with the chromogen 2,2'-azinobis (3-ethylbenzothizoline-6-sulfonic acid), diammonium salt (ABTS). The presence of oxidized proteins was measured fluorimetrically using Bodipy FL hydrazide (Molecular Probes, Eugene, OR). Degradation of fibronectin was visualized by Western blot. Relative susceptibilities of fibronectin and collagen I to free radical cleavage were measured with the Fenton reaction.


RESULTS
Redox-active iron concentration in lavage samples was found to be as high as 3.66 micromol/L. A 70-kd protein band, presumed to be albumin, was found to contain higher levels of carbonyls than peripheral serum albumin, which correlated with a greater degree of oxidative damage. Fibronectin was found to be more susceptible than collagen I to free radical degradation, and fragments of the former were found in the lavage. The TMJ lavage fluid was capable of producing free radicals in the presence of peroxide.


CONCLUSIONS
Circumstantial evidence is provided that the presence of modified and cleaved proteins isolated from lavage of symptomatic TMJs may have been subjected to oxidative stress.
Zardeneta
G
G
Department of Oral and Maxillofacial Surgery, The University of Texas Health Science Center, San Antonio 78284-7903, USA. Zardeneta@uthscsa.edu
Milam
S B
SB
Schmitz
J P
JP
D015153:Blotting, Western; D003094:Collagen / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D005353:Fibronectins / Q000378:metabolism; D005609:Free Radicals / Q000378:metabolism; D006454:Hemoglobins / Q000378:metabolism; D006801:Humans; D006861:Hydrogen Peroxide; D017665:Hydroxyl Radical / Q000378:metabolism; D007501:Iron / Q000032:analysis / Q000378:metabolism*; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D019152:Paracentesis; D011489:Protein Denaturation; D013582:Synovial Fluid / Q000737:chemistry; D013704:Temporomandibular Joint / Q000378:metabolism*; D013705:Temporomandibular Joint Disorders / Q000209:etiology* / Q000378:metabolism*; D007507:Therapeutic Irrigation
C045076:Fenton's reagent; D005353:Fibronectins; D005609:Free Radicals; D006454:Hemoglobins; D017665:Hydroxyl Radical; D003094:Collagen; D006861:Hydrogen Peroxide; D007501:Iron