Femoral Head In Rabbits Research Articles

Evaluation of Femoral Head Perfusion by Contrast-Enhanced Ultrasound in a Rabbit Model of Steroid-Induced Osteonecrosis.

To study the technical feasibility of contrast-enhanced ultrasound (CEUS) in evaluating femoral head perfusion in a rabbit model of steroid-induced femoral head osteonecrosis. Twenty rabbits were divided randomly into a control group (n = 8) and an experimental group (n = 12). Rabbits in the experimental group were induced by lipopolysaccharide and methylprednisolone to build a model of steroid-induced femoral head osteonecrosis. Contrast-enhanced ultrasound examinations were performed at 3 and 5 weeks after induction. Then, pathologic examinations and microvessel density (MVD) calculations were performed on the excised rabbit femoral heads. The MVD of the experimental group decreased significantly 3 and 5 weeks after induction compared with that of the control group. According to the CEUS examination results, significant differences existed in the ascending slope, descending slope, mean transit time, and time to peak between the groups at 5 weeks (P < .05). A correlation analysis showed that the descending slope had a certain correlation with the MVD (correlation coefficient, 0.376). A receiver operating characteristic curve was used to analyze the capacity of the CEUS parameters to predict the occurrence of osteonecrosis. The areas under the curve for the ascending slope and descending slope were 0.758 and 0.760, respectively (P < .05). Contrast-enhanced ultrasound can visualize the microcirculation in steroid-induced osteonecrosis of the femoral head in rabbits and may be a useful imaging method for the early monitoring and prediction of femoral head osteonecrosis.

Therapeutic effects of calcitonin gene-related peptide-modified bone marrow mesenchymal stem cells combined with autogenous bone grafting for treatment of osteonecrosis of the femoral head in rabbits

Calcitonin gene-related peptide (CGRP) promotes proliferation and osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs), which influence bone regeneration and reconstruction. We previously established CGRP-modified BMSCs that express the CGRP gene and demonstrated that this elevated CGRP expression promotes osteogenic differentiation of BMSCs. The current study investigated the therapeutic effects of CGRP-modified BMSCs combined with autogenous bone grafting for the treatment of osteonecrosis of the femoral head (ONFH) in rabbits. ONFH model rabbits were divided into three groups: CGRP, BMSCs/pLV5-CGRP combined with autogenous bone grafting; NC, BMSCs/pLV5-NC combined with autogenous bone grafting; and Blank, BMSCs combined with autogenous bone grafting. After autogenous bone grafting, mRNA and protein expression levels of osteogenesis-related genes (CGRP, OCN, Runx2, BMP-2, and OPG) were measured in the femoral head, as were ALP, laminin, and collagen type I contents. The fraction of trabecular bone area, percentage of empty osteocyte lacunae, maximum compressive strength, and mean elasticity modulus of the femoral head were also determined. Our results demonstrated that CGRP-modified BMSCs combined with autogenous bone grafting accelerated osteogenic differentiation of BMSCs, improved osteogenesis efficiency, and promoted the repair and reconstruction of bone tissue in the femoral head, which significantly improved the success rate of ONFH treatment.

Development of chitosan/gelatin hydrogels incorporation of biphasic calcium phosphate nanoparticles for bone tissue engineering

The chitosan/gelatin hydrogel incorporated with biphasic calcium phosphate nanoparticles (BCP-NPs) as scaffold (CGB) for bone tissue engineering was reported in this article. Such nanocomposite hydrogels were fabricated by using cycled freeze-thawing method, of which physicochemical and biological properties were regulated by adjusting the weight ratio of chitosan/gelatin/BCP-NPs. The needle-like BCP-NPs were dispersed into composites uniformly, and physically cross-linked with chitosan and gelatin, which were identified via Scanning Electron Microscope (SEM) images and Fourier Transform Infrared Spectroscopy (FT-IR) analysis. The porosity, equilibrium swelling ratio, and compressive strength of CGB scaffolds were mainly influenced by the BCP-NPs concentration. In vitro degradation analysis in simulated body fluids (SBF) displayed that CGB scaffolds were degraded up to at least 30 wt% in one month. Also, CCK-8 analysis confirmed that the prepared scaffolds had a good cytocompatibility through in culturing with bone marrow mesenchymal stem cells (BMSCs). Finally, In vivo animal experiments revealed that new bone tissue was observed inside the scaffolds, and gradually increased with increasing months, when implanted CGB scaffolds into large necrotic lesions of rabbit femoral head. The above results suggested that prepared CGB nanocomposites had the potential to be applied in bone tissue engineering.

