Treatment Of Large Bone Defects Research Articles

Biomimetic triphase composite scaffolds with antibacterial and anti-tumor potentials for bone repair

Abstract Clinically, the treatment of large bone defects caused by trauma, tumor resection and other bone diseases remains a significant challenge, especially bone defects at the load-bearing sites. To promote bone regeneration and simultaneously prevent tumor recurrence and bacterial infection after surgical resection of oteosarcoma, we designed biomimetic triphase composite scaffolds comprised of porous Ti6Al4V (pTi), chitosan (CS) and selenium (Se) doped hydroxyapatite (HAP) nanoparticles (HAP-Se), termed as pTi/CS/HAP-Se. The scaffolds promoted osteoblast proliferation while inhibited tumor cells growth and bacterial viability. The scaffolds resemble natural bone tissues with hierarchical porous structures, showing potential applications in treating bone defects resulted from surgical resection of osteosarcoma.

Regeneration of large bone defects using mesoporous silica coated magnetic nanoparticles during distraction osteogenesis

Distraction osteogenesis (DO) represents an effective but undesirably lengthy treatment for large bone defects. Both magnetic nanoparticles and silicon have been shown to induce osteogenic differentiation of mesenchymal stem cells (MSCs), the key participant in bone regeneration. We herein synthesized mesoporous silica coated magnetic (Fe3O4) nanoparticles (M-MSNs) and evaluated its potential for acceleration of bone regeneration in a rat DO model. The M-MSNs exhibited good biocompatibility and remarkable capability in promoting the osteogenic differentiation of MSCs via the canonical Wnt/β-catenin pathway in vitro. More importantly, local injection of M-MSNs dramatically accelerated bone regeneration in a rat DO model according to the results of X-ray imaging, micro-CT, mechanical testing, histological examination, and immunochemical analysis. This study demonstrates the notable potential of M-MSNs in promoting bone regeneration during DO by enhancing the osteogenic differentiation of MSCs, paving the way for clinical translation of M-MSNs in DO to repair large bone defects.

Dual delivery of encapsulated BM-MSCs and BMP-2 improves osteogenic differentiation and new bone formation.

Stem cell-based therapies provide a promising approach for bone repair. In the present work, we developed a novel 3D vehicle system for dual-delivery of encapsulated bone marrow mesenchymal stem cells (BM-MSCs) and bone morphogenetic protein-2 (BMP-2) for treatment of large bone defects. The vehicle system consists of sodium alginate microcapsules and polylactic acid (PLLA) microspheres. BM-MSCs are encapsulated in the microcapsules, and BMP-2 proteins are encapsulated in the PLLA microspheres. This vehicle system acted as a multicore structure for sustained release of BMP-2, which enabled pulsed dosing induction of osteogenic differentiation of the co-embedded BM-MSCs. in vitro experiments showed that the loaded BMP-2 was constitutively released up to 30 days. Bioactivity of the incorporated BMP-2 in the microspheres was preserved and osteogenic differentiation of the BM-MSCs in the microcapsules was improved. In vivo, osteogenesis studies demonstrated that satisfactory degree of repair of a rat calvarial defect was achieved with the delivery of either encapsulated BM-MSCs alone or encapsulated BMP-2 alone. Transplantation of encapsulated both BM-MSCs and BMP-2 exhibited the greatest repair potential following 4- or 8-weeks treatment. In conclusion, microencapsulation of BM-MSCs and BMP-2 promoted the maturity of newly generated bone and improved new bone formation. Transplantation of BM-MSCs and BMP-2 in our novel 3-D vehicle system is a promising strategy for regenerative therapies of large bone defects.

