Aim. This study describes the histologic changes seen with a gentamicin-eluting synthetic bone graft substitute (BGS)(1) in managing bone defects after resection of chronic osteomyelitis (cOM). Method. 154 patients with mean follow-up of 21.8 months (12–56) underwent treatment of cOM with an antibiotic-loaded BGS for defect filling. Nine patients had subsequent surgery, not related to infection recurrence, allowing biopsy of the implanted material. These biopsies were harvested between 19 days and two years after implantation, allowing a description of the material's remodelling over time. Samples were fixed in formalin and stained with haematoxylin-eosin. Immunohistochemistry, using an indirect immunoperoxidase technique, identified the osteocyte markers Dentine Matrix Protein-1 (DMP-1) and Podoplanin, the macrophage/osteoclast marker CD68, and the macrophage marker CD14. Results. The material was actively remodelled and was osteoconductive. There was evidence of osteoblast recruitment, leading to osteoid and intramembranous formation of woven and lamellar bone on the material's surface, seen most prominently in areas of well-vascularised fibrous tissue. Osteocytes in woven bone expressed the markers DMP-1 and Podoplanin. No cartilage or endochondral ossification was seen. There was a prominent (CD14+/ CD68+) macrophage response to the BSG and macrophages within reparative cellular and collagenous fibrous tissue. In biopsies taken between 4 and 5 months, there were bone trabeculae containing BGS of mainly woven but partly lamellar type. Giant cells on the surface of newly formed mineralised osteoid and woven bone expressed an osteoclast phenotype (CD68+/CD14-). In later biopsies (up to 2 years), larger bone trabeculae were seen more frequently within well-vascularised reparative fibrous tissue. The BGS was replaced with predominantly lamellar bone. One biopsy was taken from an extraosseous leak of BGS into the soft tissues, behind the distal tibia. The histology showed a heavy macrophage infiltrate, but notably no evidence of osteoid or bone formation in the material or surrounding soft tissues. Conclusion. There was clear evidence that this BGS is osteoconductive with first osteoid then woven and lamellar bone being formed. DMP-1 and podoplanin-expressing osteocytes present in woven and lamellar bone demonstrate osteoclastic bone remodelling. Increased lamellar bone was noted in later samples and bone formation was most prominent in well-vascularised areas. There was on-going remodelling of the material beyond one year. The BGS did not ossify in soft tissue. The hydroxyapatite scaffold in this material is probably responsible for its high osteoconductivity and potential to be transformed into bone

Aim. Open fractures with bone loss and skin lesions carry a high risk of infection and complication. Treatment options are usually a two-stage approach (debridement, temporary stabilization with external fixation followed by open reduction and stabilization with plate). We describe an experience for a single stage procedure with an antibiotic eluting bone graft substitute (BGS) for prophylaxis of implant-related infection. Method. Between December 2014 and January 2016 were analysed the data of twenty-six patients with open fractures (Gustilo and Anderson grade I and II) or with skin lesion and high risk of contamination and bone loss. They where treated with debridement of soft tissue, closed reduction of fracture, placement of a plate augmented with BGS eluting antibiotic (gentamicin (1) and/or Vancomicin (2)). Ampicillin and sulbactam 3g three times daily was used as systemic antibiotic prophylaxis minimum for one week. Clinical outcome and radiographic bone defect filling were assessed by blinded observers. Results. From 2014 to 2015 twelve male and fourteen female with mean age 53yrs (24–77) were treated with plate and BGS. Fracture locations were four distal femur (m:4; f: 1), four tibial plateau (m:3; f:1), one proximal humerus (f:1), seven calcaneus (m:4; f: 3), one talus (m:1), four forearm (m:3), one elbow (f:1) and two phalanx (m: 2). Follow up was fourteen month (range: 3 – 26 months). During follow-up no implant-related infection was observed. One patient developed sterile seroma, which was treated conservatively. The calcium sulphate phase of BS dissolved in all cases within 4–6 weeks. Bone ingrowth was assessed at 1, 2, 3, 6 and 12 months. On six patients large bone was treated with a revision surgery (autologous cancellous bone graft combined with BGS and antibiotic. No complications were reported. Conclusions. We suggest the application of poly therapy for the treatment of bone defects. BGS eluting antibiotic is easy to use and offers the opportunity for a one-stage procedure and might reduce the risk of implanted-related infection and allow early joint mobilization. Good early clinical outcomes were observed in almost all cases. More studies and larger series are necessary to confirm the potential for the prophylaxis of infection in the treatment of open fractures. (1): CERAMENT™|G. (2): CERAMENT™|V”

