Aims

The success of anterior cruciate ligament reconstruction (ACLR) depends on osseointegration at the graft-tunnel interface and intra-articular ligamentization. Our aim was to conduct a systematic review of clinical and preclinical studies that evaluated biological augmentation of graft healing in ACLR.

Objectives

The aim of this systematic literature review was to assess the clinical level of evidence of commercially available demineralised bone matrix (DBM) products for their use in trauma and orthopaedic related surgery.

Background

Re-attachment of tendon to bone is challenging with surgical repair failing in up to 90% of cases. Poor biological healing is common and characterised by the formation of weak scar tissue. Previous work has demonstrated that decellularised allogenic demineralised bone matrix (DBM) regenerates a physiologic enthesis. Xenografts offer a more cost-effective option but concerns over their immunogenicity have been raised. We hypothesised that augmentation of a healing tendon-bone interface with DBM incorporated with autologous mesenchymal stem cells (MSCs) would result in improved function, and restoration of the native enthesis, with no difference between xenogenic and allogenic scaffolds.

Demineralized bone matrix (DBM) is a natural, collagen-based, well-established osteoinductive biomaterial. Nevertheless, there are conflicting reports on the efficacy of this product. The purpose of this study was to evaluate whether DBM collagen structure is affected by particle size and can influence DBM osteoinductivity.

Sheep cortical bone was ground and particles were divided in three fractions with different sizes, defined as large (L, 1–2 mm), medium (M, 0.5–1 mm), and small (S, < 0.5 mm). After demineralization, the three DBM samples were characterized by DTA analysis, XRD, ICP-OES, and FTIR. Data clearly showed a particle size-dependent alteration in collagen structure, with DBM-M being altered but not as much as DBM-S. The in vivo study showed that only DBM-M was able to induce new bone formation in a subcutaneous ectopic mouse model. When sheep MSC were seeded onto DBM particles before implantation, all DBM particles were able to induce new bone formation with the best incidence for DBM-M and DBM-S. Gene expression analysis performed on recovered implants supports the histological results and underlines the supportive role of MSC in DBM osteoinduction through the regulation of host cells. In conclusion, our results show a relation between DBM particle size, structural modification of the collagen and in vivo osteoinductivity. The medium particles represent a good compromise between no modification (largest particles) and excessive modification (smallest particles) of collagen structure, yielding highest osteoinduction. We believe that these results can guide researchers to use DBM particles of 0.5–1 mm size range in applications aimed at inducing new bone formation, obtaining results more comparable and reliable among different research groups. Furthermore, we suggest to carefully analyze the structure of the collagen when a collagen-based biomaterial is used alone or in association with cells to induce new bone formation.

Aims. Demineralised bone matrix (DBM) is rarely used for the local delivery of prophylactic antibiotics. Our aim, in this study, was to show that a graft with a bioactive glass and DBM combination, which is currently available for clinical use, can be loaded with tobramycin and release levels of antibiotic greater than the minimum inhibitory concentration for Staphylococcus aureus without interfering with the bone healing properties of the graft, thus protecting the graft and surrounding tissues from infection. Materials and Methods. Antibiotic was loaded into a graft and subsequently evaluated for drug elution kinetics and the inhibition of bacterial growth. A rat femoral condylar plug model was used to determine the effect of the graft, loaded with antibiotic, on bone healing. Results. We found that tobramycin loaded into a graft composed of bioglass and DBM eluted antibiotic above the minimum inhibitory concentration for three days in vitro. It was also found that the antibiotic loaded into the graft produced no adverse effects on the bone healing properties of the DBM at a lower level of antibiotic. Conclusion. This antibiotic-loaded bone void filler may represent a promising option for the delivery of local antibiotics in orthopaedic surgery. Cite this article: Bone Joint J 2016;98-B:1126–31

Objectives

We performed a systematic review of the literature to determine the safety and efficacy of bone morphogenetic protein (BMP) compared with bone graft when used specifically for revision spinal fusion surgery secondary to pseudarthrosis.

The December 2014 Research Roundup³⁶⁰ looks at: demineralised bone matrix not as good as we thought?; trunk control following ACL reconstruction; subclinical thyroid dysfunction: not quite subclinical?; establishing musculoskeletal function in mucopolysaccharidosis; starting out: a first year in consultant practice under the spotlight; stroke and elective surgery; sepsis and clots; hip geometry and arthritis incidence; and theatre discipline and infection.

