The impact of low-magnitude high-frequency vibration on fracture healing is profoundly influenced by the oestrogen status in mice

Fracture healing is impaired in aged and osteoporotic individuals. Because adequate mechanical stimuli are able to increase bone formation, one therapeutical approach to treat poorly healing fractures could be the application of whole-body vibration, including low-magnitude high-frequency vibration (LMHFV). We investigated the effects of LMHFV on fracture healing in aged osteoporotic mice. Female C57BL/6NCrl mice (n=96) were either ovariectomised (OVX) or sham operated (non-OVX) at age 41 weeks. When aged to 49 weeks, all mice received a femur osteotomy that was stabilised using an external fixator. The mice received whole-body vibrations (20 minutes/day) with 0.3 g peak-to-peak acceleration and a frequency of 45 Hz. After 10 and 21 days, the osteotomised femurs and intact bones (contra-lateral femurs, lumbar spine) were evaluated using bending-testing, micro-computed tomography (μCT), histology and gene expression analyses. LMHFV disturbed fracture healing in aged non-OVX mice, with significantly reduced flexural rigidity (−81%) and bone formation (−80%) in the callus. Gene expression analyses demonstrated increased oestrogen receptor β (ERβ, encoded by Esr2) and Sost expression in the callus of the vibrated animals, but decreased β-catenin, suggesting that ERβ might mediate these negative effects through inhibition of osteoanabolic Wnt/β-catenin signalling. In contrast, in OVX mice, LMHFV significantly improved callus properties, with increased flexural rigidity (+1398%) and bone formation (+637%), which could be abolished by subcutaneous oestrogen application (0.025 mg oestrogen administered in a 90-day-release pellet). On a molecular level, we found an upregulation of ERα in the callus of the vibrated OVX mice, whereas ERβ was unaffected, indicating that ERα might mediate the osteoanabolic response. Our results indicate a major role for oestrogen in the mechanostimulation of fracture healing and imply that LMHFV might only be safe and effective in confined target populations.KEY WORDS: Whole-body vibration, LMHFV, Fracture healing, Oestrogen receptor signalling, Wnt signalling     

The Osteogenic Potential of Mesoporous Bioglasses/Silk and Non-Mesoporous Bioglasses/Silk Scaffolds in Ovariectomized Rats: In vitro and In vivo Evaluation

Silk-based scaffolds have been introduced to bone tissue regeneration for years, however, their local therapeutic efficency in bone metabolic disease condition has been seldom reported. According to our previous report, mesoporous bioactive glass (MBG)/silk scaffolds exhibits superior in vitro bioactivity and in vivo osteogenic properties compared to non-mesoporous bioactive glass (BG)/silk scaffolds, but no information could be found about their efficiency in osteoporotic (OVX) environment. This study investigated a biomaterial-based approach for improving MSCs behavior in vitro, and accelerating OVX defect healing by using 3D BG/silk and MBG/silk scaffolds, and pure silk scaffolds as control. The results of SEM, CCK-8 assay and quantitative ALP activity showed that MBG/silk scaffolds can improve attachment, proliferation and osteogenic differentiation of both O-MSCs and sham control. In vivo therapeutic efficiency was evaluated by μCT analysis, hematoxylin and eosin staining, safranin O staining and tartrate-resistant acid phosphatase, indicating accelerated bone formation with compatible scaffold degradation and reduced osteoclastic response of defect healing in OVX rats after 2 and 4 weeks treatment, with a rank order of MBG/silk > BG/silk > silk group. Immunohistochemical markers of COL I, OPN, BSP and OCN also revealed that MBG/silk scaffolds can better induce accelerated collagen and non-collagen matrix production. The findings of this study suggest that MBG/silk scaffolds provide a better environment for cell attachment, proliferation and differentiation, and act as potential substitute for treating local osteoporotic defects.     

