Osteoinductive bone substitutes are in their developmental infancy and a paucity of effective grafts options persists despite clinical demand. Bone mineral substitutes such as hydroxyapatite cause minimal biological activity when compared to osteoinductive systems present biological growth factors in order to drive bone regeneration. We have previously demonstrated the in-vitro efficacy of a bioengineered system at presenting growth factors at ultra low-doses. This study aimed to translate this growth factor delivery system towards a clinically applicable implant. Osteoinductive surfaces were engineered using plasma polymerisation of poly(ethyl acrylate) onto base materials followed by adsorption of fibronectin protein and subsequently growth factor (BMP-2). Biological activity following ethylene oxide (EO) sterilisation was evaluated using ELISAs targeted against BMP-2, cell differentiation studies and atomic force microscopy. Scaffolds were 3D printed using polycaprolactone/hydroxyapatite composites and mechanically tested using a linear compression models to calculate stress/strain. In-vivo analysis was performed using a critical defect model in 23 mice over an 8 week period. Bone formation was assessed using microCT and histological analysis. Finally, a computer modelling process was developed to convert patient CT images into surface models, then formatted into 3D-printable scaffolds to fill critical defects. Following EO sterilisation, there was no change in scaffold surface and persistent availability of growth factors. Scaffolds showed adequate porosity for cell migration with mechanical stiffness similar to cancellous bone. Finally, the in vivo murine model demonstrated rapid bone formation with evidence of trabecular remodelling in samples presenting growth factors compared to controls

Objectives. Deep bone and joint infections (DBJI) are directly intertwined with health, demographic change towards an elderly population, and wellbeing. The elderly human population is more prone to acquire infections, and the consequences such as pain, reduced quality of life, morbidity, absence from work and premature retirement due to disability place significant burdens on already strained healthcare systems and societal budgets. DBJIs are less responsive to systemic antibiotics because of poor vascular perfusion in necrotic bone, large bone defects and persistent biofilm-based infection. Emerging bacterial resistance poses a major threat and new innovative treatment modalities are urgently needed to curb its current trajectory. Materials and Methods. We present a new biphasic ceramic bone substitute consisting of hydroxyapatite and calcium sulphate for local antibiotic delivery in combination with bone regeneration. Gentamicin release was measured in four setups: 1) in vitro elution in Ringer’s solution; 2) local elution in patients treated for trochanteric hip fractures or uncemented hip revisions; 3) local elution in patients treated with a bone tumour resection; and 4) local elution in patients treated surgically for chronic corticomedullary osteomyelitis. Results. The release pattern in vitro was comparable with the obtained release in the patient studies. No recurrence was detected in the osteomyelitis group at latest follow-up (minimum 1.5 years). Conclusions. This new biphasic bone substitute containing antibiotics provides safe prevention of bone infections in a range of clinical situations. The in vitro test method predicts the in vivo performance and makes it a reliable tool in the development of future antibiotic-eluting bone-regenerating materials. Cite this article: M. Stravinskas, P. Horstmann, J. Ferguson, W. Hettwer, M. Nilsson, S. Tarasevicius, M. M. Petersen, M. A. McNally, L. Lidgren. Pharmacokinetics of gentamicin eluted from a regenerating bone graft substitute: In vitro and clinical release studies. Bone Joint Res 2016;5:427–435. DOI: 10.1302/2046-3758.59.BJR-2016-0108.R1

Abstract. Distraction Osteogenesis (DO) for the management of bone defects in long bones is an established technique. Problems with bone regeneration are a common occurrence and literature is full of different modalities to enhance regenerate formation and quality. Strontium Ranelate (SR) has a dual mode of action and enhances bone formation in addition to decreasing osteoclastic activity. Due to this dual mode of action as well as ease of administration in a suspension form, it makes an ideal drug in scenarios where realignment of bone homeostasis towards positive bone balance is desirable. We studied the relationship of administration of SR with rate of regenerate progression, docking site union and complications associated with bone transport in 48 patients undergoing bone transport for management of bone defects. The findings of our retrospective observation study indicated that compliant use of SR was associated with good regenerate progression, decreased problems with docking site union and decreased the need for additional interventions

