Estrogen Promotes Mandibular Condylar Fibrocartilage Chondrogenesis and Inhibits Degeneration via Estrogen Receptor Alpha in Female Mice

Temporomandibular joint degenerative disease (TMJ-DD) is a chronic form of TMJ disorder that specifically afflicts people over the age of 40 and targets women at a higher rate than men. Prevalence of TMJ-DD in this population suggests that estrogen loss plays a role in the disease pathogenesis. Thus, the goal of the present study was to determine the role of estrogen on chondrogenesis and homeostasis via estrogen receptor alpha (ERα) during growth and maturity of the joint. Young and mature WT and ERαKO female mice were subjected to ovariectomy procedures and then given placebo or estradiol treatment. The effect of estrogen via ERα on fibrocartilage morphology, matrix production, and protease activity was assessed. In the young mice, estrogen via ERα promoted mandibular condylar fibrocartilage chondrogenesis partly by inhibiting the canonical Wnt signaling pathway through upregulation of sclerostin (Sost). In the mature mice, protease activity was partly inhibited with estrogen treatment via the upregulation and activity of protease inhibitor 15 (Pi15) and alpha-2- macroglobulin (A2m). The results from this work provide a mechanistic understanding of estradiol on TMJ growth and homeostasis and can be utilized for development of therapeutic targets to promote regeneration and inhibit degeneration of the mandibular condylar fibrocartilage.National Institute of Dental & Craniofacial Research of the National Institutes of Health under Award Numbers R56DE020097 (SW) and F32DE026366 (JR

Estrogen/Estrogen Receptor Alpha Signaling in Mouse Posterofrontal Cranial Suture Fusion

BACKGROUND: While premature suture fusion, or craniosynostosis, is a relatively common condition, the cause is often unknown. Estrogens are associated with growth plate fusion of endochondral bones. In the following study, we explore the previously unknown significance of estrogen/estrogen receptor signaling in cranial suture biology. METHODOLOGY/PRINCIPAL FINDINGS: Firstly, estrogen receptor (ER) expression was examined in physiologically fusing (posterofrontal) and patent (sagittal) mouse cranial sutures by quantitative RT-PCR. Next, the cranial suture phenotype of ER alpha and ER beta knockout (alphaERKO, betaERKO) mice was studied. Subsequently, mouse suture-derived mesenchymal cells (SMCs) were isolated; the effects of 17-beta estradiol or the estrogen antagonist Fulvestrant on gene expression, osteogenic and chondrogenic differentiation were examined in vitro. Finally, in vivo experiments were performed in which Fulvestrant was administered subcutaneously to the mouse calvaria. Results showed that increased ERalpha but not ERbeta transcript abundance temporally coincided with posterofrontal suture fusion. The alphaERKO but not betaERKO mouse exhibited delayed posterofrontal suture fusion. In vitro, addition of 17-beta estradiol enhanced both osteogenic and chondrogenic differentiation in suture-derived mesenchymal cells, effects reversible by Fulvestrant. Finally, in vivo application of Fulvestrant significantly diminished calvarial osteogenesis, inhibiting suture fusion. CONCLUSIONS/SIGNIFICANCE: Estrogen signaling through ERalpha but not ERbeta is associated with and necessary for normal mouse posterofrontal suture fusion. In vitro studies suggest that estrogens may play a role in osteoblast and/or chondrocyte differentiation within the cranial suture complex

Up-regulation of type II collagen gene expression by 17beta-estradiol in differentiated and dedifferentiated chondrocytes.

PosterInternational audienc

Décryptage des mécanismes moléculaires des chondrocytes dans la greffe autologue de chondrocytes pour la réparation du cartilage

