BMPRII, a BMP type II receptor, plays an important role in regulating diverse biological events. BMPRII contains a long carboxyl tail domain, which is highly conserved among vertebrate species. The tail domain has been shown to regulate cytoskeleton remodeling, whereas the function in regulating canonical BMP signalling is not well studied. Here, I show that the BMPRII tail domain reduces the magnitude of BMP7-induced pSmad1 activation in the early stage, which also changes the magnitude of BMP target gene expression. Furthermore, my data also suggest that the BMPRII tail not only modulates BMP7-induced Smad1 carboxyl terminal phosphorylation, but also inhibits endogenous BMP signalling under overexpression conditions. Finally, BMP7 promotes Neuro2a neurite extension and I demonstrate that knockdown of BMPRII affects BMP7 induced neurite outgrowth. Altogether, these studies demonstrate that the BMPRII tail may play a role in regulating responsiveness to BMP7, and thereby modulates BMP7 dependent neurite extension in neuronal cells.

BMPRII, a BMP type II receptor, plays an important role in regulating diverse biological events. BMPRII contains a long carboxyl tail domain, which is highly conserved among vertebrate species. The tail domain has been shown to regulate cytoskeleton remodeling, whereas the function in regulating canonical BMP signalling is not well studied. Here, I show that the BMPRII tail domain reduces the magnitude of BMP7-induced pSmad1 activation in the early stage, which also changes the magnitude of BMP target gene expression. Furthermore, my data also suggest that the BMPRII tail not only modulates BMP7-induced Smad1 carboxyl terminal phosphorylation, but also inhibits endogenous BMP signalling under overexpression conditions. Finally, BMP7 promotes Neuro2a neurite extension and I demonstrate that knockdown of BMPRII affects BMP7 induced neurite outgrowth. Altogether, these studies demonstrate that the BMPRII tail may play a role in regulating responsiveness to BMP7, and thereby modulates BMP7 dependent neurite extension in neuronal cells.

A research report submitted to the School of Oral Health Sciences, University of the Witwatersrand, Johannesburg, in partial fulfilment of the requirements for the degree of Master of Dentistry in the branch Maxillofacial and Oral Surgery. Johannesburg 2014 / The reconstruction of osseous mandibular defects remains a significant challenge. The use of autologous bone for mandibular reconstruction is associated with numerous limitations, and alternatives to autologous bone would provide significant benefits for patients. The aim of this study was to evaluate and compare binary application of recombinant human bone morphogenetic protein 7 (rhBMP-7) and recombinant human transforming growth factor  (rhTGF-3) to solo application of recombinant human bone morphogenetic protein 7 (rhBMP-7) in full-thickness mandibular defects in the non-human primate Papio ursinus. In four baboons, a 2.5cm segmental defect was created in the mandible and stabilized with a 2.7mm titanium reconstruction plate. Two defects were implanted with rhBMP-7 solo, and the other two with binary application rhBMP-7 and rhTGF-3 at a ratio of 20:1. All four baboons were euthanazed at 180 days post implantation. All four specimens were radiographed prior to sectioning. Tissue processing and histomorphometry were done on the undecalcified sections prepared from the harvested mandible specimens. In all defects bone regeneration re-established bony continuity at six months. The mean area of the regenerate was 336 ± 107.5 mm2 (range 229-444.7) in the solo specimens, and 312 ± 63.5mm2 (range 249-376.6) in the binary specimens. Radiographic examination confirmed complete bone healing in all defects but variable restitution of defect volume. The regenerated bone had a trabecular pattern consistent with mature mandibular bone and the defect interfaces were indiscernible. Due to the small sample size no performance advantage could be identified between the two treatment groups. These results confirm that successful bone regeneration by tissue induction in surgically created mandibular defects can be achieved with osteogenic proteins of the transforming growth factor- superfamily.