5-Azacytidine and trichostatin A enhance the osteogenic differentiation of bone marrow mesenchymal stem cells isolated from steroid-induced avascular necrosis of the femoral head in rabbit

Bone marrow mesenchymal stem cells (BMSCs) play an important role in the process of bone repair. The present study investigated the effect of 5-azacytidine (AZA) and trichostatin A (TSA) on BMSC behaviors in vitro. The role of WNT family member 5A (WNT5A)/WNT family member 5A (WNT7A)/beta-catenin signaling was also investigated. BMSCs were isolated from a steroid-induced avascular necrosis of the femoral head (SANFH) rabbit model. The third-generation of BMSCs was used after identification. The results revealed obvious degeneration and necrosis in the SANFH rabbit model. AZA, TSA and TSA + AZA increased BMSC proliferation in a time-dependent fashion. AZA, TSA and TSA + AZA induced the cell cycle release from the G0/G1 phase and inhibited apoptosis in BMSCs. AZA, TSA and TSA + AZA treatment significantly decreased caspase-3 and caspase-9 activities. The treatment obviously increased the activity and relative mRNA expression of alkaline phosphatase. The treatment also significantly up-regulated the proteins associated with osteogenic differentiation, including osteocalcin and runt-related transcription factor 2 (RUNX2), and Wnt/beta-catenin signal transduction pathway-related proteins beta-catenin, WNT5A and WNT7A. The relative levels of Dickkopf-related protein 1 (an inhibitor of the canonical Wnt pathway) decreased remarkably. Notably, TSA + AZA treatment exhibited a stronger adjustment ability than either single treatment. Collectively, the present studies suggest that AZA, TSA and TSA + AZA promote cell proliferation and osteogenic differentiation in BMSCs, and these effects are potentially achieved via upregulation of WNT5A/WNT7A/b-catenin signaling.

Exploration of the Optimal Shape for Bone Tumour Biopsy.

Biopsy hole for bone tumour biopsy may cause pathological fractures. This study aimed to identify the optimal shape of bone tumour biopsy hole using the rabbit femoral head compression test to avoid pathological fracture. A compression test with no defect was performed to identify bone fracture location. Three shapes of biopsy holes (same size) were made artificially. Sixty rabbit femurs were randomly divided (n=15 each) into control (no defect), Shape 1 (round), Shape 2 (square), and Shape 3 (rectangular) groups. Twelve out of fifteen femurs were fractured on the femoral shaft; the femoral shaft was targeted for the compression test. Compressive maximum load and fracture energy were significantly higher for Shape 3 than for the other Shapes. A rectangular biopsy hole helps minimise reduction in bone strength. The defect width may be related to fragility of the affected bone.

Effects of transplantation of FGF-2-transfected MSCs and XACB on TNF-α expression with avascular necrosis of the femoral head in rabbits.

Objective: The present study aimed to investigate the effect of the transplantation of basic fibroblast growth factor (FGF-2) gene-transfected mesenchymal stem cells (MSCs) and xenogeneic antigen-cancellous bone (XACB) on tumor necrosis factor-α (TNF-α) expression with avascular necrosis of the femoral head (ANFH) in rabbits. Methods: The models of steroid-induced osteonecrosis in rabbits were randomly divided into five groups: A (model), B (XACB), C (XACB + MSCs), D (XACB + MSCs + LV), and E (XACB + MSCs + LV-FGF-2) groups. The therapeutic effect was evaluated by Hematoxylin and Eosin (H&E) staining. Immunohistochemical and RT-PCR assays were used to detect the protein and mRNA expression of TNF-α in the femoral head, respectively. Results: At 12 weeks after the operation, the defect in rabbits in group E was completely repaired, while defects in rabbits in the other groups were not completely repaired, and the area of new bone formation was higher, when compared with the other groups (P<0.05). Furthermore, the protein and mRNA expression TNF-α was lower at 3, 6, and 12 weeks after surgery, when compared with the other groups, and the difference was statistically significant (P<0.05). Conclusion: FGF-2/MSCs/XACB could promote the repair of ANFH, and may be correlated to the inhibition of TNF-α expression.

