Aims. To evaluate the effect of ultrasound-targeted simvastatin-loaded microbubble destruction (UTMDSV) for alleviation of the progression of osteoarthritis (OA) in rabbits through modulation of the peroxisome proliferator-activated receptor (PPARγ). Methods. In vitro, OA chondrocytes were treated with ultrasound (US), US-targeted microbubble destruction (UTMD), simvastatin (SV), and UTMDSV on alternate days for four weeks. Chondrocytes were also treated with PPARγ inhibitor, PPARγ inhibitor+ UTMDSV, and UTMDSV. The cholesterol efflux rate and triglyceride levels were measured using an assay kit and oil red O staining, respectively. In vivo, the OA rabbits were treated with a single intra-articular injection of UTMD, SV, and UTMDSV every seven days for four weeks. Cartilage histopathology was assessed by safranin-O staining and the Mankin score. Total cholesterol (TC) and high-density lipoprotein-cholesterol (HDL-C) in rabbit knee synovial fluid were detected by enzyme-marker assay. Aggrecan, collagen II, and PPARγ expression levels were analyzed by Western blotting (WB). Results. In vitro, UTMDSV significantly increased the cholesterol efflux rate and aggrecan, collagen II, and PPARγ levels in OA chondrocytes; these effects were blocked by the PPARγ inhibitor. In vivo, UTMD. SV. significantly increased aggrecan, collagen II, PPARγ, and HDL-C levels, while TC levels and Mankin scores were decreased compared with the UTMD, SV, OA, and control groups. Conclusion. UTMDSV promotes cartilage extracellular matrix synthesis by modulating the PPARγ-mediated cholesterol efflux pathway in OA rabbits. Cite this article: Bone Joint Res 2021;10(10):693–703

Aims. The purpose of this study was to explore a simple and effective method of preparing human acellular amniotic membrane (HAAM) scaffolds, and explore the effect of HAAM scaffolds with juvenile cartilage fragments (JCFs) on osteochondral defects. Methods. HAAM scaffolds were constructed via trypsinization from fresh human amniotic membrane (HAM). The characteristics of the HAAM scaffolds were evaluated by haematoxylin and eosin (H&E) staining, picrosirius red staining, type II collagen immunostaining, Fourier transform infrared spectroscopy (FTIR), and scanning electron microscopy (SEM). Human amniotic mesenchymal stem cells (hAMSCs) were isolated, and stemness was verified by multilineage differentiation. Then, third-generation (P3) hAMSCs were seeded on the HAAM scaffolds, and phalloidin staining and SEM were used to detect the growth of hAMSCs on the HAAM scaffolds. Osteochondral defects (diameter: 3.5 mm; depth: 3 mm) were created in the right patellar grooves of 20 New Zealand White rabbits. The rabbits were randomly divided into four groups: the control group (n = 5), the HAAM scaffolds group (n = 5), the JCFs group (n = 5), and the HAAM + JCFs group (n = 5). Macroscopic and histological assessments of the regenerated tissue were evaluated to validate the treatment results at 12 weeks. Results. In vitro, the HAAM scaffolds had a network structure and possessed abundant collagen. The HAAM scaffolds had good cytocompatibility, and hAMSCs grew well on the HAAM scaffolds. In vivo, the macroscopic scores of the HAAM + JCFs group were significantly higher than those of the other groups. In addition, histological assessments demonstrated that large amounts of hyaline-like cartilage formed in the osteochondral defects in the HAAM + JCFs group. Integration with surrounding normal cartilage and regeneration of subchondral bone in the HAAM + JCFs group were better than those in the other groups. Conclusion. HAAM scaffolds combined with JCFs promote the regenerative repair of osteochondral defects. Cite this article: Bone Joint Res 2022;11(6):349–361

