Aims: The treatment of long bone non-union now days finds its gold standard in autologous bone grafting. Since this technique is affected by a high morbidity rate of the donor site, many studies tried to find valid alternatives to this procedure, but during the last few years the advances made in tissue engineering techniques opened new frontiers. In this study BMPs/AGFs were used in posttraumatic long bone non-union and osseous defects to test their clinical and radiological effectiveness in order to find a valid alternative to autologous bone grafting. Methods: The cases selected can be divided in two groups. Group A: Patients affected by long bones Non Union, 9 months minimum duration, who are judged not to heal by simply changing the osteosynthesis device. Group B: Patients with non neoplastic, posttrauma or post-resection osseous defects of a critical size that will probably not heal using traditional surgical techniques or for which such techniques are considered to be unsuitable. Moreover, the overall recruitment period is 3 years during which 40 patients/year will be enrolled up to a total of 120 cases; half of these will be treated with rhBMP-7 and the other half with PRP. Results: Only 66 patients can be evaluated as they have completed the minimum follow-up period of 9 months, 35 of whom have been treated with rhBMP-7 and 31 treated with PRP. Advanced results indicate the RX Healing rate was 85% for BMP-7 and 68% for PRP with a Clinical Healing rate of 88.5% and 68%; therefore a higher efficiency of BMP-7 over PRP was found, confirmed by a significant failure rate of 15% versus 32,3% between BMP-7 and PRP, respectively. Conclusions: According to our results, the use of growth factors showed a similar effectiveness to autologous bone grafting with better tolerability, moreover, a relevant difference in healing/failure rate between rhBMP-7 and PRP is observed

Aims. To explore the efficacy of extracorporeal shockwave therapy (ESWT) in the treatment of osteochondral defect (OCD), and its effects on the levels of transforming growth factor (TGF)-β, bone morphogenetic protein (BMP)-2, -3, -4, -5, and -7 in terms of cartilage and bone regeneration. Methods. The OCD lesion was created on the trochlear groove of left articular cartilage of femur per rat (40 rats in total). The experimental groups were Sham, OCD, and ESWT (0.25 mJ/mm. 2. , 800 impulses, 4 Hz). The animals were euthanized at 2, 4, 8, and 12 weeks post-treatment, and histopathological analysis, micro-CT scanning, and immunohistochemical staining were performed for the specimens. Results. In the histopathological analysis, the macro-morphological grading scale showed a significant increase, while the histological score and cartilage repair scale of ESWT exhibited a significant decrease compared to OCD at the 8- and 12-week timepoints. At the 12-week follow-up, ESWT exhibited a significant improvement in the volume of damaged bone compared to OCD. Furthermore, immunohistochemistry analysis revealed a significant decrease in type I collagen and a significant increase in type II collagen within the newly formed hyaline cartilage following ESWT, compared to OCD. Finally, SRY-box transcription factor 9 (SOX9), aggrecan, and TGF-β, BMP-2, -3, -4, -5, and -7 were significantly higher in ESWT than in OCD at 12 weeks. Conclusion. ESWT promoted the effect of TGF-β/BMPs, thereby modulating the production of extracellular matrix proteins and transcription factor involved in the regeneration of articular cartilage and subchondral bone in an OCD rat model. Cite this article: Bone Joint Res 2024;13(7):342–352

Periosteum is a specialized connective tissue that surrounds bone, containing progenitor cells that develop into osteoblasts. The osteo-progenitor cells along with growth factors, such as BMPs, play critical role in development, reconstruction and bone formation. Aim: to evaluate the expression of BMPs in human periosteum and in different subrgroups, including different donor sites, gender, and smoking habits. Gene expression of BMPs 2,4,6,7 was performed in 60 periosteal samples using quantitative RT-PCR. Samples were obtained from 32 men/28 women, 22 smokers/38 non-smokers, 29 lower/31 upper extremities. BMP2 gene expression was significantly higher (median: 12.02, p< 0.05) than the mRNA levels of BMPs 4,6,7 (median: 1.36, 2.55, 0.04) in all samples. BMP2 mRNA levels were higher in large compared to small bones (median: 13.4 vs 9.48), while BMPs 4,6,7 gene expression was similar (1.3 vs 1.4, 2.7 vs 2.1, 0.04 vs 0.03, respectively). In lower extremities, BMPs mRNA levels were higher than in the upper; the same was detected in non-smokers versus smokers group (BMPs2,4,6,7: 13.9 vs 1.5, 3.1 vs 0.048, 8.7 vs 1.06, 1.6 vs 0.026, respectively). mRNA transcripts of BMP2 were higher in men than women (median: 13.1 vs 10.8). In our study, BMP2 expression is characteristically higher than that of BMP4, BMP6 and BMP7, highlighting the critical role that BMP2 plays in bone homeostasis. Furthermore, the elevated expression of BMP2 in men towards women, and of all BMPs of the lower extremity samples indicate the effect of hormones and mechanical factors in periosteal BMPs gene regulation; while the effect of smoking is reflected in the reduction of BMPs expression in smokers

