Preparation and Characterization of Nanocomposite Scaffolds (Collagen/β-TCP/SrO) for Bone Tissue Engineering

BACKGROUND: Nowadays, production of nanocomposite scaffolds based on natural biopolymer, bioceramic, and metal ions is a growing field of research due to the potential for bone tissue engineering applications. METHODS: In this study, a nanocomposite scaffold for bone tissue engineering was successfully prepared using collagen (COL), beta-tricalcium phosphate (β-TCP) and strontium oxide (SrO). A composition of β-TCP (4.9 g) was prepared by doping with SrO (0.05 g). Biocompatible porous nanocomposite scaffolds were prepared by freeze-drying in different formulations [COL, COL/β-TCP (1:2 w/w), and COL/β-TCP-Sr (1:2 w/w)] to be used as a provisional matrix or scaffold for bone tissue engineering. The nanoparticles were characterized by X-ray diffraction, Fourier transforms infrared spectroscopy and energy dispersive spectroscopy. Moreover, the prepared scaffolds were characterized by physicochemical properties, such as porosity, swelling ratio, biodegradation, mechanical properties, and biomineralization. RESULTS: All the scaffolds had a microporous structure with high porosity (~ 95–99%) and appropriate pore size (100–200 µm). COL/β-TCP-Sr scaffolds had the compressive modulus (213.44 ± 0.47 kPa) higher than that of COL/β-TCP (33.14 ± 1.77 kPa). In vitro cytocompatibility, cell attachment and alkaline phosphatase (ALP) activity studies performed using rat bone marrow mesenchymal stem cells. Addition of β-TCP-Sr to collagen scaffolds increased ALP activity by 1.33–1.79 and 2.92–4.57 folds after 7 and 14 days of culture, respectively. CONCLUSION: In summary, it was found that the incorporation of Sr into the collagen-β-TCP scaffolds has a great potential for bone tissue engineering applications.

Mechanical Properties and Biological Characteristics of the Porous 3D-Printed β-TCP Composite Scaffolds / 医用生物力学

Objective To study the mechanical properties and biological characteristics of 3D-printed porous β-tricalcium phosphate ［β-Ca3(PO4)2, β-TCP］ scaffolds, so as to provide guidance for the design of composite scaffolds in animal experimentation. Methods Poly 1,8-octanediol citrate (POC), a kind of novel biodegradable materials, was used as the adhesive. The 3D-printed porous β-TCP scaffolds were fabricated by fused deposition modeling (FDM) technology, and Gly-Arg-Gly-Asp-Ser (GRGDS), a kind of polypeptides, was added into the scaffolds to improve the adhesive property of cells. The optical microscope and scanning electron microscope (SEM) were used to observe the micro-pore architectures of those scaffolds. The material testing machine was used to conduct compressive test on the scaffolds, and the water contact angles of the scaffolds were measured. The cell adhesion rate and proliferation rate of the scaffolds were also tested by in vitro cell experiment. The model of SD rat skull defects was repaired by the scaffolds, and the osteogenic ability in vivo was further studied. Results The GRGDS, remaining active, was evenly distributed in the composite scaffolds. The micro-pore architectures of the polypeptide modified scaffolds changed, with improvement in cell adhesion rate, while the compressive modulus, water contact angle and osteogenic ability in vivo of the scaffolds were not obviously affected. Conclusions The cell adhesion capacity of β-TCP composite scaffolds modified by polypeptide improved significantly, while the mechanical properties and hydrophilicity, osteogenic ability in vivo of the scaffolds were not affected very much. These research results provide new ideas for reconstruction of scaffolds for repairing bone defects in clinic, and a laboratory basis for further clinical application of this scaffold.

Histomorphometric study of rabbit's maxillary sinus augmentation with various graft materials / 대한해부학회지

The purpose of this animal study is to evaluate, by histomorphometric analysis, bone regeneration in rabbit's maxillary sinuses with blood clots alone, Bio-Oss, β-tricalcium phosphate (β-TCP), and demineralized tooth dentin (DTD) grafting. Bilateral sinus augmentation procedures were performed in 18 adult male rabbits. Rectangular replaceable bony windows were made with a piezoelectric thin saw insert. In the group 1, blood clots were filled; group 2, anorganic bovine graft (Bio-Oss) was grafted; group 3, β-TCP was grafted; group 4, DTD was grafted, and covered by replaceable bony windows. Animals were sacrificed at 2, 4, and 8 weeks after surgical procedure. The augmented sinuses were evaluated by histomorphometric analysis using hematoxylin and eosin and Masson's trichrome stains. Histologically, new bone formation was revealed along the elevated sinus membrane and all graft materials. The new bone area of the group 2 was significantly greater than the group 1, and of the group 3 was significantly greater than the group 2, and of the group 4 was significantly greater than the group 3 at 8 weeks with P < 0.05. The bone marrow area of group 1 was significantly greater than other groups at 8 weeks. The DTD area was significantly lesser than Bio-Oss or β-TCP particles area at 8 weeks. This present study suggests that DTD can be effective graft materials for bone regeneration of the maxillary sinus augmentation.

