The enhancement of bone regeneration by a combination of osteoconductivity and osteostimulation using β-CaSiO3/β-Ca3(PO4)2 composite bioceramics

β-Tricalcium phosphate (β-TCP) is osteoconductive, while β-calcium silicate (β-CS) is bioactive with osteostimulative properties. Porous β-CaSiO₃/β-Ca₃(PO₄)₂ composite bioceramic scaffolds with various β-TCP:β-CS ratios were designed to combine both osteoconductivity and osteostimulation in order to enhance bone regeneration. The composite scaffolds were implanted in critical sized femur defects (6 × 12 mm) for 4, 12 and 26 weeks with pure β-TCP and β-CS scaffolds as the controls. The in vivo biodegradation and bone regeneration of the specimens were investigated using sequential histological evaluations, immunohistochemical examination and micro-computed tomography technology. The results showed that the scaffolds with 50 and 80 wt.% β-CS dramatically enhanced the amount of newly formed bone and reduced the degradation rate. In contrast, porous β-CS displayed poor new bone formation due to its rapid degradation, while porous β-TCP showed moderate bone regeneration starting on the surface of the implants, due to a lack of osteostimulation. More importantly, the scaffolds with 50 and 80 wt.% β-CS not only had excellent osteoconductivity, but also stimulated rapid bone formation, and they could degrade progressively at a rate matching the regeneration of new bone. In summary, our findings indicated that the degradation rate and bioactivity of β-CS/β-TCP composite bioceramic scaffolds could be adjusted by controlling the ratio of β-CS to β-TCP, suggesting the potential application of β-CS/β-TCP composite bioceramic scaffolds with 50 and 80 wt.% β-CS component in hard tissue regeneration and bone tissue engineering.     

Osteogenic and angiogenic potentials of monocultured and co-cultured human-bone-marrow-derived mesenchymal stem cells and human-umbilical-vein endothelial cells on three-dimensional porous beta-tricalcium phosphate scaffold

The use of biodegradable beta-tricalcium phosphate (β-TCP) scaffolds holds great promise for bone tissue engineering. However, the effects of β-TCP on bone and endothelial cells are not fully understood. This study aimed to investigate cell proliferation and differentiation of mono- or co-cultured human-bone-marrow-derived mesenchymal stem cells (hBMSCs) and human-umbilical-vein endothelial cells (HUVECs) on a three-dimensional porous, biodegradable β-TCP scaffold. In co-culture studies, the ratios of hBMSCs:HUVECs were 5:1, 1:1 and 1:5. Cellular morphologies of HUVECs, hBMSCs and co-cultured HUVECs/hBMSCs on the β-TCP scaffolds were monitored using confocal and scanning electron microscopy. Cell proliferation was monitored by measuring the amount of double-stranded DNA (dsDNA) whereas hBMSC and HUVEC differentiation was assessed using the osteogenic and angiogenic markers, alkaline phosphatase (ALP) and PECAM-1 (CD31), respectively. Results show that HUVECs, hBMSCs and hBMSCs/HUVECs adhered to and proliferated well on the β-TCP scaffolds. In monoculture, hBMSCs grew faster than HUVECs on the β-TCP scaffolds after 7 days, but HUVECs reached similar levels of proliferation after 14 days. In monoculture, β-TCP scaffolds promoted ALP activities of both hBMSCs and HUVECs when compared to those grown on tissue culture well plates. ALP activity of cells in co-culture was higher than that of hBMSCs in monoculture. Real-time polymerase chain reaction results indicate that runx2 and alp gene expression in monocultured hBMSCs remained unchanged at days 7 and 14, but alp gene expression was significantly increased in hBMSC co-cultures when the contribution of individual cell types was not distinguished.     