Accidents, osteoporosis or cancer can cause severe bone damage requiring grafts to heal. All current grafting methods have disadvantages including scarcity and infection/rejection risks. An alternative is therefore needed. Hydroxyapatite/calcium carbonate (HA/CC) scaffolds mimic the mineral bone composition but lack growth factors present in auto- and allografts, limiting their osteoinductive capacity. We hypothesize that this will increase the osteogenicity and osteoinductivity of scaffolds through the presence of growth factors. The objectives of this study are to develop and mass-produce grafts with enhanced osteoinductive capacity. HA/CC scaffolds were cultured together with umbilical cord mesenchymal stem cells in bioreactors so that they adhere to the surface and deposit growth factors. Cells growing on the scaffolds are confirmed by Alamar blue assays, SEM, and confocal microscopy. ELISA and IHC are used to assess the growth factor content of the finished product. It has been confirmed that cells attach to the scaffolds and proliferate over time when grown in bioreactors. Dynamic seeding of cells is clearly advantageous for cell deposits, equalizing the amount of cells on each scaffold granule. Hydroxyapatite/calcium carbonate scaffolds support cell-growth. This should be confirmed by further research, including Quantification of BMPs and other indicators of osteogenic differentiation such as Runx2, osteocalcin and ALP is pending, and amounts are expected to be increased in enhanced scaffolds and in-vivo implantation

Bone morphogenetic proteins are low molecular weight proteins which have extensive similarity in structure and function to the transforming growth factor beta factors. They bind receptors on the surface of osteoprogenitor stem cells and activate intracellular signal transduction cascades resulting in the osteoblastic differentiation of pluripotential stem cells. Bone morphogenetic proteins (BMP) are being increasingly used in orthopaedic surgery including spinal fusion. These small molecules are capable of inducing bone formation when delivered in the appropriate concentration and on the appropriate scaffold. Recombinant BMP usually is combined with an osteoconductive carrier to form a composite graft. The osteoconductive carrier not only supports cellular adhesion but restricts the diffusion of these soluble factors away from the fusion site increasing local concentration of BMP. There is currently no consensus as to the ideal carrier but the optimal carrier may be dependent upon the specific clinical application for which it is used. In addition osteogenic cells that are able to respond to these osteoinductive signals must also be present for a successful spinal fusion to occur. Not all BMPs are equally effective. Over 15 BMPs have been identified and there are currently only two Food and Drug Administration (FDA)-approved BMPs (BMP-2 with a full PMA approval and BMP-7 with an HDE approval). Recombinant BMPs have been used successfully in anterior lumbar interbody fusions. Multiple animal studies have shown recombinant human BMP to be superior to autograft in the cervical, thoracic and lumbar spine, while human clinical trials have also shown recombinant human BMP-2 to be superior to autograft for anterior fusion. Similarly, multiple animal studies and clinical trials have shown that recombinant human BMPs result in equivalent or superior fusion rates for posterior spinal fusion compared to autograft. The use of BMPs may obviate the need for decortication and overcome the negative effects of nicotine and anti-inflammatories. In all studies, the concentration of BMPs necessary to produce successful spinal fusion was substantially greater than physiological levels, raising several potential safety concerns including bony overgrowth and bone formation which may lead to neural compression or unintended extension of the fusion. There are also the risks of local toxicity and a host immunologic response. These potential complications related to off-label use of BMPs need to be understood. For this reason, it is essential to determine the appropriate dose for each clinical application and develop efficient carrier systems. There are economic concerns associated with the use of this new technology. A single treatment of recombinant human BMP is expensive but may be cost effective if clinical outcomes are improved or other costs are avoided. The increased cost of BMP may offset the complications associated with harvesting autograft bone. When used properly, these molecules have the potential to eliminate the need for iliac crest bone graft harvest and improve the speed and success of spinal fusion

