Biphasic Calcium Phosphate Scaffolds Research Articles

Evaluating osteogenic potential of a 3D-printed bioceramic-based scaffold for critical-sized defect treatment: an in vivo and in vitro investigation.

The integration of precision medicine principles into bone tissue engineering has ignited a wave of research focused on customizing intricate scaffolds through advanced 3D printing techniques. Bioceramics, known for their exceptional biocompatibility and osteoconductivity, have emerged as a promising material in this field. This article aims to evaluate the regenerative capabilities of a composite scaffold composed of 3D-printed gelatin combined with hydroxyapatite/tricalcium phosphate bioceramics (G/HA/TCP), incorporating human dental pulp-derived stem cells (hDPSCs). Using 3D powder printing, we created cross-shaped biphasic calcium phosphate scaffolds with a gelatin layer. The bone-regenerating potential of these scaffolds, along with hDPSCs, was assessed through in vitro analyses and in vivo studies with 60 rats and critical-sized calvarial defects. The assessment included analyzing cellular proliferation, differentiation, and alkaline phosphatase activity (ALP), and concluded with a detailed histological evaluation of bone regeneration. Our study revealed a highly favorable scenario, displaying not only desirable cellular attachment and proliferation on the scaffolds but also a notable enhancement in the ALP activity of hDPSCs, underscoring their pivotal role in bone regeneration. However, the histological examination of calvarial defects at the 12-wk mark yielded a rather modest level of bone regeneration across all experimental groups. The test and cell group exhibited significant bone formation compared to all other groups except the control and cell group. This underscores the complexity of the regenerative process and paves the way for further in-depth investigations aimed at improving the potential of the composite scaffolds.

3D-printed Biphasic Calcium Phosphate Scaffold to augment cytocompatibility evaluation for load-bearing implant applications

In this work, we developed and analyzed a biphasic calcium phosphate (BCP) bioceramic for bone regeneration using stereolithography (SLA). The SLA method is a promising additive manufacturing (AM) technique capable of creating BCp parts with high accuracy and efficiency. However, the ceramic suspension used in SLA exhibits significantly higher viscosity and is not environmentally friendly. Therefore, adequate preparation of a suspension with low viscosity and high solid loading is essential. In this paper, we optimized the effects of surfactant doses and solid loading on the BCp slurry, and initially examined the process parameters of photocuring, debinding, and sintering. The utilization of 9 wt % Disperbyk (BYK) with a 40 vol % loading of BCp bioceramics exhibited a reasonably low viscosity of 8.9 mPa·s at a shear level of 46.5 s−1. Functional and structural analyses confirmed that BCp was retained after photocuring and subsequent treatment, which were incorporated into the BYK dispersion. The 3D printed objects with different sintered temperatures, specifically at 1100 °C, 1200 °C, and 1300 °C, were further optimized. Additionally, the surface roughness, porosity, and mechanical properties of BCp green parts were systematically investigated. Most importantly, in vitro analysis of cell attachment, differentiation, and red alizarin analysis could support the application of bone regeneration.

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.

3D gel-printing of hierarchically porous BCP scaffolds for bone tissue engineering

In this study, a conventional technique of porous preparation was used to improve the constructive capability of direct ink writing on microstructures, and the hierarchically porous scaffolds were successfully prepared by 3D gel printing (3DGP). Micron-sized hydroxyapatite (HA) was coated with tricalcium phosphate (TCP) nanopowders synthesized by chemical co-precipitation to form biphasic calcium phosphate (BCP). The random structure of concave micropores was achieved by filling the BCP slurry with PMMA microspheres while successfully controlling the internal porosity of printed filaments. The results showed that the three-stage porous structure was successfully constructed, i.e., macroscopic pores of 1.50-2.00 mm, spherical micropores of 100-200 μm, and inter-powder interstices of 1.00-10.00 μm. Nano-TCP coated micron-HA powders improved the sintering activity of BCP particles. The compressive strength and porosity of the scaffolds sintered at 1400 °C were 2.78 MPa and 84.98%. The hierarchically porous BCP scaffolds had bright applications in bone tissue engineering.

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.

Mangiferin-Enriched Mn-Hydroxyapatite Coupled with β-TCP Scaffolds Simultaneously Exhibit Osteogenicity and Anti-Bacterial Efficacy.

