The effect of crosslinking heparin to demineralized bone matrix on mechanical strength and specific binding to human bone morphogenetic protein-2.

Demineralized bone matrix (DBM) is a collagen-based scaffold, but its low mechanical strength and limited BMP-2 binding ability restrict its application in bone repair. It is known that heparin could be immobilized onto scaffolds to enhance their binding of growth factors with the heparin-binding domain. Here, we crosslinked heparin to DBM to increase its BMP-2 binding ability. To our surprise, the mechanical strength of DBM was also dramatically increased. The compression modulus of heparin crosslinked DBM (HC-DBM) have improved (seven-fold increased) under wet condition, which would allow the scaffolds to keep specific shapes in vivo. As expected, HC-DBM showed specific binding ability to BMP-2. Additional studies showed the bound BMP-2 exerted its function to induce cell differentiation on the scaffold. Subcutaneous implantation of HC-DBM carrying BMP-2 showed higher alkaline phosphatase (ALP) activity (2 weeks), more calcium deposition (4 and 8 weeks) and more bone formation than that of control groups. It is concluded that HC-DBM has increased mechanical intensity as well as specific BMP-2 binding ability; HC-DBM/BMP-2 enhances the osteogenesis and therefore could be an effective medical device for bone repair.

Homogeneous osteogenesis and bone regeneration by demineralized bone matrix loading with collagen-targeting bone morphogenetic protein-2.

Considerable research has been focused on the development of bone morphogenetic protein-2 (BMP-2) delivery system for homologous and efficient bone regeneration. The aim of the present study was to develop a collagen-based targeting bone repair system. A collagen-binding domain (CBD) was added to the N-terminal of native BMP-2 to allow it bind to collagen specifically. We showed that the collagen-binding bone morphogenetic protein-2 (named bone morphogenetic protein2-h, BMP2-h) had maintained the full biological activity as compared to rhBMP2 lacking the CBD. In vitro functional study also demonstrated that collagen matrix could maintain higher bioactivity of BMP2-h than native BMP-2. When demineralized bone matrix (DBM) impregnated with BMP2-h was implanted subcutaneously in rats, homogeneous bone formation was observed. Moreover, in a rabbit mandible defect model, surgical implantation of collagen matrix loaded with BMP2-h exhibited remarkable osteoinductive properties and excellent homogeneous bone formation. Our studies suggested that this novel collagen-based BMP-2 targeting bone repair system induced better bone formation not only in quantity but also in quality. Similar approaches may also be used for the repair of other tissue injuries.

Crosslinked three-dimensional demineralized bone matrix for the adipose-derived stromal cell proliferation and differentiation.

Stem cell-based therapy has been a promising method for tissue regeneration and wound repair. Adult adipose-derived stromal cells (ADSCs) are often used for adipose and bone tissue reconstruction because of their abundant sources and multipotential differentiation ability. When combined with carriers, ADSCs could be useful for constructing tissue substitutes in vitro or facilitating tissue regeneration in vivo. Demineralized bone matrix (DBM) has been used for tissue reconstruction because collagen presents good cell compatibility. However, DBM degrades rapidly when used for three-dimensional ADSC culture. Here DBM was crosslinked with 1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide and N-hydroxysulfosuccinimide to investigate whether crosslinked DBM (CRL-DBM) could be used as ADSC carrier. CRL-DBM showed not only improved mechanical property and enhanced stability, but also sustained ADSC proliferation and effective differentiation into adipocytes and bone lineage cells. The results indicated that CRL-DBM may be a suitable ADSC carrier for adipose and bone tissue regeneration.

The bone-derived collagen containing mineralized matrix for the loading of collagen-binding bone morphogenetic protein-2.

Bone tissue-derived biomaterials have often been applied for bone repair because of their similarity to human bone in structure and composition. When combined with growth factors, they could accelerate bone formation. Here, we explore a collagen containing mineralized bone-derived matrix (CCMBM) from bovine bone tissues, which not only maintains proper mechanical strength but also binds to the collagen-binding recombinant human collagen-binding bone morphogenetic protein-2 (CBD-BMP(2)). By analyzing its morphology and composition, we found that CCMBM was porous and mainly composed of calcium compounds. CCMBM could provide mechanical support for bone injury repair. It also showed good biocompatibility and proper degradation rate that would be helpful for bone regeneration. In addition, the intentionally preserved collagen allowed the specific binding of CBD-BMP(2) to CCMBM, and resulted in significantly increased osteogenesis in vivo. The results indicated that the combination of CCMBM with collagen-binding BMP(2) could be emerged into an effective medical device for bone repair.

Collagen membranes loaded with collagen-binding human PDGF-BB accelerate wound healing in a rabbit dermal ischemic ulcer model.

Studies have shown that exogenous platelet-derived growth factor-BB (PDGF-BB) could accelerate the ulcer healing, but the lack of efficient growth factor delivery system limits its clinical application. Our previous work has demonstrated that the native human PDGF-BB was added a collagen-binding domain (CBD), TKKTLRT, to develop a collagen-based PDGF targeting delivery system. Here, we showed that this CBD-fused PDGF-BB (CBD-PDGF) could bind to collagen membrane efficiently. We used the rabbit dermal ischemic ulcer model to study the effects of CBD-PDGF loaded on collagen membranes. Results revealed that this system maintained a higher concentration and stronger bioactivity of PDGF-BB on the collagen membranes and promoted the re-epithelialization of dermal ulcer wounds, the collagen deposition, and the formation of capillary lumens within the newly formed tissue area. It demonstrated that collagen membranes loaded with collagen-targeting human PDGF-BB could effectively promote ulcer healing.