Two-stage Treatment of Large Bone Defects With Cerament® G and Cerament® V Plus Auto and/or Allograft Using the Masquelet Technique as Salvage Therapy After Bone Resection in Patients With Septic Pseudarthrosis

Two-stage Treatment of Large Bone Defects With Cerament® G and Cerament® V Plus Auto and/or Allograft Using the Masquelet Technique as Salvage Therapy After Bone Resection in Patients With Septic Pseudarthrosis

Multicellularity-interweaved bone regeneration of BMP-2-loaded scaffold with orchestrated kinetics of resorption and osteogenesis

Synchronization of material resorption and new bone formation is vital to achieve harmonious bone regeneration in the treatment of large bone defects. To exposit the resorption/osteogenesis properties in the guided bone repairing, rhBMP-2-loaded trimodal macro/micro/nano-porous bioactive glass scaffolds (TMS-rhBMP-2) were set as substrate model. We penetratingly investigated the particular function of hierarchical structure and incorporated rhBMP-2 in the resorption/osteogenesis, and dissected the cellular interplay throughout the regenerative procedure. The results suggested that rhBMP-2 significantly facilitated osteoclastogenesis-mediated scaffold degradation and strikingly up-regulated mesenchymal stem cells (MSCs)-involved osteogenesis in vitro. Further gene microarray and related proteins expression indicated that in the presence of rhBMP-2, MSCs rather than differentiated MSCs could exert synergistic effects on osteoclastogenesis, osteoclasts maturation and resorptive function; meanwhile, rhBMP-2-induced MSCs osteogenesis was also strengthened by the osteoclasts. In vivo micro-CT, X-ray, kinetic and histological analyses qualitatively and quantitively demonstrated the optimized coupling of bioresorption/osteogenesis and the most rapid regeneration in TMS-rhBMP-2. Consequently, with rhBMP-2 acted as ignitor and MSCs/osteoclasts interaction as booster, a harmonious bone regeneration was obtained. Besides, long-term magnetic resonance imaging (MRI) in virtue of Gd3+ suggested that the degradation products mainly distributed in liver and spleen, verifying the accumulation/discharge profiles and safety application of TMS-rhBMP-2 system in vivo. This study will not merely provide guidance for the design of clinical bone repairing materials, but shed substantial light on the multicell-mediated tissue regeneration.

Progress in clinical treatment of large segmental bone defect

Clinical treatment of large segmental bone defect has always been a major challenge for the medical community, which is due to the complex and diverse pathogenic factors of large segmental bone defect.In recent years, the clinical treatment of large segmental bone defect has made great progress, and the main treatment options include vascularized or non-vascularized autologous bone grafts, allograft bone transplantation, masquelet technology, llizarov technology and bone tissue engineering.Therefore, understanding the advantages and disadvantages of various treatment options is very important for the clinical treatment of large bone defects in long bones, laying the foundation for clinical treatment. Key words: Large bone defect; Bone transplantation; Bone tissue engineering; Treatment progress

Masquelet Induced Membrane Technique for The Surgical Treatment of Large Bone Defects: The Reasons for Successes and Failures

Masquelet Induced Membrane Technique for The Surgical Treatment of Large Bone Defects: The Reasons for Successes and Failures

Endothelial and Osteoblast Differentiation of Adipose-Derived Mesenchymal Stem Cells Using a Cobalt-Doped CaP/Silk Fibroin Scaffold.

A major problem in the treatment of large bone defects is the inability to provide an adequate blood supply to the implantation site. Therefore, a bone regeneration strategy that provides an adequate supply of vessels would address this need. Cobalt (Co2+), because of its ability to induce hypoxia, has been used to accelerate new vessel formation. In this study, we used a freeze-drying technique to fabricate a scaffold that consisted of Co2+-doped calcium phosphate (CaP) [e.g., hydroxyapatite (HA)] and natural silk fiber through an optimized alternate mineralization process. The composition and structure of the scaffold were confirmed by X-ray diffraction (XRD), Fourier transform infrared (FTIR), inductively coupled plasma (ICP), and scanning electron microscope (SEM). The data showed that the scaffolds promoted differentiation of adipose-derived mesenchymal stem cells (ADSCs) toward endothelial and osteoblast linages. We observed improved angiogenesis and bone formation with the fabricated scaffolds compared with the control groups. Computed tomography (CT) scans and radiographic imaging, in addition to histology and immunohistochemical analyses, showed the presence of angiogenesis and bone regeneration after implantation of the ADSC-seeded scaffolds in a critical size calavarial bone defect in a Wistar rat model. We obtained the best in vitro and in vivo results by doping 2% Co2+ in HA. Taken together, we propose that the Co2+-doped HA/silk fibroin (SF) scaffold would be a good candidate to induce angiogenesis and bone formation both in vitro and in vivo.

