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

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.     

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

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 β-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 β-estradiol (20 ng/ml) or relaxin plus β-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 β-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 β-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 β-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.     

Synthesis, structural and spectroscopic characterization and biomimetic properties of new copper, manganese, zinc complexes: Identification of possible superoxide-dismutase mimics bearing hydroxyl radical generating/scavenging abilities

A series of Cu(II), Zn(II) and Mn(II) coordination compounds has been synthesized by reaction of the corresponding metal salts and pyrazolyl-based ligands, i.e. the neutral 1-(2-(4-((2,2,2-tri(1H-pyrazol-1-yl)ethoxy)methyl)benzyloxy)-1,1-di(1H-pyrazol-1-yl)ethyl)-1H-pyrazole {p-C₆H₄[CH₂OCH₂C(pz)₃]₂, (L¹), and the anionic hydridotris(3-phenyl-5-methylpyrazolyl)borate (L²)⁻, bis(pyrazolyl)acetate (L³) and bis(3,5-dimethylpyrazolyl)acetate (L⁴)⁻: the species [L¹(CuCl₂)₂] (1), [L¹(Cu(OAc)₂)₂] (2), [L¹(Zn(OAc)₂)₂] (3), [(CuCl(L²)(Hpz^Ph,Me)] (4), [Mn(L³)₂]·2H₂O, (5), [ZnCl(L³)(imH)]·MeOH [CuCl(L⁴)(imH)]·2H₂O (7) have been obtained (Hpz^Ph,Me = 3-phenyl-5-methylpyrazole, imH = imidazole). Complexes 1 and 4 have been structurally characterized, also using less conventional powder diffraction methods. The superoxide scavenging activity has been characterized by indirect assays including EPR analysis. All complexes exhibit superoxide scavenging activity with IC₅₀ in the µM range and they protect against the oxidative action of peroxynitrite in different ways. 1, 4 and 7 exert both an anti- and pro-oxidant effect depending on their concentration as evaluated by EPR and fluorescence methods. The pro-oxidative effects are absent in Zn(II) and Mn(II) complexes.     

Production of cellulases and hemicellulases by an anaerobic mixed culture from lignocellulosic biomass

A comparison of different habitats, biogas plant, rumen fluid and sewage sludge, for cellulolytic organisms indicated sewage studge was the best source. Enrichment cultura gave a mixed culture which exhibited CMCase activity as well as extracellular Avicelase, xylanase, -glucosidase, -xylosidase activities and cell-bound -glucosidase, and -xylosidase production in a synthetic medium with eleven different cellulosic and lignocellulosic substrates. The activity of extracellular -glucosidase and -xylosidase production was significantly higher than endogenous activities. Hemicellulases were induced better than cellulases. The anzyme system was stable under aerobic conditions. Of the different lignocellulosic substrates, kallar grass was the best inducer of extracellular enzymes.

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Résumé La comparaison de différents habitats: digesteur méthanique, fluide du rumen ou boue de station d'épuration, pour leur contenu en organismes cellulolytiques, indiquent que la boue de station d'épuration est la meilleure source. Une culture par enrichissement a produit une culture mixte qui a exhibé aussi bien une activité CMCase que des activitiés extracellulaires avicelasique, xylanasique, -glucosidasique et -xylosidasique et qu'une production de -glucosidase et de -xylosidase liées à la cellule, dans un milieu synthétique et pour onze substrats cellulosiques et lignocellulosiques différents. L'activité de la -glucosidase extracellulaire et la production de -xylosidase sont significativement plus élevées que les activitiés endogènes. Les hemicellulases sont mieux induites que les cellulases. Le système enzymatique est stable dans des conditions aérobies. Parmi les divers substrats lignocellulosiques, l'herbe Kallar est le meilleur inducteur d'enzymes extracellufaires.