Rabbit Joints Research Articles

Can treatment with chondroitin and glucosamine sulphate prevent changes in the articular disc caused by temporomandibular joint osteoarthritis?

Chondroitin and glucosamine sulphates (CGS) are considered structure-modifying drugs and have been studied in the prevention, delay or reversal of structural morphological changes in joints caused by osteoarthritis. The aim of the present study was to investigate the action of CGS on the progression of chemically induced osteoarthritis in the temporomandibular joint (TMJ) of rabbits by evaluating the serum levels of tumour necrosis factor (TNF-α) and collagen in the articular discs. A sample of 36 male rabbits was divided into three groups: control (CG), osteoarthritis (OG) and treatment (TG). The disease was induced by intra-articular injection of sodium monoiodoacetate (10 mg/mL) in the OG and TG groups bilaterally. After 10 days, the TG animals received subcutaneous injection of chondroitin sulphates and glucosamine (7.5 mg/kg) and the OG and CG received saline solution (50 μL). Euthanasia times were subdivided into 40 and 100 days. Collagen quantification was performed by biochemical and histological analysis and for the quantification of serum levels of TNF-α, an enzyme immunoassay was used. The TG showed an increase in the collagen area of the articular disc when compared to the CG and the OG. The increase collagen concentration in the discs did not show a statistically significant difference between the groups. Post-treatment TNF-α levels were significantly lower in TG compared to OG. The results indicate that CGS treatment delayed the degeneration of the collagen in the TMJ articular disc and reduced serum TNF-α levels, indicating a preventive effect on OA progression.

An HFD negatively influences both joint and liver health in rabbits with and without an enzymatically-induced model of arthritis

Osteoarthritis (OA) is a common arthritis types in animals that causes persistent pain and reduces quality of life. Although a high-fat diet (HFD) is widely believed to induce obesity and have adverse effects on the body, the connection between HFD and joint health is not well understood. Therefore, in this study, 32 healthy male New Zealand rabbits were randomly divided into four groups: healthy rabbits fed a standard diet (NDG, n=8) or an HFD (HDG, n=8), rabbits fed a standard diet (OAG, n=8) and an HFD (HOG, n=8), and arthritis was induced by intra-articular enzyme injection. After 12 weeks of HFD feeding, articular cartilage, synovium, and subchondral bone were isolated and collected. Joint tissue damage was evaluated using histopathological and imaging tests. The results showed that there was no significant difference in body weight between rabbits fed a normal diet and those fed an HFD. However, the HFD led to an increase in joint injuries in both induced and non-induced arthritis rabbits. Specifically, the HFD induced lipid metabolism disorders and liver damage in vivo, significantly elevating the levels of serum inflammatory cytokines and bone metabolism markers. Moreover, HFD exacerbated articular cartilage damage in the joints and increased the accumulation of inflammatory cells in synovial tissue, resulting in a notable increase in synovial macrophages and inflammatory cytokines. Additionally, HFD accelerated the bone resorption process in subchondral bone, leading to the destruction of bone mass and subchondral bone microstructure. In summary, the results of this study indicate that an HFD can cause histological damage to the articular cartilage, synovium, and subchondral bone in rabbits, exacerbating arthritis in pre-existing joint damage. Notably, weight is not the primary factor in this effect.

Intra-articular injection of modified citrus pectin and hyaluronate gel induces synergistic effects in treating osteoarthritis

We previously found that modified citrus pectin (MCP), an inhibitor of pro-inflammatory factor Galectin-3 (Gal-3), has significant anti-inflammatory and chondroprotective effects. In this study, a hyaluronate (HA) gel-based sustained release system of MCP (MCP-HA) was developed as an anti-inflammatory agent for chronic inflammation for osteoarthritis (OA) treatment. The MCP-HA gel was injected into the knee joint cavities of OA rabbit models induced by anterior cruciate ligament transection (ACLT) or modified Hulth method once a week for five weeks. We found that MCP-HA could improve the symptoms and signs of OA, protect articular cartilage from degeneration, suppress synovial inflammation, and therefore alleviate OA progression. Proteomic analysis of the synovial fluid obtained from the knee joints of OA rabbits revealed that MCP-HA synergistically regulated the levels of multiple inflammatory mediators and proteins involved in metabolic pathways. Taken together, our results demonstrate that the MCP-HA shows a synergistic effect of HA and MCP by modulating both inflammation and metabolic processes, thereby alleviating OA progression. The MCP-HA sustained release system has promising potential for long-term use in OA treatment.

