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Additive manufacturing has emerged as a transformative tool in biomedical engineering, offering precise control over scaffold design for bone tissue engineering and regenerative medicine. While much attention has been focused on optimizing pore-based scaffold architectures, filament-based microarchitectures remain relatively understudied, despite the fact that the majority of 3D-printers generate filament-based structures. Here, we investigated the influence of filament characteristics on bone regeneration outcomes using a lithography-based additive manufacturing approach. Three distinct filament-based scaffolds (Fil050, Fil083, and Fil125) identical in macroporosity and transparency, crafted from tri-calcium phosphate (TCP) with varying filament thicknesses and distance, were evaluated in a rabbit model of bone augmentation and non-critical calvarial defect. Additionally, two scaffold types differing in filament directionality (Fil and FilG) were compared to elucidate optimal design parameters. Distance of bone ingrowth and percentage of regenerated area within scaffolds were measured by histomorphometric analysis. Our findings reveal filaments of 0.50 mm as the most effective filament-based scaffold, demonstrating superior bone ingrowth and bony regenerated area compared to larger size filament (i.e., 0.83 mm and 1.25 mm scaffolds). Optimized directionality of filaments can overcome the reduced performance of larger filaments. This study advances our understanding of microarchitecture's role in bone tissue engineering and holds significant implications for clinical practice, paving the way for the development of highly tailored, patient-specific bone substitutes with enhanced efficacy.

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Triply periodic minimal surface microarchitectures (TPMS) were developed by mathematicians and evolved in all kingdoms of living organisms. Renowned for their lightweight yet robust attributes, TPMS structures find application in diverse fields, such as the construction of satellites, aircrafts, and electric vehicles. Moreover, these microarchitectures, despite their intricate geometric patterns, demonstrate potential for application as bone substitutes, despite the inherent gothic style of natural bone microarchitecture. Here, we produced three TPMS microarchitectures, D-diamond, G-gyroid, and P-primitive, by 3D printing from hydroxyapatite. We explored their mechanical characterization and, further, implanted them to study their bone augmentation and osteoconduction potential. In terms of strength, the D-diamond and G-gyroid performed significantly better than the P-primitive. In a calvarial defect model and a calvarial bone augmentation model, where osteoconduction is determined as the extent of bony bridging of the defect and bone augmentation as the maximal vertical bone ingrowth, the G-gyroid performed significantly better than the P-primitive. No significant difference in performance was observed between the G-gyroid and D-diamond. Since, in real life, the treatment of bone deficiencies in patients comprises elements of defect bridging and bone augmentation, ceramic scaffolds with D-diamond and G-gyroid microarchitectures appear as the best choice for a TPMS-based scaffold in bone tissue engineering.

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Introduction: Given the ensuing increase in bone and periodontal diseases and defects, de novo bone repair and/or regeneration strategies are constantly undergoing-development alongside advances in orthopedic, oro-dental and cranio-maxillo-facial technologies and improvements in bio-/nano-materials. Indeed, there is a remarkably growing need for new oro-dental functional biomaterials that can help recreate soft and hard tissues and restore function and aesthetics of teeth/ dentition and surrounding tissues. In bone tissue engineering, HydroxyApatite minerals (HAp), the most stable CaP/Calcium Phosphate bioceramic and a widely-used material as a bone graft substitute, have been extensively studied for regenerative medicine and dentistry applications, including clinical use. Yet, limitations and challenges owing principally to its bio-mechanical strength, exist and therefore, research and innovation efforts continue to pursue enhancing its bio-effects, particularly at the nano-scale. Methods: Herein, we report on the physico-chemical properties of a novel nanoHydroxyApatite material obtained from the backbone of Salmon fish (patent-pending); an abundant and promising yet under-explored alternative HAp source. Briefly, our nanoS-HAp obtained via a modified and innovative alkaline hydrolysis-calcination process was characterized by X-ray diffraction, electron microscopy, spectroscopy, and a cell viability assay. Results and Discussion: When compared to control HAp (synthetic, human, bovine or porcine), our nanoS-HAp demonstrated attractive characteristics, a promising biomaterial candidate for use in bone tissue engineering, and beyond.

