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Relaxin's induction of metalloproteinases is associated with the loss of collagen and glycosaminoglycans in synovial joint fibrocartilaginous explants.

Naqvi T, Duong TT, Hashem G, Shiga M, Zhang Q, Kapila S - Arthritis Res. Ther. (2004)

Bottom Line: Here we determined the effects of relaxin with or without beta-estradiol on the modulation of MMPs in joint fibrocartilaginous explants, and assessed the contribution of these proteinases to the loss of collagen and glycosaminoglycan (GAG) in this tissue.None of the hormone treatments altered the synthesis of GAGs, suggesting that the loss of this matrix molecule probably resulted from increased matrix degradation.These findings show for the first time that relaxin has degradative effects on non-reproductive synovial joint fibrocartilaginous tissue and provide evidence for a link between relaxin, MMPs, and matrix degradation.

View Article: PubMed Central - HTML - PubMed

Affiliation: University of California, San Francisco, USA. sheema@hotmail.com

ABSTRACT
Diseases of specific fibrocartilaginous joints are especially common in women of reproductive age, suggesting that female hormones contribute to their etiopathogenesis. Previously, we showed that relaxin dose-dependently induces matrix metalloproteinase (MMP) expression in isolated joint fibrocartilaginous cells. Here we determined the effects of relaxin with or without beta-estradiol on the modulation of MMPs in joint fibrocartilaginous explants, and assessed the contribution of these proteinases to the loss of collagen and glycosaminoglycan (GAG) in this tissue. Fibrocartilaginous discs from temporomandibular joints of female rabbits were cultured in medium alone or in medium containing relaxin (0.1 ng/ml) or beta-estradiol (20 ng/ml) or relaxin plus beta-estradiol. Additional experiments were done in the presence of the MMP inhibitor GM6001 or its control analog. After 48 hours of culture, the medium was assayed for MMPs and the discs were analyzed for collagen and GAG concentrations. Relaxin and beta-estradiol plus relaxin induced the MMPs collagenase-1 and stromelysin-1 in fibrocartilaginous explants--a finding similar to that which we observed in pubic symphysis fibrocartilage, but not in articular cartilage explants. The induction of these proteinases by relaxin or beta-estradiol plus relaxin was accompanied by a loss of GAGs and collagen in joint fibrocartilage. None of the hormone treatments altered the synthesis of GAGs, suggesting that the loss of this matrix molecule probably resulted from increased matrix degradation. Indeed, fibrocartilaginous explants cultured in the presence of GM6001 showed an inhibition of relaxin-induced and beta-estradiol plus relaxin-induced collagenase and stromelysin activities to control baseline levels that were accompanied by the maintenance of collagen or GAG content at control levels. These findings show for the first time that relaxin has degradative effects on non-reproductive synovial joint fibrocartilaginous tissue and provide evidence for a link between relaxin, MMPs, and matrix degradation.

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Relaxin-induced collagenase activity contributes to loss of disc collagen. Conditioned medium from disc incubated with control medium (Ct), β-estradiol (Es), relaxin (R), or β-estradiol plus relaxin (Es+R) in the presence of the matrix metalloproteinase inhibitor GM6001 or its control analog was subjected to fluorescein isothiocyanate-labelled collagen degradation assay. The collagenase activity (relative fluorescence units [RFU]/ml) was standardized by the dry weight of the tissue (mg), and fold changes (means ± SD) were plotted (a, b). Disc digests from these experiments were assayed for collagen with the Sircol assay, and the results were standardized to tissue dry weight (mg). Fold changes in collagen concentration (means ± SD) were calculated and plotted (c, d). The untreated control (Ct) discs used in all experiments were exposed to control analog only. ** P < 0.01, *** P < 0.0001 by Fisher's test.
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Figure 5: Relaxin-induced collagenase activity contributes to loss of disc collagen. Conditioned medium from disc incubated with control medium (Ct), β-estradiol (Es), relaxin (R), or β-estradiol plus relaxin (Es+R) in the presence of the matrix metalloproteinase inhibitor GM6001 or its control analog was subjected to fluorescein isothiocyanate-labelled collagen degradation assay. The collagenase activity (relative fluorescence units [RFU]/ml) was standardized by the dry weight of the tissue (mg), and fold changes (means ± SD) were plotted (a, b). Disc digests from these experiments were assayed for collagen with the Sircol assay, and the results were standardized to tissue dry weight (mg). Fold changes in collagen concentration (means ± SD) were calculated and plotted (c, d). The untreated control (Ct) discs used in all experiments were exposed to control analog only. ** P < 0.01, *** P < 0.0001 by Fisher's test.

Mentions: All three hormone treatments increased the expression of procollagenase-1 in the presence of GM6001 or its control analog similarly to that shown in Fig. 1a,1b,1c. However, as shown by FITC-collagen degradation assays, collagenase activity was significantly increased only by relaxin or β-estradiol plus relaxin in the presence of the control analog (Fig. 5a). In discs incubated with GM6001, hormone-induced collagenase activity was inhibited to control levels (Fig. 5b). Conversely, Sircol assays showed the collagen content was significantly decreased (P < 0.0001, ANOVA) only in the presence of the control analog and only by relaxin (40% of control and β-estradiol alone; P < 0.0001, Fisher's test) or β-estradiol plus relaxin (60% versus control and β-estradiol alone; P < 0.0001) (Fig. 5c). In the presence of GM6001, hormone treatments did not affect collagen content (Fig. 5d).


