Anterior Cruciate Ligament Transection Group Research Articles

The infrapatellar fat pad contributes to spontaneous healing after complete anterior cruciate ligament injury.

Anterior cruciate ligament (ACL) injuries have a very low healing capacity but have recently been shown to heal spontaneously with conservative treatment. This study examined the mechanism of spontaneous ACL healing by focusing on the intra-articular tissues of the knee joint. Skeletally mature Wistar rats (n = 70) were randomly assigned to two groups: the controlled abnormal movement (CAM) and anterior cruciate ligament transection (ACLT) groups. The ACL was completely transected at the mid-portion in both groups. Only the CAM group underwent extra-articular braking to control for abnormal tibial translation. The animals were allowed full cage activity until sacrifice for histological, and molecular biology analyses. The results showed that the behavior of the stump after ACL injury differed between models 12 h after injury. The femoral stump in the ACLT group retreated posteriorly and upwardly. Macrophage polarity analysis revealed that the stump immune response in the CAM group was more activated than that in the ACLT group 6 h after injury. Microarray analysis of the ACL parenchyma and infrapatellar fat pads suggested the involvement of nuclear factor kappa B (NF-κB) signaling. Real-time polymerase chain reaction (PCR) analysis showed that NF-κB gene expression in the infrapatellar fat pad was significantly increased in the CAM group than in the ACLT group. However, there was no difference in the gene expression levels in the ACL parenchyma between models. In conclusion, the healing response of the ACL was activated within 12 h of injury, resulting in differences in the healing response between the models. It has been suggested that infrapatellar fat pads are involved in the healing process and that angiogenesis and antiapoptotic effects through NF-κB signaling may contribute to this mechanism.

Combined detection of serum CTX-II and C2C in a rat model of ACLT-induced osteoarthritis.

Osteoarthritis (OA) is a chronic degenerative bone and joint disease that often occurs in aging animals. Currently, there are still no biomarkers that can effectively diagnose OA in the early stage. To identify possible biomarkers, here we examined changes in the expression of C-telopeptide fragments of type II collagen (CTX-II) and collagenase generated carboxy-terminal neoepitope of type II collagen (C2C) in serum at different time points in an anterior cruciate ligament transection (ACLT)-induced rat OA model. The serum levels of CTX-II and C2C, and the OARSI score in the ACLT group were increased from week two until the end of the experiment. The AUC of the combined biomarkers was higher than that of CTX-II or C2C alone. Moreover, serum levels of CTX-II and C2C were positively correlated with the OARSI score. The results suggest that the combined detection of serum CTX-II and C2C concentrations may have potential for assessing and diagnosing OA at early stages.

Axial Compressive Loading Attenuates Early Osteoarthritis by Reducing Subchondral Bone Remodeling

Background: Mechanical loading and alendronate (ALN) can be used as noninvasive physical therapy methods for osteoarthritis (OA). However, the timing and efficacy for treatments are unknown. Purpose: To determine whether the timing of mechanical loading and ALN influences the pathobiological changes of OA. Study Design: Controlled laboratory study. Methods: Mice with OA induced by anterior cruciate ligament transection were subjected to early (1-3 weeks) or late (5-7 weeks) axial compressive dynamic load or intraperitoneal injection of ALN. Changes in gait were analyzed using gait analysis system, pathobiological changes in subchondral bone, cartilage, osteophyte, and synovitis were assessed using micro–computed tomography, tartrate-resistant acid phosphatase staining, pathologic section staining, and immunohistochemistry at 1, 2, 4, and 8 weeks. Results: At 1, 2, and 4 weeks, the OA limb had lower mean footprint pressure intensity, lower bone volume per tissue volume (BV/TV) in the subchondral bone, and more osteoclasts. At 4 weeks, the early loading, ALN, and load + ALN treatments induced less cartilage destruction, with a corresponding reduction in Osteoarthritis Research Society International score and increased hyaline cartilage thickness. The treatments also resulted in fewer osteoclasts and higher BV/TV and bone mineral density of subchondral bone and suppressed inflammation and interleukin 1β– and tumor necrosis factor α-positive cells in synovium. At 8 weeks, early loading or load + ALN improved the mean footprint pressure intensity and knee flexion. At 8 weeks, early load + ALN had a synergistic effect on protecting hyaline cartilage and proteoglycans. Footprint pressure intensity and cartilage destruction were worse in late loading limbs, and no differences in BV/TV, bone mineral density, osteophyte formation, and synovium inflammation were found between the late load, ALN, and load + ALN groups and the anterior cruciate ligament transection group. Conclusion: Dynamic axial mechanical loading or ALN in the early stages of knee trauma protected against OA by suppressing subchondral bone remodeling. However, late loading promoted cartilage degeneration in advanced OA, indicating that reduced loading should be performed in the late stages of OA to avoid the acceleration of OA. Clinical Relevance: Early low-level functional exercise or antiosteoporotic drugs could clearly slow or prevent the progression of early OA. For patients with mild to severe OA, loading reduction via brace protection or maintenance of joint stability via early ligament reconstruction surgery may ameliorate OA exacerbation.