The osteogenic differentiation of human bone marrow MSCs on HUVEC-derived ECM and β-TCP scaffold

Extracellular matrix (ECM) serves a key role in cell migration, attachment, and cell development. Here we report that ECM derived from human umbilical vein endothelial cells (HUVEC) promoted osteogenic differentiation of human bone marrow mesenchymal stem cells (hMSC). We first produced an HUVEC-derived ECM on a three-dimensional (3D) beta-tricalcium phosphate (β-TCP) scaffold by HUVEC seeding, incubation, and decellularization. The HUVEC-derived ECM was then characterized by SEM, FTIR, XPS, and immunofluorescence staining. The effect of HUVEC-derived ECM-containing β-TCP scaffold on hMSC osteogenic differentiation was subsequently examined. SEM images indicate a dense matrix layer deposited on the surface of struts and pore walls. FTIR and XPS measurements show the presence of new functional groups (amide and hydroxyl groups) and elements (C and N) in the ECM/β-TCP scaffold when compared to the β-TCP scaffold alone. Immunofluorescence images indicate that high levels of fibronectin and collagen IV and low level of laminin were present on the scaffold. ECM-containing β-TCP scaffolds significantly increased alkaline phosphatase (ALP) specific activity and up-regulated expression of osteogenesis-related genes such as runx2, alkaline phosphatase, osteopontin and osteocalcin in hMSC, compared to β-TCP scaffolds alone. This increased effect was due to the activation of MAPK/ERK signaling pathway since disruption of this pathway using an ERK inhibitor PD98059 results in down-regulation of these osteogenic genes. Cell-derived ECM-containing calcium phosphate scaffolds is a promising osteogenic-promoting bone void filler in bone tissue regeneration.     

Osteoimmunomodulatory properties of magnesium scaffolds coated with β-tricalcium phosphate

The osteoimmunomodulatory property of bone biomaterials is a vital property determining the in vivo fate of the implants. Endowing bone biomaterials with favorable osteoimmunomodulatory properties is of great importance in triggering desired immune response and thus supports the bone healing process. Magnesium (Mg) has been recognized as a revolutionary metal for applications in orthopedics due to it being biodegradable, biocompatible, and having osteoconductive properties. However, Mg's high rate of degradation leads to an excessive inflammatory response and this has restricted its application in bone tissue engineering. In this study, β-tricalcium phosphate (β-TCP) was used to coat Mg scaffolds in an effort to modulate the detrimental osteoimmunomodulatory properties of Mg scaffolds, due to the reported favorable osteoimmunomodulatory properties of β-TCP. It was noted that macrophages switched to the M2 extreme phenotype in response to the Mg-β-TCP scaffolds, which could be due to the inhibition of the toll like receptor (TLR) signaling pathway. VEGF and BMP2 were significantly upregulated in the macrophages exposed to Mg-β-TCP scaffolds, indicating pro-osteogenic properties of macrophages in β-TCP modified Mg scaffolds. This was further demonstrated by the macrophage-mediated osteogenic differentiation of bone marrow stromal cells (BMSCs). When BMSCs were stimulated by conditioned medium from macrophages cultured on Mg-β-TCP scaffolds, osteogenic differentiation of BMSCs was significantly enhanced; whereas osteoclastogenesis was inhibited, as indicated by the downregualtion of MCSF, TRAP and inhibition of the RANKL/RANK system. These findings suggest that β-TCP coating of Mg scaffolds can modulate the scaffold's osteoimmunomodulatory properties, shift the immune microenvironment towards one that favors osteogenesis over osteoclastogenesis. Endowing bone biomaterials with favorable osteoimmunomodulatory properties can be a highly valuable strategy for the development or modification of advanced bone biomaterials.     