The Masquelet Technique for Membrane Induction and the Healing of Ovine Critical Sized Segmental Defects

The healing of critical sized segmental defects is an ongoing clinical problem. No method has achieved pre-eminence. The Masquelet technique is a relatively new innovation involving the induction of a fibrous tissue membrane around the bone defect site taking advantage of the body’s foreign body reaction to the presence of a polymethylmethacrylate (PMMA) spacer. The aim of this study was to investigate the properties and characteristics of this induced membrane and its effectiveness when used in conjunction with allograft or an allograft/autograft mix as filler materials in an ovine critical sized defect model. The resultant induced membrane was found to be effective in containing the graft materials in situ. It was demonstrated to be an organised pseudosynovial membrane which expressed bone morphogenic protein 2 (BMP2), transforming growth factor- beta (TGFβ), vascular endothelial growth factor (VEGF), von Willerbrand factor (vWF), interleukin 6 (IL-6) and interleukin 8 (IL-8). While more new bone growth was evident in the test groups compared to the controls animals at 12 weeks, the volumes were not statistically different and no defects were fully bridged. Of the two graft material groups, the allograft/autograft mix was shown to have a more rapid graft resorption rate than the allograft only group. While the Masquelet technique proved effective in producing a membrane to enclose graft materials, its ability to assist in the healing of critical sized segmental defects when compared to empty controls remained inconclusive.     

表面改性双相陶瓷复合自体骨髓干细胞修复兔骨缺损的实验研究

目的:应用自体骨髓基质干细胞(Bone Marrow Stromal Cells,BMSCs)复合经低晶态羟基磷灰石(Low Crystalline Hydroxyap- atite,LcHA)涂层的双相陶瓷(Biphasic Calcium Phosphate,BCP)构建的组织工程化骨(LcBCP)修复兔桡骨节段性缺损。方法:体外分离培养、诱导扩增兔BMSCs,取第三代细胞复合LcBCP(实验组)后修复15只兔左侧桡骨15mm缺损;右侧桡骨缺损处植入复合BMSCs的BCP(对照组),于植入后4、8和12周处死动物,通过大体形态、组织学、影像学和扫描电镜检测骨缺损修复效果。结果:BMSCs-LcBCP复合物生长良好,随时间延长,X线显示实验组连接处骨痂形成,对照组连接处始终愈合稍差,12周大体观察实验组骨修复良好,髓腔再通;组织学显示板层骨形成,连接处骨性愈合,实验对照组连接处虽然也为骨性愈合,但尚有较多编织骨形成。结论:自体BMSCs复合LcBCP形成的组织工程化骨可修复兔桡骨节段性缺损,低晶态羟基磷灰石涂层能够增强双相陶瓷的早期成骨。     

多孔n-HA/PA66支架复合rhBMP-2修复兔桡骨缺损的实验研究

目的 ：研究多孔纳米羟基磷灰石/聚酰胺66（nHA/PA66）骨修复材料作为骨组织工程支架复合基因重组人骨形态发生蛋白2（rhBMP2）后的成骨能力的变化,探讨加速nHA/PA66人工骨与受体骨愈合的方法。方法：选用新西兰大白兔双侧桡骨制作骨缺损模型,将nHA/PA66/rhBMP2复合材料植入左侧骨缺损处,右侧骨缺损以nHA/PA66植入作为实验对照,另做不植入任何材料的骨缺损空白对照。在1、2、4、8、12周各时相点分别进行大体观察、X线照片、组织学切片、免疫组化原位杂交进行检测图象分析。结果：nHA/PA66/BMP2与nHA/PA66组骨缺损均完全修复,而空白对照组骨缺损未见修复;2周时nHA/PA66与nHA/PA66/rhBMP2两组间原位杂交阳性细胞表达有统计学意义（ P<0.05）, 4周时nHA/PA66与nHA/PA66/rhBMP2两组间原位杂交阳性细胞表达无统计学意义（ P>0.05）,2周及4周实验和实验对照两组分别与空白对照组比较均无统计学意义（ P>0.05）,nHA/PA66/rhBMP2组较nHA/PA66组可加速人工骨/植入体/受体界面骨愈合。 结论：多孔nHA/PA66作为骨组织工程支架复合具有诱导成骨活性的rhBMP2后,增强了早期成骨能力,加速了其与受体骨的愈合。     

