Critical-sized Defects Research Articles

Improved new bone formation capacity of hyaluronic acid-bone substitute compound in rat calvarial critical size defect

BackgroundBone loss of residual alveolar ridges is a great challenge in the field of dental implantology. Deproteinized bovine bone mineral (DBBM) is commonly used for bone regeneration, however, it is loose and difficult to handle in clinical practice. Hyaluronic acid (HA) shows viscoelasticity, permeability and excellent biocompatibility. The aim of this study is to evaluate whether high-molecular-weight (MW) HA combined with DBBM could promote new bone formation in rat calvarial critical size defects (CSDs).Materials and methodsRat calvarial CSDs (5 mm in diameter) were created. Rats (n = 45) were randomly divided into 3 groups: HA-DBBM compound grafting group, DBBM particles only grafting group and no graft group. Defect healing was assessed by hematoxylin-eosin staining and histomorphometry 2, 4 and 8 weeks postop, followed by Micro-CT scanning 8 weeks postop. Statistical analyses were performed by ANOVA followed by Tukey's post hoc test with P < 0.05 indicating statistical significance.ResultsAll rats survived after surgery. Histomorphometric evaluation revealed that at 2, 4 and 8 weeks postop, the percentage of newly formed bone was significantly greater in HA-DBBM compound grafting group than in the other two groups. Consistently, Micro-CT assessment revealed significantly more trabecular bone (BV/TV and Tb.N) in HA-DBBM compound group than in the other two groups, respectively (P < 0.05). Moreover, the trabecular bone was significantly more continuous (Tb.Pf) in HA-DBBM compound group than in the other two groups, respectively (P < 0.05).ConclusionHA not only significantly promoted new bone formation in rats calvarial CSDs but also improved the handling ability of DBBM.

Fabrication, physicochemical properties, and cytocompatibility of 3D-printed Sr2MgSi2O7 bioceramic scaffolds calcined at different temperature for bone repair

3D-printed bioactive ceramic scaffolds hold significant promise for addressing critical-size bone defects, owing to their biodegradable and osteogenic properties. Numerous 3D-printed silicate-based (e.g., Sr2MgSi2O7) ceramic scaffolds have been developed, but the rapid degradation and insufficient mechanical properties of which impede their application efficacy. Despite recognizing the pivotal role of calcination temperature in shaping the physicochemical and biological attributes of 3D-printed scaffolds, its specific influence on Sr2MgSi2O7 bioceramics remains ambiguous. This study elucidated the fabrication, physicochemical properties, and cytocompatibility of 3D-printed Sr2MgSi2O7 ceramic scaffolds calcined at different temperatures. The findings revealed a significant enhancement in compressive strength (up to 19.9 MPa) and a decrease in degradation rate with increasing calcination temperature. Compared to conventional tricalcium phosphate scaffolds, Sr2MgSi2O7 scaffold demonstrated superior osteogenic potential, particularly when calcined at 1250°C. With commendable mechanical strength and remarkable osteogenic capabilities, the Sr2MgSi2O7 scaffold emerges a promising avenue for repairing critical-size bone defects.

Multifunctional Bone Regeneration Membrane with Flexibility, Electrical Stimulation Activity and Osteoinductive Activity.

The use of membrane-based guided bone regeneration techniques has great potential for single-stage reconstruction of critical-sized bone defects. Here, a multifunctional bone regeneration membrane combining flexible elasticity, electrical stimulation (ES) and osteoinductive activity is developed by in situ doping of MXene 2D nanomaterials with conductive functionality and β-TCP particles into a Poly(lactic acid-carbonate (PDT) composite nano-absorbable membrane (P/T/MXene) via electrostatic spinning technique. The composite membrane has good feasibility due to its temperature sensitivity, elastic memory capacity, coordinated degradation profile and easy preparation process. In vitro experiments showed the P/T/MXene membrane effectively promoted the recruitment and osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) under ES and enhanced the angiogenic capacity of endothelial cells, which synergistically promoted bone regeneration through neovascularization. In addition, an in vivo rat model of cranial bone defects further confirmed the bone regeneration efficacy of the P/T/MXene membrane. In conclusion, the developed P/T/MXene membrane can effectively promote bone regeneration through their synergistic multifunctional effects, suggesting the membranes have great potential for guiding tissue regeneration and providing guidance for the biomaterials design.