Intra-articular administration of extra-virgin olive oil in degenerative osteoarthritis

BackgroundWe aimed to analyze the outcomes of intraarticular extra virgin olive oil (EVOO) injection on mechanically induced rabbit knee osteoarthritis (OA) by studying the morphological, histological, and radiological findings.MethodsThe study was conducted on 32 New Zealand White rabbits. The randomly numbered subjects were divided into two main groups. The rabbits numbered 1 to 16 were selected to be the group to receive EVOO, and the remaining were selected into a control group. Both groups were separated into two subgroups for short-term (five weeks) and long-term (10 weeks) follow-up. Anterior cruciate ligament transection was applied on the left knees of all the rabbits via medial parapatellar arthrotomy to simulate knee instability. Immediately after the surgical procedure, 0.2 cc of EVOO was injected into the knee joint of rabbits numbered 1–16, and the control group received 0.2 cc of sterile saline. On the 14th day, long-term group subjects were administered another dose of 0.2 cc EVOO intraarticularly.ResultsThe gross morphological scores of the control group subjects were significantly different from the EVOO group for both short-term (p = 0,055) and long-term (p = 0,041) scores. In parallel, the MRI results of the EVOO subjects were significantly different from the control group for both short-term and long-term follow-up assessment scores (p = 0.017, p = 0.014, respectively). The Mankin scoring results showed that there were statistically significant differences between the EVOO and control group in the comparison of both total scores (p = 0.001 for short-term and p = 0.004 for long-term) and subgroup scoring, including macroscopic appearance, chondrocyte cell number, staining, and Tidemark integrity in both short-term (p = 0.005, p = 0.028, p = 0.001, p = 0.005, respectively) and long-term assessments (p = 0.002, p = 0.014, p < 0.001, p = 0. 200, respectively).ConclusionsWe have observed promising outcomes of intra-articular application of extra virgin olive oil in the treatment of acute degenerative osteoarthritis in rabbit knees. Due to its potential cartilage restorative and regenerative effects, EVOO, when administered intra-articularly, may be a promising agent to consider for further research in the treatment of OA.

Effectiveness of Bilayer Scaffold Containing Chitosan/Gelatin/Diclofenac and Bovine Hydroxyapatite on Cartilage/Subchondral Regeneration in Rabbit Joint Defect Models.

Subchondral defects are often caused by trauma involving cartilage damage, leading to subsequent damage to the underlying bone, specifically the subchondral region. Bilayer scaffolds made from biomaterials, such as bovine hydroxyapatite, possess biocompatible and biodegradable properties that mimic the natural environmental conditions of target tissues so that they can support the formation of new tissues. On the other side, diclofenac as an anti-inflammatory drug potentiates to inhibit the inflammatory excess regarding the damage. This study aims to study the effectiveness of diclofenac scaffold to rabbit joint defect model. The scaffold was implanted in the rabbit femoral trochlear bone hole, which had a diameter of 5 mm and a depth of 4 mm. After 28 days of intervention, the animals were examined using macroscopic evaluation, hematoxylin-eosin (HE) staining, and immunohistochemistry (IHC) for type I collagen and type II collagen. Subsequently, the cartilage was evaluated using the International Cartilage Repair Society (ICRS) scoring system. The macroscopic ICRS scores were significantly higher (p < 0.05) in the bilayer scaffold implantation group compared to the monolayer scaffold and control groups. Histological ICRS scores were also significantly higher (p < 0.05) in the bilayer scaffold group compared to the control group. Type II collagen expression was higher (p < 0.05) in the bilayer scaffold group compared to the monolayer scaffold and control groups, although type I collagen expression was lower in comparison. In conclusion, this research suggests that the diclofenac-loaded bilayer scaffold effectively enhances cartilage and subchondral bone regeneration.

LIPUS Alleviates Knee Joint Capsule Fibrosis in Rabbits by Regulating SOD/ROS Dynamics and Inhibiting the TGF-β1/Smad Signaling Pathway.

The aim of the work described here was to investigate the efficacy and potential mechanisms of low-intensity pulsed ultrasound (LIPUS) for the treatment of arthrogenic contracture induced by immobilization in rabbits. The left knee joint of rabbits was immobilized for 6 wk to establish the model of extending knee joint contracture. The rabbits were divided into a control group (C), a group immobilized for 6 wk (IM-6w), a group remobilized for 1 wk (RM-1w), a group subjected to LIPUS intervention for 1 wk (LIPUS-1w), a group remobilized for 2 wk (RM-2w) and a group subjected to LIPUS intervention for 2 wk (LIPUS-2w). The degrees of arthrogenic contracture and joint capsule fibrosis were assessed, as were the levels of reactive oxygen species (ROS) and the activation status of the TGF-β1/Smad signaling pathway in the joint capsule. After immobilization for 6 wk, the degrees of arthrogenic contracture and joint capsule fibrosis increased. The ROS level increased, as evidenced by an increase in malondialdehyde content and a decrease in superoxide dismutase content. In addition, the TGF-β1/Smad signaling pathway was significantly activated. The degrees of knee joint contracture increased in the first week after remobilization and decreased in the second week. Furthermore, joint capsule fibrosis continued to develop during the 2 wk of remobilization, and the ROS level increased, while the TGF-β1/Smad signaling pathway was significantly activated. LIPUS effectively reduced the level of ROS in the joint capsule, which further inhibited activation of the TGF-β1/Smad signaling pathway, thereby improving joint capsule fibrosis and reducing arthrogenic contracture. The high ROS levels and overactivation of the TGF-β1/Smad signaling pathway may be reasons why immobilization induces knee joint capsule fibrosis. LIPUS can alleviate the degree of knee joint capsule fibrosis induced by immobilization by inhibiting the production of ROS and the activation of the TGF-β1/Smad signaling pathway.