Biomaterials used in regenerative medicine should be able to support and promote the growth and repair of natural tissues. Bioactive glasses (BGs) have a great potential for applications in bone tissue engineering [1, 2]. As it is well known BGs can bond to host bone and stimulate bone cells toward osteogenesis. Silicate BGs, e.g. 45S5 Bioglass® (composition in wt.%: 45 SiO. 2. , 6 P. 2. O. 5. , 24, 5 Na. 2. O and 24.5 CaO), exhibit positive characteristics for bone engineering applications considering that reactions on the material surface induce the release of critical concentrations of soluble Si, Ca, P and Na ions, which can lead to the up regulation of different genes in osteoblastic cells, which in turn promote rapid bone formation. BGs are also increasingly investigated for their angiogenic properties. This presentation is focused on cell behavior of osteoblast-like cells and osteoclast-like cells on BGs with varying sample geometry (including dense discs for material evaluation and coatings of highly porous Al. 2. O. 3. -scaffolds as an example of load-bearing implants). To obtain mechanically competent porous samples with trabecular architecture analogous to those of cancellous bone, in this study Al. 2. O. 3. scaffolds were fabricated by the well-known foam replication method and coated with Bioglass® by dip coating. The resulted geometry and porosity were proven by SEM and μCT. Originating from peripheral blood mononuclear cells formed multinucleated giant cells, i.e. osteoclast-like cells, after 3 weeks of stimulation with RANKL and M-CSF. Thus, the bioactive glass surface can be considered a promising material for bone healing, providing a surface for bone remodeling. Osteoblast-like cells and bone marrow stromal cells were seeded on dense bioactive glass substrates and coatings showing an initial inhibited cell attachment but later a strong osteogenic differentiation. Additionally, cell attachment and differentiation studies were carried out by staining cytoskeleton and measuring specific alkaline phosphatase activity. In this context, 45S5 bioactive glass surfaces can be considered a highly promising material for bone tissue regeneration, providing very fast kinetics for bone-like hydroxyapatite formation (mineralization). Our examinations revealed good results in vitro for cell seeding efficacy, cell attachment, viability, proliferation and cell penetration onto dense and porous Bioglass®-coated scaffolds. Recent in vivo investigations [3] have revealed also the angiogenic potential of bioactive glass both in particulate form and as 3D scaffolds confirming the high potential of BGs for bone regeneration strategies at different scales. Implant surfaces based on bioactive glasses offer new opportunities to develop these advanced biomaterials for the next generation of implantable devices and tissue scaffolds with desired tissue-implant interaction

Aim. Biomaterial-associated infections (BAI) present a formidable clinical challenge. Bioactive glasses (BG) have proven highly successful in diverse clinical applications, especially in dentistry and orthopaedics. In this study, we aimed to determine the effect of three commonly used BG composition and particle sizes on cell and bacterial attachment and growth. Our focus is on understanding the changes in pH and osmotic pressure in the surrounding environment during glass degradation. Method. First, three different melt-derived glasses were characterized by analyzing particle size and glass network structure using Raman and NMR. The different glasses were then tested in vitro by seeding 4x 10. 