Composite Hydrogels with the Simultaneous Release of VEGF and MCP-1 for Enhancing Angiogenesis for Bone Tissue Engineering Applications

Rapid new microvascular network induction was critical for bone regeneration, which required the spatiotemporal delivery of growth factors and transplantation of endothelial cells. In this study, the linear poly(d,l-lactic-co-glycolic acid)-b-methoxy poly(ethylene glycol) (PLGA-mPEG) block copolymer microspheres were prepared for simultaneously delivering vascular endothelial growth factor (VEGF) and monocyte chemotactic protein-1 (MCP-1). Then, vascular endothelial cells (VECs) with growth factor loaded microspheres were composited into a star-shaped PLGA-mPEG block copolymer solution. After this, composite hydrogel (microspheres ratio: 5 wt%) was formed by increasing the temperature to 37 °C. The release profiles of VEGF and MCP-1 from composite hydrogels in 30 days were investigated to confirm the different simultaneous delivery systems. The VECs exhibited a good proliferation in the composite hydrogels, which proved that the composite hydrogels had a good cytocompatibility. Furthermore, in vivo animal experiments showed that the vessel density and the mean vessel diameters increased over weeks after the composite hydrogels were implanted into the necrosis site of the rabbit femoral head. The above results suggested that the VECs-laden hydrogel composited with the dual-growth factor simultaneous release system has the potential to enhance angiogenesis in bone tissue engineering.

Effects of Zoledronic Acid and Vitamin E on Surgical- Induced Osteonecrosis of the Femoral Head in Rabbit.

Femoral head osteonecrosis is a progressive disease with disabling outcomes in hip joint if not treated. This study was designed to compare the effects of zoledronic acid plus vitamin E versus zoledronic acid alone in surgical induced femoral head osteonecrosis in rabbits. 26 Japanese white adult normal male rabbits at 28-32 weeks old were undertaken surgical femoral dislocation to devastate the femoral neck vessels; the femoral neck vessels were ligated and the hip was relocated. Next, the first 10 rabbits received zoledronic acid injections at 1st and the 4th weeks; the second group (10 rabbits) received zoledronic acid injections at 1st and the 4th week along with daily oral vitamin E for 12 weeks; and the third group was considered as non-treated control group. Radiographic and postmortem pathological assessments including the Ficat classification, epiphyseal quotient (EQ), new bone formation, and residual necrotic bone (RNB) were performed and compared after week 12. A significant difference was found between the combination therapy group and the control group in Ficat classification at 12th weeks (P=0.048), but, the difference between monotherapy and combination therapy groups at 12th weeks was nonsignificant (P=0.37). Also, both treated groups had significant difference with the control group for RNB (P=0.015). There were no significant differences between the three groups for Ficat classification at the 6th week (P=0.65); EQ at 6th (P=0.59) and 12th week (P=0.64); and NBF (P=0.55). Although zoledronic acid therapy along with vitamin E could improve some radiologic and pathological indices related to femoral head osteonecrosis, vitamin E showed a relative impact. I.

Expression of osteoprotegerin and receptor activator for the nuclear factor-κB ligand in XACB/LV-bFGF/MSCs transplantation for repair of rabbit femoral head defect necrosis.

The aim of this study is to observe the changes in osteoprotegerin (OPG) and receptor activator for the nuclear factor-κB ligand (RANKL) in a rabbit model, and to explore the therapeutic effect of tissue engineering bone on femoral head necrosis. A total of 60 rabbits were randomly divided into 5 groups. The necrosis model of femoral head defects was created by dexamethasone combined with a horse serum injection. The model of femoral head necrosis was reconstructed by tissue engineering bone. The protein expressions of OPG and RANKL were detected by Western blot analysis. The expression of OPG and the RANKL protein in group E was higher than that in the other 4 groups (P < .05); there was no significant difference between groups C and D ( P > .05). The expression of OPG protein in the rabbit femoral head necrosis group was improved by xenogeneic antigens of cancellous bone/lentiviral-basic fibroblast growth factor/mesenchymal stem cells, which were expected to be used as an effective tissue engineering material to repair the necrosis of the femoral head.

