Calcium Phosphate Scaffolds Research Articles

Controlling sintering temperature for biphasic calcium phosphate scaffolds using submicron hydroxyapatite slurries for LCD 3D printing

Liquid crystal display (LCD)-based 3D printing, a precise and surface-smooth technology, has found widespread use in dentistry due to its exceptional precision and surface quality. This research endeavors to create a scaffold through 3D printing using bone graft materials, specifically addressing the challenge of molding bone graft powders to fit defect areas in dental surgery. Our approach involved developing biphasic calcium phosphate (BCP) scaffolds by regulating the sintering temperature of an HA-based green body. Initially, by using wet ball mill technology, HA slurries with 30 %, 40 %, and 50 % HA content by weight were prepared and exhibited viscosities below 0.15 Pa s. Subsequently, these slurries were 3D printed via an LCD-based VP printer. The utilization of submicron-sized HA particles played a critical role in maintaining slurry stability and preventing sedimentation during the printing process. We investigated sintering temperatures of 1100 °C, 1200 °C, and 1300 °C, discovering that submicron-sized HA particles enabled precise alignment of the decomposition temperature with the typical HA sintering range. Consequently, we achieved the production of BCP scaffolds with adjustable properties by tuning the sintering temperature. Subsequent analysis delved into scaffold shrinkage, mechanical characteristics, degradation behavior, and biological responses. Among the sintered scaffolds, the 50 % HA slurry printed and sintered at 1200 °C exhibited the highest compressive strength. The cell viabilities of these sintered scaffolds consistently met the ISO 10993-5 standard, affirming their biocompatibility. Moreover, the biological responses of MG63 cells demonstrated that the sintered scaffolds not only exhibited cytocompatibility but also facilitated strong cell attachment and proliferation. Furthermore, scaffolds sintered at 1300 °C containing TCP showed the formation of a layer of HA with plate-like structures due to ion-induced precipitation, indicating their bioactivity. In conclusion, this study highlights the promising potential of LCD-based VP 3D printing technology for ceramic slurries, with clear implications for clinical applications in bone tissue engineering.

Cytotoxicity and mechanical properties of biphasic calcium phosphate scaffold from Tegilarca granosa due to its composition

Background: Biphasic calcium phosphate (BCP) is graft material contained HA and TCP. Tegilarca granosa shell is a natural source that may converted into BCP. This study aims to determine the composition and cytotoxicity of BCP synthesized from Tegilarca granosa shell used various hydrothermal hours and to evaluate the mechanical properties of BCP scaffold. Methods: Tegilarca granosa shell was converted into BCP using hydrothermal method at 200˚C for 6h (Group 1); 9h (Group 2); and 12h (Group 3). The composition was determined by XRD and the cell viability were evaluated using MTT Assays. Each group was added with 20% gelatin ratio 50:50 (w/v) and freeze-dried to form scaffold. Scaffolds (Ø6mm x 4mm) were prepared for diametral tensile strength (DTS) test (n=6) and scaffolds (Ø7mm x 11mm) were used for compressive strength (CS) test (n=6). All data were analyzed using Kruskall-Wallis followed by Mann-Whitney test. Results: The composition of BCP (HA/ TCP) at Group 1, Group 2, and Group 3 were 81.80%/14,10%; 87%/6%; and 72%/21%. The cell viabilities were good for all groups. The DTS and CS test showed there was a significant difference between Group 1 and Group 3 scaffold, meanwhile there was no significant differences between Group 2 and Group 3 scaffold. Group 3 scaffold showed the highest DTS and CS, 6.921 MPa and 1,233 MPa. Conclusion: The BCP composition were depent on hydrothermal hours. Although all scaffold groups were non-toxic, but BCP scaffold synthesized from Tegilarca granosa shell using hydrothermal for 12 hours showed the highest mechanical properties.