Optimized Bioactive Glass: the Quest for the Bony Graft.

Technological advances have provided surgeons with a wide range of biomaterials. Yet improvements are still to be made, especially for large bone defect treatment. Biomaterial scaffolds represent a promising alternative to autologous bone grafts but in spite of the numerous studies carried out on this subject, no biomaterial scaffold is yet completely satisfying. Bioactive glass (BAG) presents many qualifying characteristics but they are brittle and their combination with a plastic polymer appears essential to overcome this drawback. Recent advances have allowed the synthesis of organic-inorganic hybrid scaffolds combining the osteogenic properties of BAG and the plastic characteristics of polymers. Such biomaterials can now be obtained at room temperature allowing organic doping of the glass/polymer network for a homogeneous delivery of the doping agent. Despite these new avenues, further studies are required to highlight the biological properties of these materials and particularly their behavior once implanted in vivo. This review focuses on BAG with a particular interest in their combination with polymers to form organic-inorganic hybrids for the design of innovative graft strategies.

Bioprinted osteon-like scaffolds enhance in vivo neovascularization

Bone tissue engineers are facing a daunting challenge when attempting to fabricate bigger constructs intended for use in the treatment of large bone defects, which is the vascularization of the graft. Cell-based approaches and, in particular, the use of in vitro coculture of human umbilical vein endothelial cells (HUVECs) and human mesenchymal stem cells (hMSCs) has been one of the most explored options. We present in this paper an alternative method to mimic the spatial pattern of HUVECs and hMSCs found in native osteons based on the use of extrusion-based 3D bioprinting (3DP). We developed a 3DP biphasic osteon-like scaffold, containing two separate osteogenic and vasculogenic cell populations encapsulated in a fibrin bioink in order to improve neovascularization. To this end, we optimized the fibrin bioink to improve the resolution of printed strands and ensure a reproducible printing process; the influence of printing parameters on extruded strand diameter and cell survival was also investigated. The mechanical strength of the construct was improved by co-printing the fibrin bioink along a supporting PCL carrier scaffold. Compressive mechanical testing showed improved mechanical properties with an average compressive modulus of 131 ± 23 MPa, which falls in the range of cortical bone. HUVEC and hMSC laden fibrin hydrogels were printed in osteon-like patterns and cultured in vitro. A significant increase in gene expression of angiogenic markers was observed for the biomimetic scaffolds. Finally, biphasic scaffolds were implanted subcutaneously in rats. Histological analysis of explanted scaffolds showed a significant increase in the number of blood vessels per area in the 3D printed osteon-like scaffolds. The utilization of these scaffolds in constructing biomimetic osteons for bone regeneration demonstrated a promising capacity to improve neovascularization of the construct. These results indicates that proper cell orientation and scaffold design could play a critical role in neovascularization.

Three-Dimensional Co-Culture of Peripheral Blood-Derived Mesenchymal Stem Cells and Endothelial Progenitor Cells for Bone Regeneration.

Engraftment of tissue-engineered bone plays a pivotal role in the treatment of large bone defects. However, promoting thorough vascularization in the central area of tissue-engineered constructs remains a great challenge for clinical application. Here, we developed a three-dimensional (3D) co-culture system using biphasic calcium phosphate bioceramic (BCPB) scaffold seeded with rabbit peripheral blood-derived mesenchymal stem cells (PB-MSCs) and endothelial progenitor cells (EPCs) to improve new bone formation and vascularization for long bone segmental defects. In vitro studies, we identified morphology and characterization of PB-MSCs and EPCs. We also created a co-culture system of PB-MSCs and EPCs, and assessed the CD31 expression, gene expression of VEGF, PDGF and ALP, and tube formation ability of the co-culture system. Moreover, the biocompatibility of the BCPB was assessed and secretion levels of ALP, OC, PDGF and VEGF by co-cultured PB-MSC and EPCs in the 3D co-culture system were determined (ELISA). In vivo studies were performed to assess the ability of the cell-scaffold construct to repair a rabbit large bone defect model by X-ray examination, gross observation, and histological staining. With the extension of incubation time, both osteogenic- and vascular-related genes were up-regulated when EPCs co-cultured with PB-MSCs. In addition, BCPB is biocompatible and the expression levels of osteogenic- and vascular-related markers were also up-regulated in the 3D co-culture system. Seeding of PB-MSCs and EPCs within a modified BCPB and subsequently implanted gave rise to new bone and promoted vascularization in the rabbit model. These findings suggest that our vascularized tissue-engineered bone may be a potential alternative in the treatment of large bone defects.

