In vivo performance of combinations of autograft, demineralized bone matrix, and tricalcium phosphate in a rabbit femoral defect model.

Large bone defects may be treated with autologous or allogeneic bone preparations. Each treatment has advantages and disadvantages; therefore, a clinically viable option for treating large (e.g., gap) bone defects may be a combination of the two. In the present study, bone repair was determined with combinations of autografts, allografts, and synthetic bone grafts using an established rabbit femoral defect model. Bilateral unicortical femoral defects were surgically prepared and treated with combinatorial bone grafts according to one of seven treatment groups. Recipient sites were retrieved at six weeks. Cellular/tissue responses and new bone formation were assessed by histology and histomorphometry. Histological analysis images indicated neither evidence of inflammatory, immune responses, tissue necrosis, nor osteolysis. Data suggested co-integration of implanted agents with host and newly formed bone. Finally, the histomorphometric data suggested that the tricalcium phosphate-based synthetic bone graft substitute allowed new bone formation that was similar to the allograft (i.e., demineralized bone matrix, DBM).

Demineralized bone matrix in bone repair: history and use.

Demineralized bone matrix (DBM) is an osteoconductive and osteoinductive commercial biomaterial and approved medical device used in bone defects with a long track record of clinical use in diverse forms. True to its name and as an acid-extracted organic matrix from human bone sources, DBM retains much of the proteinaceous components native to bone, with small amounts of calcium-based solids, inorganic phosphates and some trace cell debris. Many of DBM's proteinaceous components (e.g., growth factors) are known to be potent osteogenic agents. Commercially sourced as putty, paste, sheets and flexible pieces, DBM provides a degradable matrix facilitating endogenous release of these compounds to the bone wound sites where it is surgically placed to fill bone defects, inducing new bone formation and accelerating healing. Given DBM's long clinical track record and commercial accessibility in standard forms and sources, opportunities to further develop and validate DBM as a versatile bone biomaterial in orthopedic repair and regenerative medicine contexts are attractive.

Bone regeneration in a rabbit critical-sized calvarial model using tyrosine-derived polycarbonate scaffolds.

Porous three-dimensional tyrosine-derived polycarbonate (TyrPC) scaffolds with a bimodal pore distribution were fabricated to mimic bone architecture using a combination of salt-leaching and phase separation techniques. TyrPC scaffolds degraded in register with bone regeneration during the 6-week study period and compressive moduli of the scaffolds were maintained >0.5 MPa at 6 weeks of incubation in PBS at 37 °C. The TyrPC scaffolds either unsupplemented or supplemented with recombinant human bone morphogenetic protein-2 (rhBMP-2) were implanted in a rabbit calvarial critical-sized defect (CSD) model and the TyrPC scaffolds treated with rhBMP-2 or TyrPC coated with calcium phosphate scaffold alone promoted bone regeneration in a rabbit calvarial CSD at 6 weeks postimplantation. A synthetic TyrPC polymeric scaffold either without a biological supplement or with a minimal dose of rhBMP-2 induced bone regeneration comparable to a commercially available bone graft substitute in a nonrodent CSD animal model.

Fracture healing in the elderly patient.

Clinical experience gives rise to the impression that there are differences in fracture healing in different age groups. It is evident that fractures heal more efficiently in children than in adults. However, minimal objective knowledge exists to evaluate this assumption. Temporal, spatial, and cellular quantitative and qualitative interrelationships, as well as signaling molecules and extracellular matrix have not been comprehensively and adequately elucidated for fracture healing in the geriatric skeleton. The biological basis of fracture healing will provide a context for revealing the pathophysiology of delayed or even impaired bone regeneration in the elderly. We will summarize experimental studies on age-related changes at the cellular and molecular level that will add to the pathophysiological understanding of the compromised bone regeneration capacity believed to exist in the elderly patient. We will suggest why this understanding would be useful for therapeutics focused on bone regeneration, in particular fracture healing at an advanced age.

Three-dimensional biocompatible ascorbic acid-containing scaffold for bone tissue engineering.