Introduction. iPSCs represent a promising cell source for bone regeneration. To generate osteoprogenitor cells, most protocols use the generation of embryoid bodies (EBs). However, these protocols give rise to heterogeneous population of different cell lineage. Hypothesis. We hypothesized that a direct plating method without EB formation step could be an efficient protocol for generating a homogeneous population of osteoprogenitor cells from iPSCs. Materials & Methods. Murine iPSC colonies were dissociated with trypsin-EDTA, and obtained single cells were cultured on gelatin-coated plates in MSC medium and FGF-2. Adherent cells obtained by this direct-plating technique were termed as direct-plated cells (DPCs). DPCs were evaluated for cell-surface protein expression using flow cytometry. Expression levels of Oct-3/4 mRNA in iPSCs and DPCs were analyzed by real-time PCR. DPCs were cultured for 14 days in osteogenic medium. Osteogenic differentiation was evaluated by alkaline phosphatase (ALP) activity, real-time PCR, and alizarin red S staining. Results. Expression level of Oct-3/4 in DPCs was robustly down-regulated compared to that in iPSCs. Flow cytometry analysis revealed DPCs had similar characteristics to MSC, suggesting DPCs lost pluripotency. Moreover, the DPCs exhibited high osteogenic potential. Discussion & Conclusion. Our novel direct plating method in the absence of EB formation step could be amenable to large-scale production of osteoprogenitor cells for bone regeneration

Introduction. We evaluated the osteogenic potential of a novel biomimetic bone paste (DBSint®), made of a combination of a human demineralized bone matrix (hDBM) and a nano-structured magnesium-enriched hydroxyapatite (Mg-HA), in a standardized clinical model of high tibial osteotomy for genu varus. Methods. A prospective, randomized, controlled study was performed and thirty patients were enrolled and assigned to three groups: DBSint® (Group I), nano-structured Mg-HA (SINTlife®) (Group II) and lyophilized-bone-chips (Group III). Six weeks after surgery, computed tomography-guided biopsies of the grafts were performed. Clinical/radiographic evaluation was performed at six weeks, twelve weeks, six months, one and 2 year after surgery, in order to verify if the graft type influenced the healing rate. Results. By histomorphometry, DBSint® was shown able to promote a quick and effective bone tissue regeneration, superior to the healing process occurred in presence of SINTlife® and lyophilized bone chips. At a mean follow up of 32,59 months, no statistical differences between the groups were found, both pre-and post-operatively, according to the Knee Society Scoring System. Mean time of ostointegration was 3,9 months in the DBSint® group, 4,2 months in the lyophilized-bone group and 4,5 in the SINTlife® group. Discussion/conclusion. Orthopedic practice may be adversely affected by an inadequate bone repair that might compromise the success of surgery. Therapy for bone regeneration with DBSint® could be particularly attractive in the treatment of patients with bone defects difficult to heal, where it could shorten the period necessary for bone regeneration, due to the higher osteogenetic potential

We hypothesise that the Masquelet induced membrane used for the reconstruction of large bone defects were likely to involve mesenchymal stem cells (MSCs), given the excellent resultant skeletal repair. This study represents the first characterisation in humans of the induced membrane formed as a result of the Masquelet technique. Methods. Induced membranes and matching periosteum were harvested from 7 patients. Cytokines (BMP2, VEGF, SDF1) and cell lineage markers (CD31, CD271, CD146) were studied by immunohistochemisty. Flow cytometry was used to measure the cellularity and cellular composition. MSCs were enumerated using a colony forming unit fibroblast assay. In expanded cultures, a 96-gene array card was used to assess their transcriptional profile. Alkaline phophatase, alizarin red and calcium assays were employed to measure their in vitro osteogenic potential. Results. Membrane was more cellular(p=0.028), had more MSC phenotype(p=0.043) compared to matched periosteum. The molecular profiles were similar, except for 2-fold abundance of SDF-1 in membrane (p=0.043)compared to periosteum. Membrane and periosteum had a similar proportion of endothelial cells and CFU-F colonies; expanded MSCs from both sources were highly osteogenic. Discussion. These results indicate that the induced membrane possesses a rich source of MSC and therefore our findings support the view that the induced membrane plays an active role in bone regeneration