A common situation presenting to the orthopaedic surgeon today is a worn acetabular liner with substantial acetabular and pelvic osteolysis. The surgeon has many options for dealing with osteolytic defects. These include allograft, calcium based substitutes, demineralised bone matrix, or combinations of these options with or without addition of platelet rich plasma. To date there are no clinical studies to determine the efficacy of using bone-stimulating materials in osteolytic defects at the time of revision surgery and there are surprisingly few studies demonstrating the clinical efficacy of these treatment options. Even when radiographs appear to demonstrate incorporation of graft material CT studies have shown that incorporation is incomplete. The surgeon, in choosing a graft material for a surgical procedure must take into account the efficacy, safety, cost and convenience of that material.

Cite this article: Bone Joint J 2014;96-B (11 Suppl A):70–2.

Summary Statement. Demineralised bone matrix augmented tendon-bone fixations in the animal model show less scar tissue and an enthesis morphology closer to the physiologic one which may lead to a more resistant repair construct. Introduction. Rotator cuff repair is one of the most common operative procedures in the shoulder. Yet despite its prevalence recurrent tear rates of up to 94% have been reported in the literature. High failure rates have been associated with tendon detachment from bone at the tendon – bone interface. Exogenous agents as biological strategies to augment tendon – bone healing in the shoulder represent a new area of focus to improve patient outcomes. Demineralised bone matrix (DBM) contains matrix bound proteins, exposed through acid demineralization step of DBM manufacture, and has long been recognised for its osteoinductive and osteoconductive properties. We hypothesised that DBM administered to the bone bed prior to the reattachment of the tendon, will upregulate healing and result in enhanced tissue morphology that more closely resembles that of a normal enthesis. An established ovine transosseous equivalent rotator cuff model was used. Methods. Following ethics approval, 10 adult wethers (18 months) were randomly allocated to control, n=4 (without DBM) or DBM, n=6 (DBM administered to bone bed) groups. The infraspinatus tendon was detached from its insertion and repaired in a transosseous equivalent fashion using PEEK suture anchors. In treatment animals 0.25cc of ovine DBM, previously prepared using a modified Urist protocol, was injected into two drill holes within the bony tendon footprint. Animals were culled at 4 weeks following surgery and processed for tissue histology and microcomputed tomography (μCT) endpoints. Results. No infection or tendon detachment following repair was noted in either group. 3D reconstructed images of μCT scans verified correct DBM and suture anchor placement. Histological images demonstrated distinct differences in tissue morphology between the two groups; however there was no evidence of the four – zoned structure characteristic of a healthy tendon bone insertion, in any specimens. In the control group specimens, the tendon midsubstance was highly disorganised with randomly arranged collagen fibres and diminutive areas of fibrocartilage. In the treatment group, large regions between tendon and bone were occupied by fibrocartilage. Within the fibrocartilage region, insertional collagen fibres appeared organised and chondrocytes were orientated in the direction of the insertional collagen fibres. Organised collagen fibre orientation within the tendon midsubstance was observed, though this was not consistent throughout all the specimens. DBM particles were resorbed and trabecular bone occupied the DBM holes. The PEEK anchors were all in direct contact with the ongrowing bone indicating good quality integration and fixation. Discussion. This study showed that DBM augmented tendon to bone repair leads to an upregulated cellular activity resulting in increased amounts of fibrocartilage between the repaired tendon and underlying bone. The upshot of this is an improved tissue organization which more closely resembles the morphology of the normal enthesis. Introduction of osteoinductive DBM at the tendon – bone interface during surgery may reduce failure rates associated with rotator cuff repair and improve clinical outcomes

The osteoinductive properties of demineralised bone matrix have been demonstrated in animal studies. However, its therapeutic efficacy has yet to be proven in humans. The clinical properties of AlloMatrix, an injectable calcium-based demineralised bone matrix allograft, were studied in a prospective randomised study of 50 patients with an isolated unstable distal radial fracture treated by reduction and Kirschner (K-) wire fixation. A total of 24 patients were randomised to the graft group (13 men and 11 women, mean age 42.3 years (20 to 62)) and 26 to the no graft group (8 men and 18 women, mean age 45.0 years (17 to 69)).

At one, three, six and nine weeks, and six and 12 months post-operatively, patients underwent radiological evaluation, assessments for range of movement, grip and pinch strength, and also completed the Disabilities of Arm, Shoulder and Hand questionnaire. At one and six weeks and one year post-operatively, bone mineral density evaluations of both wrists were performed.