Acidic peptide hydrogel scaffolds enhance calcium phosphate mineral turnover into bone tissue

Designed peptides may generate molecular scaffolds in the form of hydrogels to support tissue regeneration. We studied the effect of hydrogels comprising β-sheet-forming peptides rich in aspartic amino acids and of tricalcium phosphate (β-TCP)-loaded hydrogels on calcium adsorption and cell culture in vitro, and on bone regeneration in vivo. The hydrogels were found to act as efficient depots for calcium ions, and to induce osteoblast differentiation in vitro. In vivo studies on bone defect healing in rat distal femurs analyzed by microcomputerized tomography showed that the peptide hydrogel itself induced better bone regeneration in comparison to non-treated defects. A stronger regeneration capacity was obtained in bone defects treated with β-TCP-loaded hydrogels, indicating that the peptide hydrogels and the mineral act synergistically to enhance bone regeneration. In vivo regeneration was found to be better with hydrogels loaded with porous β-TCP than with hydrogels loaded with non-porous mineral. It is concluded that biocompatible and biodegradable matrices, rich in anionic moieties that efficiently adsorb calcium ions while supporting cellular osteogenic activity, may efficiently promote β-TCP turnover into bone mineral.     

Biomineralized porous composite scaffolds prepared by chemical synthesis for bone tissue regeneration

Scaffold design is a key factor in the clinical success of bone tissue engineering grafts. To date, no existing single biomaterial used in bone repair and regeneration fulfils all the requirements for an ideal bone graft. In this study hydroxyapatite/polycaprolactone (HA/PCL) composite scaffolds were prepared by a wet chemical method at room temperature. The physico-chemical properties of the composite materials were characterized by X-ray diffraction, Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy, while scaffold morphology was investigated by scanning electron microscopy (SEM) with energy-dispersive spectroscopy to validate the process used for synthesis. Finally, the response of bone marrow-derived human mesenchymal stem cells (hMSCs) in terms of cell proliferation and differentiation to the osteoblastic phenotype was evaluated using the Alamar blue assay, SEM and alkaline phosphatase activity. Microstructural analysis indicated that the HA particles were distributed homogeneously within the PCL matrix. The biological results revealed that the HA/PCL composite scaffolds are suitable for the proliferation and differentiation of MSCs in vitro, supporting osteogenesis after 15 days. All the results indicate that these scaffolds meet the requirements of materials for bone tissue engineering and could be used for many clinical applications in orthopaedic and maxillofacial surgery.     

Unique microstructural design of ceramic scaffolds for bone regeneration under load

During the past two decades, research on ceramic scaffolds for bone regeneration has progressed rapidly; however, currently available porous scaffolds remain unsuitable for load-bearing applications. The key to success is to apply microstructural design strategies to develop ceramic scaffolds with mechanical properties approaching those of bone. Here we report on the development of a unique microstructurally designed ceramic scaffold, strontium–hardystonite–gahnite (Sr–HT–gahnite), with 85% porosity, 500 μm pore size, a competitive compressive strength of 4.1 ± 0.3 MPa and a compressive modulus of 170 ± 20 MPa. The in vitro biocompatibility of the scaffolds was studied using primary human bone-derived cells. The ability of Sr–HT–gahnite scaffolds to repair critical-sized bone defects was also investigated in a rabbit radius under normal load, with β-tricalcium phosphate/hydroxyapatite scaffolds used in the control group. Studies with primary human osteoblast cultures confirmed the bioactivity of these scaffolds, and regeneration of rabbit radial critical defects demonstrated that this material induces new bone defect bridging, with clear evidence of regeneration of original radial architecture and bone marrow environment.     

Bone regeneration using composite non-demineralized xenogenic dentin with beta-tricalcium phosphate in experimental alveolar cleft repair in a rabbit model

Background

Alveolar cleft repair is performed via bone grafting procedure to restore the dental arch continuity. A suitable bone substitute materials should possess osteoinductive and osteoconductive properties, to promote new bone formation, along with a slowly resorbable scaffold that is subsequently replaced with functionally viable bone. Calcium phosphate biomaterials have long proved their efficacy as bone replacement materials. Dentin in several forms has also demonstrated its possibility to be used as bone graft replacement material in several studies. The purpose of this study was to evaluate bone regeneration pattern and quantify bone formation after grafting pre-established experimental alveolar clefts defects model in rabbits using composite xenogenic dentin and β-TCP in comparison to β-TCP alone.