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Objective: The aim of this study was to evaluate the osteogenic differentiation ability and proliferation of apical papilla stem cells using nanoparticles of Neo MTA and bioactive glass. Methods: Neo MTA and bioactive glass 45S5 nanoparticles were prepared and characterized using a transmission electron microscope and X-ray diffraction. Apical papilla stem cells were harvested from freshly-extracted fully-impacted wisdom teeth, cultured, and characterized using flow cytometric analysis. Tested nanomaterials were mixed and samples were classified into four equal groups as follows; Negative control group: SCAP with Dulbecco's modified eagle's medium, Positive control group: SCAP with inductive media, First experimental group: Neo MTA nanoparticles with SCAP, Second experimental group: Bioactive glass nanoparticles with SCAP. Osteoblastic differentiation was assessed using an alkaline phosphatase assay and RANKL expression using specific polyclonal antibodies by fluorescence microscope. The proliferation of SCAP was assessed using cell count and viability of Trypan Blue in addition to an MTT assay. Results: Isolated SCAP showed a non-hematopoietic origin. Neo MTA showed the highest ALP concentration followed by bioactive glass nanoparticles, and negative control. Bioactive glass nanoparticles showed the highest H score for RANKL protein expression followed by Neo MTA, and negative control. Bioactive glass nanoparticles showed the highest viable cell count. Conclusions: SCAP isolation is achievable from extracted fully impacted immature third molars. Both tested nanobiomaterials have the ability to induce osteogenic differentiation and proliferation of SCAP.

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OBJECTIVES: This preclinical model study aims to evaluate the performance and safety of a novel hydroxyapatite biomaterial (Wishbone Hydroxyapatite, WHA) on guided bone regeneration compared to a commercially available deproteinized bovine bone mineral (Bio-Oss, BO). MATERIAL AND METHODS: Twenty-four beagle dogs were allocated to three timepoint cohorts (4, 12, and 26 weeks) of eight animals each. In all animals, four critical-sized, independent wall mandibular defects were created (32 defects/cohort). Each animal received all four treatments, allocated randomly to separated defects: WHA + collagen membrane (M), BO + M, no treatment (Sham, Sh), and Sh + M. At each timepoint, the specimens were harvested for histologic and histomorphometric analyses to determine the newly formed bone and osteoconductivity. RESULTS: At 4 weeks, bone regeneration was significantly higher for WHA + M (46.8%) when compared to BO + M (21.4%), Sh (15.1%), and Sh + M (23.1%) (p < 0.05); at 12 and 26 weeks, regeneration was similar for WHA and BO. Bone-to-material contact increased over time similarly for WHA + M and BO + M. From a safety point of view, inflammation attributed to WHA + M or BO + M was minimal; necrosis or fatty infiltrate was absent. CONCLUSIONS: WHA + M resulted in higher bone regeneration rate than BO + M at 4 weeks. Both BO + M and WHA + M were more efficient than both Sh groups at all timepoints. Safety and biocompatibility of WHA was favorable and comparable to that of BO.

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The increasing prevalence of bone-related diseases has raised concern about the need for an osteoinductive and mechanically stronger scaffold-based bone tissue engineering (BTE) alternative. A mineralized microenvironment, similar to the native bone microenvironment, is required in the scaffold to recruit and differentiate local mesenchymal stem cells at the bone defect site. Further, extracellular vesicles (EVs), pre-osteoblasts' secretome, contain osteoinductive cargo and have recently been exploited in bone regeneration. This work developed a cell-free and mechanically strong interpenetrating network-based scaffold for BTE by combining the action of osteoinductive EVs with a mineralized microenvironment. The MC3T3 (a pre-osteoblast cell line) is used as a source of EVs and as the target population. The optimal concentration of MC3T3-EVs was first determined to induce osteogenesis in target cells. The osteoinductive potential of the scaffold was estimated in vitro by osteogenesis-related markers like the alkaline phosphatase (ALP) enzyme and calcium content. The MC3T3-EVs cargo was also studied for osteoinductive signals such as ALP, calcium, and mRNA. The findings of this work indicated that MC3T3-EVs at a 90 µg/mL dose had significantly higher ALP activity than 0 µg/mL (1.47-fold), 10 µg/mL (1.41-fold), and 30 µg/mL (1.39-fold) EV-concentration on day 14. Further combination of the optimum dose of EVs with a mineralized microenvironment significantly enhanced ALP activity (1.5-fold) and mineralization (3.36-fold) as compared to the control group on day 7. EV cargo analysis revealed the presence of calcium, the ALP enzyme, and the mRNAs necessary for osteogenesis and angiogenesis. ALP activity was significantly boosted in the EV-containing target cells as early as day 1, and mineralization began on day 7 because MC3T3-EVs carry ALP enzymes and calcium as cargo. When osteoinductive EVs were combined with an osteoconductive mineralized microenvironment, osteogenesis was significantly enhanced in target cells at early time points. The interaction between osteoinductive EVs and the mineralized milieu facilitates the process of osteogenesis in the target cells and suggests a potential cell-free strategy for in vivo bone repair.