Aims. The aim of our study is to investigate the effect induced by alternated mechanical loading on Notch-1 in mandibular condylar cartilage (MCC) of growing rabbits. Methods. A total of 64 ten-day-old rabbits were randomly divided into two groups according to dietary hardness: normal diet group (pellet) and soft diet group (powder). In each group, the rabbits were further divided into four subgroups by feeding time: two weeks, four weeks, six weeks, and eight weeks. Animals would be injected 5-bromo-2′-deoxyuridine (BrdU) every day for one week before sacrificing. Histomorphometric analysis of MCC thickness was performed through haematoxylin and eosin (HE) staining. Immunochemical analysis was done to test BrdU and Notch-1. The quantitative real-time polymerase chain reaction (qRT-PCR) and western blot were used to measure expression of Notch-1, Jagged-1, and Delta-like 1 (Dll-1). Results. The thickness of MCC in the soft diet group was thinner than the one in normal diet group. Notch-1 was restricted in fibrous layer, proliferative layer, and hypertrophic layer. The expression of Notch-1 increased from two weeks to six weeks and then fell down. Notch-1 in normal diet group was higher than that in soft diet group in anterior part of MCC. The statistical differences of Notch-1 were shown at two, four, and six weeks (p < 0.05). The result of western blot and quantitative real-time PCR (qRT-PCR) showed the expression of Dll-1 and Jagged-1 rose from two to four weeks and started to decrease at four weeks. BrdU distributed in all layers of cartilage and subchondral bone. The number of BrdU-positive cells, which were less in soft diet group, was decreasing along with the experiment period. The significant difference was found at four, six, and eight weeks in anterior and posterior parts (p < 0.05). Conclusion. The structure and proliferation of MCC in rabbits were sensitive to dietary loading changes. The proper mechanical loading was essential for transduction of Notch signalling pathway and development of mandibular condylar cartilage. Cite this article: Bone Joint Res 2021;10(7):437–444

Aims. Several artificial bone grafts have been developed but fail to achieve anticipated osteogenesis due to their insufficient neovascularization capacity and periosteum support. This study aimed to develop a vascularized bone-periosteum construct (VBPC) to provide better angiogenesis and osteogenesis for bone regeneration. Methods. A total of 24 male New Zealand white rabbits were divided into four groups according to the experimental materials. Allogenic adipose-derived mesenchymal stem cells (AMSCs) were cultured and seeded evenly in the collagen/chitosan sheet to form cell sheet as periosteum. Simultaneously, allogenic AMSCs were seeded onto alginate beads and were cultured to differentiate to endothelial-like cells to form vascularized bone construct (VBC). The cell sheet was wrapped onto VBC to create a vascularized bone-periosteum construct (VBPC). Four different experimental materials – acellular construct, VBC, non-vascularized bone-periosteum construct, and VBPC – were then implanted in bilateral L4-L5 intertransverse space. At 12 weeks post-surgery, the bone-forming capacities were determined by CT, biomechanical testing, histology, and immunohistochemistry staining analyses. Results. At 12 weeks, the VBPC group significantly increased new bone formation volume compared with the other groups. Biomechanical testing demonstrated higher torque strength in the VBPC group. Notably, the haematoxylin and eosin, Masson’s trichrome, and immunohistochemistry-stained histological results revealed that VBPC promoted neovascularization and new bone formation in the spine fusion areas. Conclusion. The tissue-engineered VBPC showed great capability in promoting angiogenesis and osteogenesis in vivo. It may provide a novel approach to create a superior blood supply and nutritional environment to overcome the deficits of current artificial bone graft substitutes. Cite this article: Bone Joint Res 2023;12(12):722–733

Objectives. The injured anterior cruciate ligament (ACL) is thought to exhibit an impaired healing response, and attempts at surgical repair have not been successful. Connective tissue growth factor (CTGF) is reported to be associated with wound healing, probably through transforming growth factor beta 1 (TGF-β1). Methods. A rabbit ACL injury model was used to study the effect of CTGF on ligament recovery. Quantitative real-time PCR (qRT-PCR) was performed for detection of changes in RNA levels of TGF-β1, type 1 collagen (COL1), type 2 collagen (COL2), SRY-related high mobility group-box gene9 (SOX9), tissue inhibitor of metalloproteinase-1 (TIMP-1) and matrix metallopeptidase 13 (MMP-13). Expression of related proteins was detected by Western blotting. Results. The current study showed that CTGF could promote the recovery of an injured anterior cruciate ligament. It can upregulate mRNA and expression of TGF-β1, COL1, COL2, SOX9, and tissue inhibitor of TIMP-1, and downregulate mRNA and expression of MMP-13, suggesting that the curative effect of CTGF on injured rabbit ligaments is through regulation of these cellular factors. Conclusions. This finding revealed the healing role of CTGF in injured tissues and provides new possibilities of treating injured tissues and wound healing by using CTGF. Cite this article: X. Sun, W. Liu, G. Cheng, X. Qu, H. Bi, Z. Cao, Q. Yu. The influence of connective tissue growth factor on rabbit ligament injury repair. Bone Joint Res 2017;6:399–404. DOI: 10.1302/2046-3758.67.BJR.2016-0255.R1