Methods

Unilateral alveolar cleft defects were created in 16 New Zealand rabbits according to previously described methodology. Alveolar clefts were allowed 8 weeks healing period. 8 defects were filled with β-TCP, whereas 8 defects filled with composite xenogenic dentin with β-TCP. Bone regeneration of the healed defects was compared at the 8 weeks after intervention. Quantification of bone formation was analyzed using micro-computed tomography (µCT) and histomorphometric analysis.

Results

µCT and histomorphometric analysis revealed that defects filled with composite dentin/β-TCP showed statistically higher bone volume fraction, bone mineral density and percentage residual graft volume when compared to β-TCP alone. An improved surgical handling of the composite dentin/β-TCP graft was also noted.

Conclusions

Composite xenogenic dentin/β-TCP putty expresses enhanced bone regeneration compared to β-TCP alone in the reconstruction of rabbit alveolar clefts defects.

     

A bioactive “self-fitting” shape memory polymer scaffold with potential to treat cranio-maxillo facial bone defects

While tissue engineering is a promising alternative for treating critical-sized cranio-maxillofacial bone defects, improvements in scaffold design are needed. In particular, scaffolds that can precisely match the irregular boundaries of bone defects as well as exhibit an interconnected pore morphology and bioactivity would enhance tissue regeneration. In this study, a shape memory polymer (SMP) scaffold was developed exhibiting an open porous structure and the capacity to conformally “self-fit” into irregular defects. The SMP scaffold was prepared via photocrosslinking of poly(ε-caprolactone) (PCL) diacrylate using a SCPL method, which included a fused salt template. A bioactive polydopamine coating was applied to coat the pore walls. Following exposure to warm saline at T > T_trans (T_trans = T_m of PCL), the scaffold became malleable and could be pressed into an irregular model defect. Cooling caused the scaffold to lock in its temporary shape within the defect. The polydopamine coating did not alter the physical properties of the scaffold. However, polydopamine-coated scaffolds exhibited superior bioactivity (i.e. formation of hydroxyapatite in vitro), osteoblast adhesion, proliferation, osteogenic gene expression and extracellular matrix deposition.     

Understanding the influence of MgO and SrO binary doping on the mechanical and biological properties of β-TCP ceramics

The objective of this study was to evaluate the influence of MgO and SrO doping on the mechanical and biological properties of β-tricalcium phosphate (β-TCP). β-TCP was doped with two different binary compositions, 0.25 and 1.0 wt.% SrO along with 1.0 wt.% MgO. MgO and SrO doping increased the β phase stability at a sintering temperature of 1250 °C and marginally decreased the compressive strength of β-TCP. An in vitro cell–material interaction study, using human fetal osteoblast cells (hFOB), indicated that doped β-TCP was non-toxic, and MgO/SrO dopants improved cell attachment and growth. β-TCP implants doped with 1.0 wt.% MgO and 1.0 wt.% SrO showed good in vivo biocompatibility when tested in male Sprague–Dawley rats for 16 weeks. Histomorphology analysis indicated that MgO/SrO-doped β-TCP promoted more osteogenesis than pure β-TCP. In vivo osteocalcin and type I collagen assay also revealed faster bone formation in rats with doped β-TCP implant compared to rats with pure β-TCP implant. Low Ca²⁺ concentration in the urine of rats with doped β-TCP implant confirmed slower degradation of MgO/SrO-doped β-TCP than pure β-TCP.     

Chondrogenic pre-induction of human mesenchymal stem cells on β-TCP: Enhanced bone quality by endochondral heterotopic bone formation