4. cells/well (SaOS Cell line) in a 48 well plate. After a pre-incubation period of 72 hours, the different BGs and particle sizes were added to the cells and the pH value, ion release and live/dead staining was measured every hour. The effect of BG against bacteria (S. epidermidis) was analyzed after 24 and 72 hours of treatment by using XTT viability assay and CFU counting by plating out the treated aliquot agar to estimate the viable bacteria cells. Results. All three BG compositions tested showed a significant increase in pH, which was highest in BG composition 45S5 with a value of 11 compared to the other BG compositions 10 and 9 in S53P4 and 13-93 respectively. This strong increase in the pH in all BG samples tested results in a strongly reduced cell viability rate of more than 75% compared to the untreated control and 6-fold reduction in bacterial viability compared to the untreated control. The live/ dead assay also showed an increased cell viability with increasing glass particle size (i. e smallest glass particle < 25% viable cell and largest glass particle> 65% viable cell). The ion release concentration over 50 h showed an increase in sodium ions to 0.25 mol/L, calcium to 0.003 mol/L and a decrease in phosphorus. Conclusions. These results show that the composition of the bioactive glass and the choice of particle size have a major influence on subsequent applications. In addition to the different compositions of the BG, particle size and additional medium change also influence the pH and ion release, and therefore also on cells or bacteria viability. The sizes of the bioactive glass particle are inversely proportional to it. Further tests are necessary to develop custom design BG compositions, which simultaneously stimulate osteoblasts proliferation and prevent microbial adhesion

Aim. To compare a variety of commercially available bone graft substitutes (BGS) in terms of promoting adherence, proliferation and differentiation of osteoprogenitor cells. Materials and methods. A fixed number of porcine mononuclear cells obtained from cancellous bone of the proximal femur was mixed with a standard volume of BGS and then cultured for one week in media followed by two weeks in osteogenic media. BGS included commercially available β-Tricalcium Phosphate (□-TCP), highly porous β-TCP, Hydroxyapatite/Tricalcium phosphate composite, calcium sulphate (CS), Hydroxyapatite (HA), Demineralised bone matrix (DBM), polygraft, and polymers (PGA, PLGA). Staining for live/dead cells as well as scanning electron microscopy (SEM) were carried out on all samples to determine viability and cellular binding. Further outcome measures included alkaline phosphatase assays with normalisation for DNA content to quantify osteogenic potential. Negative (BGS without cells) and positive (culture expanded osteoprogenitors) control experiments were carried out in parallel to validate the results. Results. Live/dead and SEM imaging showed higher cellular viability and attachment with β-TCP than with other BGS. In the experimental setup the average alkaline phosphatase activity in nmol/ml (normalised value for DNA content in nmol/μg DNA) per sample was 657.58 (132.03) for β-TCP, 36.22 (unable to normalise) for calcium sulphate, 19.93 (11.39) for the HA/ TCP composite, 14.79 (18.53) for polygraft, 13.98 (8.15) for the highly porous β-TCP, 5.56 (10.0) for PLGA, 3.82 (3.8) and for HA. It was not possible to analyse data for either DBM or PGA. Conclusion. Under theses experimental conditions, β-TCP has apparent favourable characteristics in terms of maintaining viability of osteoprogenitor cells and allowing proliferation and differentiation. Further work will be carried out to characterise the effect that BGS have on osteoprogenitor cells

Open fractures carry a high risk of infection. Our objective was to evaluate the effect of a resorbable bone substitute (BS) (calcium sulphate and hydroxyapatite) eluting Gentamicin (Cerament™| G) in the prevention of bone infection and nonunion after open fracture and/or skin lesion. The data of patients undergoing osteosynthesis augmented with BS and Gentamicin between December 2012 and April 2015 were retrospectively analyzed from a prospectively established database. Patients were treated for open fractures grade 1 Gustilo or skin lesion with high risk of contamination. Surgical technique included initial debridement, open reduction and internal fixation (ORIF), implantation of BS and Gentamicin, soft tissue closure, and systemic antibiotic therapy for 2 weeks in all cases. Clinical outcome and radiographic bone defect filling were assessed by blinded observers. From 12/2013 to 4/2015 nine male and six female with mean age 53yrs (24–77) were treated with ORIF and BS and Gentamicin for open fractures. Fracture locations were tibial plateau (two), tibia (two), proximal humerus (one), calcaneus (four), talus (one), forearm (three), and elbow (one) distal femur (one). at final follow-up (mean 11.1 months; range 7–13). One patient developed a sterile seroma, which was treated conservatively. No post-operative infection occurred during the follow-up period. The calcium sulphate phase of BGS dissolved within 4–6 weeks in all cases. Bone ingrowth was assessed at 1, 2, 3, 6 and 12 months and new bone formation was observed at 6 months. One patient with an exposed comminuted fracture and large bone defect showed poor bone regeneration and was treated with a revision surgery (exchange of plate, autologous cancellous bone graft combined with BGS and Gentamicin. No complications were reported. The use of this bone substitute is well documented in the literature. The new product containing 175 mg gentamicin in 10 ml shows a high release of gentamicin in in-vitro testing, comparable to the elution profile of PMMA beads that some authors suggested to use to reduce the risk of infection. However, the use of this antibiotic carrier in order to prevent bone infection after open fracture has not been studied yet. In this case series 15 patients have been treated and good early clinical outcomes were observed in almost all cases. This material is highly osteoconductive and has a potential for the prophylaxis of infection in the treatment of open fractures. Further investigations and larger series are necessary to show the prophylactic effect in detail

Introduction. Primary stability is an important factor for long-term implant survival in total hip arthroplasty. In revision surgery, implant fixation becomes especially challenging due the acetabular bone defects, which are often present. Previous studies on primary stability of revision components often applied simplified geometrical defect shapes in a variety of sizes and locations. The objectives of this study were to (1) develop a realistic defect model in terms of defect volume and shape based on a clinically existing acetabular bone defect, (2) develop a surrogate acetabular test model, and (3) exemplarily apply the developed approach by testing the primary stability of a pressfit-cup with and without bone graft substitute (BGS). Materials & Methods. Based on clinical computed tomography data and a method previously published [1], volume and shape information of a representative defect, chosen in consultation with four senior hip revision surgeons, was derived. Volume and shape of the representative defect was approximated by nine reaming procedures with hemispherical acetabular reamers, resulting in a simplified defect with comparable volume (18.9 ml original vs. 18.8 ml simplified) and shape. From this simplified defect (Defect D), three additional defect models (Defect A, B, C) were derived by excluding certain reaming procedures, resulting in four defect models to step-wise test different acetabular revision components. A surrogate acetabular model made of 20 PCF polyurethane foam with the main support structures was developed [2]. For the exemplary test, three series for Defect A were defined: Native (acetabulum without defect), Empty (defect acetabulum without filling), Filled (defect acetabulum with BGS filling). All series were treated with a pressfit-cup and subjected to dynamic axial load in direction of maximum resultant force during level walking. Minimum load was 300 N and maximum load was increased step-wise from 600 N to 3000 N. Total relative motion between cup and foam, consisting of inducible displacement and migration, was assessed with the optical measurement system gom Aramis (gom GmbH, Braunschweig, DE). Results. Total relative motion increased with increasing load, with a maximum of 0.63 mm for Native, 0.86 mm for Filled, and 1.9 mm for Empty. At load stage 1800 N, total relative motion in Empty was 11.0-fold increased in comparison to Native, but could be reduced to a 3.3-fold increase in Filled. Discussion. The objective of this study was to develop a simplified, yet realistic and modular defect model which could be used to step-wise test different treatment strategies. Applicability of the developed test setup was shown by assessing primary stability of a pressfit-cup in a native, empty, and filled situation. The presented method could potentially be used as a modular test setup to compare different acetabular revision components in a standardized way. For any figures or tables, please contact authors directly