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Autologous bone remains the gold standard bone substitute in clinical practice. Therefore, the microarchitecture of newly developed synthetic bone substitutes, which reflects the spatial distribution of materials in the scaffold, aims to recapitulate the natural bone microarchitecture. However, the natural bone microarchitecture is optimized to obtain a mechanically stable, lightweight structure adapted to the biomechanical loading situation. In the context of synthetic bone substitutes, the application of a Triply Periodic Minimum Surface (TPMS) algorithm can yield stable lightweight microarchitectures that, despite their demanding architectural complexity, can be produced by additive manufacturing. In this study, we applied the TPMS derivative Adaptive Density Minimal Surfaces (ADMS) algorithm to produce scaffolds from hydroxyapatite (HA) using a lithography-based layer-by-layer methodology and compared them with an established highly osteoconductive lattice microarchitecture. We characterized them for compression strength, osteoconductivity, and bone regeneration. The in vivo results, based on a rabbit calvaria defect model, showed that bony ingrowth into ADMS constructs as a measure of osteoconduction depended on minimal constriction as it limited the maximum apparent pore diameter in these scaffolds to 1.53 mm. Osteoconduction decreased significantly at a diameter of 1.76 mm. The most suitable ADMS microarchitecture was as osteoconductive as a highly osteoconductive orthogonal lattice microarchitecture in noncritical- and critical-size calvarial defects. However, the compression strength and microarchitectural integrity in vivo were significantly higher for scaffolds with their microarchitecture based on the ADMS algorithm when compared with high-connectivity lattice microarchitectures. Therefore, bone substitutes with high osteoconductivity can be designed with the advantages of the ADMS-based microarchitectures. As TPMS and ADMS microarchitectures are true lightweight structures optimized for high mechanical stability with a minimal amount of material, such microarchitectures appear most suitable for bone substitutes used in clinical settings to treat bone defects in weight-bearing and non-weight-bearing sites.

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One of the objectives of bone tissue engineering is to produce scaffolds that are biocompatible, osteoinductive, and mechanically equivalent to the natural extracellular matrix of bone in terms of structure and function. Reconstructing the osteoconductive bone microenvironment into a scaffold can attract native mesenchymal stem cells and differentiate them into osteoblasts at the defect site. The symbiotic relationship between cell biology and biomaterial engineering could result in composite polymers containing the necessary signals to recreate tissue- and organ-specific differentiation. In the current work, drawing inspiration from the natural stem cell niche to govern stem cell fate, the cell-instructive hydrogel platforms were constructed by engineering the mineralized microenvironment. This work employed two different hydroxyapatite delivery strategies to create a mineralized microenvironment in an alginate-PEGDA interpenetrating network (IPN) hydrogel. The first approach involved coating of nano-hydroxyapatite (nHAp) on poly(lactide-co-glycolide) microspheres and then encapsulating the coated microspheres in an IPN hydrogel for a sustained release of nHAp, whereas the second approach involved directly loading nHAp into the IPN hydrogel. This study demonstrate that both direct encapsulation and a sustained release approach showed enhanced osteogenesis in target-encapsulated cells; however, direct loading of nHAp into the IPN hydrogel increased the mechanical strength and swelling ratio of the scaffold by 4.6-fold and 1.14-fold, respectively. In addition, the biochemical and molecular studies revealed improved osteoinductive and osteoconductive potential of encapsulated target cells. Being less expensive and simple to perform, this approach could be beneficial in clinical settings.