Chondroitin Sulfate Glycosaminoglycan Scaffolds for Cell and Recombinant Protein-Based Bone Regeneration.

Bone morphogenetic protein 2 (BMP‐2)‐loaded collagen sponges remain the clinical standard for treatment of large bone defects when there is insufficient autograft, despite associated complications. Recent efforts to negate comorbidities have included biomaterials and gene therapy approaches to extend the duration of BMP‐2 release and activity. In this study, we compared the collagen sponge clinical standard to chondroitin sulfate glycosaminoglycan (CS‐GAG) scaffolds as a delivery vehicle for recombinant human BMP‐2 (rhBMP‐2) and rhBMP‐2 expression via human BMP‐2 gene inserted into mesenchymal stem cells (BMP‐2 MSC). We demonstrated extended release of rhBMP‐2 from CS‐GAG scaffolds compared to their collagen sponge counterparts, and further extended release from CS‐GAG gels seeded with BMP‐2 MSC. When used to treat a challenging critically sized femoral defect model in rats, both rhBMP‐2 and BMP‐2 MSC in CS‐GAG induced comparable bone formation to the rhBMP‐2 in collagen sponge, as measured by bone volume, strength, and stiffness. We conclude that CS‐GAG scaffolds are a promising delivery vehicle for controlling the release of rhBMP‐2 and to mediate the repair of critically sized segmental bone defects. stem cells translational medicine 2019;8:575–585

Treatment of Large Bone Defect of Long Bones: Comparison of Free Vascularized Fibular Graft with Compression-Distraction Technique

We treated 18 patients with congenital pseudarthrosis of tibia and 27 patients with acquired bone loss. Out of 18 cases of congenital pseudarthrosis of tibia the free vascularized fibular grafts (FVFGs) were performed in 16 cases and compression-distraction techniques in 2 cases. In patients with angular deformities and shortenings in the lower leg soft tissue distraction and correction were conducted. The mean soft tissue distraction period was 78.4 days, the length of bone defect 10.7 cm, the mean operative time 3.2 hours, and primary bone healing achieved in 12 cases. The mean union period was 7.3 months and the mean period of full weight bearing was 10.8 months. In 4 cases pseudarthroses and deformities developed and healed secondarily, and the treatment period ranged from 16 months to 26 months. The mean length of bone defects was 9.7 cm and the mean period of full weight bearing 14 months. The mean length of bone defects in the group of Ilizarov technique was 6.6 cm, the mean period of distraction 82 days and the mean period of full weight bearing 12.2 months. The complications include pseudarthrosis, refracture, angular deformity, equinovarus and temporary peroneal nerve palsy in the FVFG group and in the group of compression-distraction technique, refracture, equinovarus and temporary peroneal nerve palsy. The FVFG is suitable for the treatment of pseudarthrosis in young patients and the compression-distraction technique is effective option for the treatment of bone defects of patients with infection and deformity.

Modified Masquelet’s technique with nail and allograft: A case report

The large bone defect treatment is a challenge issue in modern orthopaedic trauma surgery. One of the most used technique is the Masquelet's technique. In this case report we used a modified Masquelet technique to fill a six centimeters bone gap in the proximal femur. A 18-year-old boy with a septic pseudoarthrosis was treated with a two stages procedure: in the first step we used a hollow antibiotic spacer and an intramedullary nail. In the second step, we used both omologous (6 cm of bone allograft) and autologous bone graft with a new intramedullary nail. Immediate partial weight bearing was allowed and after 3 months the patient started walking with complete weight bearing. A year later the fracture had healed uneventfully. The modification of the Masquelet Technique that we have made allows the patient to immediately weight bearing safely, speeding up the functional recovery. Further studies are needed to standardize this type of combined technique.