Incorporation of chitosan-alginate complex into injectable calcium phosphate cement system as a bone graft material

Calcium phosphate brushite type of cements have been used to replace bone graft materials because of their biocompatibility and other attractive features. Especially, injectability of cement allows easy handling of minimally invasive surgical techniques. New calcium phosphate cement (CPC) system, brushite based cement incorporated into polyelectrolyte complex, was developed in this study. Chitosan-alginate complex produced by an interaction between a cationic polymer (chitosan) and an anionic polymer (alginate) was loaded in the cement. This improved the functional properties and biocompatibility of the final cement. We optimized the liquid/solid (L/S) ratio of the cement components and investigated the compressive strength, setting time, pH change of CPC0 (with only citric acid) and CPC0.5, 1, and 1.5 (0.5, 1, and 1.5 v/v % chitosan-alginate complex in citric acid solution, respectively). The L/S ratio did not affect structural formation, while the addition of polymer complex showed new formation of macro-pores within CPC. However, a lower L/S ratio and higher amount of added polymer complex shortened the setting time and improved the compressive strength. The appropriate conditions for the injectable bone substitute were CPC1 with an L/S ratio of 0.45. To investigate the effect of the chitosan-alginate complex on CPC system in physiological conditions, CPC0 and CPC1 were implanted in a rabbit femoral head defect model for 1 and 3 months. Micro-computed tomography revealed improved bone formation in CPC1 compared to CPC0 3 months after implantation. Histological analysis revealed newly formed bone tissues around the peripheral sides of CPC0 and CPC1. The results indicate the potential value of the CPC system containing polymer complex as an injectable bone substitute. The study of the CPC-polymer complex system incorporating drugs or cells can be further developed into a controlled release system for faster bone regeneration.

Role of FGF-2 Transfected Bone Marrow Mesenchymal Stem Cells in Engineered Bone Tissue for Repair of Avascular Necrosis of Femoral Head in Rabbits

Background/Aims: Avascular necrosis of the femoral head (ANFH) is the focus and difficulty of orthopedic diseases. Recently, tissue engineering bone for this disease has shown a good therapeutic effect. The aim of the present study was to investigate the therapeutic effect of basic fibroblast growth factor (FGF-2) as cytokines transfected bone marrow mesenchymal stem cells (BMSCs) in constructing tissue-engineered bone for avascular necrosis of the femoral head. Methods: The FGF-2 gene overexpressed lentivirus-transfected rBMSCs with xenogeneic antigen-extracted cancellous bone (XACB) to construct tissue engineered bone, and the model of early avascular necrosis of the femoral head was established by lipopolysaccharide (LPS) combined with hormone. The models were randomly divided into five groups: A (control), B (XACB), C (XACB+rBMSCs), D (XACB+rBMSCs+Lv-GFP), and E (XACB+rBMSCs+Lv-FGF-2/GFP) groups. The therapeutic effect of the tissue engineered bone for the avascular necrosis of the femoral head was evaluated by gross anatomy, X-ray examination, immunohistochemistry and H&E staining. Results: The FGF-2 gene was transfected into rBMSCs (Multiplicity of infection [MOI] = 100) by lentivirus, and its efficiency was above 95%. The rBMSCs were successfully transfected overexpressing FGF-2 by qPCR and western blot. After tissue engineering bone implantation, X-ray examination and gross specimen observation revealed that the repair area in the E group was > 80% at six weeks, the defect was completely repaired at 12 weeks, and osteogenesis was stronger, when compared with the other groups. For the X-ray score, vascular area density and new bone formation area were higher, when compared with the other groups, and the difference was statistically significant (P< 0.05). Conclusion: FGF-2 gene overexpression lentivirus transfection BMSCs combined with XACB to construct tissue engineered bone can effectively promote vascular regeneration, and improve the repair effect of avascular necrosis of the femoral head.

