Effects of particle size and porosity on in vivo remodeling of settable allograft bone/polymer composites.

Established clinical approaches to treat bone voids include the implantation of autograft or allograft bone, ceramics, and other bone void fillers (BVFs). Composites prepared from lysine-derived polyurethanes and allograft bone can be injected as a reactive liquid and set to yield BVFs with mechanical strength comparable to trabecular bone. In this study, we investigated the effects of porosity, allograft particle size, and matrix mineralization on remodeling of injectable and settable allograft/polymer composites in a rabbit femoral condyle plug defect model. Both low viscosity and high viscosity grafts incorporating small (<105 µm) particles only partially healed at 12 weeks, and the addition of 10% demineralized bone matrix did not enhance healing. In contrast, composite grafts with large (105-500 µm) allograft particles healed at 12 weeks postimplantation, as evidenced by radial µCT and histomorphometric analysis. This study highlights particle size and surface connectivity as influential parameters regulating the remodeling of composite bone scaffolds.

Balancing the rates of new bone formation and polymer degradation enhances healing of weight-bearing allograft/polyurethane composites in rabbit femoral defects.

There is a compelling clinical need for bone grafts with initial bone-like mechanical properties that actively remodel for repair of weight-bearing bone defects, such as fractures of the tibial plateau and vertebrae. However, there is a paucity of studies investigating remodeling of weight-bearing bone grafts in preclinical models, and consequently there is limited understanding of the mechanisms by which these grafts remodel in vivo. In this study, we investigated the effects of the rates of new bone formation, matrix resorption, and polymer degradation on healing of settable weight-bearing polyurethane/allograft composites in a rabbit femoral condyle defect model. The grafts induced progressive healing in vivo, as evidenced by an increase in new bone formation, as well as a decrease in residual allograft and polymer from 6 to 12 weeks. However, the mismatch between the rates of autocatalytic polymer degradation and zero-order (independent of time) new bone formation resulted in incomplete healing in the interior of the composite. Augmentation of the grafts with recombinant human bone morphogenetic protein-2 not only increased the rate of new bone formation, but also altered the degradation mechanism of the polymer to approximate a zero-order process. The consequent matching of the rates of new bone formation and polymer degradation resulted in more extensive healing at later time points in all regions of the graft. These observations underscore the importance of balancing the rates of new bone formation and degradation to promote healing of settable weight-bearing bone grafts that maintain bone-like strength, while actively remodeling.

Injectable foams for regenerative medicine.

The design of injectable biomaterials has attracted considerable attention in recent years. Many injectable biomaterials, such as hydrogels and calcium phosphate cements (CPCs), have nanoscale pores that limit the rate of cellular migration and proliferation. While introduction of macroporosity has been suggested to increase cellular infiltration and tissue healing, many conventional methods for generating macropores often require harsh processing conditions that preclude their use in injectable foams. In recent years, processes such as porogen leaching, gas foaming, and emulsion-templating have been adapted to generate macroporosity in injectable CPCs, hydrogels, and hydrophobic polymers. While some of the more mature injectable foam technologies have been evaluated in clinical trials, there are challenges remaining to be addressed, such as the biocompatibility and ultimate fate of the sacrificial phase used to generate pores within the foam after it sets in situ. Furthermore, while implantable scaffolds can be washed extensively to remove undesirable impurities, all of the components required to synthesize injectable foams must be injected into the defect. Thus, every compound in the foam must be biocompatible and noncytotoxic at the concentrations utilized. As future research addresses these critical challenges, injectable macroporous foams are anticipated to have an increasingly significant impact on improving patient outcomes for a number of clinical procedures.

Effects of local delivery of D-amino acids from biofilm-dispersive scaffolds on infection in contaminated rat segmental defects.

Infectious complications of open fractures continue to be a significant factor contributing to non-osseous union and extremity amputation. The persistence of bacteria within biofilms despite meticulous debridement and antibiotic therapy is believed to be a major cause of chronic infection. Considering the difficulties in treating biofilm-associated infections, the use of biofilm dispersal agents as a therapeutic strategy for the prevention of biofilm-associated infections has gained considerable interest. In this study, we investigated whether local delivery of D-Amino Acids (D-AAs), a biofilm dispersal agent, protects scaffolds from contamination and reduces microbial burden within contaminated rat segmental defects in vivo. In vitro testing on biofilms of clinical isolates of Staphylococcus aureus demonstrated that D-Met, D-Phe, D-Pro, and D-Trp were highly effective at dispersing and preventing biofilm formation individually, and the effect was enhanced for an equimolar mixture of D-AAs. Incorporation of D-AAs into polyurethane scaffolds as a mixture (1:1:1 D-Met:D-Pro:D-Trp) significantly reduced bacterial contamination on the scaffold surface in vitro and within bone when implanted into contaminated femoral segmental defects. Our results underscore the potential of local delivery of d-AAs for reducing bacterial contamination by targeting bacteria within biofilms, which may represent a treatment strategy for improving healing outcomes associated with open fractures.