Advances in research of tissue engineering bone vascularization in treating large bone defects

In recent years, the research on tissue engineering bone in the treatment of large bone defects has shifted from animal experiments to clinical application, showing broad prospect. The blood supply of the implant plays a critical role in the osteogenic stability, so vascularization of tissue engineering bone has become the research focus of bone tissue engineering in the treatment of bone defects. Therefore, the author reviews the research progress of tissue engineering bone vascularization in treatment of large bone defect from the aspects of tissue engineering bone construction, prefabricated vascular technology, and cell technology so as to provide reference for the clinical treatment. Key words: Tissue engineering; Blood vessels; Large bone defects

Masquelet membrane induction technique for reconstruction of bone defect

There are many reasons for bone defects. Trauma, malignant tumor resection, chronic osteomyelitis and congenital factors can all lead to bone defects. The treatment methods include autogenous cancellous bone grafting, vascularized fibular grafting, distraction osteogenesis and Masquelet techniques, etc. Masquelet technique was first discovered by Masquelet et al. in 1986. It provides a simple and safe method for the treatment of large bone defects. This technique has two stages, and the first stage includes wound thoroughly debridement, maintenance of the stability of fracture, placement of (antibiotic) cement spacer, and closure of the wound without tension, then 6 to 8 weeks later, the second stage will be conducted including performing a longitudinal incision of the induced membrane, removal of the bone cement and bone graft. Bone cement includes polymethyl methacrylate (PMMA), calcium phosphate, calcium sulfate and other materials, if necessary, antibiotics could be combined at the same time. Different bone cement induced biofilmhas different characteristics.Induced membrane can secrete transforming growth factor (TGF-β1), bone morphogenetic protein 2 (BMP-2), platelet endothelial cell adhesion molecule-1 (PECAM-1), and vascular endothelial growth factor (VEGF) and other osteogenesis growth and angiogenesis related factors, then an independent space is built to isolate the bone graft from outside environment, avoiding the phenomenon of bone graft absorption and providing a stable biological environment of bone defect. Masquelet technique is simple to operate, safe, accompanied by fewer complications, less demanding to the recipient area, easy to be accepted by patients and promoted clinically. It is applicable to bone defects in all parts and has great potential for clinical application and especially suitable for large segmental bone defectwhich is ineffective in other treatment methods.

Influence of concentration and preparation of platelet rich fibrin on human bone marrow mononuclear cells (in vitro)

Large bone defects have always been a big challenge. The use of bone marrow mononuclear cells (BMCs) combined with an osteoconductive scaffold has been proved a good alternative for the treatment of large bone defects. Another autologous source for tissue engineering is platelet rich fibrin (PRF). PRF is a blood concentrate system obtained through a one-step centrifugation. The generated 3D matrix of the PRF clot serves as a reservoir of growth factors. Those growth factors might support the regenerative response of BMC, and therefore the effect of PRF, centrifuged with either high medium (208 g) or low (60 g) relative centrifugation force (RCF) on BMCs was evaluated in vitro in the present study. The two PRF matrices obtained were initially characterized and compared to human serum. Significantly increased concentrations of insulin-like growth factor (IGF), soluble intercellular adhesion molecule-1 (sICAM1) and transforming growth factor (TGF)-β were found in PRF compared to human serum whereas VEGF concentration was not significantly altered. A dose-response study revealed no further activation of BMC’s metabolic activity, if concentration of both PRF matrices exceeded 10% (v/v). Effect of both PRF preparations [10%] on BMC was analyzed after 2, 7, and 14 days in comparison to human serum [10%]. Metabolic activity of BMC increased significantly in all groups on day 14. Furthermore, gene expression of matrix metalloproteinases (MMP)-2, −7, and −9 was significantly stimulated in BMC cultivated with the respective PRF matrices compared to human serum. Apoptotic activity of BMC incubated with PRF was not altered compared to BMC cultivated with serum. In conclusion, PRF could be used as a growth factor delivery system of autologous or allogeneic source with the capability of stimulating cells such as BMC.

Bone morphogenic protein expression and bone formation are induced by bioactive glass S53P4 scaffolds in vivo.