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Background: Bone tunnel enlargement after single-bundle anterior cruciate ligament reconstruction remains an unsolved problem that complicates revision surgery. Hypothesis: Positioning of an osteoconductive scaffold at the femoral tunnel aperture improves graft-to-bone incorporation and thereby decreases bone tunnel widening. Study Design: Randomized controlled trial; Level of evidence, 1. Methods: In a 1:1 ratio, 56 patients undergoing primary anterior cruciate ligament reconstruction were randomized to receive femoral fixation with cortical suspension fixation and secondary press-fit fixation at the tunnel aperture of the tendon graft only (control) or with augmentation by an osteoconductive scaffold (intervention). Adverse events, patient-reported outcomes, and passive knee stability were recorded over 2 years after the index surgery. Three-dimensional bone tunnel widening was assessed using computed tomography at the time of surgery and 4.5 months and 1 year postoperatively. Results: The intervention group exhibited a similar number of adverse events as the control group (8 vs 10; P = .775) including 2 partial reruptures in both groups. The approach was feasible, although 1 case was encountered where the osteoconductive scaffold was malpositioned without adversely affecting the patient's recovery. There was no difference between the intervention and control groups in femoral bone tunnel enlargement, as expressed by the relative change in tunnel volume from surgery to 4.5 months (mean ± SD, 36% ± 25% vs 40% ± 25%; P = .644) and 1 year (19% ± 20% vs 17% ± 25%; P =.698). Conclusion: Press-fit graft fixation with an osteoconductive scaffold positioned at the femoral tunnel aperture is safe but does not decrease femoral bone tunnel enlargement at postoperative 1 year. Registration: NCT03462823 (ClinicalTrials.gov identifier).

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Chitosan (CS) is a natural biopolymer that shows promise as a biomaterial for bone-tissue regeneration. However, because of their limited ability to induce cell differentiation and high degradation rate, among other drawbacks associated with its use, the creation of CS-based biomaterials remains a problem in bone tissue engineering research. Here we aimed to reduce these disadvantages while retaining the benefits of potential CS biomaterial by combining it with silica to provide sufficient additional structural support for bone regeneration. In this work, CS-silica xerogel and aerogel hybrids with 8 wt.% CS content, designated SCS8X and SCS8A, respectively, were prepared by sol-gel method, either by direct solvent evaporation at the atmospheric pressure or by supercritical drying in CO2, respectively. As reported in previous studies, it was confirmed that both types of mesoporous materials exhibited large surface areas (821 m2g-1-858 m2g-1) and outstanding bioactivity, as well as osteoconductive properties. In addition to silica and chitosan, the inclusion of 10 wt.% of tricalcium phosphate (TCP), designated SCS8T10X, was also considered, which stimulates a fast bioactive response of the xerogel surface. The results here obtained also demonstrate that xerogels induced earlier cell differentiation than the aerogels with identical composition. In conclusion, our study shows that the sol-gel synthesis of CS-silica xerogels and aerogels enhances not only their bioactive response, but also osteoconduction and cell differentiation properties. Therefore, these new biomaterials should provide adequate secretion of the osteoid for a fast bone regeneration.

