BMP-6 exerts its osteoinductive effect through activation of IGF-I and EGF pathways

Abstract

We have recently shown that human recombinant BMP-6 (rhBMP-6), given systematically, can restore bone in animal models of osteoporosis. To further elucidate the underlying mechanisms of new bone formation following systemic application of BMPs, we conducted gene expression profiling experiments using bone samples of oophrectomised mice treated with BMP-6. Gene set enrichment analysis revealed enrichment of insulin-like growth factor-I and epidermal growth factor related pathways in animals treated with BMP-6. Significant upregulation of IGF-I and EGF expression in bones of BMP-6 treated mice was confirmed by quantitative PCR. To develop an in vitro model for evaluation of the effects of BMP-6 on cells of human origin, we cultured primary human osteoblasts. Treatment with rhBMP-6 accelerated cell differentiation as indicated by the formation of mineralised nodules by day 18 of culture versus 28-30 days in vehicle treated cultures. In addition, alkaline phosphatase gene expression and activity were dramatically increased upon BMP-6 treatment. Expression of IGF-I and EGF was upregulated in human osteoblast cells treated with BMP-6. These results collectively indicate that BMP-6 exerts its osteoinductive effect, at least in part, through IGF-I and EGF pathways, which can be observed both in a murine model of osteopenia and in human osteoblasts.Résumé Nous avons récemment pu mettre en évidence que la BMP-6 (rhBMP-6), administrée de façon systématique, pouvait améliorer la restauration du capital osseux de modèles animaux avec ostéoporose. Nous avons conduit une expérimentation utilisant des souris ovarieactomisées traitées par BMP-6. L'analyse a montré qu'il y avait un apport d'insulin like growth factor et d'épidermal growth factor chez les animaux traités par BMP-6. Pour développer un modèle in vitro nous avons étudié l'effet de la BMP-6 sur les cellules de type ostéoplasties d'origine humaine. Le traitement par BMP-6 accélère la différenciation cellulaire au 18ème jour alors que normalement cette différence est notée aux alentours du 28ème et 30ème jour. De plus, l'expression du gène de la phosphatase alkaline et l'activité sont augmentées par le traitement par la BMP-6, de même en ce qui concerne l'IGF-1 et l'EGF. Ces résultats nous permettent de penser que le BMP-6 a un effet ostéo conducteur notamment pour les pathologies intéressant IGF-1 et EGF. Nous avons observé ces effets dans un modèle animal avec ostéoplastie et sur les ostéoblastes humains.

Exogenous heparin binds and inhibits bone morphogenetic protein 6 biological activity

“…It is known that BMPs support differentiation of C2C12 cells towards an osteoblastic phenotype [51,52]. BMP6 in concentration of 50 ng/ml induced the expression of ALP by 180-fold and OC by four-fold after 72 h. The addition of heparin (10 μg/ml) attenuated the expression of both osteoblastic markers, which is also consistent with previous reports demonstrating that exogenous heparin can inhibit BMP2 and BMP7 binding to their receptors [10,15].…”

The clinical use of bone morphogenetic proteins revisited: a novel biocompatible carrier device OSTEOGROW for bone healing

“…To elucidate the underlying mechanism, gene expression profiling experiments revealed enrichment of IGF-1 and EGF related pathways in rats treated with BMP6. Similar results were obtained on primary human osteoblasts, suggesting that BMP6 exerts its osteoinductive effect, at least in part, through IGF-1 and EGF pathways [13]. In C2C12 premyoblasts, alteration of the plasma membrane heparan-sulfate (HS) structures with chlorate or heparinase III treatment inhibited BMP6-mediated signaling and differentiation of the cells into osteoblasts, indicating that endogenous HS plays an important role in the BMP6 activity.…”

“…Functional analysis demonstrated that PDGF-BB stimulated osteoblast chemotaxis that was reduced by assessing siRNAs of PDGFR-β in osteoblasts [10,11]. Growth factors released during bone resorption that rebound to the extracellular matrix components also include transforming growth factor β1 (TGFβ1) and insulin growth factor 1 (IGF-1) [12], stimulating osteoblast differentiation from bone marrow mesenchymal stem cells (BMSCs) [13,14]. A novel molecule named couplin has been recently identified in osteoclasts that also promotes osteoblast differentiation from BMSCs.…”

“…(a) At the endosteal surface BMP2 affects both osteoclasts and osteoblasts with a net outcome of downregulation of Runx2, collagen I and Wnt signaling [75]; at the periosteum BMPs stimulate differentiation of precursor cells (green) into osteoblasts (orange); and in surrounding muscle cells (myoblasts, pericytes and vascular satellite cells) BMPs upregulate Id genes and form new osteoblasts and prechondrocytes to form cartilage and new bone around the cortical layer from which the new bone spreads into the medullar cavity. (b) As shown in patients with high tibial osteotomy and a fibular defect, BMP7 (Osigraft) stimulates bone formation from periosteum (P) and surrounding muscles (M) spreading endostealy at six and ten weeks to rebuild the cortical bone (CB) at one year following implantation (modified from [109]) Uniqueness and exceptionality of BMP6 BMP6 has specific effects in its ability to convert stem cells to cartilage and bone forming cells [13,48,116]. BMP6 and several other BMP family members are produced and released by BMSC and hematopoietic stem cells (HSC) which both constitute the stem cell niche of the bone marrow [117].…”

The Use of Mass Spectrometry in Characterization of Bone Morphogenetic Proteins from Biological Samples

“…Next, using gene expression profiling, we found that BMP-6 exerts its osteoinductive effect at least in part through the IGF-I and epidermal growth factor (EGF) pathways. (Grasser et al, 2007 have studied impairment of osteoblasts differentiation in context of the glucocorticoid-induced osteoporosis. They treated MC3T3-E1 cells with high dose of dexamethasone, which inhibited cell differentiation and proliferation and induced apoptosis.…”