Introduction: Periosteum is a tissue with pluripotential mesenchymal cells (MSCs). During fracture repair several growth factors are released from periosteum, including bone morphogenetic proteins (BMPs), which induce the differentiation of bone marrow stromal cells towards the osteoblastic lineage, therefore increasing the pool of mature bone forming cells and enhance the differentiated function of osteoblasts. The purpose of our study is to evaluate the expression of periosteal BMPs mRNA from fracture samples, collected within 24 hours of fracture and to compare it with BMPs expression from periosteal samples of normal (non-fractured) bones. Materials and Methods: Periosteum samples were collected from 25 patients with recent fracture (during the past 24 hours) (age: 12–80) and 25 individuals without fracture (age: 10–73). BMPs (BMP2, BMP4, BMP6) mRNA levels were analysed by Real Time RT-PCR by using the Light Cycler machine and PBGD as a housekeeping gene. Results: BMP2 mRNA levels were significantly higher (p< 0.05) in normal samples (median:12.15) than in fracture (median:4.39). BMP6 and BMP4 mRNA expression followed similar pattern to that of BMP2 but in significant lower levels. In normal samples, BMP4 mRNA median levels were 1.99, while in fracture samples the levels were significantly lower (median:0.35), (p< 0.05). BMP6 mRNA levels were also higher in normal samples (median:2.21) than in fractures (median:1.87) (p> 0.05). Furthermore, the decrease of BMPs mRNA levels in fracture samples was higher for BMP4 followed by BMP2 and BMP6. Discussion: Our results indicate high BMP2 mRNA levels expressed from periosteal cells. In recent fractures there is a significant reduction of BMP2 compared to normal samples; however, the expression of BMP2 remains more elevated in comparison to the other BMPs highlighting the potential role of BMP2 at the initiation of healing process of fractures. BMP6 and BMP4 expression was similar among normal periosteal cells while levels of BMP6 were higher than BMP4 in fracture periosteal cells. The suppression of BMP6 expression was minimum and less significant than BMP2 and BMP4 suppression indicating the potential role of BMP6 at the early stages of MSCs differentiation in periosteum. On the other hand, BMP4 remains in low levels in any confrontation and seems that plays a minor role in early healing process of fracture. BMPs are considered to play central role in fracture response and bone remodelling but further investigation has to be done as much in their correlation and toward other growth factors as in their expression levels during bone fracture repair process

Background: Autologous bone graft remains the “gold standard”, but the associated morbidity and finite supply of tissue has resulted in surgeons seeking methods of enhancing healing with bone graft substitutes and bone morphogenic proteins (BMPs). There are 54 bone graft substitutes and 2 BMPs currently on sale in the UK. The aim of this study was to review the published clinical evidence in support of their use in spinal surgery. Methods: The 19 manufacturers of bone graft substitutes and BMPs were contacted asking for details of their products on the market. A systematic literature review was conducted using the ISI Web of Knowledge, EMBASE (1980–2008) and OVID databases. Publications providing clinical data were classified according to the hierarchy of clinical evidence published in the Journal of Bone and Joint Surgery Am in 2003. Level I evidence is a prospective randomised control trial with definitive results to support the use of an intervention in a clinical setting. Level V evidence is isolated case reports. A Grade of recommendation A-i was accredited to each product to assist the clinical spinal surgeon in making decisions on which product to use based on the evidence in the literature. Results: 102 clinical studies found, most of which were level IV or V evidence. Both BMPs, InductOS® and OP-1® have Level I papers and have Grade A recommendation. There is very poor evidence for the use of bone graft substitutes in spinal surgery with no products receiving Grade A recommendation and only 6 out of 54 products having Grade B recommendation. Conclusions: There is a lack of evidenced based clinical data for the use of bone graft substitutes in spinal surgery. Regarding BMPs there is good evidence for the use of these products in spinal surgery but surgeons must be aware of the reported complications

In the treatment of bone non-unions an alternative to bone autografts is the use of bone morphogenetic proteins (BMP-2, BMP-7) with powerful osteoinductive and osteogenic properties. In clinical settings, BMPs are applied using absorbable collagen sponges. Supraphysiological doses are needed and major side effects may occur as induce ectopic bone formation, chronic inflammation and excessive bone resorption. In order to increase the efficiency of the delivered for BMPs we designed cryostructured collagen scaffolds functionalized with hydroxyapatite, mimicking the structure of cortical bone (aligned porosity, anisotropic, ANI) or trabecular bone (random distributed porosity, isotropic, ISO). We hypothesize that anisotropic structure would enhance osteoconductive properties of the scaffolds increasing rhBMP-2 regenerative properties. In vitro, both scaffolds presented similar mechanical properties, rhBMP-2 retention and delivery capacity. For in vivo testing, a rat femoral critical size defect model was created. Four groups were assessed depending on the implant applied to the bone defect: ISO, unloaded isotropic sponge; ISO-BMP, isotropic sponge loaded with 5 μg of hrBMP-2; ANI, unloaded anisotropic sponge; and ANI-BMP, anisotropic sponge loaded with 5 μg of hrBMP-2. Regeneration was allowed for 10 weeks. X-ray, μCT, biomechanical testing and histology were used to evaluate repair. Independently of their structure, sponges loaded with rhBMP-2 demonstrate increased bone volume, and biomechanical properties than their controls (p<0.01 and p<0.05 respectively). Globally, ANI-BMP group demonstrated better bone regeneration outputs with increased defect bridging (p<0.05 when compared ANI-BMP vs ISO-BMP groups). In conclusion, anisotropic cryostructured collagen scaffolds improve the efficiency of rhBMP-2 in bone regeneration