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Triply periodic minimal surfaces (TPMSs) are found to be promising microarchitectures for bone substitutes owing to their low weight and superior mechanical characteristics. However, existing studies on their application are incomplete because they focus solely on biomechanical or in vitro aspects. Hardly any in vivo studies where different TPMS microarchitectures are compared have been reported. Therefore, we produced hydroxyapatite-based scaffolds with three types of TPMS microarchitectures, namely Diamond, Gyroid, and Primitive, and compared them with an established Lattice microarchitecture by mechanical testing, 3D-cell culture, and in vivo implantation. Common to all four microarchitectures was the minimal constriction of a sphere of 0.8 mm in diameter, which earlier was found superior in Lattice microarchitectures. Scanning by µCT revealed the precision and reproducibility of our printing method. The mechanical analysis showed significantly higher compression strength for Gyroid and Diamond samples compared with Primitive and Lattice. After in vitro culture with human bone marrow stromal cells in control or osteogenic medium, no differences between these microarchitectures were observed. However, from the TPMS microarchitectures, Diamond- and Gyroid-based scaffolds showed the highest bone ingrowth and bone-to-implant contact in vivo. Therefore, Diamond and Gyroid designs appear to be the most promising TPMS-type microarchitectures for scaffolds produced for bone tissue engineering and regenerative medicine. Impact Statement Extensive bone defects require the application of bone grafts. To match the existing requirements, scaffolds based on triply periodic minimal surface (TPMS)-based microarchitectures could be used as bone substitutes. This work is dedicated to the investigation of mechanical and osteoconductive properties of TPMS-based scaffolds to determine the influencing factors on differences in their behavior and choose the most promising design to be used in bone tissue engineering.

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The early phase of bone healing is a complex and poorly understood process. With additive manufacturing, we can generate a specific and customizable library of bone substitutes to explore this phase. In this study, we produced tricalcium phosphate-based scaffolds with microarchitectures composed of filaments of 0.50 mm in diameter, named Fil050G, and 1.25 mm named Fil125G, respectively. The implants were removed after only 10 days in vivo followed by RNA sequencing (RNAseq) and histological analysis. RNAseq results revealed upregulation of adaptive immune response, regulation of cell adhesion, and cell migration-related genes in both of our two constructs. However, significant overexpression of genes linked to angiogenesis, regulation of cell differentiation, ossification, and bone development was observed solely in Fil050G scaffolds. Moreover, quantitative immunohistochemistry of structures positive for laminin revealed a significantly higher number of blood vessels in Fil050G samples. Furthermore, µCT detected a higher amount of mineralized tissue in Fil050G samples suggesting a superior osteoconductive potential. Hence, different filament diameters and distances in bone substitutes significantly influence angiogenesis and regulation of cell differentiation involved in the early phase of bone regeneration, which precedes osteoconductivity and bony bridging seen in later phases and as consequence, impacts the overall clinical outcome.

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Polymeric membranes are widely used in guided bone regeneration (GBR), particularly in dentistry. In addition, bioactive glasses can be added to the polymers in order to develop a matrix that is osteoconductive and osteoinductive, increasing cell adhesion and proliferation. The bioactive glasses allow the insertion into its network of therapeutic ions in order to add specific biological properties. The addition of zinc into bioactive glasses can promote antibacterial activity and induce the differentiation and proliferation of the bone cells. In this study, bioactive glasses containing zinc (0.25, 0.5, 1 and 2 mol%) were developed and structurally and biologically characterized. The biological results show that the Zn-containing bioactive glasses do not present significant antibacterial activity, but the addition of zinc at the highest concentration does not compromise the bioactivity and promotes the viability of Saos-2 cells. The cell culture assays in the membranes (PCL, PCL:BG and PCL:BGZn2) showed that zinc addition promotes cell viability and an increase in alkaline phosphatase (ALP) production.

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Developing a scaffold for efficient and functional bone regeneration remains challenging. To accomplish this goal, a "scaffold-on-a-chip" device was developed as a platform to aid with the evaluation process. The device mimics a microenvironment experienced by a transplanted bone scaffold. The device contains a circular space at the center for scaffold insert and microfluidic channel that encloses the space. Such a design allows for monitoring of cell behavior at the blood-scaffold interphase. MC3T3-E1 cells were cultured with three different types of scaffold inserts to test its capability as an evaluation platform. Cellular behaviors, including migration, morphology, and osteogenesis with each scaffold, were analyzed through fluorescence images of live/dead assay and immunocytochemistry. Cellular behaviors, such as migration, morphology, and osteogenesis, were evaluated. The results revealed that our platform could effectively evaluate the osteoconductivity and osteoinductivity of scaffolds with various properties. In conclusion, our proposed platform is expected to replace current in vivo animal models as a highly relevant in vitro platform and can contribute to the fundamental study of bone regeneration.