Objectives. Recent studies have shown that systemic injection of rapamycin can prevent the development of osteoarthritis (OA)-like changes in human chondrocytes and reduce the severity of experimental OA. However, the systemic injection of rapamycin leads to many side effects. The purpose of this study was to determine the effects of intra-articular injection of Torin 1, which as a specific inhibitor of mTOR which can cause induction of autophagy, is similar to rapamycin, on articular cartilage degeneration in a rabbit osteoarthritis model and to investigate the mechanism of Torin 1’s effects on experimental OA. Methods. Collagenase (type II) was injected twice into both knees of three-month-old rabbits to induce OA, combined with two intra–articular injections of Torin 1 (400 nM). Degeneration of articular cartilage was evaluated by histology using the Mankin scoring system at eight weeks after injection. Chondrocyte degeneration and autophagosomes were observed by transmission electron microscopy. Matrix metallopeptidase-13 (MMP-13) and vascular endothelial growth factor (VEGF) expression were analysed by quantitative RT-PCR (qPCR).Beclin-1 and light chain 3 (LC3) expression were examined by Western blotting. Results. Intra-articular injection of Torin 1 significantly reduced degeneration of the articular cartilage after induction of OA. Autophagosomes andBeclin-1 and LC3 expression were increased in the chondrocytes from Torin 1-treated rabbits. Torin 1 treatment also reduced MMP-13 and VEGF expression at eight weeks after collagenase injection. Conclusion. Our results demonstrate that intra-articular injection of Torin 1 reduces degeneration of articular cartilage in collagenase-induced OA, at least partially by autophagy activation, suggesting a novel therapeutic approach for preventing cartilage degeneration and treating OA. Cite this article: N-T. Cheng, A. Guo, Y-P. Cui. Intra-articular injection of Torin 1 reduces degeneration of articular cartilage in a rabbit osteoarthritis model. Bone Joint Res 2016;5:218–224. DOI: 10.1302/2046-3758.56.BJR-2015-0001

Objectives. The purpose of this study was to evaluate chronological changes in the collagen-type composition at tendon–bone interface during tendon–bone healing and to clarify the continuity between Sharpey-like fibres and inner fibres of the tendon. Methods. Male white rabbits were used to create an extra-articular bone–tendon graft model by grafting the extensor digitorum longus into a bone tunnel. Three rabbits were killed at two, four, eight, 12 and 26 weeks post-operatively. Elastica van Gieson staining was used to colour 5 µm coronal sections, which were examined under optical and polarised light microscopy. Immunostaining for type I, II and III collagen was also performed. Results. Sharpey-like fibres comprised of type III collagen in the early phase were gradually replaced by type I collagen from 12 weeks onwards, until continuity between the Sharpey-like fibres and inner fibres of the tendon was achieved by 26 weeks. Conclusions. Even in rabbits, which heal faster than humans, an observation period of at least 12 to 26 weeks is required, because the collagen-type composition of the Sharpey-like fibre bone–tendon connection may have insufficient pullout strength during this period. These results suggest that caution is necessary when permitting post-operative activity in humans who have undergone intra-bone tunnel grafts

Lubricin is a proteoglycan that is a boundary lubricant in synovial joints and both a surface and collagen inter-fascicular lubricant in ligaments. The purpose of this study was to characterise the mRNA levels for lubricin in the anterior cruciate ligament (ACL), posterior cruciate ligament (PCL), medial collateral ligament (MCL), and lateral collateral ligament (LCL) in aging and surgically-induced menopausal rabbits. We hypothesised that lubricin mRNA levels would be increased in ligaments from aging and menopausal rabbits compared with ligaments from normal rabbits. All four knee ligaments (ACL, PCL, MCL, LCL) were isolated from normal (1-year-old rabbits, n=8), aging (3-year-old rabbits, n=6), and menopausal (1-year-old rabbits fourteen weeks after surgical ovariohysterectomy, n=8) female New Zealand White rabbits. RT-qPCR was used to evaluate the mRNA levels for lubricin normalised to the housekeeping gene 18S. After removing outliers, data for normal, aging, and menopausal rabbits for each knee ligament (ACL, PCL, MCL, LCL) were compared using ANOVA with linear contrasts or Kruskal-Wallis test with Conover post-hoc analysis. For ACLs, the mRNA levels for lubricin were increased in menopausal and aging rabbits compared with normal rabbits (p<0.056). For PCLs, trends for increased lubricin mRNA levels were found when comparing menopausal and aging rabbits with normal rabbits (p<0.092). For MCLs, the mRNA levels for lubricin were increased in menopausal and aging rabbits compared with normal rabbits (p<0.050). For LCLs, no differences in lubricin mRNA levels were detected comparing the three groups. For all four knee ligaments (ACL, PCL, MCL, LCL), no differences in lubricin mRNA levels were detected comparing the ligaments from menopausal rabbits with those from aging rabbits. Lubricin plays a role in collagen fascicle lubrication in ligaments (1,2). Increased lubricin gene expression was associated with mechanical changes (including decreased modulus and increased failure strain) in the aging rabbit MCL (3). Detection of similar molecular changes in the ACL, and possibly the PCL, may indicate that their mechanical properties may also change as a result of increased lubricin gene expression, thereby potentially pre-disposing these ligaments to damage accumulation. Compared to aging ligaments, aging tendons exhibited decreased lubricin gene and protein expression, and increased stiffness (4). Although opposite changes than aging ligaments, these findings support the relationship between lubricin and modulus/stiffness. The similarities between ligaments in the aging and menopausal groups may suggest that surgically-induced menopause results in a form of accelerated aging in the rabbit ACL, MCL and possibly PCL