Assessment of Novel Surgical Procedures Using Bone Morphogenic Protein-7 Infused Into Decellularised Muscle and Bioactive Ceramic: A Histological Analysis.

Reconstruction of critical bone defects is considered a challenge due to vascular reperfusion injury that may occur. The present study hypothesized that the use of decellularized muscle scaffold (DMS) and bone morphogenic protein-7 (BMP-7), along with resorbable bioactive ceramic silica calcium phosphate cement (SCPC) seeded with human bone marrow stromal cells, can expedite bone formation and maturation. Surgical bone defects were created in 20 nude transgenic mice. In experimental group 1 (n = 10), a critical-size (4mm) calvarial defect was made and grafted with DMS-BMP-7/SCPC. In situ human bone marrow stromal cells [human mesenchymal stromal cells (hMSC)] were seeded thereafter. As a control, group 2 (n = 10) was treated with DMS/SCPC seeded with hMSCs. After 8 weeks, bone regeneration was evaluated using histology and histomorphometry for both groups. Histological examination showed bone regeneration crossing the gap (experimental group 1), bone regeneration was noted at the defect periphery, and scattered islands of bone at the canters of the defects (control group 2). New bone formation and maturation were superior in the groups treated with the DMS/BMP-7/SCPC/hMSC constructs. The quantitative histological assessment revealed that the average bone surface area was 255 ± 25mm2, which was 1.5 times the surface area of group 2, which was reported at 170 ± 35mm2. The reported difference was considered statistically significant (P < 0.05). The DMS-BMP-7/SCPC scaffold induced bone regeneration and neovascularization in critical-size defects.

Fabrication of 3D PCL/PVP scaffolds using monosodium glutamate as porogen by solvent casting/particulate leaching method for oral and maxillofacial bone tissue engineering

The design of three-dimensional (3D) scaffolds should focus on creating highly porous, 3D structures with an interconnected pore network that supports cell growth. The scaffold's pore interconnectivity is directly linked to vascularization, cell seeding, guided cell migration, and transportation of nutrients and metabolic waste. In this study, different types of food flavors including monosodium glutamate, sugar, and sodium chloride were used as the porogens along with PCL/PVP blend polymer for solvent casting/particulate leaching method. The morphology, porosity, interconnectivity, chemical composition, water absorption, and mechanical properties of the fabricated scaffolds are carefully characterized. The scaffolds are biocompatible in bothin vitroandin vivoexperiments and do not trigger any inflammatory response while enhancing new bone formation and vascularization in rabbit calvaria critical-sized defects. The new bone merges and becomes denser along with the experiment timeline. The results indicate that the 3D PCL/PVP scaffolds, using monosodium glutamate as porogen, exhibited suitable biological performance and held promise for bone tissue engineering in oral and maxillofacial surgery.

Impact of Freeze-dried Corticocancellous Bone Allograft Combined with Enamel Matrix Derivative in the Treatment of Critical-sized Calvarial Bone Defects: An Animal Study.

This study compared the quality and quantity of newly formed bone in rabbits' critical-sized calvarial defects filled with enamel matrix derivative (EMD) combined with freeze-dried bone allograft (FDBA) vs FDBA alone. A total of 24 adult male white New Zealand rabbits were included. In each rabbit, three bone defects with a diameter of 8 mm were created on the calvarium bone; the first defect was left untreated, while the second was filled with FDBA, and the third was filled with EMD + FDBA. Twelve rabbits were randomly euthanized after a month, and the remaining 2 month postsurgery. Bone sections were histologically evaluated by hematoxylin and eosin and vascular endothelial growth factor (VEGF), alkaline phosphatase (ALP), osteoprotegerin (OPG), and receptor activator of NF-kappaB (RANK) immune-histochemical staining. An improvement in the newly formed bone percentage was found in the defects filled with EMD + FDBA in comparison with FDBA and control defects at 1 month and 2 months postsurgery. Additionally, the expression of VEGF, ALP, OPG, and RANK showed highly significant differences in the defects filled with EMD + FDBA compared to the FDBA and control ones at 1 month postsurgery (p = 0.001). Meanwhile, VEGF and ALP expression showed a significant decrease in defects filled with EMD + FDBA compared to the FDBA and control ones (p = 0.001), while OPG and RANK expression showed non-significant differences between treated groups at 2 months postsurgery. Enamel matrix derivative combined with FDBA has a synergistic effect on bone formation and graft substitution. This combination accelerates the expression of VEGF, ALP, OPG, and RANK. The combination of EMD and FDBA accelerates and ameliorates the quality of newly formed bone, aiding in maxillofacial reconstruction. How to cite this article: Zakri RN, Grawish ME, Mowafey B, et al. Impact of Freeze-dried Corticocancellous Bone Allograft Combined with Enamel Matrix Derivative in the Treatment of Critical-sized Calvarial Bone Defects: An Animal Study. J Contemp Dent Pract 2024;25(5):424-431.