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BACKGROUND: The Biocompatibility between osteoprogenitor cells and bone substitutes is necessary for cell differentiation and osteogenesis. The aim of this study was to assess the in vitro effect of bovine (Geistlich BioOss®), porcine (OsteoBiol Gen-Os®) and beta-tricalcium phosphate (Cerasorb M®) bone substitutes, and their combination with polyphenol epigallocatechin-3-gallate (EGCG), upon cultured dental pulp stem cells (DPSCs). METHODS: The DPSCs were isolated from third molars extracted from healthy individuals and seeded with 5 mg/ml of Bio-Oss® (BO), Gen-Os® (GO) and Cerasorb® (CE) in combination with EGCG 1 µM. The effects were evaluated based on cell viability / cytotoxicity assay (MTT, cell viability staining test), cell migration, scanning electron microscopy (SEM), and alkaline phosphatase (ALP) activity. RESULTS: BO and CE produced negative effects upon cell viability and migration, and GO and CE resulted in deficient cell adhesion. On the other hand, all the biomaterials exerted no negative effects upon ALP activity. Interestingly, the addition of EGCG reverted the cytotoxic effect and the loss of migration capacity in the BO and CE groups, and improved cell adhesion in the GO and CE groups. Furthermore, EGCG promoted an overall increased in ALP activity. CONCLUSION: The addition of EGCG to the tested biomaterials BO, GO and CE reverts their negative impact on DPSCs, and improves their biocompatibility with cultured DPSCs. The use of EGCG, thus, appears to be a promising strategy for restoring and enhancing the osteoconductive properties of BO, GO and CE in bone regeneration treatments.

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BACKGROUND: Caprine species satisfy the conditions of an ideal donor animal when compared to bovine species that has been extensively studied and commercialized for bone xenograft. Histopathological and radiological evaluations of caprine demineralized bone matrix (CDBM) were therefore carried out for fracture healing properties for its possible use in bone grafting procedures. MATERIALS AND METHODS: Twenty-four rabbits were used for this study and were divided randomly into three groups of eight (n = 8) rabbits each. Critical bone defect was created on the ulnar diaphysis under xylazine-ketamine anaesthesia for autogenous bone graft (ABG) group, CDBM group and the last group was left unfilled as negative control (NC). Immediate post-grafting radiograph was taken and repeated on days 14, 28, 42 and 56 to monitor the evidence of radiographic healing. The animals were euthanized on day 56 and defect sites were harvested for histopathology. RESULTS: There was a progressive evidence of radiographic healing and bone formation in all the groups with significance difference (P = 0.0064). When compared with ABG, NC differ significantly (P < 0.0001) whereas the CDBM did not differ significantly (P = 0.6765). The histopathology sections of ABG and CDBM showed normal bone tissue while the NC section was predominated by fibrous connective tissue. There was therefore an overall significant difference (P = 0.0001) in which CDBM did not differ from ABG (P = 0.2946) while NC did (P = 0.0005). CONCLUSION: The ABG and CDBM groups showed a similar healing effect in the critical bone defect. Therefore, CDBM could be used as an effective alternative to ABG in orthopaedics to circumvent the limitations and complications associated with it. LEVEL OF EVIDENCE: Not applicable.

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Synthetic implants are used to treat large bone defects that are often unable to regenerate, for example those caused by osteoporosis. It is necessary that the materials used to manufacture them are biocompatible and resorbable. Polymer-ceramic composites, such as those based on poly(L-lactide) (PLLA) and calcium phosphate ceramics (Ca-P), are often used for these purposes. In this study, we attempted to investigate an innovative strategy for two-step (dual) modification of composites and their components to improve the compatibility of composite components and the adhesion between PLA and Ca-P whiskers, and to increase the mechanical strength of the composite, as well as improve osteological bioactivity and prevent bone resorption in composites intended for bone regeneration. In the first step, Ca-P whiskers were modified with a saturated fatty acid namely, lauric acid (LA), or a silane coupling agent γ-aminopropyltriethoxysilane (APTES). Then, the composite, characterized by the best mechanical properties, was modified in the second stage of the work with an active chemical compound used in medicine as a first-line drug in osteoporosis-sodium alendronate, belonging to the group of bisphosphonates (BP). As a result of the research covered in this work, the composite modified with APTES and alendronate was found to be a promising candidate for future biomedical engineering applications.