Introduction. The rabbit common calcanean (Achilles) tendon is a compound apparatus frequently used in studies considering novel interventions to facilitate tendon regeneration. These studies often employ complete surgical transection of the apparatus. Due consideration of the translational relevance to human tendinopathy is often lacking and refinement of this injury model, consistent with the principles of the 3Rs, has not been forthcoming. Materials and Methods. Wild rabbit cadavers (n=10) were obtained from a licensed game dealer. For gross anatomy studies the caudal crus was dissected and transverse sections obtained every 5 mm. Ultrasongraphic examination of the entire apparatus was peformed with a 15 Hz transducer in transverse sections. Results. This study reannotates the apparatus and demonstrates that the principal structures, the superficial digital flexor tendon and medial and lateral gastrocnemius tendons, may be clearly identified by ultrasonographic examination. Discussion. Historical descriptions of the rabbit Achilles apparatus are shown to be inaccurate and follow human gross anatomical descriptions. Ultrasonographic identification of the constituent structures in the rabbit are poorly represented in the literature. Reference measurements and qualitative descriptions are provided that may facilitate the development of refined surgical techniques for in vivo studies of tendon regeneration in the rabbit beyond crude transection studies

Meniscus has many important functions in the knee joint such as load bearing, shock absorption, joint stability, joint lubrication and proprioception. In the recent years, meniscus injuries have been the focus of orthopaedic surgeons and musculoskeletal tissue engineering applications because of its avascular nature. In this study, we aimed to compare the regeneration capacities of two composite scaffolds in a New Zealand Rabbit meniscal defect model. The first scaffold consists Poly-Lactic Acid (PLA) + chitosan + loofah and the second PLA + Hydroxyapatite (HAp) + loofah. In order to produce these scaffolds; 4% chitosan, 4% PLA and 4% HAp solutions were seperately prepared. The loofah pieces were saturated with these solutions and vacuum-dried for 14 days and sterilized with ethylene oxide. There were several characterizations performed such as Fourier Transform Infrared Spectroscopy (FTIR) for the investigation of chemical structure, Scanning Electron Microscopy (SEM) for morphological analysis, thermogravimetric differential thermal analysis (TGA/DTA) for thermal properties, mechanical compression and swelling ratio analysis. Moreover, in order to investigate biocompatibility of the scaffolds, WST-1 colorimetric assay at days 3, 7, 10, 14 and 21 was conducted. After these biocompatibility analysis, a 1.5-mm cylindrical defect was created in the avascular portion of the anterior horn of the medial meniscus in 14 New Zealand rabbits (2.5–3 kg weight) which were randomly grouped in two. The scaffolds were implanted at the defect site with the help of a freshly prepared fibrin glue. 8 weeks after the operation, the rabbits were sacrificed and their tissues were kept for further mechanical, radiological and histological analysis. In conclusion, we succeeded to produce a new meniscus scaffold. The proliferation ability of PLA + chitosan + loofah scaffold is higher than PLA + HAp + loofah scaffold. However, there was no statistically significant difference among them

Objectives. Sustained intra-articular delivery of pharmacological agents is an attractive modality but requires use of a safe carrier that would not induce cartilage damage or fibrosis. Collagen scaffolds are widely available and could be used intra-articularly, but no investigation has looked at the safety of collagen scaffolds within synovial joints. The aim of this study was to determine the safety of collagen scaffold implantation in a validated in vivo animal model of knee arthrofibrosis. Materials and Methods. A total of 96 rabbits were randomly and equally assigned to four different groups: arthrotomy alone; arthrotomy and collagen scaffold placement; contracture surgery; and contracture surgery and collagen scaffold placement. Animals were killed in equal numbers at 72 hours, two weeks, eight weeks, and 24 weeks. Joint contracture was measured, and cartilage and synovial samples underwent histological analysis. Results. Animals that underwent arthrotomy had equivalent joint contractures regardless of scaffold implantation (-13.9° versus -10.9°, equivalence limit 15°). Animals that underwent surgery to induce contracture did not demonstrate equivalent joint contractures with (41.8°) or without (53.9°) collagen scaffold implantation. Chondral damage occurred in similar rates with (11 of 48) and without (nine of 48) scaffold implantation. No significant difference in synovitis was noted between groups. Absorption of the collagen scaffold occurred within eight weeks in all animals. Conclusion. Our data suggest that intra-articular implantation of a collagen sponge does not induce synovitis or cartilage damage. Implantation in a native joint does not seem to induce contracture. Implantation of the collagen sponge in a rabbit knee model of contracture may decrease the severity of the contracture. Cite this article: J. A. Walker, T. J. Ewald, E. Lewallen, A. Van Wijnen, A. D. Hanssen, B. F. Morrey, M. E. Morrey, M. P. Abdel, J. Sanchez-Sotelo. Intra-articular implantation of collagen scaffold carriers is safe in both native and arthrofibrotic rabbit knee joints. Bone Joint Res 2016;6:162–171. DOI: 10.1302/2046-3758.63.BJR-2016-0193