Hst1/Gel-MA Scaffold Significantly Promotes the Quality of Osteochondral Regeneration in the Temporomandibular Joint.

Our aim was to evaluate the capacity of the human salivary histatin-1-functionalized methacrylic gelatin scaffold to control osteochondral tissue regeneration and repair in vivo in rabbits with major temporomandibular joint dimensional abnormalities. In order to compare human salivary histatin-1-functionalized methacrylic gelatin scaffolds to the Blank and Gel-MA hydrogel groups, scaffolds were implanted into osteochondral lesions of a critical size (3 × 3 mm) in the anterior region of the condyle of the temporomandibular joint in New Zealand white rabbits. At 4 weeks after implantation, the repair was evaluated using macroscopic examination, histology, and micro-CT analysis. In the comparison of the composite scaffold group with the Blank and Gel-MA groups, analysis of the healed tissue revealed an improved macroscopic appearance in the composite scaffold group. Regeneration was induced by host cell migration in the Hst1/Gel-MA scaffold group. The current study offers a viable method for in vivo cartilage repair that does not require cell transplantation. Future clinical applications of this strategy's optimization have many potential advantages.

Establishment of an ultrasound-guided protocol for the assessment of hip joint osteoarthritis in rabbits-A sonoanatomic study.

Ultrasound (US) has emerged as one of the most applied imaging tools to diagnose musculoskeletal disorders and assist in guided intra-articular administrations. Nevertheless, in evaluating the rabbit hip joint, there is a need for an ultrasonographic approach. Therefore, this study aimed to describe the hip sonoanatomy, develop and validate a US-guided protocol to assess the hip joint in rabbits and apply this protocol in vivo. This study was carried out in three phases, phase I: a pilot cadaveric study, to assess the applicability of different US approaches in the hip of rabbits and, consequently, develop a detailed US-guided protocol (2 rabbit cadavers, n = 4 hips); phase II: validation of the established US-guided protocol through a numerical scoring system in healthy joints (11 rabbit cadavers, n = 22 hips), and, lastly, phase III: application of the US-guided protocol in vivo in osteoarthritic joints (5 rabbits, n = 5 hips). A total of six planes were validated, two in the ventral approach and four in the dorsal approach. The ventral transverse plane was deemed more informative regarding the hip joint sonoanatomy, enabling the identification of a greater number of structures when compared to the other planes. Nevertheless, this study suggested that the isolated application of a plane was deemed insufficient for a complete and detailed evaluation of the hip joint anatomy, rendering it necessary to employ other planes complementarily. Furthermore, the established US-guided protocol allowed a definitive diagnosis of OA, and osteophytes and capsular hypertrophy were among the defects most frequently detected. This novel study provided US anatomical landmarks for forthcoming therapeutic research and monitoring of OA development, granting the accurate identification of osseous and cartilaginous defects.

A sustained-release phospholipid-based phase separation gel loaded with berberine for treating rheumatoid arthritis.

Berberine (BBR) has a long history of use in the treatment of Rheumatoid arthritis (RA) and is considered one of the most promising natural product for the treatment of RA. However, oral administration of berberine has low bioavailability and requires frequent administration, resulting in poor patient compliance. In this study, we developed a BBR-loaded phospholipid-based phase separation gel (BBR-PPSG) to achieve sustained drug release and long-term therapeutic effect. The stability of BBR-PPSG was verified and it was found that it can be stored for a long time. The pharmacokinetic study on rats and rabbits showed that BBR-PPSG not only achieved 1-month of sustained release, but also significantly increased the area under the curve (AUC) by nearly 9-fold and prolonged the half-life (t1/2) by 10-fold. By constructing rat and rabbit models of RA, we also proved that BBR-PPSG administration once a month effectively alleviated joint swelling, and significantly reduce TNF-α levels in AIA rats and OIA rabbits. Histopathological analysis of rabbit joint sections revealed that after intra-articular injection of BBR-PPSG, the synovial cell layer remained intact, while in the model group, the synovial cells were significantly reduced and exhibited necrosis. MicroCT data analysis showed that the values of Tb.N and Tb. Sp in the BBR-PPSG group were significantly better than those in the model group (p < 0.05). This study addressed the limitations of frequent administration of BBR by developing a phospholipid-based phase separation gel system for berberine delivery, achieving long-term sustained release. The BBR-PPSG demonstrated good biocompatibility, simple preparation and excellent stability, thus holding potential as a novel pharmaceutical formulation for RA treatment.