Bone‐Inspired Tube Filling Decellularized Matrix of Toad Cartilage Provided an Osteoinductive Microenvironment for Mesenchymal Stem Cells to Facilitate the Radius Defect Repair of Rabbit

Toad bone not only contains the rich cartilage‐like matrix but also presents low immunogenicity. It is inferred that decellularized toad bone matrix (dBECM) may provide the more profitable osteoinductive microenvironment for mesenchymal stem cells (MSCs) to promote the repair of bone defects. Herein, a hollow bone‐inspired tube is first made from hydroxyapatite (HA) and poly (γ‐glutamic acid) (PGA), and then MSCs/dBECM hydrogel is uniformly filled to its central cavity, constructing a biomimetic bone (dBECM + MSCs ? PGA + HA). In vitro scratch and transwell experiments show that dBECM hydrogel not only effectively promotes migration and proliferation of MSCs but also induces their osteogenic differentiation. Moreover, the less inflammatory macrophages infiltrate at rat skin after subcutaneously injecting dBECM hydrogel, indicating its low potential for inflammatory attack. After implanting dBECM + MSCs ? PGA + HA to critical radius defect of rabbit, X‐ray and CT imaging shows that the cortex is effectively regenerated and the medullary cavity recanalization is completed at 20 weeks. Moreover, the expression of Collagen‐II and OCN are obviously increased in the defect after implanting dBECM + MSCs ? PGA + HA. The therapeutic mechanism of dBECM + MSCs ? PGA + HA scaffold are highly associated with the enhanced angiogenesis. Collectively, the biomimetic dBECM + MSCs ? PGA + HA scaffold may be a promising strategy to improve radius defect healing efficiency.     

Dimethyloxalylglycine Prevents Bone Loss in Ovariectomized C57BL/6J Mice through Enhanced Angiogenesis and Osteogenesis

Hypoxia-inducible factor 1-α (HIF-1α) plays a critical role in angiogenesis-osteogenesis coupling during bone development and bone regeneration. Previous studies have shown that 17β-estradiol activates the HIF-1α signaling pathway and that mice with conditional activation of the HIF-1α signaling pathway in osteoblasts are protected from ovariectomy (OVX)-induced bone loss. In addition, it has been shown that hypoxia facilitates the osteogenic differentiation of mesenchymal stem cells (MSCs) and modulates Wnt/β-catenin signaling. Therefore, we hypothesized that activation of the HIF-1α signaling pathway by hypoxia-mimicking agents would prevent bone loss due to estrogen deficiency. In this study, we confirmed the effect of dimethyloxalylglycine (DMOG), a hypoxia-mimicking agent, on the HIF-1α signaling pathway and investigated the effect of DMOG on MSC osteogenic differentiation and the Wnt/β-catenin signaling pathway. We then investigated the effect of DMOG treatment on OVX-induced bone loss. Female C57BL/6J mice were divided into sham, OVX, OVX+L-DMOG (5 mg/kg/day), and OVX+H-DMOG (20 mg/kg/day) groups. At sacrifice, static and dynamic bone histomorphometry were performed with micro computed tomography (micro-CT) and undecalcified sections, respectively. Bone strength was assessed with the three-point bending test, and femur vessels were reconstructed and analyzed by micro-CT. Serum vascular endothelial growth factor (VEGF), osteocalcin, and C-terminal telopeptides of collagen type(CTX) were measured by ELISA. Tartrate-resistant acid phosphatase staining was used to assess osteoclast formation. Alterations in the HIF-1α and Wnt/β-catenin signaling pathways in the bone were detected by western blot. Our results showed that DMOG activated the HIF-1α signaling pathway, which further activated the Wnt/β-catenin signaling pathway and enhanced MSC osteogenic differentiation. The micro-CT results showed that DMOG treatment improved trabecular bone density and restored the bone microarchitecture and blood vessels in OVX mice. Bone strength was also partly restored in DMOG-treated OVX mice. Dynamic bone histomorphometric analysis of the femur metaphysic revealed that DMOG increased the mineralizing surface, mineral apposition rate, and bone formation rate. The serum levels of VEGF and osteocalcin were higher in DMOG-treated OVX mice. However, there were no significant differences in serum CTX or in the number of tartrate-resistant acid phosphatase-stained cells between DMOG-treated OVX mice and OVX mice. Western blot results showed that DMOG administration partly rescued the decrease in HIF-1α and β-catenin expression following ovariectomy. Collectively, these results indicate that DMOG prevents bone loss due to ovariectomy in C57BL/6J mice by enhancing angiogenesis and osteogenesis, which are associated with activated HIF-1α and Wnt/β-catenin signaling pathways.     