International audienceSince the emergence in the 1990s of the autologous chondrocytes transplantation (ACT) in the treatment of cartilage defects, the technique, corresponding initially to implantation of chondrocytes, previously isolated and amplified in vitro, under a periosteal membrane, has greatly evolved. Indeed, the first generations of ACT showed their limits, with in particular the dedifferentiation of chondrocytes during the monolayer culture, inducing the synthesis of fibroblastic collagens, notably type I collagen to the detriment of type II collagen. Beyond the clinical aspect with its encouraging results, new biological substitutes must be tested to obtain a hyaline neocartilage. Therefore, the use of differentiated chondrocytes phenotypically stabilized is essential for the success of ACT at medium and long-term. That is why researchers try now to develop more reliable culture techniques, using among others, new types of biomaterials and molecules known for their chondrogenic activity, giving rise to the 4th generation of ACT. Other sources of cells, being able to follow chondrogenesis program, are also studied. The success of the cartilage regenerative medicine is based on the phenotypic status of the chondrocyte and on one of its essential component of the cartilage, type II collagen, the expression of which should be supported without induction of type I collagen. The knowledge accumulated by the scientific community and the experience of the clinicians will certainly allow to relief this technological challenge, which influence besides, the validation of such biological substitutes by the sanitary authorities.Depuis l’émergence dans les années 1990 de la transplantation de chondrocytes autologues (TCA) dans le traitement des lésions du cartilage, la technique, correspondant initialement à l’implantation de chondrocytes, préalablement isolés et amplifiés in vitro, sous une membrane périostée, a beaucoup évolué. En effet, les premières générations de TCA ont montré leurs limites, avec en particulier la dédifférenciation des chondrocytes au cours de la culture en monocouche, qui se manifeste par la synthèse de collagènes de type fibroblastique, notamment du collagène de type I au détriment du collagène de type II, le collagène phénotypique du cartilage hyalin. Au delà de l’aspect clinique avec ses résultats encourageants, de nouveaux substituts biologiques doivent être éprouvés afin d’obtenir, in vivo, du néocartilage de type hyalin. Ainsi, l’utilisation de chondrocytes différenciés stabilisés phénotypiquement est indissociable de la réussite de la TCA à moyen et long terme. C’est pourquoi se développent actuellement des techniques de culture plus complexes, utilisant entre autre de nouveaux types de biomatériaux et des molécules connues pour leur activité chondrogénique, laissant entrevoir une quatrième génération de TCA. D’autres sources de cellules, pouvant s’engager dans le lignage chondrogénique ou chondro-précurseurs, telles les cellules souches mésenchymateuses adultes, sont également à l’étude. La réussite de la médecine régénérative du cartilage repose avant tout sur le statut phénotypique du chondrocyte et sur des composants essentiels et fonctionnels du cartilage, le collagène de type II, les isotypes IX et XI, et l’agrécanne, dont les expressions doivent être soutenues sans avoir en contrepartie des collagènes qualifiés de « fibroblastiques », notamment le collagène de type I. Les connaissances accumulées par la communauté scientifique et l’expérience des cliniciens permettront certainement de relever ce défi technologique, qui conditionne par ailleurs, la validation de tels substituts biologiques par les autorités sanitaires

17Beta-oestradiol up-regulates the expression of a functional UDP-glucose dehydrogenase in articular chondrocytes: comparison with effects of cytokines and growth factors.

International audienceOBJECTIVES: To investigate the mechanisms by which cytokines and 17beta-oestradiol (17beta-E2) modulate gene expression and activity of uridine diphosphoglucose dehydrogenase (UGDH), a key enzyme of GAG synthesis in articular chondrocytes. METHODS: Rabbit articular chondrocytes (RAC) from 3-week-old animals were incubated for 24 h with TGF-beta, insulin like growth factor-I (IGF-I), IL-1beta, IL-6 and 17beta-E2. GAG synthesis was measured by [35S]-sulphate labelling and the expression of the UGDH gene was estimated by both real-time polymerase chain reaction and western blotting, whereas the enzyme activity was assayed by a spectrophotometric procedure. In addition, the transcriptional activity of several UGDH gene promoter constructs was determined in RAC transiently transfected with wild-type or deleted human oestrogen receptor-alpha gene (hER alpha66 or hER alpha46, respectively). RESULTS: 17Beta-E2 and its receptor hER alpha66 enhanced GAG neosynthesis in rabbit articular chondrocytes, as did TGF-beta1 whereas IL-1beta decreased this synthesis. 17Beta-E2 was found to exert positive regulatory effects at mRNA, protein and UGDH activity levels. In addition, the receptor hER alpha66, but not hER alpha46, increased the transcriptional activity of the UGDH gene. In contrast, no clear correlation between transcription, translation and activity of the UGDH was found under the effects of the cytokines studied. However, TGF-beta enhanced the enzyme activity, whereas IL-1beta, IL-6 and IGF-I were without significant effect. CONCLUSIONS: 17Beta-E2 enhanced GAG synthesis in chondrocytes via up-regulation of the UGDH gene expression and enzyme activity. These data provide insights into the molecular mechanisms involved in the regulation of the UGDH gene and offer new approaches to investigate its potential alteration in joint diseases