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Bone substitute materials have been successfully used for bone defects in orthopedics and trauma surgery for a long time; however, there are cases, especially in bone defects with a critical size, in which the treatment is complicated. Nowadays, multiple bone substitute materials are available. Autologous cancellous bone grafts remain the gold standard among the bone replacement materials; however, donor site morbidity and the limited availability of autologous cancellous bone represent restrictions for autologous bone grafting. Allogeneic cancellous bone grafts have also been successfully for years in the treatment of bone defects; however, infection rates of more than 10% have been described for the use of allogeneic cancellous bone. By introducing synthetic bone substitutes further alternatives are currently available to the user for the individual treatment of bone defects. The aim of this study is to demonstrate the advantages and disadvantages of various synthetic bone substitute materials.

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BACKGROUND: To support bone regeneration, 3D-printed templates function as temporary guides. The preferred materials are synthetic polymers, due to their ease of processing and biological inertness. Poly(lactide-co-trimethylene carbonate) (PLATMC) has good biological compatibility and currently used in soft tissue regeneration. The aim of this study was to evaluate the osteoconductivity of 3D-printed PLATMC templates for bone tissue engineering, in comparison with the widely used 3D-printed polycaprolactone (PCL) templates. METHODS: The printability and physical properties of 3D-printed templates were assessed, including wettability, tensile properties and the degradation profile. Human bone marrow-derived mesenchymal stem cells (hBMSCs) were used to evaluate osteoconductivity and extracellular matrix secretion in vitro. In addition, 3D-printed templates were implanted in subcutaneous and calvarial bone defect models in rabbits. RESULTS: Compared to PCL, PLATMC exhibited greater wettability, strength, degradation, and promoted osteogenic differentiation of hBMSCs, with superior osteoconductivity. However, the higher ALP activity disclosed by PCL group at 7 and 21 days did not dictate better osteoconductivity. This was confirmed in vivo in the calvarial defect model, where PCL disclosed distant osteogenesis, while PLATMC disclosed greater areas of new bone and obvious contact osteogenesis on surface. CONCLUSIONS: This study shows for the first time the contact osteogenesis formed on a degradable synthetic co-polymer. 3D-printed PLATMC templates disclosed unique contact osteogenesis and significant higher amount of new bone regeneration, thus could be used to advantage in bone tissue engineering.

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The aim of this study was to evaluate the in vivo performance of two different deproteinized bovine bone (DBB) grafting materials: DBBB (Bio-Oss®) and DBBL (Laddec®), for the regeneration of critically sized (8 mm) defects in rabbit's calvaria. Three round-shaped defects were surgically created in the calvaria of 13 New Zealand White rabbits proximal to the coronal suture in the parietal bone. Two of the defects were filled with one of the grafting materials while a third was left empty to serve as a negative control. Bone regeneration properties were evaluated at 4- and 8-weeks after implantation by means of histological and histomorphometrical analyses. Statistical analyses were performed through a mixed model analysis with fixed factors of time and material. Histological evaluation of the control group evidenced a lack of bridging bone formation across the defect sites at both evaluation time points. For the experimental groups, new bone formation was observed around the defect periphery and to progress radially inwards to the center of the defect site, regardless of the grafting material. Histomorphometric analyses at 4 weeks demonstrated higher amount of bone formation through the defect for DBBB group. However, at 8 weeks, DBBL and DBBB demonstrated osteoconductivity and low resorption rates with evidence of statistically similar bone regeneration through the complete boney defect. Finally, DBBB presented lower soft tissue migration within the defect when compared to DBBL at both evaluation time points. DBBB and DBBL presented similar bone regeneration performance and slow resorption rates. Although both materials promoted bone regeneration through the complete defect, DBBB presented lower soft tissue migration within the defects at 4- and 8-weeks.

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