Biphasic calcium phosphate (BCP) containing β-tricalcium phosphate and manganese (Mn)-substituted hydroxyapatite (HAP) was synthesized. Biomedical scaffolds were prepared using this synthesized powder on a sacrificial polyurethane sponge template after the incorporation of mangiferin (MAN). Mn was substituted at a concentration of 5% and 10% in HAP to examine the efficacy of Mn at various concentrations. The phase analysis of the as-formed BCP scaffold was carried out by X-ray diffraction analysis, while the qualitative observation of morphology and the osteoblast cell differentiation were carried out by scanning electron microscopy and confocal laser scanning microscopy techniques. Gene expressions of osteocalcin, collagen 1, and RUNX2 were carried out using qRT-PCR analyses. Significantly higher (p < 0.05) levels of ALP activity were observed with extended osteoblast induction on the mangiferin-incorporated BCP scaffolds. After characterization of the specimens, it was found that the scaffolds with 10% Mn-incorporated BCP with mangiferin showed better osteogenicity and simultaneously the same scaffolds exhibited higher anti-bacterial properties as observed from the bacterial viability test. This study was carried out to evaluate the efficacy of Mn and MAN in BCP for osteogenicity and antibacterial action.

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.

Adjusting physicochemical and cytological properties of biphasic calcium phosphate by magnesium substitution: An in vitro study

Biphasic calcium phosphate (BCP) is a highly study bone defect repair material with adjustable degradation, perfect osteoconduction and good osteoinduction. As one of the essential trace elements, magnesium (Mg) possesses the abilities of pro-osteogenesis and pro-angiogenesis. Therefore, Mg doping may further expand the application of BCP in bone defect repair, but few studies focus on promoting the osteogenesis and angiogenesis of BCP simultaneously by Mg doping, and the optimal doping amount of Mg remains to be explored. In this study, the physicochemical and biological properties of BCP scaffold affected by Mg doping were systematically study. Results showed that Mg doping enhanced the sintering of BCP scaffold, resulting in the decrease of degradation rate at the initial soaking period. However, the introduction of Mg damaged the lattice stability of BCP, leading to the increase of BCP degradation rate at the later soaking period. BCP scaffolds with Mg doping content ≥3 mol.% could achieve a long-term sustained release of Mg. The ion microenvironment created by Mg-doped scaffolds was simultaneously conducive to the osteogenic differentiation of stem cells and the enhanced angiogenic activity of endothelial cells. The scaffold doped with 5 mol.% of Mg (Mg5–S) showed the highest efficiency in promoting osteogenic differentiation. Mg-doped BCP scaffolds with a doping content ≥3 mol.%, especially Mg5–S, significantly improved the proliferation and angiogenic differentiation of endothelial cells. Based on these, we believe that the optimal doping content of Mg in BCP is 5 mol.%, and Mg5–S has great application potential in bone defect repair.

Combined Application of Dentin Noncollagenous Proteins and Odontogenic Biphasic Calcium Phosphate in Rabbit Maxillary Sinus Lifting.

Teeth can be used as a raw material for preparing bone substitutes due to their similar chemical composition to bone. The objective of our study was to evaluate the effect of odontogenic biphasic calcium phosphate (BCP) incorporating dentin noncollagenous proteins (DNCPs) on osteogenesis and stability in maxillary sinus augmentation. The composition, structure and morphology of the odontogenic BCP were tested by X-ray powder diffraction (XRD), Brunauer-Emmett-Teller, and scanning electron microscopy methods. The biocompatibility and osteoinduction of DNCPs and materials were examined in vitro and their bone regeneration capacity was verified in vivo. The results showed that the cells adhered and proliferated well on the DNCP-loaded BCP scaffold. The odontogenic BCP and DNCPs promoted osteogenic differentiation of cells, The new bone formation in the BCP groups and DNCP subgroups was significantly higher than the new bone formation in the control, and the new bone quality was better. The bone regeneration effect of odontogenic BCP was similar to the effect of deproteinized bovine bone mineral, but β-TCP did not maintain the height and volume of bone reconstruction. In conclusion, the combined application of DNCPs and odontogenic BCP is an effective strategy for tissue engineering osteogenesis in the maxillary sinus region. The biomimetic strategy could provide a new approach for patients requiring maxillary sinus lifting.

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.

DAR 16-II Primes Endothelial Cells for Angiogenesis Improving Bone Ingrowth in 3D-Printed BCP Scaffolds and Regeneration of Critically Sized Bone Defects.