The role of integrin αvβ3 in biphasic calcium phosphate ceramics mediated M2 Macrophage polarization and the resultant osteoinduction

Calcium phosphate ceramics-based biomaterials were reported to have good biocompatibility and osteoinductivity and have been widely applied for bone defect repair and regeneration. However, the mechanism of their osteoinductivity is still unclear. In our study, we established an ectopic bone formation in vivo model and an in vitro macrophage cell co-culture system with calcium phosphate ceramics to investigate the effect of biphasic calcium phosphate on osteogenesis via regulating macrophage M1/M2 polarization. Our micro-CT data suggested that biphasic calcium phosphate had significant osteoinductivity, and the fluorescence co-localization detection found increased F4/80+/integrin αvβ3+ macrophages surrounding the biphasic calcium phosphate scaffolds. Besides, our study also revealed that biphasic calcium phosphate promoted M2 polarization of macrophages via upregulating integrin αvβ3 expression compared to tricalcium phosphate, and the increased M2 macrophages could subsequently augment the osteogenic differentiation of MSCs in a TGFβ mediated manner. In conclusion, we demonstrated that macrophages subjected to biphasic calcium phosphate could polarize toward M2 phenotype via triggering integrin αvβ3 and secrete TGFβ to increase the osteogenesis of MSCs, which subsequently enhances bone regeneration.

Curcumin and resveratrol delivery from multi-functionalized calcium phosphate scaffold enhances biological properties

Natural medicinal compounds (NMCs) can assist effectively in treating bone disorders. NMC release kinetics from a ceramic bone tissue engineering scaffold can be tailored. However, inferior physicochemical properties halt their therapeutic applications and need a carrier system for delivery. We developed a multi-functionalized scaffold to understand the effect of curcumin (Cur) and resveratrol (Rsv) on in vitro biological properties. Polycaprolactone (PCL) nanoparticles encapsulated resveratrol in the polymeric matrix. Nanoparticles showed a hydrodynamic diameter of about 180 nm, −16 mV zeta potential, and up to ∼65 % encapsulation efficiency. Scaffolds made of zinc-doped tricalcium phosphate (Zn-TCP) were coated with curcumin followed by either resveratrol (Cur-Rsv) or resveratrol nanoparticles (Cur-Rsv-NP). NMC-loaded scaffolds exhibited a biphasic release pattern over 60 days. Solubility and hydrophobic-hydrophilic interactions affected the NMC release profile. Resveratrol showed rapid release as compared to curcumin. The treated scaffold increased the cell viability of human fetal osteoblast (hFOB) by 1.8-fold as compared to the control. It exhibited a 6-fold increase in cytotoxicity toward osteosarcoma (MG-63) cells as compared to the untreated scaffold. NMCs loaded scaffold effectively inhibited Staphylococcus aureus from colonizing over the scaffold. Zinc doping enhanced osteoblast growth and prevented bacterial colony formation. Such design principle provided a direction for developing multi-functionalized calcium phosphate (CaP) scaffolds against bone diseases for orthopedic applications.

Evaluating the suitability of fast sintering techniques for the consolidation of calcium phosphate scaffolds produced by DLP

Evaluating the suitability of fast sintering techniques for the consolidation of calcium phosphate scaffolds produced by DLP

Micelle encapsulated curcumin and piperine-laden 3D printed calcium phosphate scaffolds enhance in vitro biological properties

