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Implication des Bone Morphogenetic Proteins (BMPs) hypophysaires dans la régulation de la synthèse et de la libération de l'hormone folliculo-stimulante (FSH) ? / Involvement of pituitary bone morphogenetic proteins (BMPs) in the regulation of follicle stimulating hormone (FSH) synthesis and releaseSallon, Céline 10 November 2010 (has links)
Chez la brebis, les BMPs inhibent la synthèse et la libération de FSH. La détection des ARNm de certaines BMPs dans l’hypophyse et de leurs récepteurs sur les cellules gonadotropes suggère une action paracrine/autocrine des BMPs sur la production de FSH. L’objectif de cette thèse a visé à déterminer l’importance des BMPs hypophysaires, en particulier BMP-4, dans la régulation de la sécrétion de FSH chez la brebis. Nous montrons que l’expression des ARNm du système BMP-4,analysée par RT-PCR en temps réel, ne varie pas au cours du cycle oestrien ou in vitro quelque soit le temps d’incubation (6h-48h) et le traitement (GnRH, oestradiol, activine) des cellules hypophysaires.Afin de déterminer si les cellules hypophysaires ovines sécrètent des BMPs, des milieux conditionnés(MC) hypophysaires ont été soumis à un test d’activité biologique reposant sur des cellules embryonnaires transfectées avec un élément de réponse BMPs couplé au gène rapporteur luciférase.Aucun effet des MC n’a été observé sur l’activité luciférase comparé au milieu non conditionnésuggérant l’absence ou la très faible activité BMP des MC, et cela quelque soit le temps d’incubation(6h-48h) et le traitement (GnRH, oestradiol, activine) des cellules hypophysaires. Toutefois, nous détectons une activité inhibitrice de BMPs qui est augmentée par la GnRH et l’oestradiol suggérant l’implication d’inhibiteurs de BMPs dans la régulation de la production de FSH. En conclusion,l’ensemble de nos résultats n’est pas en faveur d’un rôle de BMP-4 hypophysaire dans la synthèse de FSH chez l’adulte. Un rôle des BMPs au niveau hypophysaire via la voie endocrine peut s’envisager puisque nous avons mis en évidence une bioactivité de type BMP dans le sérum ovin. Les inhibiteurs BMPs produits par l’hypophyse, qui restent à identifier, pourraient moduler la biodisponibilité des BMPs atteignant l’hypophyse par la voie sanguine. / BMPs inhibit FSH synthesis and release in ewe. The detection of BMP mRNAs in the pituitary as well as the colocalisation of the two types of BMP receptors on gonadotrope cells suggest that these BMPs can exert paracrine/autocrine actions on FSH production. The aim of this thesis work was to determine the importance of pituitary BMPs in the regulation of FSH production in the ewe. We showed that the level of mRNAs for BMP-4 system, analyzed by real-time RT-PCR, did not vary across the oestrous cycle or in vitro whatever the incubation period (6h-48h) and the treatment (GnRH, oestradiol, activin) of pituitary cells. By using a bioactivity test based on embryonic mesenchymal cells transfected with an expression construct containing a BMP-responsive element fused to firefly luciferase reporter gene,we detected no BMP activity within conditioned media from pituitary cells whatever the incubation period (6h-48h) and the treatment (GnRH, oestradiol, activin) of the pituitary cells. However, we detected a BMP inhibitory activity which is increased by GnRH and oestradiol suggesting the implication of BMP inhibitor(s) in FSH regulation. In conclusion, the results are not in favor of a role for pituitary BMP-4 in the regulation of FSH synthesis in adult ewe. An endocrine action of BMPs at pituitary level can be evoked since BMP bioactivity was detected within ovine serum. BMP inhibitors that remain to be identified can modulate the bioavailability of BMPs reaching the pituitary by the blood way.
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Reduced Burst Release of Bioactive rhBMP-2 from a Three-phase Composite ScaffoldGrant, David William 31 December 2010 (has links)
Recombinant human bone morphogenic proteins (rhBMPs) are extensively studied and employed clinically for treatment of various bone defects. Current clinical delivery vehicles suffer wasteful burst releases that mandate supra-physiological dosing driving concerns over safety and cost. It was therefore investigated whether a unique drug delivery vehicle sequestered within a composite scaffold could lower the burst release of rhBMP-2. PLGA-calcium phosphate tri-phasic composite scaffolds delivered model protein BSA with burst release of ~13% and sustained kinetics of 0.5-1.5% BSA/day up to 45 days. rhBMP-2 was delivered with zero burst release however at much lower levels, totaling 0.09% to 0.9 % release over 10 days, but had up to 6.3-fold greater bioactivity than fresh rhBMP-2 (p<0.05). In conclusion, the three-phase composite scaffold can deliver bioactive proteins with a reduced burst release and sustained secondary kinetics.
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Reduced Burst Release of Bioactive rhBMP-2 from a Three-phase Composite ScaffoldGrant, David William 31 December 2010 (has links)
Recombinant human bone morphogenic proteins (rhBMPs) are extensively studied and employed clinically for treatment of various bone defects. Current clinical delivery vehicles suffer wasteful burst releases that mandate supra-physiological dosing driving concerns over safety and cost. It was therefore investigated whether a unique drug delivery vehicle sequestered within a composite scaffold could lower the burst release of rhBMP-2. PLGA-calcium phosphate tri-phasic composite scaffolds delivered model protein BSA with burst release of ~13% and sustained kinetics of 0.5-1.5% BSA/day up to 45 days. rhBMP-2 was delivered with zero burst release however at much lower levels, totaling 0.09% to 0.9 % release over 10 days, but had up to 6.3-fold greater bioactivity than fresh rhBMP-2 (p<0.05). In conclusion, the three-phase composite scaffold can deliver bioactive proteins with a reduced burst release and sustained secondary kinetics.
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