Bone is a highly vascularized tissue and relies on the angiogenesis and response of cells in the immediate environmental niche at the defect site for regeneration. Hence, the ability to control angiogenesis and cellular responses during osteogenesis has important implications in tissue-engineered strategies. Self-assembling ionic-complementary peptides have received much interest as they mimic the natural extracellular matrix. Three-dimensional (3D)-printed biphasic calcium phosphate (BCP) scaffolds coated with self-assembling DAR 16-II peptide provide a support template with the ability to recruit and enhance the adhesion of cells. In vitro studies demonstrated prompt the adhesion of both human umbilical vein endothelial cells (HUVEC) and human mesenchymal stem cells (hMSC), favoring endothelial cell activation toward an angiogenic phenotype. The SEM-EDS and protein micro bicinchoninic acid (BCA) assays demonstrated the efficacy of the coating. Whole proteomic analysis of DAR 16-II-treated HUVECs demonstrated the upregulation of proteins involved in cell adhesion (HABP2), migration (AMOTL1), cytoskeletal re-arrangement (SHC1, TMOD2), immuno-modulation (AMBP, MIF), and morphogenesis (COL4A1). In vivo studies using DAR-16-II-coated scaffolds provided an architectural template, promoting cell colonization, osteogenesis, and angiogenesis. In conclusion, DAR 16-II acts as a proactive angiogenic factor when adsorbed onto BCP scaffolds and provides a simple and effective functionalization step to facilitate the translation of tailored 3D-printed BCP scaffolds for clinical applications.

Novel Epigenetic Modulation Chitosan-Based Scaffold as a Promising Bone Regenerative Material.

Bone tissue engineering is a complicated field requiring concerted participation of cells, scaffolds, and osteoactive molecules to replace damaged bone. This study synthesized a chitosan-based (CS) scaffold incorporated with trichostatin A (TSA), an epigenetic modifier molecule, to achieve promising bone regeneration potential. The scaffolds with various biphasic calcium phosphate (BCP) proportions: 0%, 10%, 20%, and 40% were fabricated. The addition of BCP improved the scaffolds’ mechanical properties and delayed the degradation rate, whereas 20% BCP scaffold matched the appropriate scaffold requirements. The proper concentration of TSA was also validated. Our developed scaffold released TSA and sustained them for up to three days. The scaffold with 800 nM of TSA showed excellent biocompatibility and induced robust osteoblast-related gene expression in the primary human periodontal ligament cells (hPDLCs). To evaluate in vivo bone regeneration potential, the scaffolds were implanted in the mice calvarial defect model. The excellent bone regeneration ability was further demonstrated in the micro-CT and histology sections compared to both negative control and commercial bone graft product. New bone formed in the CS/BCP/TSA group revealed a trabeculae-liked characteristic of the mature bone as early as six weeks. The CS/BCP/TSA scaffold is an up-and-coming candidate for the bone tissue engineering scaffold.

Effect of silicon or cerium doping on the anti-inflammatory activity of biphasic calcium phosphate scaffolds for bone regeneration.

Biphasic calcium phosphate (BCP) bioceramics composed of hydroxyapatite and β-tricalcium phosphate have attracted considerable attention as ideal bone substitutes for reconstructive surgery, orthopedics, and dentistry, owing to their similar chemical composition to bone mineral and biocompatibility. The addition of trace elements to BCP bioceramics, such as magnesium (Mg), cerium (Ce), and silicon (Si), can alter the physicochemical and biological properties of the resulting materials. To improve the anti-inflammatory activity of a pure BCP scaffold, this study developed a simple wet chemical precipitation and gel-casting method to fabricate microporous BCP scaffolds containing Si or Ce. The BCP scaffolds exhibited interconnected microporous structures with uniform micropores and unequiaxed grains. No changes in the phase composition and microstructure of the scaffolds with the Si or Ce doping were observed. Conversely, Si or Ce doping into the BCP crystal lattice influenced the in vitro biological activity of the scaffolds and the bone-forming ability of the cells cultured on the BCP scaffolds. The results of biological activity assays demonstrated that Ce-BCP promoted cell proliferation and osteogenic differentiation more effectively than the other scaffolds. In particular, Ce-BCP significantly suppressed the expression of bone-active cytokines via the anti-inflammatory and anti-oxidative effects. Therefore, Si- or Ce-doped BCP scaffolds can contribute to providing a new generation of bone graft substitutes.

Enhanced osteogenesis and angiogenesis of biphasic calcium phosphate scaffold by synergistic effect of silk fibroin coating and zinc doping.