Influence of Exercise on Bone Remodeling-Related Hormones and Cytokines in Ovariectomized Rats: A Model of Postmenopausal Osteoporosis

This study aims to explore the effects of exercise on postmenopausal osteoporosis and the mechanisms by which exercise affects bone remodeling. Sixty-three Wistar female rats were randomly divided into five groups: (1) control group, (2) sham-operated group, (3) OVX (Ovariectomy) group, (4) DES-OVX (Diethylstilbestrol-OVX) group, and (5) Ex-OVX (Exercise-OVX) group. The rat osteoporosis model was established through ovariectomy. The Ex-OVX rats were made to run 251.2 meters every day, 6 d/wk for 3 months in a running wheel. Trabecular bone volume (TBV%), total resorption surface (TRS%), trabecular formation surface (TFS%), mineralization rate (MAR), bone cortex mineralization rate (mAR), and osteoid seam width (OSW) were determined by bone histomorphometry. The mRNA and protein levels of interleukin-1β (IL-1β₂), interleukin-6 (IL-6), and cyclooxygenase-2 (Cox-2) were determined by in situ hybridization and immunohistochemistry, respectively. Serum levels of estrogen estradiol (E₂), calcitonin (CT), osteocalcin (BGP), and parathyroid hormone (PTH) were determined by ELISA assays. The investigation revealed that compared to the control and the sham-operated groups, the OVX group showed significantly lower levels of TBV%, E₂, and CT, but much higher levels of TRS%, TFS%, MAR, OSW, BGP, and PTH. The Ex-OVX group showed increased TBV% and serum levels of E₂ and CT compared to the OVX group. Ovariectomy also led to a significant increase in IL-1β mRNA and protein levels in the bone marrow and IL-6 and Cox-2 protein levels in tibias. In addition, the Ex-OVX group showed lower levels of IL-1 mRNA and protein, IL-6 mRNA, and Cox-2 mRNA and protein than those in the OVX group. The upshot of the study suggests that exercise can significantly increase bone mass in postmenopausal osteoporosis rat models by inhibiting bone resorption and increasing bone formation, especially in trabecular bones.     

利用聚己内酯/羟基磷灰石复合支架修复犬胸骨缺损

目的：探讨利用生物可降解支架修复动物胸骨缺损,为临床手术治疗提供新的可行性方法。方法：对于12只比格犬进行手术切除部分胸骨,并利用聚己内酯/羟基磷灰石（PCL/HA）复合支架,并制备出与临床相似的胸骨缺损模型。实验动物分成2组,分别是：空白对照组和PCL/HA支架组。分别于术后第4、12周进行胸部CT扫描,并对胸廓进行三维重建,观察胸骨缺损部位的修复情况,并在第12周取胸骨缺损部位组织进行硬组织切片,苦味酸-品红染色,观察缺损部位的骨组织修复情况,并利用软件进行骨组织比率分析,评估修复情况。结果：通过检查发现空白对照组的胸骨缺损部位未见明显骨连接,胸廓的骨性结构有明显畸形,PCL/HA支架组能很好地维持胸廓的完整性,组织学检查发现PCL/HA支架组的缺损部位有明显新生骨形成,通过软件分析可发现支架组的骨组织比率较空白组的高（P〈0.05）。结论：这些结果表明采用PCL/HA复合材料支架能很好地修复胸骨缺损。     

Bone loss in the ovariectomized baboon Papio ursinus: densitometry, histomorphometry and biochemistry