New techniques to heal bone defects include the combination of bone substitute materials with mesenchymal stem cells (MSC). To find solutions not hampered by low material resorbability or high donor variability of human MSC, the potency of such composites is usually evaluated by heterotopic bone formation assays in immunocompromised animals. The aim of this study was to investigate whether resorbable phase-pure β-tricalcium-phosphate (β-TCP) could support heterotopic bone formation by MSC comparable to partially resorbable hydroxyapatite/tricalcium-phosphate (HA/TCP). Furthermore, in light of disappointing results with osteogenic in vitro priming of MSC, we tested whether chondrogenic pre-induction of constructs may allow for enhanced bone formation by triggering the endochondral pathway. β-TCP granules of three different sizes and HA/TCP were seeded with MSC and transplanted subcutaneously into immunocompromised mice either immediately or after a chondrogenic pre-induction for 6 weeks. After 8 weeks, explants were analysed by histology. β-TCP seeded with unprimed MSC revealed intramembranous bone formation without haematopoietic marrow with 3.8-fold more bone formed with granules smaller than 0.7 mm than with 0.7–1.4 mm particles (p ? 0.018). Chondrogenic pre-induction of β-TCP/MSC composites resulted in collagen type II and proteoglycan-rich cartilage-like tissue which, after transplantation, underwent endochondral ossification, yielding ectopic bone produced by human cells while haematopoietic marrow was derived from the mouse. Transdifferentiation of MSC-derived chondrocytes to osteoblasts or direct osteogenesis of cartilage-resident MSC is postulated to explain the human origin of new bone. In conclusion, β-TCP was significantly more osteo-permissive (p = 0.004) than HA/TCP for human MSC, and chondrogenic priming of β-TCP/MSC represented a superior approach capable of supporting full bone formation, including marrow organization.     

Osteogenic differentiation of bone marrow MSCs by β-tricalcium phosphate stimulating macrophages via BMP2 signalling pathway

Immune reactions play important roles in determining the in vivo fate of bone substitute materials, either in new bone formation or inflammatory fibrous tissue encapsulation. The paradigm for the development of bone substitute materials has been shifted from inert to immunomodulatory materials, emphasizing the importance of immune cells in the material evaluation. Macrophages, the major effector cells in the immune reaction to implants, are indispensable for osteogenesis and their heterogeneity and plasticity render macrophages a primer target for immune system modulation. However, there are very few reports about the effects of macrophages on biomaterial-regulated osteogenesis. In this study, we used β-tricalcium phosphate (β-TCP) as a model biomaterial to investigate the role of macrophages on the material stimulated osteogenesis. The macrophage phenotype switched to M2 extreme in response to β-TCP extracts, which was related to the activation of calcium-sensing receptor (CaSR) pathway. Bone morphogenetic protein 2 (BMP2) was also significantly upregulated by the β-TCP stimulation, indicating that macrophage may participate in the β-TCP stimulated osteogenesis. Interestingly, when macrophage-conditioned β-TCP extracts were applied to bone marrow mesenchymal stem cells (BMSCs), the osteogenic differentiation of BMSCs was significantly enhanced, indicating the important role of macrophages in biomaterial-induced osteogenesis. These findings provided valuable insights into the mechanism of material-stimulated osteogenesis, and a strategy to optimize the evaluation system for the in vitro osteogenesis capacity of bone substitute materials.     

经表面修饰的-TCP对BMSCs细胞增殖及分化作用的研究

目的研究三种材料,材料一：β-磷酸三钙支架（β—TCP）;材料二：明胶／1氐结晶度羟基磷灰石涂层-磷酸三钙支架（gel／lc—HA．β-TCP）;材料三：明胶／高结晶度羟基磷灰石涂层-β-磷酸三钙（gel／he-HA-β-TCP）。三种材料的细胞毒性及兔骨髓基质细胞（BMSCs）与材料共培养增殖的情况。方法取兔股骨骨髓腔细胞,进行贴壁培养BMSCs。在成骨诱导液中诱导BMSCs。将诱导后的BMSCs与三种支架材料在培养板内共培养1、3、5、7、9d。采用形态学观察、MTT法及ALP检测试剂盒等方法检测材料的BMSCs细胞毒性及BMSCs细胞在材料表面的增殖和分化能力。结果光镜及电镜下观察各组无显著差异。细胞毒性在0-1级。与细胞共培养,β-TCP组在第7．9天与阴性对照组差异有统计学意义。ALP检测,β-TCP组在第7、9天与阴性对照组差异有统计学意义（P〈0．05）,gel／lc—HA-β-TCP在第9天与阴性对照组差异有统计学意义（p〈0．05）。结论新型明胶／高结晶度羟基磷灰石涂层-β-磷酸三钙支架和明胶／低结晶度羟基磷灰石涂层-β-磷酸三钙支架与BMSCs生物相容性好,适合作为支架材料负载BMSCs构建组织工程骨。     