Anti-NGF treatment worsens subchondral bone and cartilage measures while improving symptoms in floor-housed rabbits with osteoarthritis.

Objective: Osteoarthritis (OA) is a common joint disorder often affecting the knee. It is characterized by alterations of various joint tissues including subchondral bone and by chronic pain. Anti-nerve growth factor (NGF) antibodies have demonstrated improvement in pain associated with OA in phase 3 clinical trials but have not been approved due to an increased risk of developing rapidly progressive OA. The aim of this study was to investigate effects of systemic anti-NGF-treatment on structure and symptoms in rabbits with surgically induced joint instability. Methods: This was elicited by anterior cruciate ligament transection and partial resection of the medial meniscus in right knee of 63 female rabbits, housed altogether in a 56m2 floor husbandry. Rabbits received either 0.1, 1 or 3mg/kg anti-NGF antibody intra-venously at weeks 1, 5 and 14 after surgery or vehicle. During in-life phase, static incapacitance tests were performed and joint diameter was measured. Following necropsy, gross morphological scoring and micro-computed tomography analysis of subchondral bone and cartilage were performed. Results: After surgery, rabbits unloaded operated joints, which was improved with 0.3 and 3mg/kg anti-NGF compared to vehicle injection during the first half of the study. The diameter of operated knee joints increased over contralateral measures. This increase was bigger in anti-NGF treated rabbits beginning 2weeks after the first IV injection and became dose-dependent and more pronounced with time. In the 3mg/kg anti-NGF group, the bone volume fraction and trabecular thickness increased in the medio-femoral region of operated joints compared to contralateral and to vehicle-treated animals, while cartilage volume and to a lesser extent thickness decreased. Enlarged bony areas were found in right medio-femoral cartilage surfaces of animals receiving 1 and 3mg/kg anti-NGF. Alterations of all structural parameters were particularly distinct in a subgroup of three rabbits, which also exhibited more prominent symptomatic improvement. Conclusion: This study showed that anti-NGF administration exerted negative impact on structure in destabilized joints of rabbits, while pain-induced unloading of joints was improved. Our findings open up the possibility to better understand the effects of systemic anti-NGF, particularly on subchondral bone, and thus the occurrence of rapidly progressive OA in patients.

Magnetic resonance imaging, computed tomography, and gross anatomy of forelimb joints in New Zealand rabbit (Oryctolagus cuniculus L.).

The present study aimed to define the anatomical structures by comparing the transversal computed tomography (CT) and magnetic resonance (MR) images of the forelimb joints of the rabbits with the cross-sectional plastinated images. A total of 14 (seven females, seven males) one-year-old adult New Zealand rabbits were used in the study. After the CT and MR imaging procedures were completed, the forelimbs were removed from the body. The forelimbs were plastinated using the silicone plastination method and sectioned transversal. Cross-sectional plastinates were evaluated and correlated anatomically with conjugate CT and MR images. Joint and surrounding anatomical structures were defined in sections. Cross-sectional plastinated samples were highly correlated with CT and MR images in terms of bone and soft tissue, respectively. It is thought that the anatomical and radiological data obtained from the forelimb joints of rabbits will provide a basis for scientists who are involved in both experimental surgical interventions and clinical anatomy education.

Effect of heat-reinforcing needling on the inflammation and necrotizing apoptosis of synovial cells in synovial tissues of knee joint in rabbits with cold syndrome rheumatoid arthritis