Biphasic calcium phosphate (BCP) scaffold has been widely applied to bone regeneration because of its good biocompatibility and bone conduction property. However, the low mechanical strength and the lack of angiogenic and osteogenic induction properties have restricted its application in bone tissue regeneration. In this study, we combined zinc (Zn2+) doping and silk fibroin (SF) coating with expectation to enhance compressive strength, osteogenesis and angiogenesis of BCP scaffolds. The phase composition, morphology, porosity, compressive strength, in vitro degradation and cell behaviors were investigated systematically. Results showed that the scaffold coated with SF exhibited almost 3 times of compressive strength without compromising its porosity compared with the uncoated scaffold. Zn2+ doping and SF coating synergistically enhanced the alkaline phosphatase activity and osteogenesis-related genes expression of mouse bone mesenchymal stem cells (mBMSCs). Furthermore, SF coating notably improved the proliferation, cell viability and in vitro angiogenesis of human umbilical vein endothelial cells (HUVECs). This work provides a novel way to modify BCP scaffolds simultaneously with enhancing mechanical strength and biological properties.

Leukemia Inhibitory Factor Facilitates Self-Renewal and Differentiation and Attenuates Oxidative Stress of BMSCs by Activating PI3K/AKT Signaling.

Objective Transplantation of bone marrow-derived mesenchymal stem cells (BMSCs) remains a hopeful therapeutic approach for bone defect reconstruction. Herein, we investigated the effects and mechanisms of leukemia inhibitory factor (LIF) in the function and viability of hypoxic BMSCs as well as bone defect repair. Methods The effects of LIF on apoptosis (flow cytometry, TUNEL staining), mitochondrial activity (JC-1 staining), proliferation (colony formation, EdU staining), and differentiation (CD105, CD90, and CD29 via flow sorting) were examined in hypoxic BMSCs. LIF, LIFR, gp130, Keap1, Nrf2, antioxidant enzymes (SOD1, catalase, GPx-3), bone-specific matrix proteins (ALP, BSP, OCN), PI3K, and Akt were detected via immunoblotting or immunofluorescent staining. BMSCs combined with biphasic calcium phosphate scaffolds were implanted into calvarial bone defect mice, and the therapeutic effect of LIF on bone defect was investigated. Results Hypoxic BMSCs had increased apoptosis and oxidative stress and reduced mitochondrial activity. Additionally, LIF, LIFR, and gp130 were upregulated and PI3K/Akt activity was depressed in hypoxic BMSCs. Upregulated LIF alleviated apoptosis and oxidative stress and heightened mitochondrial activity and PI3K/Akt signaling in hypoxic BMSCs. Additionally, LIF overexpression promoted self-renewal and osteogenic differentiation of BMSCs with hypoxic condition. Mechanically, LIF facilitated self-renewal and differentiation as well as attenuated oxidative stress of BMSCs through enhancing PI3K/AKT signaling activity. Implantation of LIF-overexpressed BMSC-loaded BCP scaffolds promoted osteogenesis as well as alleviated oxidative stress and apoptosis through PI3K/Akt signaling. Conclusion Our findings demonstrate that LIF facilitates self-renewal and differentiation and attenuates oxidative stress of BMSCs by PI3K/AKT signaling.

Sr and Mg Doped Bi-Phasic Calcium Phosphate Macroporous Bone Graft Substitutes Fabricated by Robocasting: A Structural and Cytocompatibility Assessment.

Bi-phasic calcium phosphates (BCPs) are considered prominent candidate materials for the fabrication of bone graft substitutes. Currently, supplemental cation-doping is suggested as a powerful path to boost biofunctionality, however, there is still a lack of knowledge on the structural role of such substituents in BCPs, which in turn, could influence the intensity and extent of the biological effects. In this work, pure and Mg- and Sr-doped BCP scaffolds were fabricated by robocasting from hydrothermally synthesized powders, and then preliminarily tested in vitro and thoroughly investigated physically and chemically. Collectively, the osteoblast cell culture assays indicated that all types of BCP scaffolds (pure, Sr- or Sr–Mg-doped) delivered in vitro performances similar to the biological control, with emphasis on the Sr–Mg-doped ones. An important result was that double Mg–Sr doping obtained the ceramic with the highest β-tricalcium phosphate (β-TCP)/hydroxyapatite mass concentration ratio of ~1.8. Remarkably, Mg and Sr were found to be predominantly incorporated in the β-TCP lattice. These findings could be important for the future development of BCP-based bone graft substitutes since the higher dissolution rate of β-TCP enables an easier release of the therapeutic ions. This may pave the road toward medical devices with more predictable in vivo performance.