In vivo safety assessment of a bio-inspired bone adhesive.

A new class of materials, bone adhesives, could revolutionise the treatment of highly fragmented fractures. We present the first biological safety investigation of a bio-inspired bone adhesive. The formulation was based upon a modified calcium phosphate cement that included the amino acid phosphoserine. This material has recently been described as substantially stronger than other bioresorbable calcium phosphate cements. Four adhesive groups with the active substance (phosphoserine) and two control groups without phosphoserine were selected for in vitro and in vivo biocompatibility testing. The test groups were subject for cell viability assay and subcutaneous implantation in rats that was followed by gene expression analysis and histology assessment after 6 and 12 weeks. All adhesive groups supported the same rate of cell proliferation compared to the α-TCP control and had viability between 45-64% when compared to cell control. There was no evidence of an increased immune response or ectopic bone formation in vivo. To conclude, this bio-inspired bone adhesive has been proven to be safe, in the present study, without any harmful effects on the surrounding soft tissue.

A new bone adhesive candidate- does it work in human bone? An ex-vivo preclinical evaluation in fresh human osteoporotic femoral head bone.

INTRODUCTION: The fixation of small intraarticular bone fragments is clinically challenging and an obvious first orthopaedic indication for an effective bone adhesive. In the present study the feasibility of bonding freshly harvested human trabecular bone with OsSticR, a novel phosphoserine modified cement, was evaluated using a bone cylinder model pull-out test and compared with a commercial fibrin tissue adhesive. METHODS: Femoral heads (n=13) were collected from hip fracture patients undergoing arthroplasty and stored refrigerated overnight in saline medium prior to testing. Cylindrical bone cores with a pre-inserted bone screw, were prepared using a coring tool. Each core was removed and glued back in place with either the bone adhesive (α-tricalcium phosphate, phosphoserine and 20% trisodium citrate solution) or the fibrin glue. All glued bones were stored in bone medium at 37°C. Tensile loading, using a universal testing machine (5 kN load cell), was applied to each core/head. For the bone adhesive, bone cores were tested at 2 (n=13) and 24 (n=11) hours. For the fibrin tissue adhesive control group (n=9), bone cores were tested exclusively at 2 hours. The femoral bone quality was evaluated with micro-CT. RESULTS: The ultimate pull-out load for the bone adhesive at 2 hours ranged from 36 to 171 N (mean 94 N, SD 42 N). At 24 hours the pull-out strength was similar, 47 to 198 N (mean 123 N, SD 43 N). The adhesive failure usually occurred through the adhesive layer, however in two samples, at 167 N and 198 N the screw pulled out of the bone core. The fibrin tissue adhesive group reached a peak force of 8 N maximally at 2 hours (range 2.8-8 N, mean 5.4 N, SD 1.6 N). The mean BV/TV for femoral heads was 0.15 and indicates poor bone quality. CONCLUSION: The bone adhesive successfully glued wet and fatty tissue of osteoporotic human bone cores. The mean ultimate pull-out force of 123 N at 24 hours corresponds to ∼ 300 kPa shear stress acting on the bone core. These first ex-vivo results in human bone are a promising step toward potential clinical application in osteochondral fragment fixation.

Gluing Living Bone Using a Biomimetic Bioadhesive: From Initial Cut to Final Healing.

Osteoporotic fractures are a growing issue due to the increasing incidence of osteoporosis worldwide. High reoperation rates in osteoporotic fractures call for investigation into new methods in improving fixation of osteoporotic bones. In the present study, the strength of a recently developed bone bioadhesive, OsStictm, was evaluated in vivo using a novel bone core assay in a murine animal model at 0, 3, 7, 14, 28, and 42 days. Histology and micro-CT were obtained at all time points, and the mean peak pull-out force was assessed on days 0-28. The adhesive provided immediate fixation to the bone core. The mean peak bone core pull-out force gradually decreased from 6.09 N (σ 1.77 N) at day 0 to a minimum of 3.09 N (σ 1.08 N) at day 7, recovering to 6.37 N (σ 4.18 N) by day 28. The corresponding fibrin (Tisseel) control mean peak bone core pull-out characteristic was 0.27 N (σ 0.27 N) at day 0, with an abrupt increase from 0.37 N (σ 0.28) at day 3, 6.39 N (σ 5.09 N) at day 7, and continuing to increase to 11.34 N (σ 6.5 N) by day 28. The bone cores failed either through core pull-out or by the cancellous part of the core fracturing. Overall, the adhesive does not interrupt healing with pathological changes or rapid resorption. Initially, the adhesive bonded the bone core to the femur, and over time, the adhesive was replaced by a vascularised bone of equivalent quality and quantity to the original bone. At the 42 day time point, 70% of the adhesive in the cancellous compartment and 50% in the cortical compartment had been replaced. The adhesive outwith the bone shell was metabolized by cells that are only removing the material excess with no ectopic bone formation. It is concluded that the adhesive is not a physical and biochemical barrier as the bone heals through the adhesive and is replaced by a normal bone tissue. This adhesive composition meets many of the clinical unmet needs expressed in the literature, and may, after further preclinical assessments, have potential in the repair of bone and osteochondral fragments.