Introduction. Nonunion is a common and costly fracture outcome. Intricate reciprocity between angiogenesis and osteogenesis means vascular cell-based therapy offers a novel approach to stimulating bone regeneration. Hypothesis. The current study compared early and late outgrowth endothelial progenitor cell subtypes (EPCs vs OECs) for fracture healing potential in vitro and in vivo. Methods. Primary cell cultures were isolated and characterized by endothelial assays, immunosorbent assays, and multi-color flow cytometry. Co-cultures of EPC subtypes with/without primary osteoblasts (pObs) were analyzed for tube length and connectivity. In vivo, EPCs or OECs (1×10. 6. ) seeded on a gelfoam scaffold were implanted in a rat model of nonunion. Radiography was used to monitor callus formation. Results. OECs expressed more BMP-2 and less VEGF than EPCs (p<0.05). Analysis of surface markers showed decreased CD34+/CD133+/Flk-1+, CD133+ and CD45+ populations in OECs while CD34+/CD31+/Flk-1+ cells increased. pObs significantly inhibited the strong tubulogenesis of OECs while enhancing connectivity and sprout length of EPCs. In vivo, 0/6 scaffold-control and 1/5 OEC rats achieved union at 10 weeks. In comparison, all EPC rats achieved full or partial union. Discussion and Conclusion. Despite favorable tubulogenic and osteoconductive profiles of OECs, EPCs display enhanced fracture healing in vivo. Differences in CXCR4 expression and cell-mediated effects may contribute to this result

Aims

To clarify the effectiveness of the induced membrane technique (IMT) using beta-tricalcium phosphate (β-TCP) for reconstruction of segmental bone defects by evaluating clinical and radiological outcomes, and the effect of defect size and operated site on surgical outcomes.

Aims

The purpose of this study was to: review the efficacy of the induced membrane technique (IMT), also known as the Masquelet technique; and investigate the relationship between patient factors and technique variations on the outcomes of the IMT.

Introduction. Osteochondral defects (OCDs) of the talus are treated initially by arthroscopic bone marrow stimulation. For both large and secondary defects, current alternative treatment methods have disadvantages such as donor site morbidity or two-stage surgery. Demineralized bone matrix (DBM) was published for the treatment of OCDs of rabbit knees. Autologous platelet-rich plasma (PRP) may improve the treatment effect of DBM. We previously developed a goat model to investigate new treatment methods for OCDs of the talus. The aim of the current study was to test whether DBM leads to more bone regeneration than control OCDs, and whether PRP improves the effectiveness of DBM. Methods. A standardized 6-mm OCD was created in 32 ankles of 16 adult Dutch milk goats. According to a randomized schedule, 8 goats were treated with commercially available DBM (Bonus DBM, Biomet BV, Dordrecht, the Netherlands) hydrated with normal saline, and 8 were treated with the same DBM but hydrated with autologous PRP (DBM+PRP). The contralateral ankles (left or right) were left untreated and served as a control. The goats were sacrificed after 24 weeks and the tali were excised. The articular talar surfaces were assessed macroscopically using the international cartilage repair society (ICRS) cartilage repair assessment, with a maximum score of 12. Histologic analysis was performed using 5-μm sections, and histomorphometric parameters (bone% and osteoid%) were quantified on representative areas of the surface, center, and peripheral areas of the OCDs. Furthermore, μCT-scans of the excised tali were obtained, quantifying the bone volume fraction, trabecular number, trabecular thickness, and trabecular spacing in both the complete OCDs and the central 3-mm cylinders. Results. All goats recovered well and were able to bear full weight within 24 hours after surgery. The mean ICRS-score of the ankles treated with DBM was 8.0 ± 1.0, compared to a score of 8.4 ± 1.5 in the contralateral ankle (NS); those treated with DBM+PRP scored 6.9 ± 2.4, compared to 7.4 ± 2.0 in the contralateral ankle (NS). Histologic analysis showed four different patterns of healing, distributed evenly over the treatment and control groups: type 1 (n = 4), almost completely healed; type 2 (n = 11), restoration of the subchondral bone with a cystic lesion underneath; type 3 (n = 14), superficial defect with regeneration from the margins and bottom; type 4 (n = 3), no healing tendency. Histomorphometry and μCT revealed no statistically significant difference between treatment (DBM or DBM+PRP) and contralateral control or between both treatment groups in any of the parameters investigated. Conclusion. No treatment effect of DBM was found compared to control defects, and the addition of PRP was not beneficial

Aims

In this randomized study, we aimed to compare quality of regenerate in monolateral versus circular frame fixation in 30 patients with infected nonunion of tibia.