We have developed a versatile delivery platform comprising a novel composite of two biomaterials with proven track records: apatite and poly(lactic-co-glycolic acid) (PLGA). These composites have been tested in the delivery of a model protein, bovine serum albumin (BSA), as well as a growth factor, bone morphogenetic protein-2 (BMP-2), which is a potent inducer of bone formation. The controlled release strategy is based on the use of a polymer with acidic degradation products to control the dissolution of a basic inorganic component, resulting in protein release. The release profile can be modified systematically by changing variables that affect polymer degradation and/or apatite dissolution, such as polymer molecular weight, polymer composition, apatite loading, and apatite particle size. We have found that an increase in polymer molecular weight and polymer hydrophobicity led to slower polymer degradation, and in turn, slower apatite dissolution and protein release. Protein release was enhanced by reducing apatite particle size and by lowering the apatite content in the composites. We anticipate that this delivery platform can be extended to the controlled release of other therapeutic proteins and chemicals. / Singapore-MIT Alliance (SMA)

Bone Morphogenetic proteins (BMPs) belong to transforming growth factor beta (TGF-£]) superfamily. They regulate cell proliferation, cell differentiation, and bone morphogenesis. Previous evidence suggests that BMP-2, as an antagonist of TGF-£], may play an inhibitory role in tissue fibrogenesis. The aim of this study is to examine the expression profile of BMP-2 in fibrotic livers and to test whether BMP-2 gene delivery could alleviate or reverse the liver fibrogenesis models in mice including bile duct ligation (BDL) or carbon tetrachloride (CCl4) model. The results showed that the AST, ALT, and bilirubin levels in sera and the expression of TGF-£], £\-smooth muscle actin, type I collagen in livers were significantly up-regulated by BDL surgery or CCl4 administration. After BDL, the hepatic BMP-2 mRNA and protein levels in mice decreased at 7 and 14 days after surgery. Similarly, the hepatic BMP-2 mRNA and protein levels in mice decreased at day 14 and 28 after CCl4 administration. BMP-2 gene delivery alleviated the inflammation and the liver injury caused by BDL or CCl4 exposure. These findings strongly suggest that BMP-2 is involved in the pathogenesis of liver fibrosis. Moreover, BMP-2 supplementation may facilitate a novel strategy for treatment of liver fibrosis.

Bone morphogenetic proteins-2/4/7 (BMP-2/4/7) are important cytokines in systemic tissue morphogenesis. It has been demonstrated BMPs may have positive effects on liver repair and regeneration after hepatic injury. However, their function in the liver still remains unclear. D-galactosamine (D-gal) is a hepatotoxin used to induce hepatic failure. We employed D-gal and rat hepatoma cell line (1548) to investigate BMP-2/4/7 expression in hepatic injury induced by D-gal and probe their relations with liver repair and regeneration in hepatic injury. LDH release, mRNA and protein expression were detected. Results indicated that BMP-2/4/7 expression was activated by injury of rat hepatoma cells. It is indicative that repair and regeneration of the liver after hepatic injury and morphogenesis in early embryos seem to proceed through the same process. BMPs may be not only associated with hepatic injury after repair and regeneration, but also involved in chronic liver.

Bone morphogenetic proteins-2/4/7 (BMP-2/4/7) are important cytokines in systemic tissue morphogenesis. It has been demonstrated BMPs may have positive effects on liver repair and regeneration after hepatic injury. However, their function in the liver still remains unclear. D-galactosamine (D-gal) is a hepatotoxin used to induce hepatic failure. We employed D-gal and rat hepatoma cell line (1548) to investigate BMP-2/4/7 expression in hepatic injury induced by D-gal and probe their relations with liver repair and regeneration in hepatic injury. LDH release, mRNA and protein expression were detected. Results indicated that BMP-2/4/7 expression was activated by injury of rat hepatoma cells. It is indicative that repair and regeneration of the liver after hepatic injury and morphogenesis in early embryos seem to proceed through the same process. BMPs may be not only associated with hepatic injury after repair and regeneration, but also involved in chronic liver.