Limitations in the current clinical management of critical-sized osseous defects have driven the need for multifunctional bone constructs. The ideal bone scaffold should possess advanced microarchitecture, well-defined pore interconnectivity, and supply biological signals, which actively guide and control tissue regeneration while simultaneously preventing post-implantation complications. Here, a natural medicine-based localized drug delivery from 3D printed scaffold is presented, which offers controlled release of curcumin, piperine from nano-sized polymeric micelles, and burst release of antibacterial carvacrol from the coating endowing the scaffold with their distinct, individual biological properties. This functionalized scaffold exhibits improved osteoblast (hFOB) cell attachment, 4-folds higher hFOB proliferation, and 73% increased hFOB differentiation while simultaneously providing cytotoxicity towards osteosarcoma cells with 61% lesser viability compared to control. In vitro, early tube formation (p < 0.001) indicates that the scaffolds can modulate the endothelial cellular network, critical for faster wound healing. The scaffold also exhibits 94% enhanced antibacterial efficacy (p < 0.001) against gram-positive Staphylococcus aureus, the main causative bacteria for osteomyelitis. Together, the multifunctional scaffolds provide controlled delivery of natural biomolecules from the nano-sized micelle-loaded 3D printed matrix for significant improvement in osteoblast proliferation, endothelial formation, osteosarcoma, and bacterial inhibition, guiding better bone regeneration for post-traumatic defect repair.

Osteogenic differentiation of adipose-derived canine mesenchymal stem cells seeded in porous calcium-phosphate scaffolds.

Engineered bone graft substitutes are a promising alternative and supplement to autologous bone grafts as treatments for bone healing impairment. Advances in human medicine extend an invitation to pursue these biomimetic strategies in animal patients, substantiated by the theory that specialized scaffolds, multipotent cells, and biological cues may be combined into a bioactive implant intended for the enhancement of tissue regeneration. This proof-of-concept study was designed to evaluate and validate the feasibility of beta-tricalcium phosphate foam scaffolds seeded with canine mesenchymal stem cells derived from adipose tissue. Cell-inoculated samples and sham controls were cultured statically for 72 hours in complete growth medium to evaluate seeding capacity, while a subset of loaded scaffolds was further induced with osteogenic culture medium for 21 days. Produced implants were characterized and validated with a combination of immunofluorescence and reflection confocal microscopy, scanning electron microscopy, and polymerase chain reaction to confirm osteogenic differentiation in tridimensional-induced samples. After 72 hours of culture, all inoculated scaffolds presented widespread yet heterogeneous surface seeding, distinctively congregating stem cells around pore openings. Furthermore, at 21 days of osteogenic culture conditions, robust osteoblastic differentiation of the seeded cells was confirmed by the change of cell morphology and evident deposition of extra-cellular matrix, accompanied by mineralization and scaffold remodeling; furthermore, all induced cell-loaded implants lost specific stemness immunophenotype expression and simultaneously upregulated genomic expression of osteogenic genes Osterix and Ostecalcin. β-TCP bio-ceramic foam scaffolds proved to be suitable carriers and hosts of canine adipose-derived MSCs, promoting not only surface attachment and proliferation, but also demonstrating strong in-vitro osteogenic potential. Although this research provides satisfactory in-vitro validation for the conceptualization and feasibility of a canine bio-active bone implant, further testing such as patient safety, large-scale reproducibility, and quality assessment are needed for regulatory compliance in future commercial clinical applications.

Chitosan/collagen/Mg, Se, Sr, Zn-substituted calcium phosphate scaffolds for bone tissue engineering applications: A growth factor free approach

According to the biomimetic bone scaffold design paradigm, a scaffold resembling natural bone tissue with molecular, structural and biological compatibility is needed to allow effective regeneration of bone tissue. Continuing our previous studies regarding scaffolds with chitosan matrix containing Mg, Se, Sr, Zn-substituted calcium phosphates (CaPs), the focus of this work was to further improve the properties of these growth factor-free scaffolds. By addition of collagen into the chitosan matrix at weight ratios of 100:0, 75:25, 50:50, 25:75 and 0:100, we aimed to better resemble natural bone tissue. Highly porous composite scaffolds based on chitosan and collagen, with 30 wt% of Mg, Se, Sr, Zn-substituted CaPs, were prepared by the freeze-gelation method. The scaffolds show a highly porous structure, with interconnected pores in the range of 20–350 μm and homogeneously dispersed CaPs. The added collagen further enhanced the stability measured during 28 days in simulated biological conditions. Live/dead and CyQUANT assays confirmed good viability and proliferation of human bone marrow-derived mesenchymal stem/stromal cells, while successful osteogenic differentiation was confirmed by alkaline phosphatase quantification and type I collagen immunocytochemical staining. Results indicated that the addition of collagen into the chitosan matrix containing Mg, Se, Sr, Zn-substituted CaPs improved the physicochemical and biological properties of the scaffolds.