Calcitonin gene-related peptide-transfected bone marrow mesenchymal stem cells combined with autologous bone grafting to treat osteonecrosis of the femoral head in rabbit model

Objective To investigate the histopathological changes of calcitonin peptide-transfected bone marrow mesenchymal stem cells (BMSCs) combined with bone transplantation in the treatment of osteonecrosis of the femoral head (ONFH) in rabbit model. Methods The osteonecrosis models on one side femoral head of rabbits were established by liquid nitrogen. Forty-eight rabbit ONFH models were randomly divided into 3 groups. In human calcitonin gene-related peptide α (hCGRPα)-transfected BMSCs group (A), the BMSCs transfected with hCGRPα in combination with autologous bone graft were implanted into the femoral head. In empty plasmid group (B), the BMSCs transfected with empty plasmid (only pLNCX2 plasmid) and autologous bone graft were implanted. In irrelated sequence group (C), BMSCs transfected with irrelated gene [only pLNCX2-green fluorescent protein (GFP)] and autologous bone graft were implanted. Animals were sacrificed at 3rd and 6th month respectively to observe the histopathological changes in the femoral heads of rabbits. Results At 3rd month, the bone trabecular area fraction respectively was (40.89±1.12)%, (36.12±1.17)%, (36.33±1.53)% and (42.11±1.91)% in groups A, B and C and normal femoral head. The percentage of empty osteocyte lacunae respectively was (11.34±1.58)%, (16.33±1.45)%, (17.21±1.23)% and (10.47±1.41)% in groups A, B and C and normal femoral head. In group A, there were proliferation and growth of a large number of osteocytes, more mature bone trabeculae and higher area fraction, less empty osteocyte lacunae, and stronger biomechanical strength in the femoral head, which were nearly close to those in the normal control group and obviously superior to those in groups B and C. In groups B and C, there were thin and irregular bone trabeculae, more empty osteocyte lacunae, lower area fraction and imperfect bone repair. At 6th month, the bone trabeculae were mature, densification and satiation, and neatly arranged rules were seen in the femoral head of group A. The area fraction of bone trabeculae and the number of empty bone lacunae in group A were superior to those in groups B and C. Conclusion The implantation of hCGRPα-transfected BMSCs combined with autologous bone graft into the femoral head of ONFH animal models can significantly promote cells proliferation, accelerate osteogenesis, enhance bone tissue repair, and obtain good effect in the treatment of ONFH. Key words: Calcitonin gene related peptide; Bone marrow mesenchymal stem cells; Osteonecrosis of the femoral head necrosis; Model, animal

Exosomes secreted from mutant-HIF-1α-modified bone-marrow-derived mesenchymal stem cells attenuate early steroid-induced avascular necrosis of femoral head in rabbit.

Mesenchymal stem cells (MSCs)-derived exosomes exhibit protective effects on damaged or diseased tissues. Hypoxia-inducible factor 1α (HIF-1α) plays a critical role in bone development. However, HIF-1α is easily biodegradable under normoxic conditions. The bone-marrow-derived mesenchymal stem cells (BMSCs) were transfected with adenovirus carrying triple point-mutations (amino acids 402, 564, and 803) in the HIF-1α coding sequence (CDS). The mutant HIF-1α can efficiently express functional proteins under normoxic conditions. To date, no study has reported the role of exosomes secreted by mutant HIF-1α modified BMSCs in the recovery of the early steroid-induced avascular necrosis of femoral head (SANFH). In this study, we firstly analyzed exosomes derived from BMSCs modified by mutant (BMSC-ExosMU ) or wild-type HIF-1α (BMSC-ExosWT ). In vitro, we investigated the osteogenic differentiation capacity of BMSCs modified by BMSC-ExosMU or BMSC-ExosWT , and the angiogenesis effects of BMSC-ExosMU and BMSC-ExosWT on human umbilical vein endothelial cells (HUVECs). Besides, the healing of the femoral head was also assessed in vivo. We found that the potential of osteogenic differentiation of BMSCs treated with BMSC-ExosMU was higher than the wild-type group in vitro. In addition, BMSC-ExosMU stimulated the proliferation, migration, and tube formation of HUVECs in a dose-dependent manner. Compared with the BMSC-ExosWT or PBS control group, the injection of BMSC-ExosMU into the necrosis region markedly accelerated the bone regeneration and angiogenesis, which were indicated by the increased trabecular reconstruction and microvascular density. Taken together, our data suggest that BMSC-ExosMU facilitates the repair of SANFH by enhancing osteogenesis and angiogenesis.