An established rabbit model was used to preliminarily investigate the effect of acellular triphase, namely bone-cartilage-tendon, scaffold (ATS) sandwiched with autologous bone mesenchymal stem cells (BMSCs) sheets on tendon-bone interface healing. Bone, fibrocartilage and tendon tissue were harvested from the rabbits and sectioned into a book-type scaffold. The scaffolds were decellularized and their characterization was presented. BMSCs were isolated and co-cultured with the scaffolds to verify their cytocompatibility. BMSCs sheets were fabricated and inserted into the book page of the scaffold to construct an autologous BMSCs-sheets/book-type ATS complex. The complex was implated in the right knee of rabbits which operated standard partial patellectomy for TBI regeneration using Imaging, histological and biomechanical examinations. The bone, fibrocartilage and tendon tissue were sectioned into a book-type scaffold before decellularization. Then we decellularized the above tissue and mostly preserved their microstructure and composition of the natural extracellular matrix, including collagen and proteoglycan. After the physicochemical and biological properties of the book-type ATS were evaluated, autologous BMSCs sheets were inserted into the book page of the scaffold to construct an autologous BMSCs-sheets/book-type ATS implants for TBI regeneration. In addition, the ATS has the advantages of non-toxicity, suitable for cell adhesion and growth as well as low immunogenicity while co-cultured with the BMSCs. At the same time, different scaffolds has the ability to induce the osteogenic, chondrogenic and tenogenic differentiation of BMSCs by immunofluorescence, reverse transcription-polymerase chain reaction and western blot analysis. To determine the efficacy of the tissue-engineered implants for TBI regeneration, we transplanted it into a rabbit patella-patellar tendon (PPT) injury model, and the rabbits were sacrificed at postoperative week 8 or 16 for the radiological, histological, and mechanical evaluation. Radiologically, Synchrotron radiation micro-computed tomography (SR-μCT) showed that BMSCs/ATS group significantly increased bone area, BV/TV, trabecular thickness and trabecular number at the healing interface as compared with other groups at postoperative week 8 or 16. Histologically, the BMSCs/ATS group showed more woven bone, and a more robust fibrocartilaginous junction with a characteristic matrix rich in proteoglycans was seen at the PPT healing interface in comparison with other groups after 8 weeks. At week 16, the healing interface in 3 groups displayed better remodeling with respect to postoperative week 8. Healing and remodeling at the PPT junction were almost complete, with a resemblance to a healthy BTI consisting of the characteristic 4 zones in all groups. At last, we used biomechanical test as functional parameters to evaluate the quality of tendon-bone healing. Biomechanical testing indicated that BMSCs/ATS group showed significantly higher failure load and stiffness than other groups at postoperative week 8 and 16. The complex composed of acellular triphase, namely bone-cartilage-tendon, scaffold (ATS) sandwiched with autologous bone mesenchymal stem cells (BMSCs) sheets can simulate the gradient structure of tendon-bone interface, inducing stem cell directional differentiation, so as to promote patella-patellar tendon interface healing effectively after injury