In Vitro Evaluation of Biphasic Calcium Phosphate Scaffolds Derived from Cuttlefish Bone Coated with Poly(ester urea) for Bone Tissue Regeneration.

This study investigates the osteogenic differentiation of umbilical-cord-derived human mesenchymal stromal cells (hUC-MSCs) on biphasic calcium phosphate (BCP) scaffolds derived from cuttlefish bone doped with metal ions and coated with polymers. First, the in vitro cytocompatibility of the undoped and ion-doped (Sr2+, Mg2+ and/or Zn2+) BCP scaffolds was evaluated for 72 h using Live/Dead staining and viability assays. From these tests, the most promising composition was found to be the BCP scaffold doped with strontium (Sr2+), magnesium (Mg2+) and zinc (Zn2+) (BCP-6Sr2Mg2Zn). Then, samples from the BCP-6Sr2Mg2Zn were coated with poly(ԑ-caprolactone) (PCL) or poly(ester urea) (PEU). The results showed that hUC-MSCs can differentiate into osteoblasts, and hUC-MSCs seeded on the PEU-coated scaffolds proliferated well, adhered to the scaffold surfaces, and enhanced their differentiation capabilities without negative effects on cell proliferation under in vitro conditions. Overall, these results suggest that PEU-coated scaffolds are an alternative to PCL for use in bone regeneration, providing a suitable environment to maximally induce osteogenesis.

ANTIBIOTIC-LOADED CALCIUM PHOSPHATE SCAFFOLD FOR TREATING NONUNION BONE INFECTION IN A RABBIT MODEL

Treatment of bone infection often includes a burdensome two-stage revision. After debridement, contaminated implants are removed and replaced with a non-absorbable cement spacer loaded with antibiotics. Weeks later, the spacer is exchanged with a bone graft aiding bone healing. However, even with this two-stage approach infection persists. In this study, we investigated whether a novel 3D-printed, antibiotic-loaded, osteoinductive calcium phosphate scaffold (CPS) is effective in single-stage revision of an infected non-union with segmental bone loss in rabbits.A 5 mm defect was created in the radius of female New Zealand White rabbits. The bone fragment was replaced, stabilized with cerclage wire and inoculated with Staphylococcus aureus (MSSA). After 4 weeks, the infected bone fragment was removed, the site debrided and a spacer implanted. Depending on group allocation, rabbits received: 1) PMMA spacer with gentamycin; 2) CPS loaded with rifampin and vancomycin and 3) Non-loaded CPS. These groups received systemic cefazolin for 4 weeks after revision. Group 4 received a loaded CPS without any adjunctive systemic therapy (n=12 group1-3, n=11 group 4). All animals were euthanized 8 weeks after revision and assessed by quantitative bacteriology or histology. Covariance analysis (ANCOVA) and multiple regression were performed.All animals were culture positive at revision surgery. Half of the animals in all groups had eliminated the infection by end of study. In a historical control group with empty defect and no systemic antibiotic treatment, all animals were infected at euthanasia. There was no significant difference in CFU counts between groups at euthanasia.Our results show that treating an osteomyelitis with segmental bone loss either with CPS or PMMA has a similar cure rate of infection. However, by not requiring a second surgery, the use of CPS may offer advantages over non-resorbable equivalents such as PMMA.