Avascular necrosis (AVN) is a disorder leading to femoral head (FH) destruction, while BMPs are known for their osteogenic ability. In this study we analyzed BMP-2, BMP-4, BMP-6 and BMP-7 expression at the RNA and protein level in the normal and necrotic sites of the FHs. Quantitative RT-PCR for BMP-2,-4,-6,-7 genes was performed in samples from the normal and necrotic sites of 52 FHs with AVN. Protein levels of BMP-2,-4,-6 were estimated by Western Blot analysis. Statistical analysis was performed using the t-test (p< 0.05). BMP-2 and BMP-6 mRNA levels were higher in the normal than the necrotic site (BMP-2 and BMP-6, normal vs necrotic: 16.8 vs 7.5 and 2 vs 1.66, respectively). On the contrary, BMP-4 mRNA levels were higher in the necrotic (1.2) than the normal site (0.97), while BMP-7 mRNA levels were low in both sites. At the protein level, BMP-2 expressed higher in the normal (0.63) than the necrotic region (0.58), while BMP-4 and BMP-6 detected at higher levels in the necrotic site (BMP-4 and BMP-6, normal vs necrotic: 0.51 vs 0.61 and 0.52 vs 0.57, respectively). Different mRNA levels between the normal and necrotic site, as well as discrepancies between the gene and protein BMPs expression levels suggest a different regulation mechanism between the two regions. Better understanding of the expression pattern of BMPs could lead to a more successful use of these molecules in the prevention and treatment of AVN

*Winner of ISFR Young Investigator Award. Purpose: The aim of this study was to compare the temporal expression pattern of factors related to cartilage and bone formation and endochondral ossification during standard and delayed bone healing for a more in-depth understanding of the molecular basis of disturbed bone healing and to elucidate suitable timing for substitution of factors to stimulate the healing process. Methods: A tibial osteotomy was performed in two groups of sheep (n=30 each) and stabilized with either a rigid external fixator leading to standard healing or with a mechanically critical one leading to delayed healing. Hematoma/callus tissue was harvested 4, 7, 14, 21 and 42 days postop. qPCR was employed to determine the expression patterns of BMPs and other molecules. Results: Gene expressions of BMP2, BMP4, BMP7, Noggin, MMP9 and MMP13 were distinctly lower in the delayed compared to the standard healing group at several time points from day 14, whilst no differential gene expression of Coll II and Coll X was found between both groups. Among the BMPs, BMP7 showed the most markedly differential expression. The first evident difference in BMP7 expression between both groups was found at day 14 suggesting that exogen substitution in the context of a therapeutic approach should be postponed. The differential expression pattern of both MMP9 and MMP13 suggests that there might be a failure or delay in endochondral ossification in delayed bone healing. Conclusion: Downregulation in gene expression of osteogenic BMPs and cartilage matrix degrading MMPs may account for a considerable delay of bone healing

BMPs, among which BMP-7 or OP-1, unlike several growth factors involved in new bone formation, are the only proteins able to start the whole process. That is BMPs are the only factors with osteoinduction ability. Contrary to other growth factors, BMPs on the market are drugs. RhOP-1, carried by collagen type 1, is the first osteo-inductive drug approved in the world for the clinical usage: in long-bone non-unions in US, Australia and Canada and in tibia non-unions, recalcitrant to autograft, in Europe (Osigraft). We report data related to a retrospective observation on some patients treated in Italy with rhOP-1. 90 patients (66 with long-bone non-union diagnosis, 8 with delayed union, 7 with bone defect /bone cyst and the remaining with other pathologies) are reported, and efficacy results are showed on 60 patients with follow-up > 6 months. Radiographic analysis shows that rhOP-1 is effective in 86,6% of patients. Unions have been reported in 34,8% at 4–5 months, and in 69,1% at 6–8 months. Failure: 8/60 (13,4%). No adverse event has been reported. These data are similar to those reported in literature in randomised and not randomised studies