We describe the natural history of a rabbit knee model of permanent post-traumatic joint contractures. Twenty-four skeletally mature female NZW rabbits had five-mm-squares of cortical bone removed from both femoral condyles. An extra-articular K-wire immobilized the knee joint in flexion. The K-wire was removed eight weeks later and the rabbits were divided into four groups depending on the remobilization time. The average extension loss for the experimental knees in the zero, eight, sixteen and thirty-two weeks remobilization groups was thirty-eight, thirty-two, twenty-one and twenty degrees, respectively. The motion loss stabilized in the later time intervals suggesting permanent contractures had developed. The contralateral unoperated knees average extension loss was nine degrees. The purpose of this study was to develop a rabbit knee model of post-traumatic contractures. A simulated intra-articular fracture plus eight weeks of immobilization leads to a permanent joint contracture even after thirty-two weeks of remobilization. This animal model of human post-traumatic joint contractures will allow further studies investigating mechanisms underlying the process. Twenty-four skeletally mature female NZW rabbits had 5 mm squares of cortical bone removed from both femoral condyles. An extra-articular Kirschner wire (K-wire) immobilized the knee in flexion. A second operation was performed eight weeks later to remove the K-wire. The rabbits were divided into four groups. Hind limbs were dissected, preserving the joint capsule. A device allowing six degrees-of-freedom coupled to a material testing system which applied a 0.2 Nm torque measured joint angles. Statistical analysis was performed using ANOVA with a posthoc Student-Newman-Keuls test. Data are presented as mean ± SD. The loss of extension for the experimental knees in the zero and eight weeks remobilization groups was significantly greater than the values of all contralateral unoperated knees. The loss of extension for the experimental knees in the sixteen and thirty-two weeks remobilization groups was also greater than the contralateral knees, although it was not statistically significant (p = 0.07). With this model, the severity of the contracture decreased with time of remobilization. However, the degree of contracture stabilized between sixteen and thirty-two weeks of remobilization, suggesting that the joints had developed a permanent contracture. This mimics the human scenario of permanent post-traumatic joint contractures. Funding: has not been received from a commercial party. This work was supported by The Alberta Heritage Foundation for Medical Research. Please contact author for tables and/or graphs

Summary Statement. The implantation of scaffold-free CTE from suspension culture into growth-plate defects resulted in a significant reduction in growth arrest of the rabbit tibia. Introduction. In childhood and adolescence, the growth plate injury can cause partial premature arrest of growth plate, which can make problems such as leg length discrepancy and angular deformity. Bone bridge resection and variable implantation materials such as fat, bone wax, silastic and craniopalst has been investigated. However, those procedures may show limitations including the control of bone growth and long term safety of implant materials in vivo. As an alternative, homogeneous or heterogeneous cartilage cells and stem cell transplants have been tried. In this method, scaffold for cell transplantation is needed. But, so far the most suitable scaffold has not been established. Recently, some authors generated a cartilage tissue equivalent (CTE) using a suspension culture with biophysical properties similar to native hyaline cartilage. Therefore we are able to transplant the CTE without scaffold to the physeal defect. The purpose of this study was to investigated the effects of a transplantation of a vitro-generated scaffold-free tissue-engineered cartilage tissue equivalent (CTE) using a suspension chondrocyte culture in a rabbit growth arrest model. Material and Method. Cartilage tissue equivalent culture. The CTE was generated by the suspension culture of chondrocytes (2 × 10. 7. /well/1 mL) which was isolated from articular cartilage of 5 weeks New Zealand white rabbit on a 24-well plate (2.4 cm. 2. /well) treated with poly HEMA (nunc, Roskide, Denmark) for up to 8 and 16 weeks. (2)Partial growth arrest animal model. An experimental model for growth arrest was created by excising the growth plate at the proximal medial side of tibia with the 4 mm in diameter and 4 mm in depth from 6-week-old New Zealand white rabbits. Two experimental groups were set to evaluate CTE implantation; group I, no implantation as controls; group II, implantation of CTE. (3) Evaluation of effect of the transplantation of CTE. Serial plain radiographs were performed at one week. The medial proximal tibial angle (MPTA) was measured for assessing the degree of angular deformity. Histologic examination using HE stain, Alcian bule and immunohistochemistry was done at 4 and 8 weeks after surgery. Results. Radiographic results: In group I, all damaged growth plates were arrested and angular deformities appeared 4 weeks later. In groups II, angular deformities were much less than in the control group. Histologic result: In group I, bone bridge formation was shown at the damaged growth plate at 4 weeks after surgery. In group II, regeneration of growth plates was recognised at 4 and 8 week after surgery. However, the thickness of regenerated growth plate at 8 weeks specimen was thinner than that of 4 weeks specimen. Discussion and Conclusion. The implantation of scaffold-free CTE from suspension culture into growth-plate defects resulted in a significant reduction in growth arrest of the rabbit tibia