Hierarchically porous calcium phosphate scaffold with degradable PLGA microsphere network

Calcium phosphate cement (CPC) is widely used in orthopedics, dentistry and spine surgery because of its excellent biocompatibility, osteoconductivity, arbitrary shaping and self-setting ability. However, slow degradation rate of CPC decreases its bone regeneration efficacy. Herein, poly (lactic co-glycolic acid) microspheres (PLGAm) and wollastonite (WS) were mixed with CPC powder to prepare CPC composite pastes, and then the composite pastes were perfused into poly (lactic co-glycolic acid) network (PLGAnw) to construct composite bone repair materials. The degradation of PLGAnw generated interconnected macropores with the short side of about 468 μm along horizontal axis and the long side of about 785 μm along longitudinal axis, while PLGAm degradation generated isolated or partially connected relative smaller spherical macropores with the size range of 53–106 μm and 106–150 μm, respectively, which was same with PLGAm; in addition, CPC formed micro/nano pores after hydration, thus constructing in-situ hierarchically porous CPC composite scaffolds. Meanwhile, PLGAnw and PLGAm also significantly promoted the degradation of CPC due to the release of acidic by-products. Therefore, the introduction of PLGA network and microspheres not only can evidently accelerate degradation of CPC, but also in-situ form connected macropores and hierarchically porous after their degradation, which are expected to improve bone repair effect of CPC.

Multifunctional Three-Dimensional Printed Copper Loaded Calcium Phosphate Scaffolds for Bone Regeneration

Bone regeneration using inorganic nanoparticles is a robust and safe approach. In this paper, copper nanoparticles (Cu NPs) loaded with calcium phosphate scaffolds were studied for their bone regeneration potential in vitro. The pneumatic extrusion method of 3D printing was employed to prepare calcium phosphate cement (CPC) and copper loaded CPC scaffolds with varying wt% of copper nanoparticles. A new aliphatic compound Kollisolv MCT 70 was used to ensure the uniform mixing of copper nanoparticles with CPC matrix. The printed scaffolds were studied for physico-chemical characterization for surface morphology, pore size, wettability, XRD, and FTIR. The copper ion release was studied in phosphate buffer saline at pH 7.4. The in vitro cell culture studies for the scaffolds were performed using human mesenchymal stem cells (hMSCs). The cell proliferation study in CPC-Cu scaffolds showed significant cell growth compared to CPC. The CPC-Cu scaffolds showed improved alkaline phosphatase activity and angiogenic potential compared to CPC. The CPC-Cu scaffolds showed significant concentration dependent antibacterial activity in Staphylococcus aureus. Overall, the CPC scaffolds loaded with 1 wt% Cu NPs showed improved activity compared to other CPC-Cu and CPC scaffolds. The results showed that copper has improved the osteogenic, angiogenic and antibacterial properties of CPC scaffolds, facilitating better bone regeneration in vitro.

Regenerative Potential of Bone Morphogenetic Protein 7-Engineered Mesenchymal Stem Cells in Ligature-Induced Periodontitis.