Bone Regeneration of Rat Critical-Sized Calvarial Defects by the Combination of Photobiomodulation and Adipose-Derived Mesenchymal Stem Cells.

Introduction: This study explored the synergistic effects of low-level laser therapy (LLLT) and adipose-derived stem cells (ADSCs) on cranial bone regeneration in rats, addressing the limitations of autogenous grafts and advancing bone tissue engineering with innovative photobiomodulation (PBM) applications. Methods: Sixty Wistar rats were allocated to 5 separate groups randomly; (1) natural bovine bone mineral (NBBM); (2) NBBM+LLLT; (3) NBBM+allogenic ADSCs; (4) NBBM+allogenic ADSCs+LLLT; (5) Only defects. 8-mm calvarial defects were made in each rat in the surgical procedure. A diode laser was applied with the following parameters (continuous mode, power of 100mW, wavelength of 808nm, and 4 J/cm2 energy density) immediately after the procedure and every other day. Bone samples were obtained and assessed histomorphometrically and histologically after staining with hematoxylin and eosin (H&E). Results: Different volumes of bony material were observed in two weeks; 2.94%±1.00 in group 1, 5.1%±1.92 in group 2, 7.11%±2.82 in group 3, 7.34%±2.31 in group 4, and 2.01%±0.83 in group 5 (P<0.05). On the other hand, foreign body residuals were up by 23% in the groups with scaffolding by the end of 2 weeks. Four weeks of observation led to 6.74 %±1.95, 13.24%±1.98, 15.76%±1.19, 15.92%±3.4, and 3.11%±1.00 bone formation in groups 1 to 5, respectively (P<0.05). Generally, the difference between groups 2-4 was not statistically significant based on different types of bone and the extent of inflammation. Conclusion: Bearing in mind the limitations of our research, it was demonstrated that ADSCs in combination with PBM have promising effects on bone tissue regeneration in sizeable bony defects. Furthermore, this study also showed that PBM usage improved the newly regenerated bone quality.

Boron substitution in silicate bioactive glass scaffolds to enhance bone differentiation and regeneration

Commercially available bioactive glasses (BAGs) are exclusively used in powder form, due to their tendency to crystallize. Silicate BAG 1393 was developed to allow fiber drawing and scaffold sintering, but its slow degradation limits its potential. To enable scaffold manufacturing while maintaining glass dissolution rate close to that of commercially available BAGs, the borosilicate glass 1393B20 was developed. This study investigates the potential of 1393B20 scaffolds to support bone regeneration and mineralization in vitro and in vivo, in comparison to silicate 1393. Both scaffolds supported human adipose stem cells proliferation, either in direct contact for the 1393, or mainly around for the 1393B20. Similarly, both BAGs induced osteogenesis and angiogenesis in vitro, with a better pro-angiogenic influence of the 1393B20. In addition, these scaffolds supported bone regeneration and osteoclast/osteoblast activity in vivo in critical-sized rat calvarial defect. Nevertheless, mineralization and collagen formation were significantly enhanced for the 1393B20, at 3-months post-implantation, assigned to faster and more complete dissolution of the scaffolds. Thus, 1393B20 demonstrates greater promise for bone tissue engineering certainly due to its time-controlled release of boron and silicon. Statement of significanceBioactive glasses (BAGs) show great promise in bone tissue engineering as they effectively bond with bone tissue, fostering integration and regeneration. Silicate BAG 1393 was developed to allow fiber drawing and scaffold sintering, but its slow degradation limits its potential. To enable scaffold manufacturing while maintaining glass dissolution rate close to that of commercially available BAGs, the borosilicate glass 1393B20 was developed. Both BAGs induced osteogenesis and angiogenesis in vitro, with a better pro-angiogenic influence of the 1393B20. The presence of boron in the 1393B20 enhanced mineralization and collagen formation in vivo compared to 1393, probably due to its faster dissolution rate. Here, 1393B20 demonstrated greater promise for bone tissue engineering compared to the well-known 1393 BAG.