Chez le poulet, le déterminisme du sexe est génétique (ZZ/ZW) mais à la différence des mammifères le déterminant majeur du sexe n’a pas été identifié. Néanmoins, un certain nombre d’acteurs moléculaires impliqués en aval dans la différenciation du testicule (DMRT1, AMH, SOX9…) ou de l’ovaire (CYP19A1, FOXL2, RSPO1…) ont été identifiés. D’autre part, le modèle poulet présente deux particularités que sont l’asymétrie du développement ovarien et la sensibilité aux stéroïdes donnant la possibilité d’inversions du sexe par des hormones exogènes. Par une analyse par PCR en temps réel à moyen débit, nous avons identifié des gènes dont l’expression est sexuellement dimorphique et/ou asymétrique au cours de la différenciation gonadique. Parmi ces gènes plusieurs membres de la famille des bone morphogenetic protein sont préférentiellement exprimés dans l’ovaire en comparaison au testicule. L’étude des effets de BMP4 sur la culture organotypique d’ovaires ou de testicules a montré qu’il était un inhibiteur de la stéroïdogénèse basale et induite par la FSH et d’autre part qu’il est un inhibiteur de l’expression de l’AMH. Résultats qui nous ont amené à émettre l’hypothèse que BMP4 était un facteur « anti-testiculaire ». / In chicken, sex is determined by a ZZ/ZW sex chromosome system, where the female is heterogametic (ZW). However, the mechanism involved in the sex determination is still unknown. Several genes involved in testicular (DMRT1, AMH, SOX9…) or in ovarian (CYP19A1, FOXL2, RSPO1...) differentiation have been identified. Furthermore, gonadal development in chicken embryos presents two particularities which are the asymmetrical ovarian development and the sensitivity to exogenous hormones leading to sex reversal. By real-time PCR analysis, we have identified several genes whose expression is sexually dimorphic and/or asymmetric. In particularly, we showed that several members of bone morphogenetic protein family are preferentially expressed in the ovary compared to the testis. Using organotypic culture of embryonic ovaries and testes, we showed that BMP4 inhibits the basal and FSH induced steroidogenesis and is an inhibitor of AMH mRNA expression. These findings lead us to propose that BMP4 is an “anti-testicular” factor.

No / Bone morphogenetic proteins (BMPs) and their receptors (BMPRs) play roles in embryonic development and postnatal remodelling of the skin. Many indications suggest that BMP signalling regulates keratinocyte proliferation and differentiation. Chronic wounds have been shown to exhibit high levels of BMP ligands; however, the effect of BMP pathway modulation on human skin healing remains undefined. A human ex vivo skin wound healing model was used to analyse the expression of BMP signalling pathway components during healing and to investigate the effects of BMPs and the BMP antagonist Noggin on skin repair. Additionally, the effects of BMP signalling on keratinocyte proliferation, apoptosis and migration were tested using in vitro flow cytometry and ‘scratch’ migration assays, respectively. BMP receptor-1B (BMPR-1B) and downstream signalling protein phosphorylated-Smad-1/5/8 were highly expressed in healing epidermis. Treatment of human skin with exogenous BMPs impaired wound closure by reducing keratinocyte proliferation and increasing apoptosis. The BMP antagonist Noggin negated the inhibitory effects of BMP ligands, and when used alone, Noggin reduced keratinocyte apoptosis in the wound bed. In vitro, BMP ligands suppressed keratinocyte proliferation whilst Noggin stimulated proliferation. Keratinocyte migration was slowed following BMP treatment; in contrast, migration was significantly accelerated due to inhibition of BMP activity by either Noggin or BMPR-1B silencing. BMP signalling is inherently involved in wound healing. BMPs slow skin repair by suppressing keratinocyte proliferation and migration. Thus, modulation of BMP signalling using BMP inhibitors such as Noggin may serve as a new approach to promote cutaneous wound repair. Level of evidence: Not ratable.

Demineralized bone matrix (DBM) is an allograft material used in orthopedics that promotes endochondral bone formation. While the placement of DBM on either the periosteal surface of a bone or within a skeletal muscle promotes the recruitment of stem cells that can form skeletal tissues through the temporal progression of endochondral bone development, it remains unclear to what degree these processes are different between the two sites. In this study, we utilize a comparative in vivo model of endochondral ossification by implanting the DBM on the periosteum and in the muscle. Within the muscle we further compared the effects of DBM with and without Bone morphogenetic protein-2 (BMP-2), a primary morphogenetic factor involved in the differentiation of skeletal stem cells. The mice were harvested at various time points after DBM implantation in order to analyze the development of the bone. Analysis included X-ray imaging, microCT imaging, and mRNA expression. Plain x-ray and micro-CT imaging analysis showed mineralized bone formation in the implant on the periosteum and in the muscle with BMP-2, but no growth in the muscle when BMP-2 was not added to the DBM. The mechanisms for bone development were further analyzed by qRT-PCR to determine temporal patterns and levels of expression of various stem cell and differentiated skeletal cell associated genes. The stem cell gene expression varied between implant placement locations suggesting different mechanisms for stem cell recruitment. Interesting, while DBM implants in the muscle without BMP did not induce mineralized tissue specific mRNA expression; specific stem cell and early skeletal cell lineage commitment genes were present. These results suggest that while DBM in muscle is capable of recruiting stem cells that higher BMP-2 levels are needed to promote the progression of cartilage to mineralized bone in muscle tissues.