A biodegradable, biocompatible, ascorbic acid-containing three-dimensional polyurethane matrix was developed for bone tissue-engineering scaffolds. This matrix was synthesized with lysine-di-isocyanate (LDI), ascorbic acid (AA), glycerol, and polyethylene glycol (PEG). LDI-glycerol-PEG-AA prepolymer when reacted with water foamed with the liberation of CO(2) to provide a pliable, spongy urethane polymer with pore diameters of 100 to 500 microm. The LDI-glycerol-PEG-AA matrix degraded in aqueous solution and yielded lysine, glycerol, PEG, and ascorbic acid as breakdown products. The degradation products did not significantly affect the solution pH. The LDI-glycerol-PEG-AA matrix can be fabricated into diverse scaffold dimensions and the physicochemical properties of the polymer network supported in vitro cell growth. Green fluorescent protein-transgenic mouse bone marrow cells (GFP-MBMCs) attached to the polymer matrix and remained viable, and the cells became confluent cultures. Furthermore, ascorbic acid released from LDI-glycerol-PEG-AA matrix stimulated cell proliferation, type I collagen, and alkaline phosphatase synthesis in vitro. Cells grown on LDI-glycerol-PEG-AA matrix did not differ phenotypically from cells grown on tissue culture polystyrene plates as assessed by cell growth, expression of mRNA for collagen type I, and transforming growth factor beta(1). These observations suggest that AA-containing polyurethane may be useful in bone tissue-engineering applications.

A biodegradable polyurethane-ascorbic acid scaffold for bone tissue engineering.

A novel, nontoxic, biodegradable, sponge-like polyurethane scaffold was synthesized from lysine-di-isocyanate (LDI) and glycerol. Ascorbic acid (AA) was copolymerized with LDI-glycerol. Our hypothesis was that the AA-containing polymer foam would enhance the biological activity of the osteoblastic precursor cell (OPCs). The LDI-glycerol-AA matrix degraded in aqueous solution to the nontoxic products of lysine, glycerol, and AA. The degradation products did not significantly affect the solution pH. The physical properties of the polymer network supported the cell growth in vitro. Mouse OPCs attached to the polymer matrix and remained viable. OPCs produced multilayered confluent cultures, a characteristic typical of bone cells. Furthermore, AA release stimulated cell proliferation, type I collagen, and alkaline phosphatase synthesis. Cells grown on the LDI-glycerol-AA matrix also showed an enhancement of mRNA expression for pro-alpha1 (I) collagen and transforming growth factor-alpha1 after 1 week. Data were tested for significance with an analysis of variance model and multiple comparison test (Fisher's Protected Least Significant Difference) at p < or = 0.05. The observations suggest that AA-containing polyurethane may be useful in bone tissue engineering applications.

Zinc ligands in an astacin family metalloprotease meprin A.

A conserved tyrosine residue in the 'astacin family' of metalloproteases is one of five ligands proposed to coordinate zinc at the active site. Site-directed mutagenesis of the conserved Tyr (Y226) of recombinant mouse meprin alpha was used to test the hypothesis that this residue is essential for zinc binding and enzymatic activity. In addition, another proposed zinc binding ligand, H167, in the conserved (HEXXH) zinc binding motif of the meprin alpha protease domain was replaced by an alanine residue. Both mutants were expressed and secreted with the same subunit mass as wild type (90 kDa). The Y226F mutant retained the capacity to oligomerize to higher covalently and noncovalently-linked oligomers as the wild type, whereas H167A was predominantly a monomer. The kcat/Km for Y226F against a fluorgenic bradykinin substrate analog was approximately 15% of the wild type, while the H167A mutant had no detectable activity. Both Y226F and H167A were more susceptible to extensive degradation by trypsin compared with the wild-type protein. The zinc content in the wild-type and Y226F mutant proteins were similar, one molecule of zinc per subunit. The results indicate that Y226 is not essential for zinc binding, but Y226 and H167 are essential for full enzymatic activity and stability of the metalloproteinase.