Bioactive poly(ethylene glycol)-chondroitin sulfate-triple helical recombinant collagen hydrogel for enhanced cranial defect repair

The reconstruction of critical-size calvarial defects remains a fundamental challenge. Recombinant collagen has gained significant attention in bone tissue engineering owing to its remarkable bioactivity and non-immunogenicity. Herein, we have for the first time developed a bioactive poly(ethylene glycol)-chondroitin sulfate-triple helical recombinant collagen (PEG-ChS-THRC) hydrogel for enhanced bone regeneration in cranial defects. A simple and mild crosslinking reaction of two-arm polyethylene glycol active ester (NHS-PEG-NHS), adipic dihydrazide modified chondroitin sulfate (ChS-ADH) and triple helical recombinant collagen (THRC) leads to the formation of the PEG-ChS-THRC hydrogel. The hydrogel demonstrates interconnected porous structures, enhanced mechanical strength, diminished swelling ratios and adjustable biodegradability. It possesses exceptional biocompatibility and bioactivity, significantly facilitating cell proliferation, adhesion, migration, and osteogenic differentiation of BMSCs. Micro-computed tomography (micro-CT), magnetic resonance imaging (MRI) and histological characterization of rat models with critical-size cranial defects have consistently demonstrated that the PEG-ChS-THRC hydrogel significantly promotes bone tissues regeneration. The innovative bioactive scaffold provides a remarkably improved remedy for critical-size cranial defects, holding greatly promising applications in the fields of bone tissue regeneration.Graphical

Electrospun polycaprolactone-chitosan nanofibers on a zinc mesh as biodegradable guided bone-regeneration membranes with enhanced mechanical, antibacterial, and osteogenic properties for alveolar bone-repair applications

Guided bone-regeneration membrane (GBRM) is commonly used in bone-repair surgery because it blocks fibroblast proliferation and provides spatial support in bone-defect spaces. However, the need for removal surgery and the lack of antibacterial properties of conventional GBRM limit its therapeutic applicability for alveolar bone defects. Here we developed a GBRM for alveolar bone-repair and -regeneration applications through double-sided electrospinning of polycaprolactone and chitosan layers on a Zn mesh surface (denoted DSZM). The DSZM showed a UTS of ∼25.6 MPa, elongation of ∼16.1%, strength-elongation product of ∼0.413 GPa%, and ultrahigh spatial maintenance ability, and the UTS was over 6 times higher than that of commercial Bio-Gide membrane. The DSZM exhibited a corrosion rate of ∼17 µm/y and a Zn ion concentration of ∼0.23 µg/ml after 1 month of immersion in Hanks’ solution. The DSZM showed direct and indirect cytocompatibility with exceptional osteogenic differentiation and calcium deposition toward MC3T3-E1 cells. Further, the DSZM showed strongly sustained antibacterial activity against S. aureus and osteogenesis in a rat critical-sized maxillary defect model. Overall, the DSZM fits the requirements for alveolar bone-repair and -regeneration applications as a biodegradable GBRM material due to its spatial support, suitable degradability, cytocompatibility, and antibacterial and osteogenic capabilities. Statement of significanceThis work reports the mechanical properties, antibacterial ability and osteogenic properties of electrospun PCL-CS nanofiber on Zn mesh as biodegradable guided bone-regeneration membrane for alveolar bone-repair applications. Our findings demonstrate that the DSZM prepared by double-sided electrospinning of PCL-CS layers on Zn mesh showed a UTS of ∼25.6 MPa, elongation of ∼16.1%, strength-elongation product of ∼0.413 GPa%, and ultrahigh spatial maintenance ability, and the UTS was over 6 times greater than that of commercial Bio-Gide® membrane. The DSZM showed direct and indirect cytocompatibility with exceptional osteogenic differentiation and calcium deposition toward MC3T3-E1 cells. Further, the DSZM showed strongly sustained antibacterial activity against S. aureus and osteogenesis in a rat critical-sized maxillary defect model.