Quantitative evaluation of very early cartilage damage in experimental osteoarthritis using scanning electron microscopy

ObjectiveTo describe parametric changes observed using scanning electron microscopy (SEM) in very early stages in posttraumatic osteoarthritis (OA) models in mice.MethodsMice (5/group) had their knees subjected to anterior cruciate ligament transection (ACLT), ACLT plus meniscectomy (MNCT) or sham surgery, sacrificed after 3, 7 or 14 days, had the articular cartilage evaluated under optical microscopy using Osteoarthritis Research Society International (OARSI) parameters as well as cartilage thickness, roughness, and a damage index using SEM.ResultsAlterations of the cartilage under optical microscopy were not significantly relevant among groups. SEM analysis revealed reduction of femoral and tibial cartilage thickness in ACLT and MNCT groups at 7 and 14 days, with increased cartilage roughness in MNCT group as early as 3 days postsurgery, being sustained up to 14 days. Articular damage index was significantly higher at 14 days post surgery in ACLT and MNCT vs control groups.ConclusionThis is the first demonstration of very early quantitative changes in the cartilage of mice subjected to posttraumatic experimental OA using SEM, revealing increased roughness and thickness as early as 3 days post surgery. These changes may be used as early surrogates for later joint damage in experimental OA.

The Traditional Chinese Medicine "Hu-Qian-Wan" Attenuates Osteoarthritis-Induced Signs and Symptoms in an Experimental Rat Model of Knee Osteoarthritis.

Background Knee osteoarthritis (KOA) is a chronic, low-grade inflammatory disease that affects knee joints and causes functional disability in the elderly. KOA is typically treated with oral NSAIDs, which are commonly associated with gastrointestinal side effects or cardiovascular complications. Traditional Chinese medicine (TCM) is widely used by patients with KOA in Taiwan; the Hu-Qian-Wan (HQW) formula is typically prescribed. We investigated the therapeutic role of a modified version of the HQW decoction in Sprague-Dawley rats with KOA induced by anterior cruciate ligament transection (ACLT) of the right knee. Materials and Methods Thirty rats were randomly assigned to five groups (six animals each): arthrotomy alone (sham surgery, controls), ACLT only, ACLT + low-dose (1,000 mg/kg) HQW, ACLT + high-dose (3,000 mg/kg) HQW, and ACLT + celecoxib (30 mg/kg). All study groups underwent weight-bearing behavioral testing, micro-computed tomography (CT), and histological examinations of the knee joint cartilage, as well as immunohistochemical analyses of levels of interleukin (IL) 1β and tumor necrosis factor (TNF) α expression in articular cartilage. Results At 6 weeks, compared with ACLT group only, ACLT rats administered high-dose HQW or celecoxib exhibited the fewest weight-bearing deficits, the greatest improvements from baseline in articular cartilage architecture, and the lowest amounts of TNF-α and IL-1β staining in cartilage and synovial sections (all values were significant compared with the ACLT-only group). The only values that were significantly increased by ACLT + low-dose HQW compared with ACLT alone were bone mineral density and trabecular numbers. Conclusion Our findings suggest that high-dose HQW improves weight-bearing asymmetry, decreases bone loss, and reduces levels of TNF-α and IL-1β in the affected joint in ACLT-induced KOA rats. More evidence is needed to support our findings.

Assessment of osteoarthritis functional outcomes and intra-articular injection volume in the rat anterior cruciate ligament transection model.

The rat surgical anterior cruciate ligament transection (ACLT) model is commonly used to investigate intra-articular osteoarthritis (OA) therapies,and histological assessment is often the primary outcome measure. However, histological changes do not always correlate well with clinical outcomes. Therefore, this study evaluated functional outcomes in the rat surgical ACLT model and compared intra-articular injection volumes ranging from 20 to 50 μl. Unilateral ACLT was surgically inducedand static weight-bearing, mechanical allodynia, motor function, and gait were assessed in four groups of male, Sprague-Dawley rats (n = 6 per group). Intra-articular injections of 20 µl Dulbecco's phosphate-buffered saline (DPBS), 50 µl DPBS, or 50 µl of synthetic biomimetic boundary lubricant were administered once weekly for 3 weeks postoperatively. Structural changes were evaluated histologically at 20 weeks. Rat cadaver knees were injected with 20, 30, 40, or 50 µl of gadolinium solutions and were imaged using magnetic resonance imaging (MRI). Static weight-bearing, mechanical allodynia, and gait parameters in ACLT groups revealed differences from baseline and naïve controls for 4 weeks post-ACLT; however, these differences did not persist beyond 6 weeks. Different intra-articular DPBS injection volumes did not result in functional or histological changes; however, peri-articular leakage was documented via MRI following 50, 40, and 30 µl but not 20 µl gadolinium injections.Statement of clinical significance:Differences in functional parameters were predominantly restricted to early, postoperative changes in the rat surgical ACLT model despite evidence of moderate histologic OA at 20 weeks. Injection volumes of 20-30 µl are more appropriate for investigating intra-articular therapies in the rat knee.