Sildenafil improves blood perfusion in steroid-induced avascular necrosis of femoral head in rabbits via a protein kinase G-dependent mechanism

ObjectiveThe aim of the study were to evaluate the effect of sildenafil against avascular necrosis of femoral head (ANFH) in a rabbit model, and to study the role of protein kinase G (PKG) pathway and vascular endothelial growth factor (VEGF) in ANFH. MethodsThree weeks after inducing ANFH with methylprednisolone injection, 45 female adult New Zealand white rabbits were divided into three groups and treated as follows: group SI received daily intraperitoneal sildenafil with a dose of 10 mg/kg per day; group SD received daily sildenafil identically to group SI plus auricular vein injection DT3 (a specific PKG inhibitor); group NS received only normal saline. The blood perfusion function in the femoral head was measured by perfusion MRI and ink artery infusion. Bilateral femora heads were examined histopathologically for the presence of osteonecrosis; VEGF of tissue was examined by Western blot analysis; cGMP level and PKG activity were also measured. ResultsThe incidence of ANFH in SI group was significantly lower than that observed in NS and SD groups (p < 0.05). VEGF in SI group was increased compared to NS group. cGMP level and PKG activity were also significantly different between NS and SI group (p < 0.05). However, these effects of sildenafil in SD group were all markedly inhibited by the administration of DT3 compared to SI group. ConclusionSildenafil appear to increase the perfusion of femoral head by up-regulating VEGF through PKG pathway. The increased perfusion of femoral head could prevent ANFH.

A histomorphometric study of necrotic femoral head in rabbits treated with extracorporeal shock waves.

[Purpose] This study aimed to determine the effectiveness and mechanisms of extracorporeal shock wave therapy in the treatment of femoral head osteonecrosis. [Subjects and Methods] Histomorphometric analysis of necrotic femoral head in rabbits treated with shock waves was performed. Bilateral osteonecrosis of femoral heads was induced with methylprednisolone and lipopolysaccharide in eight rabbits. The left limb (study side) received shock waves to the femoral head. The right limb (control side) received no shock waves. Biopsies of the femoral heads were performed at 12 weeks after shock wave therapy. [Results] Necrotic femoral heads treated with shock waves, compared with controls, had higher bone volume per tissue volume, trabecular thickness, trabecular number, osteoblast surface/bone surface, osteoid surface/bone surface, osteoid thickness, mineralizing surface/bone surface, mineralizing apposition rate, and bone formation rate. However, trabecular separation was lower in shock wave-treated femoral heads than in controls. Eroded surface/bone surface and osteoclast surface/bone surface did not differ significantly between groups. [Conclusion] The bone mass of necrotic femoral heads treated with shock waves increases. Extracorporeal shock wave may promote bone repair in necrotic femoral heads through the proliferation and activation of osteoblasts.

Combined Treatment with an Anticoagulant and a Vasodilator Prevents Steroid-Associated Osteonecrosis of Rabbit Femoral Heads by Improving Hypercoagulability.

Steroid-associated osteonecrosis of the femoral head remains a challenging problem in orthopedics worldwide. One pathomechanism is ischemia of the femoral head, as a result of thrombus formation and vasoconstriction. The present study investigates the effects of combination prevention with enoxaparin and EGb 761 on steroid-associated ONFH in rabbits. Rabbits were randomly divided into 5 groups (control group, model group, enoxaparin group, ginkgo group, and combination group). With the exception of the control group, the groups of rabbits were modeled with lipopolysaccharide and methylprednisolone acetate. Starting with modeling, the enoxaparin group and ginkgo group were injected with 1 μg/kg/day enoxaparin subcutaneously and orally given 40 mg/kg/day EGb 761 for 4 weeks, respectively; the combination group received both treatments. After modeling for 6 weeks, the hematology data indicated prolonged PT and APTT in the three prevention groups. The micro-CT examination revealed higher bone density and better structure; histomorphometry revealed significant pathological changes. Immunohistochemistry revealed higher expression of BMP-2 and VEGF, thus revealing better osteogenesis and angiogenesis activities. Among the three prevention groups, the combination group had the most efficient results. In conclusion, the combined prevention with an anticoagulant and a vasodilator has the potential to decrease the incidence of steroid-associated ONFH in rabbits.