The management of critical bone defects: outcomes of a systematic approach

BackgroundThe reconstruction of segmental long bone defects remains one of ‘The holy grails of orthopaedics’. The optimal treatment of which remains a topic of great debate. This study aimed to evaluate the outcomes following the management of critical-sized bone defects using a classification-based treatment algorithm.MethodsA retrospective review of all patients undergoing treatment for segmental diaphyseal defects of long bones at a tertiary-level limb reconstruction unit between January 2016 and December 2021, was performed. The management of the bone defect was standardised as per the classification by Ferreira and Tanwar (2020).ResultsA total of 96 patients (mean age 39.8, SD 15.2) with a minimum six months follow-up were included. Most bone defects were the result of open fractures (75/96) with 67% associated with Gustilo-Anderson IIIB injuries. There was a statistical difference in the likelihood of union between treatment strategies with more than 90% of cases undergoing acute shortening and bone transport achieving union and only 72% of cases undergoing the induced membrane technique consolidating (p = 0.049). Of those defects that consolidated, there was no difference in the time to bone union between strategies (p = 0.308) with an overall median time to union 8.33 months (95% CI 7.4 – 9.2 months). The induced membrane technique was associated with a 40% risk of sepsis.ConclusionThis study reported the outcomes of a standardised approach to the management of critical-sized bone defects. Whilst overall results were supportive of this approach, the outcomes associated with the induced membrane technique require further refinement of its indications in the management of critical-sized bone defects.Level of evidence4.

The effect of injectable platelet-rich fibrin in combination with autogenous bone grafts on bone healing in an ovariectomized osteoporotic rat model with critical-sized defects.

To assess the impact of autogenous graft materials, including autogenous bone graft (ABG) and injectable platelet-rich fibrin (I-PRF) clots, on bone defect regeneration in ovariectomized osteoporotic rats. Wistar rats (6-8 weeks old) were ovariectomized, and surgery began after 8 weeks. A 5-mm defect was created bilaterally in the parietal bones of 16 rats, which were divided into 4 groups. Group A (blank) had untreated defects; group B had defects filled with 0.1 ml of I-PRF, group C had defects filled with 0.1 ml of ABG, and group D had defects filled with a combination of 0.1 ml each of I-PRF and ABG. Four weeks post-surgery, the rats were euthanized. Bone regeneration was evaluated through histopathologic analysis and microcomputed tomography (micro-CT). In ovariectomized rats treated with ABG or I-PRF, bone regeneration was enhanced, with increased periosteal activity, osteoblast count, and new bone volume, as determined histologically. The ABG+I-PRF group had the highest periosteal vascularity, but the difference compared to the ABG group was not statistically significant (p>0.05). Osteoblast numbers were significantly higher in the ABG+I-PRF group than in the blank group (p<0.05). Micro-CT showed the highest mean new bone volume ratio in the ABG+I-PRF group, followed by the ABG group. The combined use of ABG and I-PRF enhances bone formation in osteoporotic rats following ovariectomy.

Reactive oxygen-scavenging polydopamine nanoparticle coated 3D nanofibrous scaffolds for improved osteogenesis: Toward an aging in vivo bone regeneration model.

Tissue engineered scaffolds aimed at the repair of critical-sized bone defects lack adequate consideration for our aging society. Establishing an effective aged in vitro model that translates to animals is a significant unmet challenge. The in vivo aged environment is complex and highly nuanced, making it difficult to model in the context of bone repair. In this work, 3D nanofibrous scaffolds generated by the thermally-induced self-agglomeration (TISA) technique were functionalized with polydopamine nanoparticles (PD NPs) as a tool to improve drug binding capacity and scavenge reactive oxygen species (ROS), an excessive build-up that dampens the healing process in aged tissues. PD NPs were reduced by ascorbic acid (rPD) to further improve hydrogen peroxide (H2O2) scavenging capabilities, where we hypothesized that these functionalized scaffolds could rescue ROS-affected osteoblastic differentiation in vitro and improve new bone formation in an aged mouse model. rPDs demonstrated improved H2O2 scavenging activity compared to neat PD NPs, although both NP groups rescued the alkaline phosphatase activity (ALP) of MC3T3-E1 cells in presence of H2O2. Additionally, BMP2-induced osteogenic differentiation, both ALP and mineralization, was significantly improved in the presence of PD or rPD NPs on TISA scaffolds. While in vitro data showed favorable results aimed at improving osteogenic differentiation by PD or rPD NPs, in vivo studies did not note similar improvements in ectopic bone formation an aged model, suggesting that further nuance in material design is required to effectively translate to improved in vivo results in aged animal models.