The difference in joint instability affects the onset of cartilage degeneration or subchondral bone changes

It has been debated whether the onset of knee osteoarthritis is initiated in cartilage or subchondral bone. The purpose of this study was to clarify the effects of increasing or decreasing joint instability on cartilage degeneration and subchondral bone changes in knee OA by comparing different models of joint instability. We used the anterior cruciate ligament transection (ACL-T) model and the destabilization of the medial meniscus (DMM) model. In addition, we created a controlled abnormal tibial translation (CATT) model and a controlled abnormal tibial rotation (CATR) model. We performed joint instability analysis, micro-computed tomography analysis, histological and immunohistological analysis in 4 and 6 weeks. The CATT group suppressed joint instability in the ACL-T group (6 weeks; P=0.032), and the CATR group suppressed joint instability in the DMM group (6 weeks; P=0.032). Chondrocyte hypertrophy in the ACL-T and DMM groups was increased compared to the Sham group (6 weeks; [ACL-T vs Sham], P=0.002, 95%CI [5.983-33.025]; [DMM vs Sham], P=0.022, 95%CI [1.691-28.733]). In the subchondral bone, the BV/TV in the DMM and CATR groups was increased compared to the ACL-T and CATT groups (6 weeks; [DMM vs ACL-T], P=0.002, 95%CI [7.404-37.582]; [DMM vs CATT], P=0.014, 95%CI [2.881-33.059]; [CATR vs ACL-T], P=0.006, 95%CI [4.615-34.793]; [CATR vs CATT], P=0.048, 95%CI [0.092-30.270]). This study showed that joint instability promotes chondrocyte hypertrophy, but subchondral bone changes were influenced by differences in ACL and meniscus function.

Comparison of bone and articular cartilage changes in osteoarthritis: a micro-computed tomography and histological study of surgically and chemically induced osteoarthritic rabbit models

BackgroundOsteoarthritis (OA) is a multifaceted condition that affects both the subchondral bones and the articular cartilage. Animal models are widely used as an effective supplement and simulation for human OA studies in investigating disease mechanisms and pathophysiology. This study is aimed to evaluate the temporal changes of bone and cartilage in surgically and chemically induced osteoarthritis using micro-computed tomography and histology.MethodsThirty rabbits underwent either anterior cruciate ligament transection (ACLT) procedure or injected intraarticularly with monosodium iodoacetate (MIA, 8 mg) at the right knee joint. The subchondral bones were scanned via micro-CT, and articular cartilage was assessed histologically at 4-, 8- and 12-week post-induction.ResultsBased on bone micro-architecture parameters, the surgically induced group revealed bone remodelling processes, indicated by increase bone volume, thickening of trabeculae, reduced trabecular separation and reduced porosity. On the other hand, the chemically induced group showed active bone resorption processes depicted by decrease bone volume, thinning of trabeculae, increased separation of trabecular and increased porosity consistently until week 12. Histologically, the chemically induced group showed more severe articular cartilage damage compared to the surgically induced group.ConclusionsIt can be concluded that in the ACLT group, subchondral bone remodelling precedes articular cartilage damage and vice versa in the MIA group. The findings revealed distinct pathogenic pathways for both induction methods, providing insight into tailored therapeutic strategies, as well as disease progression and treatment outcomes monitoring.

Treadmill Exercise after Controlled Abnormal Joint Movement Inhibits Cartilage Degeneration and Synovitis.

Cartilage degeneration is the main pathological component of knee osteoarthritis (OA), but no effective treatment for its control exists. Although exercise can inhibit OA, the abnormal joint movement with knee OA must be managed to perform exercise. Our aims were to determine how controlling abnormal joint movement and treadmill exercise can suppress cartilage degeneration, to analyze the tissues surrounding articular cartilage, and to clarify the effect of treatment. Twelve-week-old ICR mice (n = 24) underwent anterior cruciate ligament transection (ACL-T) surgery on their right knees and were divided into three groups as follows: ACL-T, animals in the walking group subjected to ACL-T; controlled abnormal joint movement (CAJM), and CAJM with exercise (CAJM + Ex) (n = 8/group). Walking-group animals were subjected to treadmill exercise 6 weeks after surgery, including walking for 18 m/min, 30 min/day, 3 days/week for 8 weeks. Safranin-O staining, hematoxylin-eosin staining, and immunohistochemical staining were performed. The OARSI (Osteoarthritis research Society international) score was lower in the CAJM group than in the ACL-T group and was even lower in the CAJM + Ex group. The CAJM group had a lower meniscal injury score than the ACL-T group, and the CAJM + Ex group demonstrated a less severe synovitis than the ACL-T and CAJM groups. The observed difference in the perichondrium tissue damage score depending on the intervention method suggests different therapeutic effects, that normalizing joint motion can solve local problems in the knee joint, and that the anti-inflammatory effect of treadmill exercise can suppress cartilage degeneration.