No significant difference in wrist function and speed of recovery, rate of union, complications or bone mineral density was found between the two groups. The operating time was significantly higher in the graft group (p = 0.004). Radiologically, the reduction parameters remained similar in the two groups and all AlloMatrix extraosseous leakages disappeared after nine weeks.

This prospective randomised controlled trial did not demonstrate a beneficial effect of AlloMatrix demineralised bone matrix in the treatment of this category of distal radial fractures treated by K-wire fixation.

Cite this article: Bone Joint J 2013;95-B:1514–20.

We reviewed 59 bone graft substitutes marketed by 17 companies currently available for implantation in the United Kingdom, with the aim of assessing the peer-reviewed literature to facilitate informed decision-making regarding their use in clinical practice. After critical analysis of the literature, only 22 products (37%) had any clinical data. Norian SRS (Synthes), Vitoss (Orthovita), Cortoss (Orthovita) and Alpha-BSM (Etex) had Level I evidence. We question the need for so many different products, especially with limited published clinical evidence for their efficacy, and conclude that there is a considerable need for further prospective randomised trials to facilitate informed decision-making with regard to the use of current and future bone graft substitutes in clinical practice.

Cite this article: Bone Joint J 2013;95-B:583–97.

Autologous bone graft has been used in the treatment of complex bone defects for more than a century. Morbidity associated with the harvest of this bone graft has led orthopaedic surgeons to seek alternative therapies in the treatment of long bone non-unions. The aim of this study was to determine whether the use of demineralised bone matrix as a bone healing adjunct improves clinical outcomes in adult patients with long bone non-union. A systematic search was carried out of the peer-reviewed English language literature to identify all relevant studies. The search strategy returned a total of 47 studies. Five of these studies were relevant to the research question. The studies were critically assessed and where appropriate combined in a meta-analysis. 4 non-comparative studies and one comparative study were reviewed. An overall estimate of the rate of union for the five studies was 86% (95%CI: 71–94%). The one comparative study demonstrated the relative risk (RR) of healing was not significantly better than in patients treated with autologous bone graft; RR=1.03 (95%CI 0.96–1.12). There are limited data to support the use of demineralised bone matrix in the treatment of long bone non-union. Demineralised bone matrix is likely to be similarly effective to other treatments in the management of non-union. This study confirms the clinical and ethical requirements to proceed with a randomised controlled trial to test the effectiveness of this intervention

Introduction. Demineralised Bone Matrix (DBM) is widely used in Orthopaedics and dentistry as a bone graft substitute and may be used to augment bone formation in load bearing applications. In this study we examine the effect of gamma irradiation and freeze drying on the tensile strength of Demineralised Cortical Bone (DCB). Methods. Tibias were harvested from mature ewes and cut into bony strips. Demineralisation was done using 0.6M HCL and confirmed by X-ray. Specimens were washed until a pH of 7.0 +/_ 0.2 was achieved in the washing solutions. Specimens were allocated into 4 groups; group (A) non freeze dried non gamma irradiated, group (B) freeze dried non gamma irradiated, group (C) non freeze dried gamma irradiated mention the level of gamma irradiation and group (D) freeze dried and gamma irradiated. The maximum tensile force and stress were measured. Statistical analysis using the Mann-Whitney U test was carried out. Results. The Median of maximum tensile force for group (A) was 218N, group (B) was 306N, group (C) was 263N and for group (D) was 676N. Group (D) results were statistically higher (p=< 0.05) compared to group (A) and (C), while there was no statistical significance compared to group (B). Conclusion. Previously published studies suggested the possibility of using DCB as ACL graft substitute. We examined the effect of gamma radiation as the most common sterilisation technique in medical field and the freeze drying as a possible technique for long term storage on the tensile strength of the DCB. Freeze drying significantly increases the tensile strength of the DCB while gamma irradiation has no significant effect. Our results indicate that freeze dried gamma irradiated DCB can be used as a ligament substitute

The purpose of this study was to understand the effects of terminal sterilisation and residual calcium on human demineralised bone matrix (DBM) in ectopic bone formation in nude rat.