3D-printed titanium scaffolds loaded with gelatin hydrogel containing strontium-doped silver nanoparticles promote osteoblast differentiation and antibacterial activity for bone tissue engineering.

Bone tissue engineering offers a promising alternative to stimulate the regeneration of damaged tissue, overcoming the limitations of conventional autografts and allografts. Recently, titanium alloy (Ti) implants have garnered significant attention for treating critical-sized bone defects, especially with the advancement of 3D printing technology. Although Ti alloys have impressive versatility, their lack of cellular adhesion, osteogenic and antibacterial properties are significant factors that contribute to their failure. Hence, to overcome these obstacles, this study aimed to incorporate osteoinductive and antibacterial cue-loaded hydrogels into 3D-printed Ti (3D-Ti) scaffolds. 3D-Ti scaffolds were synthesized using the direct metal laser sintering method and loaded with a gelatin (Gel) hydrogel containing strontium-doped silver nanoparticles (Sr-Ag NPs). Compared with Ag NPs, Sr-doped Ag NPs increased the expression of Runx2 mRNA, which is a key bone transcription factor. We subjected the bioactive 3D-hybrid scaffolds (3D-Ti/Gel/Sr-Ag NPs) to physicochemical and material characterization, followed by cytocompatibility and osteogenic evaluation. The microporous and macroporous topographies of the scaffolds with Sr-Ag NPs showed increased Runx2 expression and matrix mineralization, with potent antibacterial properties. Therefore, the 3D-Ti scaffolds incorporated with Sr-Ag NP-loaded Gel hydrogels favored osteoblast differentiation and antibacterial activity, indicating their potential for orthopedic applications.

Micro-Nanostructured Polymeric Scaffolds for Bone Tissue Engineering

While bone tissue allograft and autograft are commonly used in bone healing, their application is limited by factors such as availability, donor site morbidity, and immune response to the grafted tissue. Tissue-engineered implants, such as acellular or cellular polymeric structures, offer a promising alternative, and are a current trend in tissue engineering. Leveraging recent advancements in bone tissue engineering (BTE), we utilize 3D printing to develop biodegradable scaffolds that combine mechanical strength and bioactivity to facilitate bone repair and regeneration. This study focuses on the design and fabrication of mechanically competent 3D printed poly (L-lactic acid) (PLLA) micro-structured scaffolds. These scaffolds are enhanced with collagen type I nanofibrils to create bioactive scaffolds that promote tissue regeneration. The performance of these mechanically competent, micro-nanostructured polymeric matrices, in combination with bone marrow stromal cells (BMSCs), is evaluated in PLLA and PLLA-collagen scaffolds. The resulting micro-nanostructured PLLA-Collagen scaffolds mimic trabecular bone architecture, mechanical strength, and the extracellular matrix environment found in native bone tissue. The composite PLLA-collagen scaffolds exhibit mechanical properties in the mid-range of human trabecular bone. Both PLLA and PLLA-Collagen scaffolds support human BMSCs adhesion, proliferation, and osteogenic differentiation. A significantly higher number of implanted host cells are distributed in the PLLA-Collagen scaffolds with greater bone density, more uniform cell distribution, and attachment compared to the PLLA microstructure. Additionally, the biomimetic collagen nanostructure potently induces osteogenic transcription evidenced by increased alkaline phosphatase activity and upregulation of bone markers such as sialoprotein and collagen type I, ultimately guiding stem cell-mediated formation of a mature, mineralized bone matrix throughout the interconnected scaffold pores. This study underscores the benefits of micro-nanostructured scaffolds in successfully generating the inductive microenvironment of native bone extracellular matrix, triggering the cascade of cellular events required for functional bone regeneration, repairing critical-sized bone defects, and ultimately serving as an alternative material platform for bone regeneration, thereby instilling confidence in the potential of our research.