The two-stage induced-membrane (IM) technique is increasingly used for treatment of large bone defects. In stage one, the bone defect is filled with polymethylmethacrylate (PMMA), which induces a membrane around the implant. In stage two, PMMA is replaced with bone graft. Bioactive glasses (BAGs) are bone substitutes with bone-stimulating and angiogenic properties. We have previously shown that a certain type of BAG can also induce a foreign-body membrane similar to PMMA. The aim of this study was to evaluate the bone-forming capacity of sintered BAG-S53P4 and poly(lactide-co-glycolide) (PLGA)-coated BAG-S53P4 scaffolds for potential use as bone substitutes in a single-stage IM technique. Sintered porous rods of BAG-S53P4, BAG-S53P4-PLGA, or PMMA were implanted in rabbit femurs for 2, 4, or 8 weeks. The expression of bone morphogenic protein (BMP)-2, -4, and -7 in the IMs of implanted materials were analyzed with real-time quantitative polymerase chain reaction. Micro-computed tomography imaging was used to evaluate bone growth and further verified with scanning electron microscopy. BAG-S53P4 and BAG-S53P4-PGLA scaffold IMs show similar or superior expression of BMP-2, -4, and -7 compared with PMMA IM. Bone ingrowth into BAG scaffolds increased over time. Active bone formation occurred inside the BAG scaffolds and the respective BMP expressions were similar or superior for the BAG IMs compared with PMMA, thus making BAGs a promising device for single-stage treatment of bone defects. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res B Part B: Appl Biomater, 2018. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 107B: 847-857, 2019.

Masquelet induced membrane technique for treatment of rat chronic osteomyelitis.

The application value of Masquelet induced membrane technique for the treatment of rat chronic osteomyelitis was explored. A total of 180 male Sprague-Dawley (SD) rats were randomly divided into sham operation group (control group), chronic osteomyelitis model group (model group) and Masquelet induced membrane therapy + chronic osteomyelitis model group (observation group); 60 rats in each group. A rat model of traumatic osteomyelitis was established using a modified blunt trauma method in model and observation group. All rats in observation group were treated with membrane induction technology while in the control group they were not. Postoperative complications, bone healing time and functional status of the affected limbs were recorded. After surgery, no obvious perioperative complications occurred in most of the rats, except that 5 rats experienced partial necrosis at the edge of the flap, and 3 experienced superficial infection around the incision site in the ilium. Primary bone healing was achieved in 50 rats with an average healing time of 15 weeks. Among them, 38 rats restored weight-bearing function after 20 weeks. Seven rats experienced infection after surgery, but complete bone healing was achieved after treatment with induced membrane technique again. Following treatment with induced membrane technique, the rat body weight, body temperature and the number of bacteria in the wound decreased over time. In conclusion, Masquelet induced membrane technique is effective for the treatment of large bone defects in rats after traumatic osteomyelitis. This technique significantly shortens the treatment cycle. The surgical procedure is simple, and the incidence of postoperative complications is low.

(Keynote) 3D-Printed Biodegradable Implants for Orthopedic Applications

This study developed a new biodegradable polymeric cage using 3D printing technique to convert corticocancellous bone chips into a structured strut graft for treating segmental bone defects. A biodegradable truncated-pyramid cage was designed and manufactured to serve as the bone graft. Commercially available PLA filaments with a diameter of 3.0 mm were used as the material for the bone cage 3D printing. The cage was fabricated using a fused-deposition-modeling (FDM) type 3D printer with a printing resolution of 200 mm. The experimental results show that higher leg length ratios and 3-point bending strengths demonstrated improved final bony ingrowths within the bone defects for rabbits implanted with the 3D printed cages. The present experimental results indicate that using 3D printing PLA can be remodeled into a cage of any geometric shape to fill the bone. With this 3D printed cage, the orthopedic surgeons will be able to convert small corticocancellous bone chips into a structured bone graft to avail both the osteoconductive and mechanical properties. Through this bone graft converting technique, the surgeons can merge different sources of bone graft to serve as an adequate strut bone graft for the treatment of large bone defects without experiencing donor site morbidity and graft resource limitations.