Lingzhi and San-Miao-San with hyaluronic acid gel mitigate cartilage degeneration in anterior cruciate ligament transection induced osteoarthritis.

SummaryObjectiveTo investigate the mitigate efficacy of Chinese medicine Lingzhi (LZ) and San-Miao-San (SMS) combined with hyaluronic acid (HA)-gel in attenuating cartilage degeneration in traumatic osteoarthritis (OA).MethodsThe standardized surgery of anterior cruciate ligament transection (ACLT) was made from the medial compartment of right hind limbs of 8-week-old female SD rats and resulted in a traumatic OA. Rats (n ​= ​5/group) were treated once intra-articular injection of 50 ​μl HA-gel, 50 ​μl HA-gel+50 ​μg LZ-SMS, 50 ​μl of saline+50 ​μg LZ-SMS and null (ACLT group) respectively, except sham group. Limbs were harvested for μCT scan and histopathological staining 3-month post-treatment. Inflammatory cytokines from plasma and synovial fluid were detected using Immunology Multiplex Assay kit. The putative targets of active compounds in LZ-SMS and known therapeutic targets for OA were combined to construct protein–protein interaction network. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis was adopted to predict the potential targets and signaling pathway of LZ-SMS in OA through the tool of DAVID Bioinformatics.ResultsIn vivo, HA-gel ​+ ​LZ-SMS treatment resulted in a higher volume ratio of hyaline cartilage (HC)/calcified cartilage (CC) and HC/Sum (total volume of cartilage), compared to ACLT and HA-gel groups. In addition, histological results showed the elevated cartilage matrix, chondrogenic and osteoblastic signals in HA-gel ​+ ​LZ-SMS treatment. Treatment also significantly altered subchondral bone (SCB) structure including an increase in BV/TV, Tb.Th, BMD, Conn.Dn, Tb.N, and DA, as well as a significant decrease in Tb.Sp and Po(tot), which implied a protective effect on maintaining the stabilization of tibial SCB microstructure. Furthermore, there was also a down-regulated inflammatory cytokines and upregulated anti-inflammatory cytokine IL-10 in HA+LZ-SMS group. Finally, 64 shared targets from 37 active compounds in LZ-SMS related to the core genes for the development of OA. LZ-SMS has a putative role in regulating inflammatory circumstance through influencing the MAPK signaling pathway.ConclusionOur study elucidated a protective effect of HA-gel ​+ ​LZ-SMS in mitigating cartilage degradation and putative interaction with targets and signaling pathway for the development of traumatic OA.The translational potential of this articleOur results provide a biological rationale for the use of LZ-SMS as a potential candidate for OA treatment.

Discoidin Domain Receptors 1 Inhibition Alleviates Osteoarthritis via Enhancing Autophagy.

We recently reported that the chondrocyte-specific knockout of discoidin domain receptors 1 (Ddr1) delayed endochondral ossification (EO) in the growth plate by reducing the chondrocyte hypertrophic terminal differentiation, and apoptosis. The biologic and phenotypic changes in chondrocytes in the articular cartilage with osteoarthritis (OA) are similar to the phenomena observed in the process of EO. Additionally, autophagy can promote chondrocyte survival and prevent articular cartilage from degradation in OA. On this basis, we explored the effect of Ddr1 inhibition on OA prevention and further investigated the roles of autophagy in treating OA with a Ddr1 inhibitor (7 rh). The anterior cruciate ligament transection (ACLT)–OA model was used to investigate the role of 7 rh in vivo. Forty 8-week-old mice were randomly assigned to four groups, including the sham group, ACLT group, and two treated groups (ACLT with 7 rh 6.9 nM or 13.8 nM). According to the study design, normal saline or 7 rh were intra-articular (IA) injected into studied knees 3 times per week for 2 weeks and then once per week for 4 weeks. The results showed that 7 rh treatment significantly improved the functional performances (the weight-bearing ability and the running endurance), decreased cartilage degradation, and also reduced the terminal differentiation markers (collagen type X, Indian hedgehog, and matrix metalloproteinase 13). Moreover, 7 rh decreased chondrocyte apoptosis by regulating chondrocyte autophagy through reducing the expression of the mammalian target of rapamycin and enhancing the light chain 3 and beclin-1 expression. These results demonstrated that the IA injection of 7 rh could reduce the chondrocyte apoptosis and promote chondrocyte autophagy, leading to the attenuation of cartilage degradation. Our observations suggested that the IA injection of 7 rh could represent a potential disease-modifying therapy to prevention OA progression.