The intramuscular implantation of human DBM prepared by the Queensland Bone Bank (QBB) from four donors into eight male athymic rats was used to assess osteoinductivity. The DBM contained different levels of residual calcium and treated with or without gamma-irradiation at 11kGy. At 6 weeks post-implantation, calcium deposition was assessed by manual palpitation and radiological imaging. Tissue morphology and cellular interactions was analysed using various histological staining methods whilst protein expression of anabolic and catabolic biomarkers were examined through immunohistochemistry. All results were then analysed in qualitative, semi-quantitative and quantitative manners and tested for statistical significance.

Bone formation was observed in all specimens at the gross level. This was confirmed by histology which revealed bony capsules surrounded by soft tissue in the muscle pockets and differences in tissue components. On a cellular level, variations in osteoclast expression were found between the two groups as well as amongst individual donors through statistical analysis which resulted in an imbalance of the expression of anabolic and catabolic markers. Furthermore, a positive relationship between residual calcium and new bone formation in gamma irradiated DBM samples was found. To date, no studies have compared the effect of calcium in gamma irradiated DBM.

Our results suggest that gamma irradiation even at low doses and residual calcium may affect new bone formation. Taken together, this study stresses the importance of selecting ideal conditions for graft processing and the need to identify an optimal level of irradiation and remaining calcium levels that confers a balance between osteoinductivity and sterility.

A literature review of bone graft substitutes for spinal fusion was undertaken from peer reviewed journals to form a basis for guidelines on their clinical use. A PubMed search of peer reviewed journals between Jan 1960 and Dec 2009 for clinical trials of bone graft substitutes in spinal fusion was performed. Emphasis was placed on RCTs. Small and duplicated RCTs were excluded. If no RCTs were available the next best clinical evidence was assessed. Data were extracted for fusion rates and complications. Of 929 potential spinal fusion studies, 7 RCTs met the inclusion criteria for BMP-2, 3 for BMP-7, 2 for Tricalcium Phosphate and 1 for Tricalcium Phosphate/Hydroxyapatite (TCP/HA). No clinical RCTs were found for Demineralised Bone Matrix (DBM), Calcium Sulphate or Calcium Silicate. There is strong evidence that BMP-2 with TCP/HA achieves similar or higher spinal fusion rates than autograft alone. BMP-7 achieved similar results to autograft. 3 RCTs support the use of TCP or TCP/HA and autograft as a graft extender with similar results to autograft alone. The best clinical evidence to support the use of DBMs are case control studies. The osteoinductive potential of DBM appears to be very low however. There are no clinical studies to support the use of Calcium Silicate. The current literature supports the use of BMP-2 with HA/TCP as a graft substitute. TCP or HA/TCP with Autograft is supported as a graft extender. There is not enough clinical evidence to support other bone graft substitutes. This study did not require ethics approval and no financial support was received

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

Background: A common problem achieving lumbar spinal fusion is developing a pseudarthrosis. The current gold standard in achieving fusion is the use of autograft from pelvis or posterior elements of the spine. However the potential limitations of insufficient quantity and donor site morbidity have led to the use of bone graft alternatives such as DBM which contains osteoinductive BMPs. Aims & Methods: A prospective randomized control trial comparing the effectiveness of Demineralised Bone Matrix (DBM Putty)/autograft composite with autograft in lumbar postero-lateral or 270 degree spinal fusion. 35 patients were required for the study. They were randomized to have DBM and autograft on one side of the posterior approach and autograft alone on other side of the same approach. Patients were followed up with interval radiographs for total of 24mons. To date 32 patients have been recruited and with an average follow up a15.3 months. The mineralization of fusion mass lateral to the instrumentation on each side was graded as Absent, Mild (< 50%), Moderate (> 50%) or Complete fusion (100%). The assessment was made by independent orthopaedic consultant and a musculoskeletal radiologist who were blinded to graft assignment. Results: The sex distribution was 17:15 male to females with a mean age of 55.2 (21–87years) and an average follow up of 15.3mons (3–24mons). 50% of patients had single level fusion and the remainder had more than one level fusion. At 12months, on the side of DBM 28% had complete fusion, 65% had moderate fusion, and 7% had no fusion mass. During the same period on the other side (non DBM side) approx 25% did not show any sign of fusion. There was no correlation with number of levels, age or sex. Conclusions: Osteoinductive properties of DBM would appear to enhance the consolidation of the lumbar spinal fusion. DBM reduces the amount of harvested autograft graft and also minimises the morbidity of donor site complications