Relaxin's induction of metalloproteinases is associated with the loss of collagen and glycosaminoglycans in synovial joint fibrocartilaginous explants.

Naqvi T, Duong TT, Hashem G, Shiga M, Zhang Q, Kapila S - Arthritis Res. Ther. (2004)

Relaxin-induced collagenase activity contributes to loss of disc collagen. Conditioned medium from disc incubated with control medium (Ct), β-estradiol (Es), relaxin (R), or β-estradiol plus relaxin (Es+R) in the presence of the matrix metalloproteinase inhibitor GM6001 or its control analog was subjected to fluorescein isothiocyanate-labelled collagen degradation assay. The collagenase activity (relative fluorescence units [RFU]/ml) was standardized by the dry weight of the tissue (mg), and fold changes (means ± SD) were plotted (a, b). Disc digests from these experiments were assayed for collagen with the Sircol assay, and the results were standardized to tissue dry weight (mg). Fold changes in collagen concentration (means ± SD) were calculated and plotted (c, d). The untreated control (Ct) discs used in all experiments were exposed to control analog only. ** P < 0.01, *** P < 0.0001 by Fisher's test.
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Related In: Results  -  Collection

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Figure 5: Relaxin-induced collagenase activity contributes to loss of disc collagen. Conditioned medium from disc incubated with control medium (Ct), β-estradiol (Es), relaxin (R), or β-estradiol plus relaxin (Es+R) in the presence of the matrix metalloproteinase inhibitor GM6001 or its control analog was subjected to fluorescein isothiocyanate-labelled collagen degradation assay. The collagenase activity (relative fluorescence units [RFU]/ml) was standardized by the dry weight of the tissue (mg), and fold changes (means ± SD) were plotted (a, b). Disc digests from these experiments were assayed for collagen with the Sircol assay, and the results were standardized to tissue dry weight (mg). Fold changes in collagen concentration (means ± SD) were calculated and plotted (c, d). The untreated control (Ct) discs used in all experiments were exposed to control analog only. ** P < 0.01, *** P < 0.0001 by Fisher's test.
Mentions: All three hormone treatments increased the expression of procollagenase-1 in the presence of GM6001 or its control analog similarly to that shown in Fig. 1a,1b,1c. However, as shown by FITC-collagen degradation assays, collagenase activity was significantly increased only by relaxin or β-estradiol plus relaxin in the presence of the control analog (Fig. 5a). In discs incubated with GM6001, hormone-induced collagenase activity was inhibited to control levels (Fig. 5b). Conversely, Sircol assays showed the collagen content was significantly decreased (P < 0.0001, ANOVA) only in the presence of the control analog and only by relaxin (40% of control and β-estradiol alone; P < 0.0001, Fisher's test) or β-estradiol plus relaxin (60% versus control and β-estradiol alone; P < 0.0001) (Fig. 5c). In the presence of GM6001, hormone treatments did not affect collagen content (Fig. 5d).

Bottom Line: Here we determined the effects of relaxin with or without beta-estradiol on the modulation of MMPs in joint fibrocartilaginous explants, and assessed the contribution of these proteinases to the loss of collagen and glycosaminoglycan (GAG) in this tissue.None of the hormone treatments altered the synthesis of GAGs, suggesting that the loss of this matrix molecule probably resulted from increased matrix degradation.These findings show for the first time that relaxin has degradative effects on non-reproductive synovial joint fibrocartilaginous tissue and provide evidence for a link between relaxin, MMPs, and matrix degradation.

View Article: PubMed Central - HTML - PubMed

Affiliation: University of California, San Francisco, USA. sheema@hotmail.com

ABSTRACT
Diseases of specific fibrocartilaginous joints are especially common in women of reproductive age, suggesting that female hormones contribute to their etiopathogenesis. Previously, we showed that relaxin dose-dependently induces matrix metalloproteinase (MMP) expression in isolated joint fibrocartilaginous cells. Here we determined the effects of relaxin with or without beta-estradiol on the modulation of MMPs in joint fibrocartilaginous explants, and assessed the contribution of these proteinases to the loss of collagen and glycosaminoglycan (GAG) in this tissue. Fibrocartilaginous discs from temporomandibular joints of female rabbits were cultured in medium alone or in medium containing relaxin (0.1 ng/ml) or beta-estradiol (20 ng/ml) or relaxin plus beta-estradiol. Additional experiments were done in the presence of the MMP inhibitor GM6001 or its control analog. After 48 hours of culture, the medium was assayed for MMPs and the discs were analyzed for collagen and GAG concentrations. Relaxin and beta-estradiol plus relaxin induced the MMPs collagenase-1 and stromelysin-1 in fibrocartilaginous explants--a finding similar to that which we observed in pubic symphysis fibrocartilage, but not in articular cartilage explants. The induction of these proteinases by relaxin or beta-estradiol plus relaxin was accompanied by a loss of GAGs and collagen in joint fibrocartilage. None of the hormone treatments altered the synthesis of GAGs, suggesting that the loss of this matrix molecule probably resulted from increased matrix degradation. Indeed, fibrocartilaginous explants cultured in the presence of GM6001 showed an inhibition of relaxin-induced and beta-estradiol plus relaxin-induced collagenase and stromelysin activities to control baseline levels that were accompanied by the maintenance of collagen or GAG content at control levels. These findings show for the first time that relaxin has degradative effects on non-reproductive synovial joint fibrocartilaginous tissue and provide evidence for a link between relaxin, MMPs, and matrix degradation.

Show MeSH
Related in: MedlinePlus