Bioactive polymer composite scaffolds fabricated from 3D printed negative molds enable bone formation and vascularization

Scaffolds for bone defect treatment should ideally support vascularization and promote bone formation, to facilitate the translation into biomedical device applications. This study presents a novel approach utilizing 3D-printed water-dissolvable polyvinyl alcohol (PVA) sacrificial molds to engineer polymerized High Internal Phase Emulsion (polyHIPE) scaffolds with microchannels and distinct multiscale porosity. Two sacrificial mold variants (250 µm and 500 µm) were generated using fused deposition modeling, filled with HIPE, and subsequently dissolved to create polyHIPE scaffolds containing microchannels. In vitro assessments demonstrated significant enhancement in cell infiltration, proliferation, and osteogenic differentiation, underscoring the favorable impact of microchannels on cell behavior. High loading efficiency and controlled release of the osteogenic factor BMP-2 were achieved, with microchannels facilitating release of the growth factor. Evaluation in a mouse critical-size calvarial defect model revealed enhanced vascularization and bone formation in microchanneled scaffolds containing BMP-2. This study not only introduces an accessible method for creating multiscale porosity in polyHIPE scaffolds but also emphasizes its capability to enhance cellular infiltration, controlled growth factor release, and in vivo performance. The findings suggest promising applications in bone tissue engineering and regenerative medicine, and are expected to facilitate the translation of this type of biomaterial scaffold. Statement of significanceThis study holds significance in the realm of biomaterial scaffold design for bone tissue engineering and regeneration. We demonstrate a novel method to introduce controlled multiscale porosity and microchannels into polyHIPE scaffolds, by utilizing 3D-printed water-dissolvable PVA molds. The strategy offers new possibilities for improving cellular infiltration, achieving controlled release of growth factors, and enhancing vascularization and bone formation outcomes. This microchannel approach not only marks a substantial stride in scaffold design but also demonstrates its tangible impact on enhancing osteogenic cell differentiation and fostering robust bone formation in vivo. The findings emphasize the potential of this methodology for bone regeneration applications, showcasing an interesting advancement in the quest for effective and innovative biomaterial scaffolds to regenerate bone defects.

Biodegradable magnesium and zinc composite microspheres with synergistic osteogenic effect for enhanced bone regeneration

Biodegradable polymer microspheres in bone tissue engineering have become appealing as their non-invasive advantages in irregular damage bone repair. However, current microspheres used in BTE still lack sufficient osteogenic capacity to induce effective bone regeneration. In this study, we developed osteogenic composite microspheres concurrently loaded with magnesium oxide (MgO) and zinc oxide (ZnO), both of which are osteogenic active substances, using a facile and scalable emulsification method. The osteogenic composite microspheres exhibited a sequential yet complementary release profile characterized by a rapid release of Mg2+ and a gradual release of Zn2+ in a physiological environment, thereby maintaining the concentration of bioactive ions at a sustained high level. As a result, the combination of Mg2+ and Zn2+ in the composite microspheres led to a synergistic enhancement in biomimetic mineralization and the upregulation in the expression of osteogenic-related genes and proteins at the cellular level. Through a critical-sized calvarial rate defect model, the osteogenic composite microspheres were demonstrated to have strong osteogenic ability to promote new bone formation via ultrasonic imaging, histological and immunohistochemical evaluations. In sum, these osteogenic composite microspheres as microcarriers of Mg2+ and Zn2+ have great potential in the delivery of therapeutic ions for treating bone defects.