The clinical application of bone morphogenetic proteins (BMPs) offers solutions to many challenging problems in orthopaedics. However, a practical clinical problem is to obtain a controlled release of the BMPs. The attachment of heparin to biomaterials may result in an appropriate matrix for the binding, and sustained release of BMPs. Binding of growth factors to heparin stabilizes these growth factors, protects them from proteolytic degradation, and prolongs the half-life of BMPs in culture media 20-fold. We created a carrier based delivery system with a localized sustained release by loading a tricalciumphosphate/hydroxyapatite (TCP/HA) bone substitute coated with cross-linked collagen and heparin, with BMP-7. TCP/HA granules (BoneSave. ™. , Stryker Orthopaedics) were coated with collagen, and subsequently the collagen was cross-linked in the presence (TCP/HA-Col-Hep) and absence (TCP/HA-Col) of heparin. BMP-7 was loaded onto the coated TCP/HA granules. Morphology of the coated collagen with and without heparin, and release kinetics of BMP-7 from the granules were analyzed. TCP/HA granules without coating were used as controls. Analysis showed a highly porous collagen network on both TCP/HA-Col and TCP/HA-Col-Hep granules. Immersion of the granules in BMP-7 solution, resulted in the binding of 54±3% (62.9±5.4 ng BMP-7/mg granule) to the TCP/HA granules, 64±8% (69.0±9.6 ng BMP-7/mg granule) to the TCP/HA-Col granules, and 78±1% (92.9±4.8 ng BMP-7/mg granule) to the TCP/HA-Col-Hep granules. TCP/HA granules showed a burst release of BMP-7 within the first 4 h. TCP/HA-Col granules showed an initial burst release, followed by a more gradual release. In contrast, BMP-7 release from the TCP/HA-Col-Hep granules was sustained up to 21 days. The sustained delivery system for BMP-7 developed in this study may provide a powerful tool for bone regeneration. This system could probably also be applied to deliver multiple growth factors that have affinities for heparin, which could for instance synergistically enhance osteogenesis by increasing vascularity

As cartilage has poor intrinsic repair capacity, in vitroexpansion of human articular chondrocytes (HACs) is required for cell-based therapies to treat cartilage pathologies. During standard expansion culture (i.e. plasma osmolarity, 280 mOsm) chondrocytes inevitably lose their specific phenotype and de-differentiate, which makes them inappropriate for autologous chondrocyte implantation. It has been shown that physiological osmolarity (i.e. 380 mOsm) increases collagen type II (COL2) expression in vitro, but the underlying molecular mechanism is unknown. Transforming growth factor beta (TGFβ) super family members are accepted key regulators of chondrocyte differentiation and known to stimulate COL2 production. In this study we aimed to elucidate the role of TGFβ superfamily member signalling as a molecular mechanism potentially driving the COL2 expression under physiological (380 mOsm) culture conditions. HACs from OA patients (p1) were cultured in cytokine-free medium of 280 or 380 mOsm, under standard 2D in vitroconditions, with or without lentiviral TGFβ2 knockdown (RNAi). Expression of TGFβ isoforms, BMPs and chondrocyte marker genes was evaluated by QPCR. TGFβ2 protein secretion was evaluated using ELISA and bioactivity was determined using an established reporter cell line. Involvement of BMP signaling was investigated by culturing OA HACs (p1) in the presence or absence of dorsomorphin (10 µM). Physiological osmolarity increased TGFβ2 and TGFβ3 mRNA expression, TGFβ2 protein secretion as well as general TGFβ activity by 380 mOsm. Upon TGFβ2 isoform-specific knockdown COL2 mRNA expression was induced. TGFβ2 RNAi induced expression of several BMPs (e.g. BMP2,-4,-6) and this induction was enhanced in culture conditions with physiological osmolarity. Dorsomorphin inhibited physiological osmolarity induced COL2 mRNA expression. TGFβ2 knockdown under 380 mOsm increases COL2 expression in human osteoarthritic chondrocytes in vitromost likely through a regulatory feedback loop via BMP signaling, which is involved in osmolarity-induced COL2 expression. Future studies will further elucidate the BMP-mediated regulatory feedback loop after TGF β2 knockdown and its influence on COL2 expression

Osteoarthritis is a slowly progressive disease which includes the intervention of several cytokines and macrophage metalleinoproteinases reaction, leading to the degradation of the local cartilage but also having an impact on the serum acute phase proteins (APPs). Subsequently, biomarkers seem to be essential to estimate its progression and the need for any surgical intervention such as total arthroplasty, but also can be used as therapeutic agents. Recently, among APPs, fetuin-A drew attention regarding its possible anti-inflammatory role in animal models but also as a therapeutic agent in the inflammatory joint disease in clinical trials. The purpose of this study is to investigate the possible attenuating role of the intra-articular administration of Fetuin-A in post-traumatic induced secondary osteoarthritis in rats, and also its effect on the systematic levels of IL-2,4,7, BMPs 2,4,7, CRP and Fetuin-A. 30 male Sprague Dawley rats were separated in two groups where post-traumatic osteoarthritis was induced surgically by Anterior Cruciate Ligament Transection and the transection of the Medial Collateral Ligament of the right knee. In the Control Group, only surgical intervention took place. In Fetuin Group, along with the induction of osteoarthritis, a single dose of bovine fetuin was administrated intra-articularly intra-operatively in 5 and 8 weeks of the experimental protocol. Both groups were examined for 8 weeks. The levels of interleukins, bone morphogenetic proteins, Fetuin-A and C-Reactive Protein were evaluated by ELISA of peripheral blood in three time periods: preoperatively, 5 and 8 weeks post-operatively. Knee osteoarthritic lesions were classified according to Osteoarthritis Research Society International Grading System and Modified Mankin Score, by histologic examination. IL-2 levels were significantly decreased in the Fetuin Group. No statistical difference was signed on the levels of IL-7, BMP-2,4,7 and Fetuin-A between the two groups. CRP levels were significantly increased in the Fetuin Group in 5 weeks of the experiment. Fetuin Group signed better scores according to the OARSI classification system and Modified Mankin Score, without any statistical significance. Intra-articular administration of Fetuin-A restrictively affected the progression of post-traumatic arthritis in rats, as only the levels of IL-2 were decreased as well as limited osteoarthritic lesions were observed on the Fetuin Group