Introduction. Bisphosphonates are among the most commonly prescribed drugs in Osteoporotic Patients. Their mode of action is anti-resorptive. Since remodeling is a key step in fracture healing, there has been concern regarding the effect of bisphosphonates on fracture healing. Objectives. To assess the effect of alendronate on fracture healing in the rabbit ulna osteotomy model. Materials and methods. 16 New Zealand white rabbits were divided into 2 equal groups. Bilateral ulnar osteotomies were performed in the first week. Group 1 was the control group and group 2 was gavaged with alendronate solution (human equivalent dose). 2 rabbits were euthanised at 3 and 6 weeks and the remaining 4 rabbits were euthanised at 8 weeks. Fracture healing was assessed radiologically, with mechanical testing using the Instron 4302 materials testing machine and histologically, in that order. Results. The fractures healed satisfactorily in all the control group animals. However, in the alendronate treated group, there was an abundance of woven bone and little lamellar bone in the callus. However there was no significant difference in mechanical testing. In addition we did not find any evidence of Osteonecrosis in the Bisphosphonate treated group. Conclusion. Bone remodelling in the alendronate treated group is slower but a larger amount of bone callus is formed around the fracture, thus giving the fracture callus a higher ultimate load to failure at an earlier stage

Aim. Antibiotic prophylaxis is critical for the prevention of fracture related infection (FRI) in trauma patients, particularly those with open wounds. Administration of prophylactic antibiotics prior to arrival at the hospital (e.g. by paramedics) may reduce intraoperative bacterial load and has been recommended; however scientific evidence for pre-hospital administration is scarce. Methods. The contaminated rabbit humeral osteotomy model of Arens was modified to resemble the sequence of events in open fractures. In an initial surgery representing the “accident”, a 2mm mid-diaphyseal hole was created in the humerus and the wound was contaminated with a clinical Staphylococcus aureus strain (mean 1.6×106 Colony Forming Units, CFU). The animals were allowed recover for 4 hours mimicking the period from trauma to debridement. At this time, a second procedure was performed in order to debride and irrigate the wound, and to fix a complete osteotomy that was made through the initial defect. Three test groups were included (n=8 rabbits per group): 1) no antibiotic therapy; 2) standard “in-hospital” antibiotic prophylaxis (24 hours therapy starting 30 minutes before surgery); 3) “pre-hospital” antibiotics (single dose 15 minutes after the “accident”). The antibiotic used was cefuroxime and was administered in a weight-adjusted dosage. Results. In the absence of any antibiotic administration (group 1), high bacterial counts were identified at fixation (1.89×106 CFU) and at euthanasia (day 7, 7.70×107 CFU) in all rabbits. When 24 hours of antibiotics were administered commencing “in hospital” (group 2), the bacterial load during fixation surgery was slightly reduced (CFU 9.88×105) and 50% of animals were infected at euthanasia. When one single shot of antibiotics was administered in the “pre-hospital” setting (group 3), the bacterial load during fixation surgery was significantly lower than for both groups 1 and 2(CFU 2.34×103) yet all animals were infected at euthanasia. Conclusions. Early pre-hospital administration of antibiotics significantly reduced the bacterial load in the operative field at the time of debridement compared to regular prophylaxis. However, continuation of systemic antibiotics is necessary in order to prevent infection in this model

In the pubertal growth plate, sex hormones play important roles for the regulation of the proliferation, differentiation, maturation and programmed death of chondrocytes. Many studies have been reported on the regulation of oestrogen in long bone growth, however, some of the mechanisms have remained unclarified to date including its role for cell kinetics in the growth plate chondrocytes. The aim of this study was to clarify the effect of the deficiency of oestrogen on growth plate chondrocytes. We obtained the growth plates of femoral head from the normal and ovariectomized Japanese white rabbits at 10, 15, 20 and 25 weeks. Ovariectomy was performed at 8 weeks. The cell kinetics of chondrocytes as defined by the numbers of proliferating and programmed dying cells was investigated using immunohistological methods. The lengths of the femur were almost same both in the ovariectomised and normal rabbits. The height of the growth plate was larger in the former. The total number of chondrocytes in the ovariectomised rabbits was less than that of normal rabbits of the same age. Immunostaining of proliferating cell nucleous antigen (PCNA) showed a decrease number of proliferating chondrocytes and that of caspase-3 indicated a little increased number of apoptotic chondrocytes. Oestrogen regulates endochondral bone formation through several pathways. It directly binds oestrogen receptor alpha and beta, and the former accelerates longitudinal bone growth whereas the latter represses it. Another pathway is through the GH-IGF-I axis: it closely interacts with GH and IGF-I for the control of longitudinal bone growth. In addition, there might be other mediators including transforming growth factor-beta, other IGFs and still unknown paracrine or auto-crine factors as IHH PTHrP. Our study suggests that in the rabbit growth plate during puberty, oestrogen mainly acts through the GH-IGF-I axis since its defi-ciency declined the proliferating ability of chondrocytes, which led the decrease of the number of chondrocytes