The effects of β tricalcium phosphate/collagen scaffold loaded with human bone morphogenetic protein 2 plasmid on the osteogenesis ability of MC3T3-E1 cells / 实用口腔医学杂志

Objective:To study the effects of beta tricalcium phosphate(β-TCP)/collagen scaffold loaded with human bone morphogenetic protein 2 (hBMP2) plasmid on the osteogenesis ability of MC3T3-E1 cells.Methods:hBMP2 DNA plasmid-modified β-TCP/collagen scaffold and the naked plasmid(control) were constructed.MC3T3-E1 cells were respectively in vitro cultured onto the β-TCP/collagen scaffold with hBMP2(Z) and with control plasmid(Z0),on peace dish with the saffold and hBMP2(M) and with the control plasmid(M0).The surface morphology of the samples was observed by SEM.Osteogenesis of the cells was examined by alkaline phosphatase activity(ALP) test,real-time fluorescent quantitative PCR for the detection of Runx2,OCN,ALP and OPN mRNA expression.Data were statistically analyzed.Results:The composite sample surface of plasmid DNA containing hBMP2 modified β-TCP/collagen was porous;group Z and M showed highter ALP activity and higher mRNA expression of Runx2,OCN,ALP and OPN than group Z0 and M0;so did group Z than group M.Conclusion:Porous β-TCP/collagen scaffold loaded with BMP2 DNA is potential for osteoinduction.

Study on the effect of β-TCP/PLGA scaffolds seeded BMSCs combing with hyperbaric Oxygen on bone defect with seawater immersing in rabbit radius / 重庆医学

Objective To investigate therapeutic effects of β-TCP/PLGA scaffolds seeded bone marrow mesenchymal stem cells (BMSCs) combing with hyperbaric oxygen (HBO) on bone defect with seawater immersing in rabbit radius.Methods BMSCs were seeded into the β-TCP/PLGA scaffolds to construct tissue engineering bone.60 New Zealand rabbits were created 1.5 cm bone defect in bilateral radius,and then bilateral wound limbs were immersed in seawater for 3 h.After debridement,all rabbits were randomly divided into 4 groups.In group A nothing was implanted into the bone defect.Only BMSCs was implanted in group B.BMSCs + HBO was given in group C.β-TCP/PLGA BMSCs + HBO was given in group D.At postoperative 4,8,12 weeks rabbits were sacrificed after radiography radius.X-ray radiography,HE staining,immunohistochemical observation were used to evaluate repairing effect of bone defect with seawater immersing.Results Radiographic analysis demonstrated that the bone defects were completely repaired with recanalization of the medullary cavity in group D,bone defects were partially repaired with partal recanalization of the medullary cavity in group C,bone defects were incompletely repaired in group B;bone defect repair unfinished and the broken ends were sclerous in group A.Each time point group D＞group C＞group B＞group A in callus grey value (P＜0.05).HE staining indicated after 12 weeks,a small amount of lainellar bone formated in group A;a small amount of lamellar bone formated in group B;a large amount of lamellar bone formated in group C;a large amount of lamellar bone formated in group D.Immunohistoehemical result indicated at postoperative 4 weeks,the expressions of osteocalcin(OCN) in each group were at high levels,the expressions decreased significantly at Postoperative 8 weeks,the expressions were at a low level at postoperative 12 weeks.At postoperative 4 and 8 weeks,the expression levels of OCN were group D＞ group C＞ group B＞group A (P＜0.05),there was no significant difference at postoperative 12 weeks between the 4 groups (P＞0.05).Conelusion the β-TCP/PLGA composite BMSCs combined hyperbaric oxygen is the effective method to repair seawater immersed bone defect in rabbit.

PCL/β-TCP Composite Scaffolds Exhibit Positive Osteogenic Differentiation with Mechanical Stimulation

We investigated the use of Polycaprolactone (PCL)/β-tricalcium phosphate (β-TCP) composites with applied mechanical stimulation as scaffold for bone tissue engineering. PCL-based three-dimensional (3D) structures were fabricated in a solvent-free process using a 3D-printing technique. The mass fraction of β-TCP was varied in the range 0–30%, and the structure and compressive modulus of the specimens was characterized. The shape and interconnectivity of the pores was found to be satisfactory, and the compressive modulus of the specimens was comparable with that of human trabecular bone. Human mesenchymal stem cells were seeded on the composites, and various biological evaluations were performed over 9 days. With a mass fraction of β-TCP of 30%, differentiation began earlier; however, the cell proliferation rate was lower. Through the use of mechanical stimulation, however, the proliferation rate recovered, and was comparable with that of the other groups. This stimulation effect was also observed in ECM generation and other biological assays. With mechanical stimulation, expression of osteogenic markers was lower on samples with a β-TCP content of 10 wt% than without β-TCP; however, with mechanical stimulation, the sample with a β-TCP content of 30 wt% exhibited significantly greater expression of those markers than the other samples. We found that mechanical stimulation and the addition of β-TCP interacted closely, and that a mass fraction of β-TCP of 30% was particularly useful as a bone tissue scaffold when accompanied by mechanical stimulation.