Biocompatibility and osteogenic potential of human fetal femur-derived cells on surface selective laser sintered scaffolds

For optimal bone regeneration, scaffolds need to fit anatomically into the requisite bone defects and, ideally, augment cell growth and differentiation. In this study we evaluated novel computationally designed surface selective laser sintering (SSLS) scaffolds for their biocompatibility as templates, in vitro and in vivo, for human fetal femur-derived cell viability, growth and osteogenesis. Fetal femur-derived cells were successfully cultured on SSLS-poly(d,l)-lactic acid (SSLS-PLA) scaffolds expressing alkaline phosphatase activity after 7 days. Cell proliferation, ingrowth, Alcian blue/Sirius red and type I collagen positive staining of matrix deposition were observed for fetal femur-derived cells cultured on SSLS-PLA scaffolds in vitro and in vivo. SSLS-PLA scaffolds and SSLS-PLA scaffolds seeded with fetal femur-derived cells implanted into a murine critical-sized femur segmental defect model aided the regeneration of the bone defect. SSLS techniques allow fabrication of biocompatible/biodegradable scaffolds, computationally designed to fit any defect, providing a template for cell osteogenesis in vitro and in vivo.     

The enhancement of bone regeneration by gene activated matrix encoding for platelet derived growth factor

Gene therapy using non-viral vectors that are safe and efficient in transfecting target cells is an effective approach to overcome the shortcomings of protein delivery of growth factors. The objective of this study was to develop and test a non-viral gene delivery system for bone regeneration utilizing a collagen scaffold to deliver polyethylenimine (PEI)-plasmid DNA (pDNA) [encoding platelet derived growth factor-B (PDGF-B)] complexes. The PEI-pPDGF-B complexes were fabricated at amine (N) to phosphate (P) ratio of 10 and characterized for size, surface charge, and in vitro cytotoxicity and transfection efficacy in human bone marrow stromal cells (BMSCs). The influence of the complex-loaded collagen scaffold on cellular attachment and recruitment was evaluated in vitro using microscopy techniques. The in vivo regenerative capacity of the gene delivery system was assessed in 5 mm diameter critical-sized calvarial defects in Fisher 344 rats. The complexes were ∼100 nm in size with a positive surface charge. Complexes prepared at an N/P ratio of 10 displayed low cytotoxicity as assessed by a cell viability assay. Confocal microscopy revealed significant proliferation of BMSCs on complex-loaded collagen scaffolds compared to empty scaffolds. In vivo studies showed significantly higher new bone volume/total volume (BV/TV) % in calvarial defects treated with the complex-activated scaffolds following 4 weeks of implantation (14- and 44-fold higher) when compared to empty defects or empty scaffolds, respectively. Together, these findings suggest that non-viral PDGF-B gene-activated scaffolds are effective for bone regeneration and are an attractive gene delivery system with significant potential for clinical translation.     

Effect of bioactive borate glass microstructure on bone regeneration,angiogenesis, and hydroxyapatite conversion in a rat calvarial defect model