INTRODUCTION. The hip arthroplasty implant is currently growing up both in orthopedic and trauma practice. This increases the frequency of prosthesis revision due to implant loosening often associated with periprosthetic osteolysis that determine the failure and lead to a loss of bone substance. Nowadays there are numerous biotechnologies seeking to join or substitute the autologous or omologous bone use. These biotechnologies (mesenchymal stromal cells, growth factors and bone substitutes) may be used in such situations, however, the literature doesn't offer class 1 clinical evidences in this field of application. MATERIALS AND METHODS. We performed a literature review using the universally validated search engines in the biomedical field: PubMed / Medline, Google Scholar, Scopus, EMBASE. The keywords used were: “Growth Factors”, “Platelet Rich Plasma”, “OP-1”, “BMP”, “BMP-2”, “BMP-7”, “Demineralized Bone Matrix”, “Stem Cell”, “Bone Marrow”, “Scaffold”, “Bone Substitutes” were crossed with “hip”, “revision”, “replacement” / “arthroplasty”, “bone loss” / “osteolysis.”. RESULTS. The search led to 321 items, of these were considered relevant: as regards the growth factors 21 articles related to in vivo animal studies and 2 articles of human clinical use of BMPs and 1 single article on the use of PRP; as regards the mesenchymal stromal cells 2 items of application in animals; as regards the use of bone substitutes we have analyzed a review of this application. DISCUSSION. The use of biotechnologies in hip prosthetic revisions has produced conflicting results: autologous growth factors (PRP) have definitely been proven effective in maxillofacial surgery, in animal studies the results of BMPs are inconsistent with articles that validate their use and others that don't recommend it. Clinical application has demonstrated, today, the limited use of BMP-7 in revisions with even an increased risk of early re-mobilization, PRP appears to be rather effective only in the early stages of peri-prosthetic osteolysis. The mesenchymal cells can increase the chances of recovery and integration of the grafts but an important variable is the number of cells that are still alive after the impaction of the graft which affects their vitality. The bone substitutes appear to be safe and very useful, particularly if applied in order to implement the omologous bone, which is still the most scaffolds used in this surgery. CONCLUSIONS. The systematic review of the literature has shown an important lack of clinical studies regarding the use of biotechnologies for prosthetic revisions. It is therefore difficult to draw guidelines that regulate the application, prospective randomized clinical studies are therefore needed to validate its effectiveness

Fracture healing involves many local and systemic regulatory factors. Progress in identifying signaling events downstream has been made with the discovery of a novel family of proteins, the Smad, as TGF-ß/activins/BMPs signal transducers. Smads are the vertebrate homologs of Mad (Mothers against decapentaplegic) gene from Drosophila and Sma genes from Caenorhabditis elegans. Smad-1, -2, -3, -5, -8 and -9 belong to the receptor-regulated class (R-Smad) which are activated by the TGF-ß type I and II receptors, forming heteromers with the common-mediator class (Co-Smad): Smad-4. Smad-6 and -7 (Anti-Smad) perform a negative regulatory or balancing role. Smad-2 and -3 regulate TGF-ß/activin effects, whilst Smad-1 and -5 work with BMPs. This study investigated the expression and localization of Smad proteins (Smad 1–6) and BMP-4 and -7 during fracture healing. Eighteen 3-month old female CD-COB rats were used. A standard closed fracture was made in the mid-shaft of right femur using a 3-point bending device. The left limb served as the non-fracture control. The rats were divided into 3 groups (6 per group) and sacrificed at day 3, 10 and 28 after fracture. The femurs were harvested, fixed in buffered formalin for 48 hours and decalcified with 10% formic acid-formalin solution. The decalcified tissues were embedded in paraffin and 5μm sections were cut onto silane-coated slides. Representative slides from each block were stained with routine haematoxylin and eosin (H& E). Sections were cut for immunohistochemistry for protein marker expression by a standard procedure for Smads and BMP 4 and 7. Sections were viewed and analysed by colour video image analysis using a 40x objective, a 10x eyepiece, and a fixed frame of 128 × 128 pixels (49152.0 μm. 2. ). Ten fields per slide were examined. Smad proteins (Smads 1, 4, and 6) were expressed during the early stages (day 3) of fracture healing by bone marrow stromal cells, osteoblasts, fibroblasts and chondrocytes located in the intramembranous and endochondral ossification regions around the fracture site. Differential expressions of individual Smads, particularly Smad 1 and Smad 6, at different time-points (Smad-1 was higher than Smad-6 at day 3, whilst Smad-6 was much higher than Smad-1 at day 10) suggest that Smad proteins are not simply BMP signal transducers. Smads may also be responsible for up- and/or down-regulation of transcriptional events during the intramembranous and endochondral ossification. Smad-4, a Co-SMAD, expression newly formed bone and cartilage suggests an additional function beyond the signal transduction in rat fracture healing. BMP-4 and BMP-7 were highly expressed at day 3 and 10. BMP-7 expression was greater than BMP-4 at day 3 but switched by day 10 (BMP-4 > BMP-7). Smads represent a new level where specific therapeutic strategies can be targeted considering the interactions with a number of BMPs