The cell membrane as biofunctional material for accelerated bone repair

The cell (plasma) membrane is enriched with numerous receptors, ligands, enzymes, and phospholipids that play important roles in cell-cell and cell-extracellular matrix interactions governing, for instance, tissue development and repair. We previously showed that plasma membrane nanofragments (PMNFs) act as nucleation sites for bone formation in vivo, and induce in vitro mineralization within 1 day. In this study, we optimized the methods for generating, isolating, and applying PMNFs as a cell-free therapeutic to expedite bone defect repair. The PMNFs were isolated from different mouse cell lines (chondrocytes, osteoblasts, and fibroblasts), pre-conditioned, lyophilized, and subsequently transplanted into 2 mm critical-sized calvarial defects in mice (n = 75). The PMNFs from chondrocytes, following a 3-day pre-incubation, significantly accelerated bone repair within 2 weeks, through a coordinated attraction of macrophages, endothelial cells, and osteoblasts to the healing site. In vitro experiments confirmed that PMNFs enhanced cell adhesion. Comparison of the PMNF efficacy with phosphatidylserine, amorphous calcium phosphate (ACP), and living cells confirmed the unique ability of PMNFs to promote accelerated bone repair. Importantly, PMNFs promoted nearly complete integration of the regenerated bone with native tissue after 6 weeks (% non-integrated bone area = 15.02), contrasting with the partial integration (% non-integrated bone area = 56.10; p < 0.01, Student's test) with transplantation of ACP. Vickers microhardness tests demonstrated that the regenerated bone after 6 weeks (30.10 ± 1.75) exhibited hardness similar to native bone (31.07 ± 2.46). In conclusion, this is the first study to demonstrate that cell membrane can be a promising cell-free material with multifaceted biofunctional properties that promote accelerated bone repair. Statement of significance•For the first time, the cell (plasma) membrane is used as material for bone repair.•The plasma membrane from chondrocytes promoted rapid bone repair within 2 weeks, with complete integration of the newly formed bone with the surrounding native tissue.•The plasma membrane from chondrocytes presented multifaceted effects, strongly promoting the attraction of macrophages, endothelial cells and osteoblasts to the healing site.

Osteoclast-Driven Polydopamine-to-Dopamine Release: An Upgrade Patch for Polydopamine-Functionalized Tissue Engineering Scaffolds.

Polydopamine, a mussel-inspired self-adherent polymer of dopamine, has impressive adhesive properties and thus is one of the most versatile approaches to functionalize tissue engineering scaffolds. To date, many types of polydopamine-functionalized scaffolds have been manufactured and extensively applied in bone tissue engineering at the preclinical stage. However, how polydopamine is biodegraded and metabolized during the bone healing process and the side effects of its metabolite remain largely unknown. These issues are often neglected in the modern manufacture of polydopamine-functionalized materials and restrict them from stepping forward to clinical applications. In this study, using our bioinspired polydopamine-laced hydroxyapatite collagen calcium silicate material as a representative of polydopamine-functionalized tissue engineering scaffolds, we discovered that polydopamine can be metabolized to dopamine specifically by osteoclasts, which we termed "osteoclast-driven polydopamine-to-dopamine release". The released dopamine showed an osteoinductive effect in vitro and promoted bone regeneration in calvarial critical-sized defects. The concept of "osteoclast-driven polydopamine-to-dopamine release" has considerable application potential. It could be easily adopted by other existing polydopamine-functionalized scaffolds: just by recruiting osteoclasts. Once adopted, scaffolds will obtain a dopamine-releasing function, which enables their modulation of osteoblast activity and hence elevates the osteoinductive effect. Thus, "osteoclast-driven polydopamine-to-dopamine release" serves as an upgrade patch, which is useful for many existing polydopamine-functionalized materials.

Enhanced vascularity in gelatin scaffolds via copper-doped magnesium–calcium silicates incorporation: In-vitro and ex-ovo insights

Addressing a critical challenge in current tissue-engineering practices, this study aims to enhance vascularization in 3D porous scaffolds by incorporating bioceramics laden with pro-angiogenic ions. Specifically, freeze-dried gelatin-based scaffolds were infused with sol-gel-derived powders of Cu-doped akermanite (Ca2MgSi2O7) and bredigite (Ca7MgSi4O16) at various concentrations (10, 20, and 30 wt%). The scaffolds were initially characterized for their structural integrity, biodegradability, swelling behavior, impact on physiological pH, and cytocompatibility with human umbilical vein endothelial cells (HUVECs). The silicate incorporation effectiveness in promoting vascularity was then assessed through HUVEC attachment, capillary tube formation, and ex-ovo chick embryo chorioallantoic membrane assays. The findings revealed significant improvements in both in-vitro and ex-ovo vascularity of the gelatin scaffolds upon the addition of Cu-doped akermanite. The most effective concentrations were determined to be 10 and 20 %, which led to notable HUVEC metabolic activity, a well-spread morphology with extensive peripheral filopodia and lamellipodia at 10 % and a cobblestone phenotype indicative of in-vivo endothelium at 20 % during cell attachment, the formation of complex networks of tubular structures, and robust vascularization in chick embryo development. Moving forward, the incorporation of Cu-doped akermanite into tissue-engineering scaffolds shows great potential for addressing the limitations of vascularization, especially for critical-sized bone defects, by facilitating the controlled release of pro-angiogenic and pro-osteogenic ions.