The Influences of Different Ratios of Biphasic Calcium Phosphate and Collagen Augmentation on Posterior Lumbar Spinal Fusion in Rat Model

PURPOSE: To determine the influence of different ratios of hydroxyapatite (HA)/beta tricalcium phosphate (β-TCP) and collagen augmentation for posterior lumbar fusion in a rat model. MATERIALS AND METHODS: We generated a posterior lumbar fusion model in 50 rats and divided it into five groups of equal number as follows; 1) autologous bone graft as group A, 2) 70% HA+30% β-TCP as group B, 3) 70% HA+30% β-TCP+collagen as group C, 4) 30% HA+70% β-TCP as group D, and 5) 30% HA+70% β-TCP+collagen as group E. Rats were euthanized at 12 weeks after surgery and fusion was assessed by manual palpation, quantitative analysis using microCT and histology. RESULTS: The score of manual palpation was significantly higher in group C than group E (3.1±1.1 vs. 1.8±0.8, p=0.033). However, in terms of microCT analysis, group D showed significantly higher scores than group B (5.5±0.8 vs. 3.1±1.1, p=0.021). According to quantitative volumetric analysis, 30% HA+70% β-TCP groups (group D and E) showed significantly reduced fusion mass at 12 weeks after surgery (123±14.2, 117±46.3 vs. 151±27.3, p=0.008, 0.003, respectively). Collagen augmentation groups revealed superior results in terms of both microCT score and histologic grade. CONCLUSION: A 7:3 HA/β-TCP ratio with collagen augmentation rather than a 3:7 HA/β-TCP ratio could be a more favorable graft substitute for lumbar spinal fusion. There was positive role of collagen as an adjunct for spinal bone fusion process.

Preparation and in vitro release behavior of adiponectin microsphere-scaffold composite / 实用口腔医学杂志

Objective:To prepare a bone substitute using microsphere scaffold containing adiponectin(APN)and to investigate the release behavior of the scaffold in vitro.Methods:Chitosan microsphere was developed by an emulsion-ionic cross-linking method. Poly (L-lactic-co-glycolic)acid (PLGA)and β-tricalcium phosphate (β-TCP)were used to prepare microsphere scaffold containing APN.The morphology,particle size,drug loading,incorporation efficiency and release behavior of the microsphere were examined. Results:The APN containing microsphere showed good spherical geometry,suitable size and microporosity under scanning electron microscope.The average diameter of the milipore was 20 -200 μm;the drug loading and incorporation efficiency were 1 .3% and 70.3% respectively.The controled-release process continued for 91 days.The extract solution from the APN microsphere-scaffold promoted MC3T3 cell proliferation without cytotoxicity.Conclusion:The APN microsphere-scaffold has sustained release function and may promote osteoblast proliferation.

Effect of beta-tricalcium phosphate/poly-l-lactide composites on radial bone defects of rabbit

OBJECTIVE@#To explore the effect ofβ-TCP/PLLA scaffold in repairing rabbit radial bone defects.@*METHODS@#Thirty New Zealand rabbits were divided into β-TCP /PLLA group (group A), pure PLLA group (group B) and contrast group (group C) randomly. The rabbits were sacrificed respectively after 4, 8, 12, 24 weeks and the X-ray film was performed at the same time to evaluate the repair effect in different groups.@*RESULTS@#X-ray film showed there was uneven low density bone callus development in defect region after 4 weeks in group A. The defect region was filled with neonate osseous tissue completely during 12-24 weeks. X-ray score revealed that repair of bone defect results significantly better than group B and group C.@*CONCLUSIONS@#The β-TCP /PLLA composite is capable of repairing radial bone bone defects. β-TCP/PLLA scaffold is significant because of rapid degradation ability, good histocompatibility and osteogenic action.

Study on the setting property and compressive strength of β-TCP/α-CSH combined bone graft / 医用生物力学

Objective To investigate the setting property and compressive strength of β-TCP (β-tricalcium phosphate)/α-CSH (α-calcium sulphate hemihydrate) combined bone graft. Method β TCP/α CSH combined bone graft and distilled water were mixed with different ratio: 1 g∶0.1 mL、1 g∶0.2 mL、1 g∶0.3 mL、1 g∶0.4 mL、1 g∶0.5 mL, respectively. The initial setting time (Ti), final setting time (TF), compressive strength were recorded, and the X-ray diffraction (XRD) and the scanning electron microscope (SEM) were examined. Results The initial and final setting time of combined bone graft increased along with the increase of solid/liquid ratio. The Ti and TF were (4.6±1.3) min and (13.1±2.9) min respectively when the solid/liquid ratio was 1 g∶0.2 mL. The average compressive strength was 7.86 MPa in one day after setting, which was about two times higher than that of β-TCP alone. The XRD results showed that no other materials were produced except that α-CSH was transformed to CSD in the setting process. The SEM results indicated that thick CSD granules were adhered to the porous surfaces of β-TCP granules after setting. Conclusions The setting time and compressive strength of β-TCP/α-CSH combined bone graft could be adjusted by the different solid/liquid ratio.