Biomechanical changes in the osteonecrosis of the femoral head of rabbits treated by neuropeptide gene-transfected stem cells combined with autologous bone grafting

Objective To investigate the biomechanical changes in the osteonecrosis of the femoral head (ONFH) of rabbits treated by neuropeptide gene (calcitonin gene-related peptide α, hCGRPα)-transfected stem cells combined with autologous bone grafting. Methods The ONFH models on one side in rabbits were established by liquid nitrogen. Forty-eight rabbits ONFH models were randomly divided into 3 groups: group A [hCGRPα-transfected bone marrow mesenchymal stem cell (BMSCs) group], the hCGRPα was transfected into BMSCs combined with autologous bone graft into the femoral head; group B (BMSCs group), BMSCs were transfected with empty plasmid (only pLNCX2 plasmid) combined with autologous bone graft; group C (irrelated sequence group), BMSCs were transfected with irrelated gene combined with autologous bone graft. Animals were sacrificed at 3rd and 6th month respectively to observe the biomechanical changes in the femoral head of rabbits. Results At 3rd month after surgical treatment, in groups A, B, and C the maximum compressive strength and mean elastic modulus of the subchondral bone of femoral head on surgically treated side respectively were (29.13±1.95) and (59.61±2.18), (24.63±1.35) and (48.87±2.15), (25.05±1.34) and (50.13±2.28) mPa, and those of the cancellous bone of femoral head on surgically treated side respectively were (16.41±1.13) and (24.11±2.32), (11.43±1.55) and (17.63±2.45), (12.51±1.85) and (18.22±2.32) mPa. The biomechanical strength was the best in the femoral head in group A, which was nearly close to that in the normal side (P>0.05) and was significantly greater than that in both group B and group C (P 0.05). Conclusion hCGRPα-transfected BMSCs combined with autologous bone grafting into the femoral head of ONFH animal model can accelerate osteogenesis, enhance bone tissue repair, and then effectively recover the normal biomechanical properties of the femoral head. Key words: Calcitonin gene-related peptide; Bone marrow mesenchymal stem cells; Bone grafting; Osteonecrosis of the femoral head; Biomechanics; Rabbit

Local Administration of Bisphosphonate-soaked Hydroxyapatite for the Treatment of Osteonecrosis of the Femoral Head in Rabbit.

Background:Systemic administration of bisphosphonates has shown promising results in the treatment of osteonecrosis of the femoral head (ONFH). However, few studies have evaluated the efficacy of local zoledronate (ZOL) administration in the treatment of ONFH. The purpose of this study was to investigate whether local administration of bisphosphonate-soaked hydroxyapatite (HA) could improve bone healing in an experimental rabbit model of ONFH.Methods:This experimental study was conducted between October 2014 and June 2015. Forty-five rabbits underwent simulated ONFH surgery. Immediately following surgery, they were divided into three groups: model (untreated, n = 15), HA (treated with HA alone, n = 15), and HA + ZOL (treated with HA soaked in a low-dose ZOL solution, n = 15). Histological, immunohistochemical, and quantitative analyses were performed to evaluate bone formation and resorption 2, 4, and 8 weeks after surgery.Results:Gross bone matrix and hematopoietic tissue formation were observed in the HA + ZOL group 4 weeks after surgery. The immunohistochemical staining intensities for 5-bromodeoxyuridine, runt-related transcription factor 2, osteocalcin, osteopontin, and osteoprotegerin were significantly higher in the HA + ZOL group than that in the model (P < 0.001, P < 0.001, P < 0.001, P < 0.001, and P = 0.018, respectively) and HA groups (P = 0.003, P = 0.049, P < 0.001, P = 0.020, and P = 0.019, respectively), whereas receptor activator of the nuclear factor-κB ligand staining intensity was significantly lower in the HA + ZOL group than that in the model and HA groups (P = 0.029 and P = 0.015, respectively) 4 weeks after surgery. No significant differences in bone formation or bone resorption marker expression were found between the three groups 2 or 8 weeks after surgery (P > 0.05).Conclusions:Local administration of HA soaked in a low-dose ZOL solution increased new bone formation while inhibiting bone resorption in an animal model of ONFH, which might provide new evidence for joint-preserving surgery in the treatment of ONFH.