To observe the effect of heat-reinforcing needling (HRN) on inflammatory factors and necrotizing apoptosis of synovial cells in synovial tissues of knee joint in rabbits with cold syndrome rheumatoid arthritis (RA), so as to explore its underlying mechanisms in treating RA. By using the random number table method, 40 New Zealand rabbits were randomly divided into normal, model, antagonist(AG), twist-reinforceing needling (TRN) and HRN groups, with 8 rabbits in each group. The model of cold syndrome RA was established by ovalbumin induction combined with Freund's complete adjuvant injection and cryogenic freezing method. In the AG group, the antagonist TAK-632 (25 mg/kg) was administered intragastrically, once every 2 days, for a total of 7 times. Rabbits of TRN and HRN groups were treated with corresponding acupuncture techniques on bilateral "Zusanli" (ST36) for 30 min, once a day for 14 days. After intervention, the changes of knee skin temperature and circumference were measured. Color Doppler ultrasound was used to observe the joint cavity effusion, synovial thickness and internal blood flow signal. The histomorphological changes of synovial tissues were observed after HE staining. ELISA was used to detect the contents of tumor necrosis factor (TNF)-α, interleukin (IL)-1β and IL-6 in serum. Transmission electron microscope was used to observe the ultrastructure, necrosis and apoptosis of synovial cells. Western blot was used to detect the protein expressions of receptor-interacting protein kinase1 (RIPK1), RIPK3, mixed lineage kinase domain-like protein (MLKL), and phosphorylation （p）-MLKL in synovial tissues. Compared with the normal group, the synovial was diffusely hyperplasia, joint cavity effusion and abnormal blood flow signal were obvious, inflammatory cells were clustered, arranged closely and disordered in the model group. The findings of transmission electron microscopy showed disruption of cell membrane integrity, swollen or ruptured mitochondria, obviously ruptured nucleus, condensed and pyknotic chromatin and nucleolus in the model group. Also, the skin temperature of the knee joint was significantly decreased (P<0.01), while the circumference of the knee joint, the contents of TNF-α, IL-1β and IL-6 in serum, the protein expressions of RIPK1, RIPK3, p-MLKL and MLKL in synovial tissues were significantly increased (P<0.01) in the model group. Compared with the model group, synovial tissue hyperplasia, joint cavity effusion, abnormal blood flow signals, synovial cell proliferation, inflammatory cell infiltration, disruption of cell membrane integrity, cell swelling, cell rupture, and nuclear pyknosis were reduced to different degrees in the AG, TRN and HRN groups. Additionally, the skin temperature of the knee joint was increased (P<0.01, P<0.05), while the circumference of the knee joint, the contents of TNF-α, IL-1β and IL-6 in serum, the expressions of RIPK1, RIPK3, p-MLKL and MLKL in synovial tissues were decreased (P<0.01, P<0.05) in the AG, TRN and HRN groups. The effects of HRN and AG were notably superior to that of TRN in up-regulating skin temperature of the knee joint, and down-regulating the circumference of the knee joint, the contents of TNF-α, IL-1β and IL-6 in serum, the expressions of RIPK1, RIPK3, p-MLKL and MLKL in synovial tissues (P<0.01, P<0.05). HRN can reduce synovial inflammation of knee joint in rabbits with cold syndrome RA, which may be related to its function in inhibiting the necrotizing apoptosis of synovial cells.

Protective effect of additional cathelicidin antimicrobial peptide PR-39 on prosthetic-joint infections.

Prosthetic-joint infection (PJI) is one of the severest complications after arthroplasty. However, antibiotics are not effective in the bacteria in biofilm outside the prosthetic-joint. Antimicrobial peptides have an efficient antimicrobial activity in staphylococcus aureus compared with conventional antibiotics. Bone marrow stem cells (BMSCs) were isolated, cultured and transfected with cathelicidins antimicrobial peptides proline-arginine-rich 39 amino acid peptide (PR-39) lentivirus. The expression of PR-39 gene in BMSCs was detected by RT-PCR, and the antibacterial activity of PR-39 was measured by agar diffusion method. The transfection efficiency was detected by fluorescence microscopy. The infection model of artificial knee joint in rabbits were established. Kirschner wire was used as the knee joint implant to implant the distal femur through the femoral intercondylar fossa of rabbits. 24 rabbits were randomly divided into 2 groups for the above operations: group A was inoculated 0.5mL into the joint cavity immediately after the incision was sutured 1 × 107 Staphylococcus aureus of colony forming unit (CFU), group B was inoculated with Staphylococcus aureus and PR-39. After operation, the wound conditions and histological changes were observed by X-ray and optical microscope respectively, CRP and erythrocyte sedimentation rate were measured by test assay. The transfection efficiency of lentivirus vectortransfected BMSCs was 74.09%. The supernatant of lentivirus vector had obvious inhibitory effect on Staphylococcus aureus, and the antibacterial rate was 98.43%. 100% infection observed in group A while few infection observed in group B; serum CRP and ESR at a high level in group A while decreased in group B after operation. There were no significant difference in CRP and ESR between the pLV/PR-39 group and pLV/EGFP group at day 1 and 3 respectively after surgery. However, CRP and ESR in the pLV/PR-39 groupwere significantly lower than the pLV/EGFP group at day 7 and 14 respectively after operation. Rabbits planted BMSCs expressing PR-39 were significantly increased resistance to Staphylococcus aureus in PJI than control group thus showing great potential for preventing implant-associated infection. It will provide a potential new therapeutic agent for implant-associated infection.