Borate bioactive glasses are biocompatible and enhance new bone formation, but the effect of their microstructure on bone regeneration has received little attention. In this study scaffolds of borate bioactive glass (1393B3) with three different microstructures (trabecular, fibrous, and oriented) were compared for their capacity to regenerate bone in a rat calvarial defect model. 12 weeks post-implantation the amount of new bone, mineralization, and blood vessel area in the scaffolds were evaluated using histomorphometric analysis and scanning electron microscopy. The amount of new bone formed was 33%, 23%, and 15%, respectively, of the total defect area for the trabecular, oriented, and fibrous microstructures. In comparison, the percent new bone formed in implants composed of silicate 45S5 bioactive glass particles (250–300 μm) was 19%. Doping the borate glass with copper (0.4 wt.% CuO) had little effect on bone regeneration in the trabecular and oriented scaffolds, but significantly enhanced bone regeneration in the fibrous scaffolds (from 15 to 33%). The scaffolds were completely converted to hydroxyapatite within the 12 week implantation. The amount of hydroxyapatite formed, 22%, 35%, and 48%, respectively, for the trabecular, oriented, and fibrous scaffolds, increased with increasing volume fraction of glass in the as-fabricated scaffold. Blood vessels infiltrated into all the scaffolds, but the trabecular scaffolds had a higher average blood vessel area compared with the oriented and fibrous scaffolds. While all three scaffold microstructures were effective in supporting bone regeneration, the trabecular scaffolds supported more bone formation and may be more promising in bone repair.     

Preparation,characterization and in vitro analysis of novel structured nanofibrous scaffolds for bone tissue engineering

In a previous study, a three-dimensional nanofibrous spiral scaffold for bone tissue engineering was developed, which showed enhanced human osteoblast cell attachment, proliferation and differentiation compared with traditional cylinder scaffolds, owing to the incorporation of spiral structures and nanofiber. However, the application of these scaffolds to bone tissue engineering was limited by their weak mechanical strength. This limitation triggered the design for novel structured scaffolds with reinforced physical characteristics. In this study, spiral polycaprolactone (PCL) nanofibrous scaffolds were inserted into poly(lactide-co-glycolide) (PLGA) microsphere sintered tubular scaffolds to form integrated scaffolds to provide mechanical properties and bioactivity appropriate for bone tissue engineering. Four experiment groups were designed: PLGA cylinder scaffold; PLGA tubular scaffold; PLGA tubular scaffold with PCL spiral structured inner core; PLGA tubular scaffold with PCL nanofiber containing spiral structured inner core. The morphology, porosity and mechanical properties of the scaffolds were characterized. Furthermore, human osteoblastic cells were seeded on these scaffolds, and the cell attachment, proliferation, differentiation and mineralized matrix deposition on the scaffolds were evaluated. The integrated scaffolds had Young’s modulus 250–300 MPa, and compressive strength 8–11 MPa under uniaxial compression. With the addition of an inner highly porous insert to the tubular shell, human osteoblast cells seeded on the integrated scaffolds showed slightly higher cell proliferation, 20–25% more alkaline phosphatase expression and twofold higher calcium deposition than those on the cylinder and tubular scaffolds. Furthermore, compared with sintered PLGA cylinder scaffolds, the integrated scaffolds allowed better cellular infiltration Therefore, this design demonstrates great potential for integrated scaffolds in bone tissue engineering applications.     

Repair of large articular osteochondral defects using hybrid scaffolds and bone marrow-derived mesenchymal stem cells in a rabbit model

In order to evaluate the repair potential in large osteochondral defects on high load-bearing sites, a hybrid scaffold system was made of three-dimensional porous Polycaprolactone (PCL) scaffold for the cartilage and tricalcium phosphate-reinforced PCL for the bone portion. Osteochondral defects of 4-mm diameter x 5.5-mm depth were created in the medial femoral condyle of adult New Zealand White rabbits. The defects were treated with hybrid scaffolds without cells (control group) or seeded with allogenic bone marrow-derived mesenchymal stem cells (BMSC) in each part (experimental group) by press-fit implantation. Implanted cells were tracked using Adeno-LacZ labeling. Repair tissues were evaluated at 3 and 6 months after implantation. Overall, the experimental group showed superior repair results as compared to the control group using gross examination, qualitative and quantitative histology, and biomechanical assessment. With BMSC implantation, the hybrid scaffolds provided sufficient support to new osteochondral tissues formation. The bone regeneration was consistently good from 3 to 6 months with firm integration to the host tissue. Cartilage layer resurfacing was more complicated. All of the samples showed cartilage tissues mixed with PCL scaffold filaments at 3 months. Histology at 6 months revealed some degradation phenomenon in several samples whereas others had a good appearance; however, the Young's moduli from the experimental group (0.72 MPa) approached that of normal cartilage (0.81 MPa). In vivo viability of implanted cells was demonstrated by the retention for 6 weeks in the scaffolds. This investigation showed that PCL-based hybrid scaffolds with BMSC may be an alternative treatment for large osteochondral defects in high-loading sites.     