Thigh pain appears often after THA used of the cement-less femoral components, but the appearance mechanism of thigh pain does not have been elucidated. As one factor of manifestation of thigh pain, it has been guessed that the pressure from the inside of medullary cavity of bone by the stem. The purpose of this study is confirming whether the flexor reflex is caused, by using the femur of a rabbit that applied the pressure from the inside of medullary cavity of bone. Japanese white rabbits with weight of about 3kg were used. Evaluation of the appearance of the pain by the pressure was performed by measurement of the hind leg flexor activity produced by the flexor reflex. After confirming that appearance of the muscles activity by the pain reflex from adding the pain stimulus to the hind leg skin of rabbits, we loaded of the pressure into the inside of medullary cavity of bone and observed whether the muscles activity appears. As the laboratory animals model, we prepared two kinds of rabbits by the difference in the amount of reaming. And we tested how the differences show up between these two kinds of rabbits. In the rabbits with few amounts of reaming, the flexor reflex appeared in low pressure. But, in the rabbits with many amounts of reaming, the flexor reflex did not appear in high pressure, either. It is known that the somatic sensory nerves are distributed in the bone, and it is known that the sensory nerve ends exist in the medullary cavity of bone. It was suggested that the pain is induced, when the sensory nerve ends remained in the inside of medullary cavity of bone and the pressure in whicha reaction is possible was carried out there

Introduction: Despite the present routine treatment of septic arthritis with antibacterial agents, articular damage is persistent and frequently leads to loss of joint function. The aim of this study was to assess the effect of intra-articular corticosteroid added to systemic antibiotics in the treatment of experimental staphylococcal knee joint infection in rabbits. Material and Methods: Thirty rabbits were injected in their knees by staphylococcus aureus. The rabbits were divided into 3 equal groups. In group A, rabbits received no treatment. In group B, rabbits were treated with systemic antibiotics alone. Group C, received systemic antibiotics and intra-articular corticosteroid. After 16 days animals were killed and the knee joint X-Ray as well as histopathological-histochemical parameters were assessed. Results: All rabbits survived the experiment; the treated groups (B, C) had better histopathological-histochemical scores in comparison with the untreated group (A). Group C had significantly better scores in joint sections in comparison with group B (mean SD=6.7 ± 2.3 v 4.0 ± 2.4, P= 0.019). Lower damage in the former group was expressed in lesser clustering of chondrocytes, proteoglycan depletion, and severity of synovitis. Radiological soft tissue scoring was significantly better in group C in comparison with group B. Three peri-articular abcesses were observed in group C but none in group B. Conclusion: Addition of intra-articularly administered corticosteroid to antibiotic treatment of septic arthritis improved histopathological-histochemical parameters in this experimental setting, although on account of the clinical observation of three cases with peri-articular abcesses in this group, caution is warranted in the interpretation of these results

Our aim was to determine the effect of denervation on repair-associated mRNA levels in the MCL after partial tear. Cohorts of rabbits underwent partial MCL tear with or without concomitant femoral nerve transection. Ligaments were harvested, RNA extracted and RT-PCR was performed using rabbit-specific primers for repair-associated molecules at three days, two wks, six wks and sixteen wks post-injury. Angiogenesis genes MMP3, MMP13, matrix components Collagen I and III and growth factors TGF-ß and NGF mRNA levels were increased in the denervated group at two-weeks post-injury (p< 0.05). Denervation significantly alters mRNA levels during the early stages of rabbit MCL healing. To determine the effect of denervation on repair-associated mRNA levels in the injured medial collateral ligament (MCL). Previous experiments revealed that denervation impairs healing of the MCL. We hypothesized that denervation would decrease repair-associated mRNA levels in the injured MCL when compared with normally innervated injured MCL. Adult, skeletally mature female rabbits were assigned to one of four groups: unoperated control, femoral nerve transection alone (denervated controls), MCL partial tear and denervated MCL partial tear. At three days, two weeks, six weeks or sixteen weeks post-surgery, cohorts of six rabbits from each experimental group were killed. Control rabbits were assessed at two weeks. Ligaments were harvested, RNA extracted and RT-PCR was performed using rabbit-specific primers. In the denervated injury group, mRNA levels of angiogenesis genes MMP-3 and MMP-13, matrix components Collagen I and III and growth factors TGF-ß and NGF had all increased at two-weeks post-injury, in comparison to non-denervated (p< 0.05). We also found increased levels of MMP-3 and NGF mRNA in the denervated group at sixteen weeks post injury (p< 0.05). The mRNA levels of the housekeeping gene GAPDH were increased in the denervated group only at three days post injury (p< 0.05). Of note, TGF-ß mRNA levels were significantly decreased in the denervated group at three days post injury (p< 0.05). Contrary to our initial hypothesis, denervation increases mRNA levels for many important molecules during the early stages of MCL healing. Additional research will be required to explain how and why denervation impairs ligament healing. No previous study has shown that innervation regulates mRNA levels in healing ligament