To develop a non-human primate model of systemic bone loss after ovariectomy, 24 ovariectomized (OVX) and eight control (non-OVX) female baboons Papio ursinus were investigated over a period of 48 months using bone mineral density (BMD), iliac crest bone histomorphometry, bone turnover markers, and variables of calcium metabolism. Lumbar spine (L1-L4) BMD measured by dual energy X-ray absorptiometry (DXA) decreased in OVX animals in the first 12 months (-7.6%) and showed a slow trend towards recovery after 24 months. Controls showed a slow increase in spinal BMD over 4 years (+9.7%). Total hip BMD decreased slowly up to 48 months in all animals (OVX -12.6%versus controls -10%); this indicated that OVX had a limited effect on total hip BMD. Forearm BMD did not change. The significant decrease in trabecular bone volume (TBV) of the iliac crest from baseline to 12 months was followed by some recovery. Microarchitectural deterioration of trabecular bone in OVX animals was demonstrated by a decline in trabecular number and an increase in trabecular spacing. These changes were also evident on sections of whole vertebrae, proximal femora and iliac crests. Changes in iliac TBV reflected spinal but not hip BMD changes in the OVX animals. Static and dynamic histomorphometric variables indicated that bone turnover was increased for 36 months following OVX. Controls showed no changes in histomorphometric variables. Bone specific alkaline phosphatase (ALPs) in OVX animals remained elevated throughout the study; osteocalcin (OC) was significantly elevated only at 6 and 12 months, and deoxypyridinoline (Pyr-D) was elevated at 12 months but declined after 24 months. ALPs was thus more sensitive to the long-term effects of OVX than were OC or Pyr-D. Controls showed no changes in bone turnover markers. This study showed consistent deleterious changes in lumbar BMD, bone histomorphometry with microarchitectural deterioration together with altered biochemical markers of bone turnover in the first 12 months after OVX. Since these changes resemble those in post-menopausal women, the non-human primate Papio ursinus is suitable for the study of bone loss in post-menopausal women.     

Osteogenic Properties of PBLG-g-HA/PLLA Nanocomposites

New development of biomaterial scaffolds remains a prominent issue for the regeneration of lost or fractured bone. Of these scaffolds, a number of bioactive polymers have been synthesized and fabricated for diverse biological roles. Although recent evidence has demonstrated that composite scaffolds such as HA/PLLA have improved properties when compared to either HA or PLLA alone, recent investigations have demonstrated that the phase compatibility between HA and PLLA layers is weak preventing optimal enhancement of the mechanical properties and making the composites prone to breakdown. In the present study, poly (γ-benzyl-L-glutamate) modified hydroxyapatite/(poly (L-lactic acid)) (PBLG-g-HA/PLLA) composite scaffolds were fabricated with improved phase compatibility and tested for their osteogenic properties in 18 Wistar female rats by analyzing new bone formation in 3 mm bilateral femur defects in vivo. At time points, 2, 4 and 8 weeks post surgery, bone formation was evaluated by µ-CT and histological analysis by comparing 4 treatment groups; 1) blank defect, 2) PLLA, 3) HA/PLLA and 4) PBLG-g-HA/PLLA scaffolds. The in vivo analysis demonstrated that new bone formation was much more prominent in HA/PLLA and PBLG-g-HA/PLLA groups as depicted by µ-CT, H&E staining and immunohistochemistry for collagen I. TRAP staining was also utilized to determine the influence of osteoclast cell number and staining intensity to the various scaffolds. No significant differences in either staining intensity or osteoclast numbers between all treatment modalities was observed, however blank defects did contain a higher number of osteoclast-like cells. The results from the present study illustrate the potential of PBLG-g-HA/PLLA scaffolds for bone tissue engineering applications by demonstrating favorable osteogenic properties.     