MicroRNA‑1 regulates the development of osteoarthritis in a Col2a1‑Cre‑ERT2/GFPfl/fl‑RFP‑miR‑1mouse model of osteoarthritis through the downregulation of Indian hedgehog expression.

The present study assessed the effects of microRNA-1 (miR-1) on the development of osteoarthritis using human tissues and a Col2a1-Cre-ERT2/GFPfl/fl-RFP-miR-1 mouse model of osteoarthritis. Human cartilage tissues (n=20) were collected for reverse transcription-quantitative polymerase chain reaction (RT-qPCR), histological analysis and immunohistochemistry experiments. A transgenic mouse model of osteoarthritis was established by subjecting Col2a1-Cre-ERT2/GFPfl/fl-RFP-miR-1 transgenic mice to anterior cruciate ligament transection (ACLT). Mice were subjected to radiography and in vivo fluorescence molecular tomography (FMT), while mouse tissues were collected for histological analysis, RT-qPCR and Safranin O staining. It was found that the miR-1 level was downregulated, whereas the levels of Indian hedgehog (Ihh), as well as those of its downstream genes were upregulated in human osteoarthritic cartilage. In the transgenic mice, treatment with tamoxifen induced miR-1, as well as collagen, type II (Col2a1) and Aggrecan (Acan) expression; however, it decreased Ihh, glioma-associated oncogene homolog (Gli)1, Gli2, Gli3, smoothened homolog (Smo), matrix metalloproteinase (MMP)-13 and collagen type X (Col10) expression. Safranin O staining revealed cartilage surface damage in the non-tamoxifen + ACLT group, compared with that in the tamoxifen + ACLT group. Histologically, an intact cartilage surface and less fibrosis were observed in the tamoxifen + ACLT group. Immunohistochemistry revealed that the protein expression of Ihh, Col10, and MMP-13 was significantly higher in the joint tissues of the non-tamoxifen + ACLT group than in those of the tamoxifen + ACLT group. However, Col2a1 expression was lower in the joint tissues of the non-tamoxifen + ACLT group than in those of the tamoxifen + ACLT group. The results of RT-qPCR and FMT further confirmed these findings. On the whole, the findings of the present study demonstrate that miR-1 expression protects against osteoarthritis-induced cartilage damage and gene expression by inhibiting Ihh signaling.

Salidroside Alleviates Cartilage Degeneration Through NF-κB Pathway in Osteoarthritis Rats.

IntroductionOsteoarthritis (OA) is the most common disease, which seriously affects the daily life of the elderly. Currently, no traditional or drug therapy has been shown to explicitly block the progression of OA. Salidroside (Sal) is a bioactive component of Rhodiola rosea, which has many beneficial effects on human health. However, the role and mechanism of Sal in OA have not been reported.MethodsWe established an anterior cruciate ligament transection (ACLT)-induced OA Rat model. The rats were divided into five groups (n = 10): Control group; ACLT group; ACLT + Sal (12.5 mg/kg) group; ACLT + Sal (25 mg/kg) group; ACLT + Sal (50 mg/kg) group.ResultsThe study showed that Sal could significantly promote the proliferation of chondrocytes in OA rats induced by ACLT and restore the histological alteration of cartilage. Besides, Sal upregulated the levels of Collagen II and Aggrecan, and downregulated the level of MMP-13. Furthermore, Sal could reduce the number of CD4+IL-17+ cells and decrease the levels of IL-17, IKBα and p65, while elevating the number of CD4+IL-10+ cells and the level of IL-10. The decrease of IL-17 further inhibited the dissociation of IKBα to p65, thus reducing the release of TNF-α and VCAM-1. Taken together, Sal alleviates cartilage degeneration through promoting chondrocytes proliferation, inhibiting collagen fibrosis, and regulating inflammation and immune responses via NF-κB pathway in ACLT-induced OA Rats.DiscussionCollectively, our study investigates the role and mechanism of Sal in OA, which lays a foundation for the application of Sal in OA.

Synovial inflammation in anterior cruciate ligament injury knees in mice: surgical vs non-surgical models