Periodontitis is an oral disease caused by bacterial infection that has stages according to the severity of tissue destruction. The advanced stage of periodontitis presents irreversible destruction of soft and hard tissues, which finally results in loss of teeth. When conventional treatment modalities show limited results, tissue regeneration therapy is required in patients with advanced periodontitis. In the present study, we aimed to evaluate the effect of bone marrow-derived mesenchymal stem cells (BM-MSCs) delivering bone morphogenetic protein 7 (BMP7) on tissue regeneration in a periodontitis model. BMP7 is a member of the BMP family that shows bone-forming ability; however, BMPs rapid clearing and degradation and unproven efficacy make it difficult to apply it in clinical dentistry. To overcome this, we established BMP7-expressing engineered BM-MSCs (BMP7-eBMSCs) that showed superior osteogenic differentiation potential when subcutaneously transplanted with a biphasic calcium phosphate scaffold into immunocompromised mice. Furthermore, the efficacy of BMP7-eBMSC transplantation for periodontal tissue regeneration was evaluated in a rat ligature-induced periodontitis model. Upon measuring two-dimensional and three-dimensional amounts of regenerated alveolar bone using microcomputed tomography, the amounts were found to be significantly higher in the BMP7-eBMSC transplantation group than in the eBMSC transplantation group. Most importantly, fibrous periodontal ligament (PDL) tissue regeneration was also achieved upon BMP7-eBMSC transplantation, which was evaluated by calculating the modified relative connective tissue attachment. The amount of connective tissue attachment in the BMP7-eBMSC transplantation group was significantly higher than that in the ligature-induced periodontitis group, although the increase was comparable between the BMP7-eBMSC and human PDL stem cell transplantation groups. Taken together, our results suggested that sustainable release of BMP7 induces periodontal tissue regeneration and that transplantation of BMP7-eBMSCs is a feasible treatment option for periodontal regeneration. Impact Statement Periodontitis is the second most common human dental disease affecting chronic systemic diseases. Despite the tremendous efforts trying to cure the damaged periodontal tissues using tissue engineering technologies, a definitive regenerative method has not been in consensus. Researchers are seeking more feasible and abundant source of mesenchymal stem cells (MSCs), and furthermore, how to use reliable growth factors under more efficient control are the issues to be solved. In this study, we aimed to evaluate the effect of bone morphogenetic protein 7 (BMP7) gene delivering bone marrow-derived MSCs on periodontal tissue regeneration to evaluate the efficacy of BMP7 and engineered BMSCs for periodontal tissue regeneration.

Enhancing osteoinduction and bone regeneration of biphasic calcium phosphate scaffold thought modulating the balance between pro-osteogenesis and anti-osteoclastogenesis by zinc doping

Biphasic calcium phosphate (BCP) is a promising bone regeneration material with an adjustable degradation rate, and ions doping is an effective method to improve its bone repair effect. However, few studies have focused on the role of ions doping-mediated osteogenesis and osteoclastogenesis balance in bone regeneration. In order to improve the bone repair effect of BCP and emphasize the role of osteogenesis and osteoclastogenesis balance regulated by ions doping to promote bone regeneration. Here, zinc (Zn), an essential trace element with pro-osteogenesis and anti-osteoclastogenesis, was dopped into BCP scaffolds, and its effects on the physicochemical properties, in vitro cytological responses, in vivo osteoinduction, and bone defect regeneration of BCP were systematically investigated. Results showed that Zn was successfully introduced into the crystal lattice of BCP. The differentiation of both osteoblasts and osteoclasts showed a doping content-dependent behavior. By increasing the doping content (0–5 mol.%), the anti-osteoclastogenesis effect of BCP gradually increased, while its pro-osteogenesis performance first increased and then decreased. ThoughBCP with a doping content of 5 mol.% could promote osteogenic differentiation of stem cells and reduced fibrous capsule encapsulation, it could not enhance ectopic bone formation or bone regeneration due to its excessive anti-osteoclastogenesis. BCP with a doping content of 2.5 mol.% remarkably enhanced the osteoinduction activity and accelerated new bone formation, which were associated with its highest pro-osteogenesis and appropriate anti-osteoclastogenesis activity. This study highlights on regulating the balance between pro-osteogenesis and anti-osteoclastogenesis by Zn doping and paves the way for the development of a more osteoinductive calcium phosphate-based material for bone regeneration.

Preparation and Properties of Self‐Setting Calcium Phosphate Scaffolds: Effect of Pore Architecture

Self‐setting calcium phosphate cement (CPC) scaffold with interconnected macropores is hard to prepare without compromising its hydration reaction. Herein, an indirect 3D printing method is using to prepare CPC scaffolds. Detailedly, polycaprolactone (PCL)‐sacrificed models with different strut sizes are first printed by 3D plotting technique, and then the CPC pastes are perfused into the PCL models and then self‐setting. After the removal of the PCL models, the CPC scaffolds with different pore sizes are obtained. It is showed in the results that the prepared CPC scaffolds had uniform shape and 3D interconnected macropore structure. Meanwhile, the compressive strength of CPC scaffolds all meets the requirements of repairing cancellous bone. More interestingly, the CPC scaffold with about 300 μm in macropore size is more conducive to the ingrowth and osteogenesis differentiation of mouse bone marrow mesenchymal stem cells. As a whole, the indirect 3D printing method is a promising strategy for preparing CPC scaffolds, which are hard to form directly by 3D printing technique.