Effect of needle-knife on chondrocyte apoptosis of knee joint in rabbits with knee osteoarthritis based on CircSERPINE2-miR-1271-5P-ERG axis

To observe the effect of needle-knife on the chondrocyte apoptosis of knee joint in rabbits with knee osteoarthritis (KOA) based on the CircSERPINE2-miR-1271-5P-E26 specific transformation-related gene (ERG) axis, and to explore the mechanism of needle-knife for KOA. Thirty-six New Zealand white rabbits were randomly divided into a normal group, a model group, a needle-knife group and a sham needle-knife group, 9 rabbits in each group. The rabbits in the model group, the needle-knife group and the sham needle-knife group were treated with modified Videman method to prepare KOA model. After successful modeling, the rabbits in the needle-knife group were treated with needle-knife at cord adhesion and nodules near quadriceps femoris tendon and internal and external collateral ligament on the affected knee joint; the rabbits in the sham needle-knife group were treated with sham needle-knife baside the needle insertion point of the needle-knife group (needle-knife was only inserted, without any operation). The treatment was given once a week, 3 times in total. The Lequesne MG behavioral score was used to evaluate the knee joint damage in each group before and after intervention. After intervention, HE staining and transmission electron microscopy were used to observe the cartilage tissue morphology and ultrastructure of chondrocytes in the knee joint in each group; TUNEL method was used to detect the level of chondrocyte apoptosis in the knee joint; real-time fluorescence quantitative PCR was used to detect the expression of CircSERPINE2, miR-1271-5P and ERG mRNA in knee cartilage tissue in each group. After intervention, compared with the normal group, the Lequesne MG behavioral score in the model group was increased (P<0.01). Compared with the model group and the sham needle-knife group, the Lequesne MG behavioral score in the needle-knife group was decreased (P<0.01). In the model group and the sham needle-knife group, the number of chondrocytes and organelles was decreased, the cell nucleus was shrunk, mitochondria was swelling or disappeared; in the needle-knife group, the number of chondrocytes and organelles was increased, the cell nucleus was not obviously shrunk and the mitochondria was not obviously swelling. Compared with the normal group, the level of chondrocyte apoptosis in the model group was increased (P<0.01); compared with the model group and the sham needle-knife group, the level of chondrocyte apoptosis in the needle-knife group was decreased (P<0.01, P<0.05). Compared with the normal group, the expression of CircSERPINE2 and ERG mRNA in the model group was decreased (P<0.01), and the expression of miR-1271-5P mRNA was increased (P<0.01); compared with the model group and the sham needle-knife group, the expression of CircSERPINE2 and ERG mRNA in the needle-knife group was increased (P<0.01), and the expression of miR-1271-5P mRNA was decreased (P<0.01). Needle-knife could reduce the knee joint damage and chondrocyte apoptosis in KOA rabbits, which may be related to up-regulating the expression of CircSERPINE2 and ERG mRNA, and inhibiting the expression of miR-1271-5P mRNA.

Application of functionalized hyaluronic acid hydrogel with the activity of regulating the behaviors of stem cells in repairing rabbit knee articular cartilage.

Using injectable hydrogels loaded with mesenchymal stem cells (MSCs) to repair chondral defects is a new trend of cartilage tissue engineering in recent years. In this study, hyaluronic acid (HA) hydrogels containing the system of sustained-release Kartogenin (KGN) and modified by RGD and HAV peptides were used to facilitate repair of cartilage defect in the knee joint of rabbits. Different groups of implants were injected into osteochondral defects, and samples were taken 4weeks after operation. Through the qualitative and quantitative analysis of Micro-CT, it can be seen that both FH (unloaded cell group) and R + FH groups (allogeneic cell group) can repair osteochondral defects well, and the amount of bone formation is high, which is close to the intact cartilage groups. Macroscopic observation and histological staining analysis showed that except for the intact cartilage group, FH group obtained the highest score. The morphology of the cartilage tissue in the FH groups was more regular and continuous than that in R + FH and H + FH (xenogeneic cell group) groups, approaching that of native cartilage. Immunohistochemical staining of Collagen II (Col II) showed that the expression and morphology of Col II in FH groups were similar to those in intact cartilage tissue. Interestingly, through in vivo experiments, this functionalized hyaluronic acid hydrogel can effectively promote the rapid repair of rabbit knee cartilage defects within one month.

Effects of heat-reinforcing needling on synovial inflammation and miR-155/TLR4/NF-κB signaling axis in rheumatoid arthritis rabbits with cold syndrome