Therapeutic effect of osteogenically induced adipose derived stem cells on vascular deprivation-induced osteonecrosis of the femoral head in rabbits

ObjectiveTo explore the therapeutic effect of osteogenically induced adipose-derived stem cells (ADSCs) on vascular deprivation-induced osteonecrosis of the femoral head (ONFH) in rabbit model. MethodsVascular deprivation-induced ONFH was established by intramuscular injection of methylpre-dnisolone, and vascular occlusion of the capital femoral epiphysis by electrocoagulation in adult New Zealand white rabbits. Eight weeks after the establishment of vascular deprivation-induced ONFH, animals were randomly divided into three equal groups. In Group A (control), no therapy was given. In Group B, core decompression was performed by drilling a hole (1.2 mm in diameter) from the outer cortex 2.5 cm distal to the proximal end of the greater trochanter. In Group C, 1> <107 osteogenically induced ADSCs were resus-pended in 0.5 ml PBS, and then injected directly into the femoral head. Femoral head specimens were obtained at postoperative 8 weeks. The bone formation and three-dimensional microstructure of the femoral head was evaluated by micro-computed tomography scans (u-CT). Immu-nohistochemical analysis was performed to detect the expression of osteocalcin. Angiogenesis and repair of the femoral head were observed histologically. ResultsIn trabecular bone at the proximal femur region, the trabecular volume was higher in Group C (130.70 mm3± 4.33 mm3) than that in Groups A (101.07 mm3±7.76 mm3) and B (107.89 mm3±8.68 mm3, P<0.01). Bone volume was significantly increased in Group C (40.09 mm3±6.35 mm3) than in Groups A (29.65 mm3±4.61 mm3) and B (31.80 mm3± 4.01 mm3, P<0.01). The trabecular number was higher in Groups C (1.58±0.25) than other two groups (1.15±0.18, 1.16± 0.21, P<0.01). Bone mineral density showed statistically significant difference between Groups C and A or B (375.38± 23.06) mg HA/ccm, vs (313.73 ±19.30) mg HA/ccm and (316.09± 16.45) mg HA/ccm, P<0.01). Histological examination indicated that there was more new bone formation in Group C than in other groups. ConclusionTreatment with autologous osteogenically induced ADSCs transplantation results in an enhanced osteogenesis and microstructure of the vascular deprivation-induced osteonecrosis in rabbits.

Tao-Hong-Si-Wu Decoction ameliorates steroid-induced avascular necrosis of the femoral head by regulating the HIF-1α pathway and cell apoptosis.

The aim of this study was to corroborate the hypothesis that Tao-Hong-Si-Wu Decoction (THSWD) affects steroid-induced avascular necrosis of the femoral head (SANFH) by regulating the hypoxia-inducible factor 1α (HIF-1α) pathway. Forty-eight New Zealand rabbits were randomly divided into a normal control group (NC group), a model group (SANFH group), a THSWD group, and a dimethyloxalylglycine group (DMOG group). Rabbits in the SANFH group were injected with both horse serum and methylprednisolone. Rabbits in the THSWD group were gavaged with THSWD in addition to receiving the same treatment as the SANFH group. Rabbits in the DMOG group were injected with extra DMOG in conjunction with the same treatment as the SANFH group. Rabbits in the NC group received the same amount of normal saline. Eight weeks after steroid treatment, the femoral heads of rabbits were removed to examine HIF-1α, vascular endothelial growth factor (VEGF), caspase-3, and bcl-2. Results indicated that THSWD significantly promoted the expression of HIF-1α and VEGF in the femoral head tissue of rabbits and markedly inhibit the apoptosis of osteocytes, chondrocytes, and bone marrow cells. In addition, THSWD suppressed caspase-3 expression and induced bcl-2 expression in femoral head tissues. In conclusion, THSWD can suppress SANFH by regulating the HIF-1α pathway and cell apoptosis.