INTRODUCTION. The hip arthroplasty implant is currently growing up both in orthopedic and trauma practice. This increases the frequency of prosthesis revision due to implant loosening often associated with periprosthetic osteolysis that determine the failure and lead to a loss of bone substance. Nowadays there are numerous biotechnologies seeking to join or substitute the autologous or omologous bone use. These biotechnologies (mesenchymal stromal cells, growth factors and bone substitutes) may be used in such situations, however, the literature doesn't offer class 1 clinical evidences in this field of application. MATERIALS AND METHODS. We performed a literature review using the universally validated search engines in the biomedical field: PubMed / Medline, Google Scholar, Scopus, EMBASE. The keywords used were: “Growth Factors”, “Platelet Rich Plasma”, “OP-1”, “BMP”, “BMP-2”, “BMP-7”, “Demineralized Bone Matrix”, “Stem Cell”, “Bone Marrow”, “Scaffold”, “Bone Substitutes” were crossed with “hip”, “revision”, “replacement” / “arthroplasty”, “bone loss” / “osteolysis.”. RESULTS. The search led to 321 items, of these were considered relevant: as regards the growth factors 21 articles related to in vivo animal studies and 2 articles of human clinical use of BMPs and 1 single article on the use of PRP; as regards the mesenchymal stromal cells 2 items of application in animals; as regards the use of bone substitutes we have analyzed a review of this application. DISCUSSION. The use of biotechnologies in hip prosthetic revisions has produced conflicting results: autologous growth factors (PRP) have definitely been proven effective in maxillofacial surgery, in animal studies the results of BMPs are inconsistent with articles that validate their use and others that don't recommend it. Clinical application has demonstrated, today, the limited use of BMP-7 in revisions with even an increased risk of early re-mobilization, PRP appears to be rather effective only in the early stages of peri-prosthetic osteolysis. The mesenchymal cells can increase the chances of recovery and integration of the grafts but an important variable is the number of cells that are still alive after the impaction of the graft which affects their vitality. The bone substitutes appear to be safe and very useful, particularly if applied in order to implement the omologous bone, which is still the most scaffolds used in this surgery. CONCLUSIONS. The systematic review of the literature has shown an important lack of clinical studies regarding the use of biotechnologies for prosthetic revisions. It is therefore difficult to draw guidelines that regulate the application, prospective randomized clinical studies are therefore needed to validate its effectiveness

