Sulfated Polysaccharides Enhance the Biological Activities of Bone Morphogenetic Proteins

Abstract

Bone morphogenetic proteins (BMPs), which have been shown to be heparin-binding proteins, induce osteoblast differentiation in mesenchymal cells. In the present study, we examined the effects of heparin on the BMP activities in C2C12 myoblasts. Heparin dose dependently enhanced the osteoblast differentiation induced by not only homodimers of BMP-2 or BMP-4 but also heterodimers of BMP-2/6 or BMP-2/7. However, the osteoblast differentiation induced by the constitutively active BMPR-IA, a functional BMP type I receptor, was not affected by heparin. Heparan sulfate and dextran sulfate also enhanced the BMP-2 activity, although the chemically desulfated heparin-derivatives have lost this stimulatory capacity. Heparin dose-dependently suppressed the accumulation of BMP-2 from the culture media into the cell layer or BMPR-IA, and retained a large amount of BMP-2 in the culture media. The biological activity of BMP-2, which was evaluated using a BMP-responsive reporter gene expression, was prolonged in the presence of heparin. Taken together, these results suggest that sulfated polysaccharides enhance the biological activity of both homodimers and heterodimers of BMPs by continuously serving the ligands to their signaling receptors expressed on cell membranes. Bone morphogenetic proteins (BMPs)1 were originally identified as unique in demineralized bone matrix that could induce ectopic bone formation when implanted into muscular tissues (1, 2). More than 15 members of BMPs have been identified, and they are members of the TGF-␤ superfamily (3-7). They are classified into several BMP subfamilies based on their homology within the mature domains; the BMP-2 and BMP-4 subfamily, the BMP-5, BMP-6, BMP-7 (also called OP-1) and BMP-8 subfamily, the GDF-5, GDF-6 (BMP-13), and GDF-7 (BMP-12) subfamily, and the BMP-3 and GDF-10 (BMP-3b) subfamily (3-7). Most BMPs are first synthesized in large inactive preproproteins. They form homodimers or heterodimers via a disulfide bond in the mature domain, and then are secreted as active dimers after proteolytic processing (3-7). Several recombinant proteins of BMP dimers were reported to be active in an ectopic bone formation assay. Some heterodimers were shown to be more potent than each homodimer (8 -11). Several lines of evidence suggest that BMPs are key molecules for normal skeletal development in vertebrates (3, 5-7, 12). We previously reported that BMP-2 inhibits myogenic differentiation of C2C12 myoblasts, and converts their differentiation pathway into that of osteoblast lineage cells (13). Both the ectopic bone-inducing activity and the osteoblast differentiation-inducing activity appear to be unique for BMPs among members of the TGF-␤ superfamily.Signaling of BMPs is initiated by binding to the specific transmembrane receptors, type I and type II serine/threonine kinase receptors (6, 12, 14 -16). The type I receptors are activated by the ligand bound-type II receptors, and then phosphorylate Smad proteins as substrates in the cytoplasm. The phosphorylated Smad forms a complex ...

Sustained release and osteogenic potential of heparan sulfate-doped fibrin glue scaffolds within a rat cranial model

“…The interaction of BMPs with their threonine-serine kinase BMP receptors is also thought to be mediated by HS (Irie et al 2003, Takada et al 2003, Coombe et al 2005. And furthermore HS is thought to prevent diffusion of BMP2 in vivo, away from the regions where is it likely to be required (Depprich et al 2005).…”

Characterization of Sugar Binding by Osteoclast Inhibitory Lectin

“…Furthermore, mOCIL binds chondroitin sulfate A with higher affinity compared with chondroitin sulfate C, which is significantly increased in proliferative tissue in contact with the mineralization zone during calcification (25). It has been reported that bone morphogenic proteins bind heparin and that sulfated polysaccharides, including heparin, heparin sulfate, and dextran sulfate (but not desulfated heparin) enhance the osteoblast differentiation induced by homodimers and heterodimers of bone morphogenic proteins (26). Therefore, although mOCIL has no requirement for carbohydrate recognition in inhibiting osteoclastogenesis, the possibility that OCIL could require sugar recognition in other bone-specific roles cannot be excluded.…”

Cell biology of osteochondromas: Bone morphogenic protein signalling and heparan sulphates

“…HSPGs can sequester BMPs at the cell surface, directing their movement across the surface through restricted diffusion [14]. HS enhances BMP activity by continuously serving the ligands to their signalling receptors on the cell surface [38]. Furthermore, HSPGs regulate BMP-2 induction of osteogenic markers and internalisation in vitro [14].…”

“…Membrane bound and extracellular HS is known to bind to BMPs and is involved in BMP-mediated morphogenesis by regulating their gradient formation and activity [14,38]. HSPGs can sequester BMPs at the cell surface, directing their movement across the surface through restricted diffusion [14].…”