Periostin Accelerates Bone Healing Mediated by Human Mesenchymal Stem Cell-Embedded Hydroxyapatite/Tricalcium Phosphate Scaffold

Background

Periostin, an extracellular matrix protein, is expressed in bone, more specifically, the periosteum and periodontal ligaments, and plays a key role in formation and metabolism of bone tissues. Human adipose tissue-derived mesenchymal stem cells (hASCs) have been reported to differentiate into osteoblasts and stimulate bone repair. However, the role of periostin in hASC-mediated bone healing has not been clarified. In the current study, we examined the effect of periostin on bone healing capacity of hASCs in a critical size calvarial defect model.

Methods and Results

Recombinant periostin protein stimulated migration, adhesion, and proliferation of hASCs in vitro. Implantation of either hASCs or periostin resulted in slight, but not significant, stimulation of bone healing, whereas co-implantation of hASCs together with periostin further potentiated bone healing. In addition, the number of Ki67-positive proliferating cells was significantly increased in calvarial defects by co-implantation of both hASCs and periostin. Consistently, proliferation of administered hASCs was stimulated by co-implantation with periostin in vivo. In addition, co-delivery of hASCs with periostin resulted in markedly increased numbers of CD31-positive endothelial cells and α-SMA-positive arterioles in calvarial defects.

Conclusions

These results suggest that recombinant periostin potentiates hASC-mediated bone healing by stimulating proliferation of transplanted hASCs and angiogenesis in calvarial defects.     

Functional Adaptation in Female Rats: The Role of Estrogen Signaling

Background

Sex steroids have direct effects on the skeleton. Estrogen acts on the skeleton via the classical genomic estrogen receptors alpha and beta (ERα and ERβ), a membrane ER, and the non-genomic G-protein coupled estrogen receptor (GPER). GPER is distributed throughout the nervous system, but little is known about its effects on bone. In male rats, adaptation to loading is neuronally regulated, but this has not been studied in females.

Methodology/Principal Findings

We used the rat ulna end-loading model to induce an adaptive modeling response in ovariectomized (OVX) female Sprague-Dawley rats. Rats were treated with a placebo, estrogen (17β-estradiol), or G-1, a GPER-specific agonist. Fourteen days after OVX, rats underwent unilateral cyclic loading of the right ulna; half of the rats in each group had brachial plexus anesthesia (BPA) of the loaded limb before loading. Ten days after loading, serum estrogen concentrations, dorsal root ganglion (DRG) gene expression of ERα, ERβ, GPER, CGRPα, TRPV1, TRPV4 and TRPA1, and load-induced skeletal responses were quantified. We hypothesized that estrogen and G-1 treatment would influence skeletal responses to cyclic loading through a neuronal mechanism. We found that estrogen suppresses periosteal bone formation in female rats. This physiological effect is not GPER-mediated. We also found that absolute mechanosensitivity in female rats was decreased, when compared with male rats. Blocking of adaptive bone formation by BPA in Placebo OVX females was reduced.

Conclusions

Estrogen acts to decrease periosteal bone formation in female rats in vivo. This effect is not GPER-mediated. Gender differences in absolute bone mechanosensitivity exist in young Sprague-Dawley rats with reduced mechanosensitivity in females, although underlying bone formation rate associated with growth likely influences this observation. In contrast to female and male rats, central neuronal signals had a diminished effect on adaptive bone formation in estrogen-deficient female rats.     

Evaluation of new porous β-tri-calcium phosphate ceramic as bone substitute in goat model

The present study was carried out to evaluate the porous β-tri-calcium phosphate (TCP) (prepared by aqueous solution combustion technique) as bone substitute and compared with normal healing in 12 adult Black Bengal goats on the basis of clinical and radiographic findings, histological studies, oxytetracycline labeling, angiography studies (on day 90). Bone defects created in the diaphysis of radius were left unfilled in control animals (group I); while in treated (group II) animals the defects were filled with porous TCP blocks. The three months study showed no marked acute inflammatory reactions in all animals, wound healing was uneventful and the implants were clinically stable in the bone. Radiological studies showed presence of unabsorbed implants which acted as a scaffold for new bone growth across the defect whereas in control animals the defect was more or less same except that the newly formed bony tissue was less organized. Histological section showed moderately differentiated lamellar bone in the cortical part with presence of woven bone at peripheral cortex whereas control animals showed moderate fibro-collagenisation and good amount of marrow material, fat cells and blood vessels. Oxytetracycline labeling study showed crossing over of new bony trabeculae along with presence of resorption cavities within the new osteoid tissues whereas in group I, the process of new bone formation was active from both the ends; the defect site appeared as a homogenous non-fluorescent area. Angiogram of the animals in control showed uniform angiogenesis in the defect site with establishment of trans transplant angiogenesis, whereas in group II there was complete trans transplant shunting of blood vessels communication. The results of this study pointed out that the porous TCP promoted extensive bone formation over the entire extension of the defect in comparison to control group, thus conforming their biological osteoconductive property.     