Arthrodesis of the spine is the preferred surgical treatment for a number of pathological disorders. This process is dependent on three primary components: osteogenic cells with osteoblastic potential, osteoinductive growth factors and an osteoconductive scaffold that facilitates bone formation and vascular ingrowth. Several systemic and local factors are known to affect the rate of spinal fusion. Autogenous bone graft remains the gold standard graft material for spinal fusion. It is the only graft material that supplies the three primary components necessary for a solid fusion. Unfortunately autogenous bone is only available in limited quantities and the procurement of autograft is associated with significant donor site morbidity. A number of different bone graft materials have been developed as alternatives to autograft. These materials may be classified into two major groups, bone graft extenders used to augment autograft, or bone graft substitutes. Several different bone graft materials have been developed including allograft, osteoconductive matrices, demineralised bone matrices, bone marrow aspiration, autologous platelet concentration, growth factors and gene therapy. Allograft is currently the most widely used substitute for autogenous bone. Because any osteogenic cells are eradicated during the tissue processes, allograft is primary osteoinductive with minimal osteoinductive potential. Processing may affects the structural and biological characteristics of a graft. The incorporation of allograft occurs by a process similar to that observed with autograft but more slowly and is less complete. Osteoconductive scaffolds do not contain any osteogenic cells or osteoinductive factors and are used as a composite graft as a carrier for either osteogenic cells or osteoinductive growth factors. They are biocompatible and do not illicit a response. There is also no inherent risk of infection and availability is unlimited. These materials are brittle with poor mechanical properties and need to be protected from excessive biomechanical forces until fully incorporated. A number of osteoconductive scaffolds have been developed including ceramics, calcium sulfate, mineralized collagen, bioactive glasses, and porous metals. Dematerialized bone matrices (DMPs) are osteoinductive with variable osteoconductive properties. DMPs consist of Type I collagen and non-collagenous proteins including multiple signaling proteins. The osteoinductive activity of DMPs is due to a small fraction of bone morphogenic proteins. There is significant variability in the osteoinductive potentials and clinical efficacy of DBMs. DBMs are most effective when combined with autograft or bone marrow aspirate. Bone marrow aspiration provides osteogenetic cells and osteoinductive growth factors but must be combined with an osteoconductive carrier to form a composite graft. It is associated with minimal morbidity compared to the use of autograft and is easily obtained. Unfractionated bone marrow contains only moderate osteogenic potential. Selective retention technology can increase the number of osteogenic cells then combined with an osteoconductive carrier such as a collagen sponge or DBM. Activated platelets release multiple factors that may enhance bone formation by promoting chemotaxis, cellular proliferation and differentiation of stem cells. Platelets do not release BMPs so this autologous platelet concentrate is not inductive. Concentrated platelet rich plasma gel is combined with an osteoconductive scaffold or osteogenic cells to form a composite graft for implantation. The capacity for fusion by this technique may be inferior to autologous graft. Bone morphogenetic proteins are low molecular weight proteins related to the transforming growth factor beta superfamily. They bind receptors on the surface of osteoprogenitor stem cells and activate intracellular signal transduction cascades resulting in the osteoblastic differentiation of pluripotential stem cells. Recombinant BMPs are typically combined with an osteoconductive carrier to form a composite graft. Recombinant BMPs have been used successfully in spinal fusions and may be superior to autograft. Gene therapy involves the transfer of specific DNA sequence into target cells that express the protein of interest. Gene therapy may provide a more potent osteoinductive signal than recombinant growth factors because the sustained local release of osteogentic proteins may be more physiologic than the administration of a single large dose of recombinant factors. There are potential safety concerns and economic issues. Autogenous bone remains the gold standard of graft material; however composite grafts consisting of multiple materials may prove to be efficacious for stimulating a spinal fusion

Common in vitro protocols for TGF-β driven chondrogenic differentiation of MSC lead to hypertrophic differentiation of cells. This might cause major problems for articular cartilage repair strategies based on tissue engineered cartilage constructs derived from these cells. BMPs have been described as alternate inductors of chondrogenesis while PTHrP and FGF-2 seem promising for modulation of chondrogenic hypertrophy. The aim of this study was to identify chondrogenic culture conditions avoiding cellular hypertrophy. We analyzed the effect of a broad panel of growth factors alone or in combination with TGF-β3 on MSC pellets cultured in vitro and after transplantation in SCID mice in vivo. Chondrogenic differentiation in vitro was successful after supplementation of the chondrogenic medium with TGF-β3 as confirmed by positive collagen type II and alcian blue staining. None of the other single growth factors (BMP-2, -4, -6, -7, FGF-1, IGF-1) led to sufficient chondrogenesis as indicated by negative collagen type II and alcian blue staining. Each of these factors, however, allowed chondrogenesis in combination with TGF-β without suppressing collagen type X expression. Combination of TGF-β with PTHrP or FGF-2 suppressed ALP activity, induced MMP13 expression, and prevented differentiation to chondrocyte-like cells when added from day 0. Delayed addition of PTHrP or FGF-2 stopped chondrogenesis at the reached level and repressed ALP activity. The treatment of MSC constructs with FGF-2 or PTHrP in the last 3 weeks before transplantation did not prevent hypertrophy and calcification in vivo. FGF-2 and PTHrP were potent inhibitors for early and late chondrogenic differentiation in contrast to BMPs. As soon as a developmental window of collagen type II positive and collagen type X negative pellet cultures can be created in this model, both seem to be potent factors to suppress hypertrophy and to generate stable chondrocytes for transplantation purposes

Osteoarthritis (OA) is a chronic degenerative joint disease characterized by progressive cartilage degradation, synovial membrane inflammation, osteophyte formation, and subchondral bone sclerosis. Pathological changes in cartilage and subchondral bone are the main processes in OA. In recent decades, many studies have demonstrated that activin-like kinase 3 (ALK3), a bone morphogenetic protein receptor, is essential for cartilage formation, osteogenesis, and postnatal skeletal development. Although the role of bone morphogenetic protein (BMP) signalling in articular cartilage and bone has been extensively studied, many new discoveries have been made in recent years around ALK3 targets in articular cartilage, subchondral bone, and the interaction between the two, broadening the original knowledge of the relationship between ALK3 and OA. In this review, we focus on the roles of ALK3 in OA, including cartilage and subchondral bone and related cells. It may be helpful to seek more efficient drugs or treatments for OA based on ALK3 signalling in future.