Enhanced bone tissue regeneration by antibacterial and osteoinductive silica-HACC-zein composite scaffolds loaded with rhBMP-2

Next-generation orthopedic implants with both osteoinductivity and antibacterial ability are greatly needed. In the present study, biodegradable rhBMP-2 loaded zein-based scaffolds with a macroporous structure were synthesized, and SBA-15 nanoparticles and hydroxypropyltrimethyl ammonium chloride chitosan (HACC) were incorporated into the scaffolds to produce an anti-infective composite scaffold for delivery of osteogenic factors that facilitate the functional repair of bone defects. The silica/HACC/zein scaffolds developed here showed bioactivity, biocompatibility, and effective antibacterial activity. Confocal laser scanning microscopy (CLSM) was used to quantitatively measure the bactericidal efficacy with respect to bacterial adhesion. Results showed that the sample zein-HACC-S20 exhibited long-lasting antibacterial activity against Escherichia coli and Staphylococcus aureus up to 5 d. At a low dosage of rhBMP-2 (ca. 80 μg), the scaffolds released rhBMP-2 protein efficiently at a relatively slow rate, even after 27 d. An ALP activity and ECM mineralization assay showed that the zein-HACC-S20 scaffolds exhibited significant early osteogenic differentiation by generating enhanced ALP product on day 14 and ECM mineralization on day 21. In a mouse model of thigh muscle pouches, zein-S20 and zein-HACC-S20 groups resulted in obvious bone formation and gave more extensive mineralization to the implants than silica free groups, indicating effective bone induction in vivo. In a rabbit model of critical-sized radius bone defects (20 mm in length and 5 mm in diameter), the bone defects were almost fully repaired and bone marrow cavity recanalization was detectable by 3D micro-CT technique and histological analysis after 12 weeks. In this way, the zein-HACC-S20 scaffolds were proven to significantly promote the bone repair. They also demonstrated considerable promise for tissue engineering. Silica/HACC/zein scaffolds with both antibacterial activity and the ability to induce osteogenesis have immense potential in orthopedics and other biomedical applications.     

Repairing a critical-sized bone defect with highly porous modified and unmodified baghdadite scaffolds

This is the first reported study to prepare highly porous baghdadite (Ca₃ZrSi₂O₉) scaffolds with and without surface modification and investigate their ability to repair critical-sized bone defects in a rabbit radius under normal load. The modification was carried out to improve the mechanical properties of the baghdadite scaffolds (particularly to address their brittleness) by coating their surfaces with a thin layer (～400 nm) of polycaprolactone (PCL)/bioactive glass nanoparticles (nBGs). The β-tricalcium phosphate/hydroxyapatite (TCP/HA) scaffolds with and without modification were used as the control groups. All of the tested scaffolds had an open and interconnected porous structure with a porosity of ～85% and average pore size of 500 μm. The scaffolds (six per scaffold type and size of 4 mm × 4 mm × 15 mm) were implanted (press-fit) into the rabbit radial segmental defects for 12 weeks. Micro-computed tomography and histological evaluations were used to determine bone ingrowth, bone quality, and implant integration after 12 weeks of healing. Extensive new bone formation with complete bridging of the radial defect was evident with the baghdadite scaffolds (modified/unmodified) at the periphery and in close proximity to the ceramics within the pores, in contrast to TCP/HA scaffolds (modified/unmodified), where bone tended to grow between the ulna adjacent to the implant edge. Although the modification of the baghdadite scaffolds significantly improved their mechanical properties, it did not show any significant effect on in vivo bone formation. Our findings suggest that baghdadite scaffolds with and without modification can serve as a potential material to repair critical sized bone defects.