Yeast-Incorporated Gallium Promotes Fracture Healing by Increasing Callus Bony Area and Improving Trabecular Microstructure on Ovariectomized Osteopenic Rats

The purpose of this study was to analyze the impact of yeast-incorporated gallium on fracture healing in ovariectomized osteopenic rats. Forty Wistar female rats used were divided into three groups: sham-operated rats (SHAM), ovariectomized (OVX) rats, and ovx rats treated with yeast-bound gallium (YG). A standardized fracture-healing model with stable plate fixation was established for rat femoral. After 4-week stable fixation, animals were killed to prepare bones for Micro-CT, biomechanical testing, and histomorphometry. In addition, bone samples were obtained to evaluate the content of mineral substances in bones. Quantitative analysis of the bones from animals in the organic gallium group revealed significantly increased mineral contents compared to bones from OVX and SHAM groups. Micro-CT showed that treatment with yeast-incorporated gallium increased BV/TV and trabecular thickness and decreased trabecular separation in ovx animals. Histomorphometric evaluation demonstrated that YG increased callus area and callus bone formation. Yeast-bound gallium also improved the biomechanical properties of bone healing. In conclusion, this study suggests that yeast-incorporated gallium could promote fracture healing in ovariectomized rats.     

Comparison of the osteogenic potentials of autologous cultured osteoblasts and mesenchymal stem cells loaded onto allogeneic cancellous bone granules

We compared the bone regeneration potentials of autologous cultured osteoblasts and of bone-marrow-derived autologous MSCs in combination with allogeneic cancellous bone granules in a rabbit radial defect model. Radial shaft defects over 15 mm were made in 26 New Zealand white rabbits. The animals underwent insertion of allogeneic cancellous bone granules containing autologous osteoblasts into right-side defects (the experimental group) and of allogeneic cancellous bone granules with autologous MSCs into left-side defects (the control group). To quantitatively assess bone regeneration, radiographic evaluations as well as BMD and BMC measurements were performed 3, 6, 9 and 12 weeks post-implantation and histology as well as micro-CT image analysis were performed at 6 and 12 weeks. Radiographic evaluations 3 weeks post-implantation showed that the experimental group had a higher mean bone quantity index (p < 0.05) and micro-CT image analysis showed that experimental sides had a greater mean total regenerated bone volume and surface area than the control sides (p < 0.05). Histologic evaluations obtained at 6 and 12 weeks revealed distinctly greater granule resorption and new bone formation in the experimental group. This in vivo study demonstrates that a combination of autologous osteoblasts and small-sized, allogeneic cancellous bone granules leads to more rapid bone regeneration than autologous MSCs and small-sized, allogeneic cancellous bone granules.     

Asprin‐loaded strontium‐containing α‐calcium sulphate hemihydrate/nano‐hydroxyapatite composite promotes regeneration of critical bone defects