Research in to tendon-bone healing techniques focus on increasing bone growth at the interface such as cell or growth factor (e.g. BMP-2) augmentation. Demineralised bone matrix (DBM) is osseoinductive and is in use clinically. Hypothesis: DBM augmentation of a healing tendon-bone interface will result in improved function at 3, 6, 9 and 12 weeks, and a morphology that more closely resembles that of a normal enthesis at 12 weeks. Materials and methods: An ovine patellar tendon model was used. 19 skeletally mature ewes were allocated to the control group or DBM group. In both groups the patellar tendon was detached, and following tibial tubercle osteotomy, was re-attached using 3 suture anchors. In the DBM group a piece of DBM was placed between the tendon and bone. 2 animals were sacrificed at 6 weeks and 6 animals at 12 weeks. Animals underwent force plate analysis at 3, 6, 9 and 12 weeks. The tendon-bone interface length which was fibrous or fibrocartilaginous, and the area of fibrocartilage, mineralised fibrocartilage and new bone was quantified. Results: 3 control group animals (33%) failed within 6 weeks. None failed in the DBM group. The DBM group was significantly better than the control at all time points (p< 0.05). DBM produced a significantly more fibrocartilaginous enthesis than the control group (p< 0.05). Controls were significantly more fibrous than the DBM group (p< 0.05). DBM produced significantly more fibrocartilage (p< 0.05), and mineralised fibrocartilage (p< 0.05). Discussion: 33% of the control group failed within 6 weeks, whilst no failures were observed in the DBM group. DBM animals mobilised earlier and had significantly better function at all time points. Histologically, the DBM group showed a more mature direct type enthesis at earlier time points. Conclusion: DBM augmentation of a healing tendon-bone interface enhances functional and morphological recovery at earlier time points

We reviewed 15 consecutive patients, 11 women and four men, with a mean age of 48.7 years (37.3 to 62.6), who between July 2004 and August 2007 had undergone percutaneous sacroiliac fusion using hollow modular anchorage screws filled with demineralised bone matrix.

Each patient was carefully assessed to exclude other conditions and underwent pre-operative CT and MR scans. The diagnosis of symptomatic sacroiliac disease was confirmed by an injection of local anaesthetic and steroid under image intensifier control.

The short form-36 questionnaire and Majeed’s scoring system were used for pre- and post-operative functional evaluation. Post-operative radiological evaluation was performed using plain radiographs.

Intra-operative blood loss was minimal and there were no post-operative clinical or radiological complications. The mean follow-up was for 17 months (9 to 39). The mean short form-36 scores improved from 37 (23 to 51) to 80 (67 to 92) for physical function and from 53 (34 to 73) to 86 (70 to 98) for general health (p = 0.037). The mean Majeed’s score improved from 37 (18 to 54) pre-operatively to 79 (63 to 96) post-operatively (p = 0.014). There were 13 good to excellent results. The remaining two patients improved in short form-36 from a mean of 29 (26 to 35) to 48 (44 to 52). Their persistent pain was probably due to concurrent lumbar pathology.

We conclude that percutaneous hollow modular anchorage screws are a satisfactory method of achieving sacroiliac fusion.

We have used human Embryonic Stem cells (hESC) and human Mesenchymal Stem Cells (hMSC) in rat models of bone repair in order to assess the efficacy of these cells for treatments of trauma and skeletal diseases. Graft survival is considered to be of key importance to efficacy of these treatments. Therefore the aim of this study was to develop a technique for identifying implanted cells in histological preparations without the need for genetic engineering of the implanted cells. Methods: In our experiments hES and hMSC were pre-differentiated during cell culture towards the osteoblast lineage, and then implanted in a Demineralised Bone Matrix (DBM) carrier into an experimentally created full thickness calvarial bone lesion. The animals were sampled seven days and fourteen days after implantation into either immune deficient (RNU-Foxn1rnu) or immune competent (wild type) Sprague Dawley rats. Fluorescent In Situ Hybridisation (FISH) using whole human genome probes identified the human cells within the host lesion site. Results: Our results have demonstrated that hESC and hMSC derived cells survive in both immune competent (wild type) and immune compromised (nude) animals for the initial seven days post implantation. On the other hand while both the hESC and hMSC derived cells are capable of surviving for at least 14 days in immune compromised animals they do not survive for this period of time in immune competent animals. Discussion: It appears that the cell/DBM graft is not rejected within seven days even when exposed to the wild type hosts T cell response. However longer term survival required an immune deficient model that is lacking in a T cell response. This data points to interesting future studies regarding which components of the host response are responsible for xenogenic stem cell implant rejection