Synthesis of Phospho-Amino Acid Analogues as Tissue Adhesive Cement Additives.

In this paper we report the synthesis of a library of phospho-amino acid analogues, via a novel single-step allyl-phosphoester protection/Pd-mediated deprotection strategy. These phosphoserine and phosphotyrosine analogues were then applied as additives to create adhesive calcium phosphate cements, allowing us to probe the chemical origins of the increased surface binding strength. We demonstrate the importance of multiple calcium binding motifs in mediating adhesion, as well as highlighting the crucial role played by substrate hydrophobicity and orientation in controlling binding strength.

Adhesive Cements That Bond Soft Tissue Ex Vivo.

The aim of the present study was to evaluate the soft tissue bond strength of a newly developed, monomeric, biomimetic, tissue adhesive called phosphoserine modified cement (PMC). Two types of PMCs were evaluated using lap shear strength (LSS) testing, on porcine skin: a calcium metasilicate (CS1), and alpha tricalcium phosphate (αTCP) PMC. CS1 PCM bonded strongly to skin, reaching a peak LSS of 84, 132, and 154 KPa after curing for 0.5, 1.5, and 4 h, respectively. Cyanoacrylate and fibrin glues reached an LSS of 207 kPa and 33 kPa, respectively. αTCP PMCs reached a final LSS of ≈110 kPa. In soft tissues, stronger bond strengths were obtained with αTCP PMCs containing large amounts of amino acid (70-90 mol%), in contrast to prior studies in calcified tissues (30-50 mol%). When αTCP particle size was reduced by wet milling, and for CS1 PMCs, the strongest bonding was obtained with mole ratios of 30-50% phosphoserine. While PM-CPCs behave like stiff ceramics after setting, they bond to soft tissues, and warrant further investigation as tissue adhesives, particularly at the interface between hard and soft tissues.

A biomechanical test model for evaluating osseous and osteochondral tissue adhesives.

BACKGROUND: Currently there are no standard models with which to evaluate the biomechanical performance of calcified tissue adhesives, in vivo. We present, herein, a pre-clinical murine distal femoral bone model for evaluating tissue adhesives intended for use in both osseous and osteochondral tissue reconstruction. RESULTS: Cylindrical cores (diameter (Ø) 2 mm (mm) × 2 mm depth), containing both cancellous and cortical bone, were fractured out from the distal femur and then reattached using one of two tissue adhesives. The adhesiveness of fibrin glue (Tisseeltm), and a novel, biocompatible, calcium phosphate-based tissue adhesive (OsStictm) were evaluated by pullout testing, in which glued cores were extracted and the peak force at failure recorded. The results show that Tisseel weakly bonded the metaphyseal bone cores, while OsStic produced > 30-fold higher mean peak forces at failure (7.64 Newtons (N) vs. 0.21 N). The failure modes were consistently disparate, with Tisseel failing gradually, while OsStic failed abruptly, as would be expected with a calcium-based material. Imaging of the bone/adhesive interface with microcomputed tomography revealed that, for OsStic, failure occurred more often within cancellous bone (75% of tested samples) rather than at the adhesive interface. CONCLUSIONS: Despite the challenges associated with biomechanical testing in small rodent models the preclinical ex-vivo test model presented herein is both sensitive and accurate. It enabled differences in tissue adhesive strength to be quantified even for very small osseous fragments (<Ø4mm). Importantly, this model can easily be scaled to larger animals and adapted to fracture fragment fixation in human bone. The present model is also compatible with other long-term in vivo evaluation methods (i.e. in vivo imaging, histological analysis, etc.).

A Novel Class of Injectable Bioceramics that Glue Tissues and Biomaterials.

Calcium phosphate cements (CPCs) are clinically effective void fillers that are capable of bridging calcified tissue defects and facilitating regeneration. However, CPCs are completely synthetic/inorganic, unlike the calcium phosphate that is found in calcified tissues, and they lack an architectural organization, controlled assembly mechanisms, and have moderate biomechanical strength, which limits their clinical effectiveness. Herein, we describe a new class of bioinspired CPCs that can glue tissues together and bond tissues to metallic and polymeric biomaterials. Surprisingly, alpha tricalcium phosphate cements that are modified with simple phosphorylated amino acid monomers of phosphoserine (PM-CPCs) bond tissues up to 40-fold stronger (2.5⁻4 MPa) than commercial cyanoacrylates (0.1 MPa), and 100-fold stronger than surgical fibrin glue (0.04 MPa), when cured in wet-field conditions. In addition to adhesion, phosphoserine creates other novel properties in bioceramics, including a nanoscale organic/inorganic composite microstructure, and templating of nanoscale amorphous calcium phosphate nucleation. PM-CPCs are made of the biocompatible precursors calcium, phosphate, and amino acid, and these represent the first amorphous nano-ceramic composites that are stable in liquids.