Our laboratory originally synthesized strontium(Sr)‐containing α‐calcium sulphate hemihydrate/nano‐hydroxyapatite composite (Sr‐α‐CSH/n‐HA) and demonstrated its ability to repair critical bone defects. This study attempted to incorporate aspirin into it to produce a better bone graft material for critical bone defects. After 5% Sr‐α‐CSH was prepared by coprecipitation and hydrothermal methods, it was mixed with aspirin solution of different concentrations (50 μg/ml, 200 μg/ml, 800 μg/ml and 3200 μg/ml) at a fixed liquid‐solid ratio (0.54 v/w) to obtain aspirin‐loaded Sr‐α‐CSH/n‐HA composite. In vitro experiments were performed on the composite extracts. The tibial defects (3 mm*5 mm) in SD rat model were filled with the composite for 4 weeks and 12 weeks to evaluate its osteogenic capacity in vivo. Our results showed its capability of proliferation, migration and osteogenesis of BMSCs in vitro got improved. In vivo treatment with 800 μg/ml aspirin–loaded Sr‐α‐CSH/n‐HA composite led to significantly more new bone formation in the defects compared with Sr‐α‐CSH/n‐HA composite and significantly promoted the expression of osteogenic‐related genes and inhibited osteoclast activity. In general, our research suggests that aspirin‐loaded Sr‐α‐CSH/n‐HA composite may have a greater capacity of repairing tibial defects in SD rats than simple Sr‐α‐CSH/n‐HA composite.     

Distributional Variations in Trabecular Architecture of the Mandibular Bone: An In Vivo Micro-CT Analysis in Rats

Purpose

To evaluate the effect of trabecular thickness and trabecular separation on modulating the trabecular architecture of the mandibular bone in ovariectomized rats.

Materials and Methods

Fourteen 12-week-old adult female Wistar rats were divided into an ovariectomy group (OVX) and a sham-ovariectomy group (sham). Five months after the surgery, the mandibles from 14 rats (seven OVX and seven sham) were analyzed by micro-CT. Images of inter-radicular alveolar bone of the mandibular first molars underwent three-dimensional reconstruction and were analyzed.

Results

Compared to the sham group, trabecular thickness in OVX alveolar bone decreased by 27% (P = 0.012), but trabecular separation in OVX alveolar bone increased by 59% (P = 0.005). A thickness and separation map showed that trabeculae of less than 100μm increased by 46%, whereas trabeculae of more than 200μm decreased by more than 40% in the OVX group compared to those in the sham group. Furthermore, the OVX separation of those trabecular of more than 200μm was 65% higher compared to the sham group. Bone mineral density (P = 0.028) and bone volume fraction (p = 0.001) were also significantly decreased in the OVX group compared to the sham group.

Conclusions

Ovariectomy-induced bone loss in mandibular bone may be related to the distributional variations in trabecular thickness and separation which profoundly impact the modulation of the trabecular architecture.     

Dimethyloxaloylglycine Improves Angiogenic Activity of Bone Marrow Stromal Cells in the Tissue-Engineered Bone

One of the big challenges in tissue engineering for treating large bone defects is to promote the angiogenesis of the tissue-engineered bone. Hypoxia inducible factor-1α (HIF-1α) plays an important role in angiogenesis-osteogenesis coupling during bone regeneration, and can activate a broad array of angiogenic factors. Dimethyloxaloylglycine (DMOG) can activate HIF-1α expression in cells at normal oxygen tension. In this study, we explored the effect of DMOG on the angiogenic activity of bone mesenchymal stem cells (BMSCs) in the tissue-engineered bone. The effect of different concentrations of DMOG on HIF-1a expression in BMSCs was detected with western blotting, and the mRNA expression and secretion of related angiogenic factors in DMOG-treated BMSCs were respectively analyzed using qRT-PCR and enzyme linked immunosorbent assay. The tissue-engineered bone constructed with β-tricalcium phosphate (β-TCP) and DMOG-treated BMSCs were implanted into the critical-sized calvarial defects to test the effectiveness of DMOG in improving the angiogenic activity of BMSCs in the tissue-engineered bone. The results showed DMOG significantly enhanced the mRNA expression and secretion of related angiogenic factors in BMSCs by activating the expression of HIF-1α. More newly formed blood vessels were observed in the group treated with β-TCP and DMOG-treated BMSCs than in other groups. And there were also more bone regeneration in the group treated with β-TCP and DMOG-treated BMSCs. Therefore, we believed DMOG could enhance the angiogenic activity of BMSCs by activating the expression of HIF-1α, thereby improve the angiogenesis of the tissue-engineered bone and its bone healing capacity.