Purpose: Anterior-cruciate-ligament (ACL) rupture is one of the most common knee injuries, resulting in pain, disability, and increased risk of developing OA. However, only ∼50% of individuals with ACL rupture develop OA, and the factors underlying this variable risk remain unclear. Changes in joint stability as a result of ACL deficiency drive mechanobiological dysfunction and the development of OA. However, specific factors are known to increase OA risk such as concurrent injury to the meniscus, cartilage, and subchondral bone. Also, the degree of initial joint inflammation and its resolution are implicated in long-term outcomes. Surgical and nonsurgical models of ACL rupture in mice result in similar joint instability but distinct risks or trajectories of OA. We used these two models to investigate if differences in synovial inflammatory response are associated with this risk of OA disease progression Methods: Ten-week-old, male C57BL/6 mice were randomly assigned to ACL transection (ACLT), ACL rupture (ACLR), sham-surgery or uninjured groups. Mice in the ACLT and sham-operated groups were subjected to anaesthesia, right unilateral medial arthrotomy, patella luxation, and fat-pad elevation to visualise the ACL, at which time the surgeon was informed to transect or spare the ACL before joint closure. Uninjured and ACLR animals were anaesthetised, the latter having a single compressive load (9-12N) to posteriorly displace the femur and traumatically rupture the ACL. Animals were housed in mixed injury groups (up to 5/cage) and allowed unrestricted activity. At 1, 2, 4 and 8 weeks post-injury, mice were euthanised and joints harvested for fixation, paraffin embedding, sectioning and OA histopathology scoring (n=7/group/timepoint); or synovial tissue (en bloc fat-pad, synovial lining and joint capsule anterior to the collateral ligaments) isolation for RNA extraction and quantitative RT-PCR analysis of cytokines (IL1, IL6, TNF), inflammatory cell markers (CD11b, CD4, CD8), and enzymes implicated in OA pathophysiology (ADAMTS4, ADAMTS5, MMP3, MMP9, MMP13) (n=6/group/time point). Results: ACL injury resulted in progressive OA pathology with cartilage pathology significantly greater in ACLR compared with ACLT at 2 and 4 weeks (p<0.001), but by 8 weeks was equivalent in the two models; posterior osteochondral erosion significantly worse in ACLR than ACLT at 4 and 8 weeks (p<0.01); and osteophyte size (p<0.001) and maturity (p<0.01) greater at 1 and 2 weels in ACLR compared with ACLT. Synovitis was increased acutely in both ACL-injuries and then decreased with time, with ACLT>ACLR (p=0.003) at week-1 but similar thereafter; both still greater than uninjured joints through to week 8 (p<0.001). Sham and ACLT had similar synovitis scores until week 8 when the latter was more severe. Despite histomorphological similarities in synovitis in ACLT and ACLR, there were differences in gene expression patterns. Synovial expression of TNF, CD11b and CD8 was unchanged at any time when comparing between groups. Surgery alone (ACLT and Sham) increased synovial IL6 (week-1: ACLT p=0.02, Sham p<0.001 vs uninjured) while ACLR did not. While CD4 was also increased by surgery (week-1 and -2: ACLT p<0.01, Sham p<0.05 vs uninjured), it was also increased in ACLR versus uninjured at these times (p<0.05; Figure 1). ADAMTS5 expression was not detected in synovial tissue, and MMP9 mRNA did not differ between groups. In contrast, ADAMTS4, MMP3, and MMP13 expression were increased by surgery (ACLT/Sham) and ACLR at week-1 and -2 compared with uninjured joint, although both surgeries>ACLR (Table 1). Conclusions: While both models caused joint instability, OA trajectory and ultimate severity were greater in ACLR. In contrast, surgery induced greater histo-pathological synovitis acutely but was similar in both injuries thereafter. There was a difference in the “phenotype” of the synovial inflammation in the two injury models. In ACLT there was upregulation of IL6 not evident in ACLR but similar to Sham, suggesting this was a response to surgery and wound healing. While helper T-cell (CD4) increase was greater in ACLT and Sham, this also occurred in ACLR suggesting a role of adaptive immunity in OA after nonsurgical injury. Expression of ADAMTS4, MMP3 and MMP13 were similarly increased acutely in both injury models, suggesting synovial tissues as a source of these enzymes implicated in the structural pathology of OA. While their expression was not correlated with ultimate pathological severity they may be broad-spectrum target across varying phenotypes of post-traumatic OA. Our data suggest that synovitis severity per se was not associated with differential OA risk with ACL injury; it likely still contributes to long-term OA structural pathology.View Large Image Figure ViewerDownload Hi-res image Download (PPT)

Comparison between two rabbit models of posttraumatic osteoarthritis: A longitudinal tear in the medial meniscus and anterior cruciate ligament transection

Animal osteoarthritis (OA) models have been developed to understand OA progression and evaluate new OA therapies. However, individual variations in joint lesions remain a critical problem in most current OA models. We established a novel rabbit model by creating a longitudinal tear in the medial meniscus body that was reproducible and similar to posttraumatic biomechanical disturbances in human OA. New Zealand rabbits underwent surgery and were assessed for 9 weeks. The rabbits were randomized into the sham control, medial meniscal tear (MMT), and anterior cruciate ligament transection (ACLT) groups. The animals were sacrificed at 4, 6, and 9 weeks posttreatment. The knee joints were harvested for histological and gene expression assessments. Both the MMT and ACLT procedures led to time-dependent degenerative changes in the femoral condyle cartilage. At each time point, the MMT group cartilage showed more severe degenerative changes than did the ACLT group cartilage. Consistently, inflammatory cytokine and catabolic gene expression were significantly higher, and anabolic gene expression was significantly lower in the MMT group than in the ACLT group. MMT treatment caused more severe structural damage to the cartilage and higher catabolic gene expression levels than the ACLT model at each time point. The MMT model may be highly beneficial in investigating posttraumatic OA (PTOA) development, especially PTOA from a meniscal injury. The MMT model replicated key features of human PTOA, including meniscal lesions, inflammatory responses, and the progression to osteoarthritic cartilage degeneration, thereby providing an exciting new avenue for translating promising treatments to clinical practice.