To observe the effects of heat-reinforcing needling on synovial inflammation and microRNA-155 (miR-155)/Toll-like receptor 4 (TLR4)/nuclear transcription factor-κB (NF-κB) signaling axis, so as to investigate its anti-inflammatory mechanism in rabbits with cold syndrome of rheumatoid arthritis (RA). A total of 36 rabbits were randomly divided into normal, model, agonist, inhibitor, heat-reinforcing needling (HRN) and agonist+heat-reinforcing needling (A+HRN) groups, with 6 rabbits in each group. The RA with cold syndrome model was induced by injecting ovalbumin dry powder and Freund's complete adjuvant combined with cold freezing. Rabbits in agonist group were intraperitoneally injected with miR-155 agomir 4.5 OD; rabbits in the inhibitor group were intraperitoneally injected with miR-155 antagomir 6.1 OD; rabbits in HRN group received heat-reinforcing needling at bilateral "Zusanli" (ST36) for 30 min；rabbits in A+HRN group received the same treatment as agonist group, and 30 min later, received the same treatment as the HRN group; rabbits in the normal and model groups were grasped and fixed in the same way, all groups received continuous treatment once a day for 7 d. After modeling, the knee joints of rabbits were examined by ultrasound, the pain threshold and the circumference were determined. After the interventions, the pain threshold and knee circumference were measured; the pathological morphology of synovial tissue of the knee joints were observed by HE staining; the mRNA levels of miR-155 and suppressor of cytokine signaling protein 1 (SOCS1), the expression levels of SOCS1, TLR4, NF-κB p65, interleukin (IL)-1β and IL-17A proteins in synovial tissue of knee joints were detected by real-time PCR and Western blot respectively. Compared with the normal group, the pain threshold was significantly decreased (P<0.05), and the knee circumference was significantly increased (P<0.05); the synovial tissue of knee joints showed significant hyperplasia, abundant blood flow signal, joint cavity effusion and obvious inflammatory invasion, the pathological score was significantly increased (P<0.05), the expressions of miR-155 mRNA and IL-1β, IL-17A, TLR4, NF-κB p65 proteins were significantly increased (P<0.05), the expressions of SOCS1 mRNA and protein were significantly decreased (P<0.05) in the model group. Compared with model group, the pain threshold was significantly increased (P<0.05), the circumference of knee joint was significantly decreased (P<0.05); in synovial tissue, the pathological score was decreased (P<0.05), the expression levels of miR-155 mRNA and IL-1β, IL-17A, TLR4, NF-κB p65 proteins were significantly decreased (P<0.05), and the expressions of SOCS1 mRNA and protein were significantly increased (P<0.05) in inhibitor group and HRN group, while the above changes in agonist group were reversed (P<0.05). Compared with the agonist group, the pain threshold was significantly increased (P<0.05), the knee circumference was significantly decreased (P<0.05), the synovial pathological score was significantly decreased (P<0.05), the expressions of miR-155 mRNA and IL-1β, IL-17A, TLR4, NF-κB proteins in synovial tissue were significantly decreased (P<0.05), and the expression levels of SOCS1 mRNA and protein were significantly increased (P<0.05) in A+HRN group. The heat-reinforcing needling can increase the pain threshold, reduce the knee circumference and inhibit the inflammatory response in rabbits with RA cold syndrome. The possible mechanism is related to the regulation of miR-155/TLR4/NF-κB signaling axis.

Analysis of Regeneration Tissue Formed in the Treatment of Cartilage Defects

Joint function losses that occur as a result of injury, healing and scarring of the cartilage tissue are varied. In this study, the contralateral joints of rabbits with complete cartilage loss were defined as the control group. Self-healing of similar cartilage losses and popular surgical methods were compared, and the results were examined histopathologically. As a result, it was determined that while autogenous full-thickness osteochondral cartilage transfer resulted in the best cartilage healing, the healed cartilage tissue without any intervention was superior to methods such as drilling and carbon fiber. We concluded that cartilage tissue must be “no men land” if lower than full thickness cartilage lesions. Cartilage must be protected any surgical impact instead of main mechanical articular problems must be focused.

CHANGES IN THE CELLULAR COMPOSITION OF SYNOVIAL FLUID IN EXPERIMENTAL OSTEOARTHRITIS OF THE KNEE JOINT IN RABBITS

Pathology of the locomotion system in animals of various species is one of the key issues. Osteoarthritis is common among large breed dogs and accounts for up to 85% of the total number of examined animals with "chronic" lameness. German and Eastern European Shepherds are most susceptible to osteoarthritis, accounting for 25% and 15% of the total number of dogs examined, respectively. At the age of 4 to 8 years, 90 % of dogs with osteoarthritis were found. The prevalence of pathology in males was found to be 4 times higher than in females. The study presents the peculiarities of changes in the cellular composition of synovial fluid taken from the knee joint of rabbits with experimentally modelled osteoarthritis of the knee joint in laboratory conditions of the joint using intra-articular Yellow peel 2*5 ml (manufacturer Medicare, Germany). The studies showed that synovial fluid plays an important role in the metabolic process inside the joint and contains many nutrients, and changes in the indicators on the 7th day were significantly different from those of the control group of animals that were intra-articularly injected with an isotonic 0.9% NaCl solution. It was noted that on the 7th day there was a clouding of the synovial fluid, the appearance of a pinkish tint and a more liquid character of the fluid. We found an increase in the level of eosinophils, macrophages, neutrophils and other cells compared to the control group of animals. The number of synovial cells decreased by 1.2 times compared to the control group. In animals of the experimental group on the 14th day of the study, the level of lymphocytes and macrophages increased, and the level of neutrophils remained constant, while the level of eosinophils decreased. In the experimental group of animals, on the 21st day of the experimental study, the pink tint of the punctured synovial fluid disappears, the colour becomes straw, the fluid becomes more viscous and remains cloudy. The level of macrophages and neutrophils decreases, but remains above the level of the control group of animals. We also noted the appearance of Tuton cells in the cytological samples, indicating the development of degenerative changes in the cartilage tissue in the joint. It was noted that in the animals of the experimental group on the 28th day of the study, the colour of the synovial fluid disappeared, but it remained cloudy. The level of neutrophils, eosinophils, lymphocytes and macrophages remained almost unchanged.