Biocompatibility and chemical reaction kinetics of injectable, settable polyurethane/allograft bone biocomposites.

Injectable and settable bone grafts offer significant advantages over pre-formed implants due to their ability to be administered using minimally invasive techniques and to conform to the shape of the defect. However, injectable biomaterials present biocompatibility challenges due to the potential toxicity and ultimate fate of reactive components that are not incorporated in the final cured product. In this study the effects of stoichiometry and triethylenediamine (TEDA) catalyst concentration on the reactivity, injectability, and biocompatibility of two component lysine-derived polyurethane (PUR) biocomposites were investigated. Rate constants were measured for the reactions of water (a blowing agent resulting in the generation of pores), polyester triol, dipropylene glycol (DPG), and allograft bone particles with the isocyanate-terminated prepolymer using an in situ attenuated total reflection Fourier transform infrared spectroscopy technique. Based on the measured rate constants, a kinetic model predicting the conversion of each component with time was developed. Despite the fact that TEDA is a well-known urethane gelling catalyst, it was found to preferentially catalyze the blowing reaction with water relative to the gelling reactions by a ratio >17:1. Thus the kinetic model predicted that the prepolymer and water proceeded to full conversion, while the conversions of polyester triol and DPG were <70% after 24h, which was consistent with leaching experiments showing that only non-cytotoxic polyester triol and DPG were released from the reactive PUR at early time points. The PUR biocomposite supported cellular infiltration and remodeling in femoral condyle defects in rabbits at 8weeks, and there was no evidence of an adverse inflammatory response induced by unreacted components from the biocomposite or degradation products from the cured polymer. Taken together, these data underscore the utility of the kinetic model in predicting the biocompatibility of reactive biomaterials.

Injectable reactive biocomposites for bone healing in critical-size rabbit calvarial defects.

Craniofacial injuries can result from trauma, tumor ablation, or infection and may require multiple surgical revisions. To address the challenges associated with treating craniofacial bone defects, an ideal material should have the ability to fit complex defects (i.e. be conformable), provide temporary protection to the brain until the bone heals, and enhance tissue regeneration with the delivery of biologics. In this study, we evaluated the ability of injectable lysine-derived polyurethane (PUR)/allograft biocomposites to promote bone healing in critical-size rabbit calvarial defects. The biocomposites exhibited favorable injectability, characterized by a low yield stress to initiate flow of the material and a high initial viscosity to minimize the adverse phenomena of extravasation and filter pressing. After injection, the materials cured within 10-12 min to form a tough, elastomeric solid that maintained mechanical integrity during the healing process. When injected into a critical-size calvarial defect in rabbits, the biocomposites supported ingrowth of new bone. The addition of 80 µg mL(-1) recombinant human bone morphogenetic protein-2 (rhBMP-2) enhanced new bone formation in the interior of the defect, as well as bridging of the defect with new bone. These observations suggest that injectable reactive PUR/allograft biocomposites are a promising approach for healing calvarial defects by providing both mechanical stability as well as local delivery of rhBMP-2.

Synthesis and characterization of an injectable allograft bone/polymer composite bone void filler with tunable mechanical properties.

In recent years, considerable effort has been expended toward the development of synthetic bone graft materials. Injectable biomaterials offer several advantages relative to implants due to their ability to cure in situ, thus conforming to irregularly shaped defects. While Food and Drug Administration-approved injectable calcium phosphate cements have excellent osteoconductivity and compressive strengths, these materials have small pore sizes (e.g., 1 mum) and are thus relatively impermeable to cellular infiltration. To overcome this limitation, we aimed to develop injectable allograft bone/polyurethane (PUR) composite bone void fillers with tunable properties that support rapid cellular infiltration and remodeling. The materials comprised particulated (e.g., >100 microm) allograft bone particles and a biodegradable two-component PUR, and had variable (e.g., 30%-70%) porosities. The injectable void fillers exhibited an initial dynamic viscosity of 220 Pa.s at clinically relevant shear rates (40 s(-1)), wet compressive strengths ranging from < 1 to 13 MPa, working times from 3 to 8 min, and setting times from 10 to 20 min, which are comparable to the properties of calcium phosphate bone cements. When injected in femoral plug defects in athymic rats, the composites supported extensive cellular infiltration, allograft resorption, collagen deposition, and new bone formation at 3 weeks. The combination of both initial mechanical properties suitable for weight-bearing applications as well as the ability of the materials to undergo rapid cellular infiltration and remodeling may present potentially compelling opportunities for injectable allograft/PUR composites as biomedical devices for bone regeneration.