Prevent action of magnoflorine with hyaluronic acid gel from cartilage degeneration in anterior cruciate ligament transection induced osteoarthritis

According to the Chinese medicine, magnoflorine exerted significant anti-inflammatory effects and potentially promoted synthesis of proteoglycans in chondrocytes to reverse the progression of rheumatoid arthritis. However, the latent beneficial effect of magnoflorine for the treatment of traumatic osteoarthritis (OA) is still unknown. Therefore, we aim to demonstrate the efficacy of magnoflorine combined with HA-gel in attenuating cartilage degeneration in anterior cruciate ligament transection (ACLT) induced OA rat model. We found that the histological results showed the elevated cartilage matrix, chondrogenic signals and chondroprogenitor cells in HA-gel + magnoflorine treatment. HA-gel + magnoflorine treatment resulted in a decreased modified Mankin’s score, and a higher volume ratio of hyaline cartilage (HC)/calcified cartilage (CC) and HC/Sum (whole cartilage), compared to ACLT and HA-gel groups. Furthermore, both the volume ratios of HC/Sum and HC/CC were negatively correlated with modified Mankin’s scores. Finally, HA-gel + magnoflorine could significantly increase the BV/TV, Tb.Th, and decrease the Tb.Pf, Po(tot), Conn.Dn and Tb.Sp. In vitro, 50 μg/ml magnoflorine treatment could significantly increase the viability, S-phase, migration rate and chondrogenesis of chondroprogenitor cells. There were significant downregulations of MAPK/NF-κB signaling, and upregulations of chondrogenic signals in 50 μg/ml magnoflorine treatment. There were significant downregulations of proinflammatory cytokines and upregulation of IL-10 in HA-gel + magnoflorine treated group. Therefore, our study elucidated a protective effect of HA-gel + magnoflorine on attenuating cartilage degradation and maintaining SCB stabilization in ACLT induced OA.

The COX-2 inhibitor etoricoxib reduces experimental osteoarthritis and nociception in rats: The roles of TGF-β1 and NGF expressions in chondrocytes.

Osteoarthritis (OA) is the most common joint disease, especially affecting the knee joint. Etoricoxib, a highly selective cyclooxygenase (COX)-2 inhibitor which can reduce postoperative pain after orthopaedic surgery. The aim of this study was to investigate the effects of oral etoricoxib on the development of OA and to examine concomitant changes in the nociceptive behaviour of rats. OA was induced in wistar rats by anterior cruciate ligament transection (ACLT) of the right knee. The ACLT+etoricoxib groups received 6.7 or 33.3mg/kg of oral etoricoxib three times a week for 12 consecutive weeks, starting at week 8 after ACLT. Nociceptive behaviours and changes in knee joint width during OA development were analyzed. Histopathological studies were then performed on the cartilage. Immunohistochemical analysis was performed to examine the effect of etoricoxib on the expression of transforming growth factor-beta (TGF-β) and nerve growth factor (NGF) in articular cartilage chondrocytes. OA rats receiving etoricoxib showed a significantly lower degree of cartilage degeneration than the rats receiving placebo. Nociceptive behaviour studies showed significant improvement in the ACLT+etoricoxib groups compared to that in the ACLT group. Moreover, etoricoxib attenuated NGF expression, but increased TGF-β expression, in OA-affected cartilage. Oral etoricoxib in a rat OA model (a) attenuates the development of OA, (b) concomitantly reduces nociception, and (c) modulates chondrocyte metabolism, possibly by inhibiting NGF expression and increasing TGF-β expression. Oral administration of etoricoxib can attenuate the development of OA, with an associated attenuation of nociceptive behaviour in an experimental rat OA model. Moreover, etoricoxib attenuated NGF expression, but enhanced TGF-β expression in OA-affected chondrocytes. These findings may pave the way for further investigations of etoricoxib as a potential therapeutic target for the treatment of the inflammatory component in OA.

Early Alterations of Subchondral Bone in the Rat Anterior Cruciate Ligament Transection Model of Osteoarthritis.