Development of Neovasculature in Axially Vascularized Calcium Phosphate Cement Scaffolds

Augmenting the vascular supply to generate new tissues, a crucial aspect in regenerative medicine, has been challenging. Recently, our group showed that calcium phosphate can induce the formation of a functional neo-angiosome without the need for microsurgical arterial anastomosis. This was a preclinical proof of concept for biomaterial-induced luminal sprouting of large-diameter vessels. In this study, we investigated if sprouting was a general response to surgical injury or placement of an inorganic construct around the vessel. Cylindrical biocement scaffolds of differing chemistries were placed around the femoral vein. A contrast agent was used to visualize vessel ingrowth into the scaffolds. Cell populations in the scaffold were mapped using immunohistochemistry. Calcium phosphate scaffolds induced 2.7-3 times greater volume of blood vessels than calcium sulphate or magnesium phosphate scaffolds. Macrophage and vSMC populations were identified that changed spatially and temporally within the scaffold during implantation. NLRP3 inflammasome activation peaked at weeks 2 and 4 and then declined; however, IL-1β expression was sustained over the course of the experiment. IL-8, a promoter of angiogenesis, was also detected, and together, these responses suggest a role of sterile inflammation. Unexpectedly, the effect was distinct from an injury response as a result of surgical placement and also was not simply a foreign body reaction as a result of placing a rigid bioceramic next to a vein, since, while the materials tested had similar microstructures, only the calcium phosphates tested elicited an angiogenic response. This finding then reveals a potential path towards a new strategy for creating better pro-regenerative biomaterials.

Processing of gelatine coated composite scaffolds based on magnesium and strontium doped hydroxyapatite and yttria-stabilized zirconium oxide

Limited bone bank capacity and risk of infection are some of the main drawbacks of autologous and allogenic grafts, giving rise to synthetic materials for bone tissue implants. The aim of this study was to process and evaluate the mechanical properties and bioactivity of magnesium and strontium doped hydroxyapatite (HAp) scaffolds and investigate the effect of adding zirconium oxide and gelatine coating the scaffolds. Doped nanosized hydroxyapatite powder was synthesized by the hydrothermal method and the scaffolds were made by the foam replica technique and sintered at different temperatures. Yttria-stabilized zirconium oxide (YSZ), synthesized by plasma technology, was used as reinforcement of calcium phosphate scaffolds. Element analysis, phase composition, morphology of the powders and microstructure of the scaffolds were investigated, as well as the compressive strength of the coated and uncoated scaffolds and bioactivity in simulated body fluid (SBF). A microporous structure was achieved with interconnected pores and bioactivity in SBF was confirmed in all cases. The best mechanical properties were given by the coated composite HAp/YSZ scaffolds, withstanding average stresses of over 1019 kPa. These results encourage the idea of use of these scaffolds in bone regenerative therapy and bone tissue engineering.

Nanoparticles based composite coatings with tunable vascular endothelial growth factor and bone morphogenetic protein-2 release for bone regeneration.

Bone healing is a complex cascade involving precisely coordinated spatiotemporal presentation of multiple growth factors (GFs), including osteogenic and angiogenic GFs, and each stage of bone healing requires varying types and content of GFs. In this study, we fabricated a composite nanocoating with tunable vascular endothelial growth factor (VEGF) and bone morphogenetic protein-2 (BMP-2) that was coated on the surface of a polydopamine (PDA)-decorated tertiary calcium phosphate (TCP) scaffold using VEGF-loaded chitosan/bovine serum albumin nanoparticles (CS/BSA-NPs) and BMP-2-loaded poly-L-lysine/oxidized alginate nanoparticles (PLL/OALG-NPs). It was found that VEGF could be efficiently released to promote vascularization in early bone repair stages due to the rapid biodegradation of CS/BSA-NPs, while bone formation can be promoted by a sustained release of BMP-2 from the slowly degrading PLL/OALG-NPs. The composite coating and TCP scaffold can be conjugated due to the excellent adhesive property of PDA. The composite coating can achieve the rapid release of VEGF and sustained release of BMP-2, which can activate GFs for accelerating bone healing.