Influence of gamma and electron beam sterilization on the stability of a premixed injectable calcium phosphate cement for trauma indications.

Premixed calcium phosphate cements (CPC's) are becoming the material of choice for injectable cements as a result of their effective delivery to the target implantation site. For orthopaedic use, it is of vital importance that the attributes of these CPC's are not compromised by irradiation sterilization. Therefore, the aim of this study is to determine the influence of irradiation sterilization on a range of premixed CPC's, with an emphasis on improving product shelf life through the use of optimal packaging configurations and annealing steps. Electron spin resonance (ESR) confirmed the presence of free radicals in the inorganic phase of the CPC paste following irradiation. The inclusion of a 24-h annealing step was the only successful method in reducing the degree of free radical formation. Based on the results of injectability force testing, it was revealed that an annealing step greater than 24-h significantly altered the viscosity, however; at 24-h the key attributes of the CPC paste were minimally effected. Overall, it was established that vacuum packing the CPC paste, placing the contents into a foil pouch, gamma irradiating at the minimal dose required and using an annealing step of ≤ 24-h, has the potential to extend the shelf life of the cement.

Bone Substitute Materials and Minimally Invasive Surgery: A Convergence of Fracture Treatment for Compromised Bone.

This article focuses on the understanding of the biochemistry and surgical application of bone substitute materials (BSMs) and particularly the newer calcium phosphate materials that can form a structural orthobiologic matrix within the metaphyseal components of the periarticular bone. Six characteristics of BSMs are detailed that can be used as a guide for the proper selection and application of the optimal BSM type for periarticular fracture repair. These 6 characteristics of BSMs are divided into 2 pillars. One pillar details the 3 biochemical features of BSMs and the other pillar details the 3 surgical application properties.

Long-term wear of ceramic matrix composite materials for hip prostheses under severe swing phase microseparation.

The purpose of this study was to evaluate the long-term wear performance of alumina matrix composite (AMC) heads against alumina matrix composite inserts and alumina matrix composite heads against alumina (Al) inserts with the use of a hip-joint simulator incorporating severe swing phase joint microseparation. The wear of AMC on Al produced an average wear rate of 0.61 mm3/million cycles over the 5-million-cycle test duration. The wear of AMC on AMC produced an average wear rate of 0.16 mm3/million cycles over the 5-million-cycle test duration. Both the AMC on alumina and AMC on AMC produced significantly lower wear than previously tested HIPed alumina, where an average wear rate of 1.84 mm3/million cycles was reported over 5 million cycles. The wear mechanisms and wear debris of AMC on AMC and AMC on Al were similar to those observed in previous alumina retrieval studies with stripe wear caused by intragranular fracture and wear debris consisting of predominantly uniform 10-20-nm-sized particles and a few irregular particles up to 3 microm in size.

Edge loading in third generation alumina ceramic-on-ceramic bearings: stripe wear.

Alumina ceramic-on-ceramic bearings perform exceptionally well under standard hip simulator conditions, but in vivo some retrieved bearings have shown an unusual stripe pattern of wear. We studied 16 bearings retrieved from a series of 1,588 cementless hip arthroplasties with third generation alumina ceramic-on-ceramic bearings to characterize the mechanism of stripe wear formation. None of these bearings were retrieved for bearing failure. The average wear volume was 0.4 mm(3) per year in the heads and 0.3 mm(3) per year in the liners. Mapping of wear stripes on the heads and liners showed that the majority do not occur with normal walking; instead they probably occur with edge loading when the hip is flexed, such as with rising from a chair or with climbing a high step.

Severe wear and fracture of zirconia heads against alumina inserts in hip simulator studies with microseparation.

The wear of zirconia femoral heads against alumina acetabular inserts with swing-phase microseparation was investigated in a hip joint simulator. Under mild microseparation conditions, the wear was very low, with an average wear rate of 0.05 mm(3)/million cycles reported over 5 million cycles of testing. However, under severe microseparation conditions representative of greater joint laxity, the wear rate of zirconia against alumina increased by 2 orders of magnitude, producing severe wear and, in one case, femoral head fracture. The adverse results of this study indicate that the combination of a zirconia femoral head articulating against an alumina acetabular insert is not recommended for clinical use. The results further raise concerns over the suitability of conventional simulators in evaluating the wear of ceramic hip prostheses.