R399E, A Mutated Form of Growth and Differentiation Factor 5, for Disease Modification of Osteoarthritis.

To preclinically characterize a mutant form of growth and differentiation factor 5, R399E, with reduced osteogenic properties as a potential disease-modifying osteoarthritis (OA) drug. Cartilage, synovium, and meniscus samples from patients with OA were used to evaluate anabolic and antiinflammatory properties of R399E. In the rabbit joint instability model, 65 rabbits underwent transection of the anterior cruciate ligament plus partial meniscectomy. Three intraarticular (IA) R399E doses were administered biweekly 6 times, and static incapacitance was determined to assess joint pain. OA was evaluated 13 weeks after surgery. In sheep, medial meniscus transection was performed to induce OA, dynamic weight bearing was measured in-life, and OA was assessed after 13 weeks. Intermittent exposure to R399E (1 week per month) was sufficient to induce cell proliferation and release of anabolic markers in 3-dimensional chondrocyte cultures. R399E also inhibited the release of interleukin-1β (IL-1β), IL-6, and prostaglandin E2 from cartilage with synovium, meniscal cell, and synoviocyte cultures. In rabbits, the mean difference (95% confidence interval [95% CI]) in weight bearing for R399E compared to vehicle was -5.8 (95% confidence interval [95% CI] -9.54, -2.15), -7.2 (95% CI -10.93, -3.54), and -7.7 (95% CI -11.49, -3.84) for the 0.6, 6, and 60 μg doses, respectively, 6 hours after the first IA injection, and was statistically significant through the entire study for all doses. Cartilage surface structure improved with the 6-μg dose. Structural and symptomatic improvement with the same dose was confirmed in the sheep model of OA. R399E influences several pathologic processes contributing to OA, highlighting its potential as a disease-modifying therapy.

Effect of heat-reinforcing needling on expression of P2X7R, NLRP3 and Caspase-1 in synovial tissues of knee joint in rabbits with cold syndrome rheumatoid arthritis

To observe the effects of heat-reinforcing needling on the expression of purinergic ligand-gated ion channel 7 receptor(P2X7R)，Nodlike receptor protein 3(NLRP3) and Caspase-1 in synovium tissues of knee joint of rabbits with cold syndrome rheumatoid arthritis(RA)，so as to explore the anti-inflammatory mechanism of heat reinforcing needling in the treatment of RA. Thirty male New Zealand white rabbits were randomly divided into normal group, model group, reinforcing-reducing needling group(RRG), heat-reinforcing needling group(HRG), and antagonist group(AG), with 6 rabbits in each group.The model of cold syndrome RA was established by ovalbumin combined with Freund's adjuvant and cryogenic freezing. Rabbits of RRG and HRG were treated with corresponding acupuncture techniques on both sides of "Zusanli"(ST36) for 30 min, once a day for 7 days; Rabbits of the AG was intraperitoneally injected with A438079(2.5 mg/kg), once a day for 7 days. After intervention, color Doppler ultrasound was used to observe the joint cavity effusion, synovial thickness and internal blood flow signal.The histomorphological changes of synovial tissues were observed by HE staining. Quantitative real-time PCR method was used to detect the mRNA expression levels of P2X7R, NLRP3, and Caspase-1 in synovial tissues. Western blot was used to detect the protein expression of interleukin(IL)-1β, IL-6 and tumor necrosis factor(TNF)-α in synovial tissues. Compared with the normal group, the synovial tissues in the model group was thickened, linear and punctate blood flow signals were increased, joint effusion was obvious, synovial coating cells were enlarged, the synovial matrix was severely hyperplasia, inflammatory cells infiltration was obvious.The mRNAs expression levels of P2X7R, NLRP3, Caspase-1 and IL-1β, IL-6, TNF-α proteins in synovial tissues were significantly increased(P<0.01). Compared with model group, abnormal blood flow signals, synovial thickness, joint effusion, proliferation of synovial matrix, inflammatory cell infiltration and synovial coating cells in the RRG, HRG and AG were improved. The mRNAs expression levels of P2X7R, NLRP3, Caspase-1 and IL-1β, IL-6, TNF-α proteins in synovial tissues were decreased in the RRG, HRG and AG (P<0.05, P<0.01). Compared with the RRG, the above indexes were lower in the HRG and AG (P<0.05, P<0.01).There was no significant difference in all indexes above between the HRG and AG (P<0.01, P<0.05). Heat reinforcing needling can improve synovial inflammation of RA, which may be related to regulating the expressions of P2X7R/NLRP3/Caspase-1 signaling pathway.