Construction of Vascularized Tissue Engineered Bone with nHA-Coated BCP Bioceramics Loaded with Peripheral Blood-Derived MSC and EPC to Repair Large Segmental Femoral Bone Defect

The regenerative repair of segmental bone defect (SBD) is an urgent problem in the field of orthopedics. Rapid induction of angiogenesis and osteoinductivity after implantation of scaffold is critical. In this study, a unique tissue engineering strategy with mixture of peripheral blood-derived mesenchymal stem cells (PBMSC) and endothelial progenitor cells (PBEPC) was applied in a 3D-printed biphasic calcium phosphate (BCP) scaffold with highly bioactive nano hydroxyapatite (nHA) coating (nHA/BCP) to construct a novel vascularized tissue engineered bone (VTEB) for rabbit femoral SBD repair. The 2D coculture of PBMSC and PBEPC showed that they could promote the osteogenic or angiogenic differentiation of the cells from each other, especially in the group of PBEPC/PBMSC = 75:25. Besides, the 3D coculture results exhibited that the nHA coating could further promote PBEPC/PBMSC adhesion, proliferation, and osteogenic and angiogenic differentiation on the BCP scaffold. In vivo experiments showed that among the four groups (BCP, BCP-PBEPC/PBMSC, nHA/BCP, and nHA/BCP-PBEPC/PBMSC), the nHA/BCP-PBEPC/PBMSC group induced the best formation of blood vessels and new bone and, thus, the good repair of SBD. It revealed the synergistic effect of nHA and PBEPC/PBMSC on the angiogenesis and osteogenesis of the BCP scaffold. Therefore, the construction of VTEB in this study could provide a possibility for the regenerative repair of SBD.

Graphene oxide coated three-dimensional printed biphasic calcium phosphate scaffold for angiogenic and osteogenic synergy in repairing critical-size bone defect

The custom-tailored medicine requires a developmental strategy that integrates excellent osteogenesis with mechanical stability to enhance the reconstruction of the critical-size bone defect (CSBD) and the healing process in weight-bearing bone. We prepared three-dimensional (3D) printed biphasic calcium phosphate (BCP) scaffolds composited with nano-graphene oxide (GO). The biological effects of the GO/BCP composite scaffolds could induce the differentiation of rat bone marrow stem cells (BMSCs) and the migration of human umbilical vein endothelial cells (HUVECs) for bone repair. The proper ratio of GO in the composite scaffold regulated the composites’ surface roughness and hydrophilicity to a suitable range for the adhesion and proliferation of BMSCs and HUVECs. Besides, the GO/BCP composite scaffold increased osteogenesis and angiogenesis by activating BMP-2, RUNX-2, Smad1/4, and VEGF. The customized intramedullary nail combined with GO/BCP scaffold was applied to repair CSBD (2.0 cm in length) in a beagle femur model. This fixation strategy was confirmed by finite element analysis. In vivo, the results indicated that the custom-made internal fixation provided sufficient stability in the early stage, ensuring bone healing in a considerable mechanical environment. At 9 months postoperatively, longitudinal bony union and blood vessels in osteon were observed in the CSBD area with partial degradation in the 0.3% GO/BCP group. In the three-point bending test, the ultimate load of 0.3% GO/BCP group reached over 50% of the normal femur at 9 months after repair. These results showed a promising application of osteogenic GO/BCP scaffold and custom-made intramedullary nails in repairing CSBD of the beagle femur. This effective strategy could provide an option to treat the clinical CSBD in weight-bearing bones.