Advances in research have shown that the subchondral bone plays an important role in the propagation of cartilage loss and progression of osteoarthritis (OA), but whether the subchondral bone changes precede or lead to articular cartilage loss remains debatable. In order to elucidate the subchondral bone and cartilage changes that occur in early OA, an experiment using anterior cruciate ligament transection (ACLT) induced posttraumatic OA model of the rat knee was conducted. Forty-two Sprague Dawley rats were divided into 2 groups: the ACLT group and the nonoperated control group. Surgery was conducted on the ACLT group, and subsequently rats from both groups were sacrificed at 1, 2, and 3 weeks postsurgery. Subchondral bone was evaluated using a high-resolution peripheral quantitative computed tomography scanner, while cartilage was histologically evaluated and scored. A significant reduction in the subchondral trabecular bone thickness and spacing was found as early as 1 week postsurgery in ACLT rats compared with the nonoperated control. This was subsequently followed by a reduction in bone mineral density and bone fractional volume at week 2, and finally a decrease in the trabecular number at week 3. These changes occurred together with cartilage degeneration as reflected by an increasing Mankin score over all 3 weeks. Significant changes in subchondral bone occur very early in OA concurrent with surface articular cartilage degenerative change suggest that factors affecting bone remodeling and resorption together with cartilage matrix degradation occur very early in the disease.

Anterior cruciate ligament transection alters the n-3/n-6 fatty acid balance in the lapine infrapatellar fat pad

BackgroundThe infrapatellar fat pad (IFP) of the knee joint has received lots of attention recently due to its emerging role in the pathogenesis of osteoarthritis (OA), where it displays an inflammatory phenotype. The aim of the present study was to examine the infrapatellar fatty acid (FA) composition in a rabbit (Oryctolagus cuniculus) model of early OA created by anterior cruciate ligament transection (ACLT).MethodsOA was induced randomly in the left or right knee joint of skeletally mature New Zealand White rabbits by ACLT, while the contralateral knee was left intact. A separate group of unoperated rabbits served as controls. The IFP of the ACLT, contralateral, and control knees were harvested following euthanasia 2 or 8 weeks post-ACLT and their FA composition was determined with gas chromatography–mass spectrometry.ResultsThe n-3/n-6 polyunsaturated FA (PUFA) ratio shifted in a pro-inflammatory direction after ACLT, already observed 2 weeks after the operation (0.20 ± 0.008 vs. 0.18 ± 0.009). At 8 weeks, the FA profile of the ACLT group was characterized with increased percentages of 20:4n-6 (0.44 ± 0.064 vs. 0.98 ± 0.339 mol-%) and 22:6n-3 (0.03 ± 0.014 vs. 0.07 ± 0.015 mol-%) and with decreased monounsaturated FA (MUFA) sums (37.19 ± 1.586 vs. 33.20 ± 1.068 mol-%) and n-3/n-6 PUFA ratios (0.20 ± 0.008 vs. 0.17 ± 0.008). The FA signature of the contralateral knees resembled that of the unoperated controls in most aspects, but had increased proportions of total n-3 PUFA and reduced MUFA sums.ConclusionsThese findings provide novel information on the effects of early OA on the infrapatellar FA profile in the rabbit ACLT model. The reduction in the n-3/n-6 PUFA ratio of the IFP is in concordance with the inflammation and cartilage degradation in early OA and could contribute to disease pathogenesis.

Icariin Reduces Cartilage Degeneration in a Mouse Model of Osteoarthritis and is Associated with the Changes in Expression of Indian Hedgehog and Parathyroid Hormone-Related Protein.

BackgroundThe aim of this study was to determine the role of icariin, a Chinese traditional herbal medicine extracted from Epimedium, in osteoarthritis (OA), using the murine anterior cruciate ligament transection (ACLT)-induced model of OA and micromass culture of murine chondrocytes.Material/MethodsTwenty-four three-month-old C57/6J mice were randomly divided into three groups: the sham group (no surgery and joint injection with normal saline) (N=8); the ACLT + ICA group (ACLT surgery and icariin treatment) (N=8); and the ACLT group (ACLT surgery and joint injection with normal saline) (N=8). At 12 weeks after ACLT surgery, murine articular cartilage was harvested from all mice for histological evaluation of any differences in cartilage degeneration. In vitro micromass culture of mouse chondrocytes was used to study the effects of icariin on chondrocyte differentiation and growth from the three mouse groups.ResultsIcariin treatment (mice in the ACLT + ICA group) significantly reduced degeneration of cartilage in OA with increased cartilage thickness, associated with increased expression of collagen type II alpha 1 (COL2A1), decreased chondrocyte hypertrophy, and decreased expression of collagen type X (ColX) and matrix metalloproteinase 13 (MMP13). In vitro, icariin promoted chondrocyte differentiation by upregulating the expression of agrrecan, Sox9 and parathyroid hormone-related protein (PHrP) and down-regulation of Indian hedgehog (Ihh) and genes regulated by Ihh.ConclusionsIn a mouse model of OA icariin treatment reduced destruction of cartilage, promoted chondrocyte differentiation, upregulated expression of PHrP and down-regulated the expression of Ihh.