Severity Of Cartilage Degradation Research Articles

Activation of Notch Signaling Pathway is involved in Extracellular Matrix Degradation in human induced pluripotent stem cells chondrocytes induced by HT-2 toxin

Notch signaling regulates cartilage formation and homeostasis. Kashin-Beck Disease (KBD), an endemic osteochondropathy, is characterized by severe cartilage degradation. The etiology of KBD is related to the exposure of HT-2 toxin, a mycotoxin and primary metabolite of T-2 toxin. This study aims to explore the role of HT-2 toxin in the Notch signaling regulation and extracellular matrix (ECM) metabolism of hiPSCs-Chondrocytes. Immunohistochemistry and qRT-PCR were employed to investigate the expression of Notch pathway molecules in KBD articular cartilage and primary chondrocytes. hiPSCs-Chondrocytes, derived from hiPSCs, were treated with 100 ng/mL HT-2 toxin and the γ-secretase inhibitor (DAPT) for 48h, respectively. The markers related to the Notch signaling pathway and ECM were assessed using qRT-PCR and Western blot. Notch pathway dysregulation was prominent in KBD cartilage. HT-2 toxin exposure caused cytotoxicity in hiPSCs-Chondrocytes, and activated Notch signaling by increasing the mRNA and protein levels of NOTCH1 and HES1. HT-2 toxin also upregulated ECM catabolic enzymes and downregulated ECM components (COL2A1 and ACAN), indicating ECM degradation. DAPT-mediated Notch signaling inhibition suppressed the mRNA and protein level of ADAMTS5 expression while enhancing ECM component expression in hiPSCs-Chondrocytes. This study suggests that HT-2 toxin may induce ECM degradation in hiPSCs-Chondrocytes through activating Notch signaling.

Methyl Canthin-6-one-2-carboxylate Restrains the Migration/Invasion Properties of Fibroblast-like Synoviocytes by Suppressing the Hippo/YAP Signaling Pathway.

Rheumatoid arthritis (RA) is an inflammatory condition that causes severe cartilage degradation and synovial damage in the joints with multiple systemic implications. Previous studies have revealed that fibroblast-like synoviocytes (FLSs) play a pivotal role in the pathogenesis of RA. The appropriate regulation of FLS function is an efficient approach for the treatment of this disease. In the present study, we explored the effects of methyl canthin-6-one-2-carboxylate (Cant), a novel canthin-6-one alkaloid, on the function of FLSs. Our data showed that exposure to Cant significantly suppressed RA-FLS migration and invasion properties in a dose-dependent manner. Meanwhile, pre-treatment with Cant also had an inhibitory effect on the release of several pro-inflammatory cytokines, including IL-6 and IL-1β, as well as the production of MMP1 and MMP3, which are important mediators of FLS invasion. In further mechanistic studies, we found that Cant had an inhibitory effect on the Hippo/YAP signaling pathway. Treatment with Cant suppressed YAP expression and phosphorylation on serine 127 and serine 397 while enhancing LATS1 and MST1 levels, both being important upstream regulators of YAP. Moreover, YAP-specific siRNA or YAP inhibition significantly inhibited wound healing as well as the migration and invasion rate of FLS cells, an impact similar to Cant treatment. Meanwhile, the over-expression of YAP significantly reversed the Cant-induced decline in RA-FLS cell migration and invasion, indicating that YAP was required in the inhibitory effect of Cant on the migration and invasion of RA-FLS cells. Additionally, supplementation of MMP1, but not MMP3, in culture supernatants significantly reversed the inhibitory effect of Cant on RA-FLS cell invasion. Our data collectively demonstrated that Cant may suppress RA-FLS migration and invasion by inhibiting the production of MMP1 via inhibiting the YAP signaling pathway, suggesting a potential of Cant for the further development of anti-RA drugs.

PPAR\u03b1\u2212ACOT12 axis is responsible for maintaining cartilage homeostasis through modulating de novo lipogenesis

Here, in Ppara−/− mice, we found that an increased DNL stimulated the cartilage degradation and identified ACOT12 as a key regulatory factor. Suppressed level of ACOT12 was observed in cartilages of OA patient and OA-induced animal. To determine the role and association of ACOT12 in the OA pathogenesis, we generated Acot12 knockout (KO) (Acot12−/−) mice using RNA-guided endonuclease. Acot12−/− mice displayed the severe cartilage degradation with the stimulation of matrix MMPs and chondrocyte apoptosis through the accumulation of acetyl CoA. Delivery of acetyl CoA-conjugated chitosan complex into cartilage stimulated DNL and cartilage degradation. Moreover, restoration of ACOT12 into human OA chondrocytes and OA-induced mouse cartilage effectively rescued the pathophysiological features of OA by regulating DNL. Taken together, our study suggested ACOT12 as a novel regulatory factor in maintaining cartilage homeostasis and targeting ACOT12 could contribute to developing a new therapeutic strategy for OA.

Subchondral Bone Microarchitectural and Mineral Properties and Expression of Key Degradative Proteinases by Chondrocytes in Human Hip Osteoarthritis.

Background: The purpose of this study was to investigate the relationship between the expression of key degradative enzymes by chondrocytes and the microarchitectural and mineral properties of subchondral bone across different stages of cartilage degradation in human hip osteoarthritis (OA). Methods: Osteochondral samples at different stages of cartilage degradation were collected from 16 femoral heads with OA. Osteochondral samples with normal cartilage were collected from seven femoral heads with osteoporosis. Microcomputed tomography was used for the investigation of subchondral bone microarchitecture and mineral densities. Immunohistochemistry was used to study the expression and distribution of MMP13 and ADAMTS4 in cartilage. Results: The microarchitecture and mineral properties of the subchondral plate and trabecular bone in OA varied with the severity of the degradation of the overlying cartilage. Chondrocytes expressing MMP13 and ADAMTS4 are mainly located in the upper zone(s) of cartilage regardless of the histopathological grades. The zonal expression of these enzymes in OA (i.e., the percentage of positive cells in the superficial, middle, and deep zones), rather than their overall expression (the percentage of positive cells in the full thickness of the cartilage), exhibited significant variation in relation to the severity of cartilage degradation. The associations between the subchondral bone properties and zonal and overall expression of these enzymes in the cartilage were generally weak or nonsignificant. Conclusions: Phenotypic changes in chondrocytes and remodelling of subchondral bone proceed at different rates throughout the process of cartilage degradation. Biological influences are more important for cartilage degradation at early stages, while biomechanical damage to the compromised tissue may outrun the phenotypic change of chondrocytes and is critical in the advanced stages.

The microbiome does not influence cartilage degradation following joint destabilisation in mice when performed under stringently controlled conditions

Purpose: The influence of gut bacteria in the modulation of disease severity of OA is poorly understood. Reports to date on the role of the microbiome in germ free (GF) models of OA are conflicting and haven’t always been stringently controlled. In the present study, surgical induction of OA by destabilisation of the medial meniscus (DMM) was used to examine the potential influence of the microbiome on cartilage damage severity. Methods: Male C57BL/6J mice used in the present study were bred and housed in the gnotobiotic facility at the Kennedy Institute. Microbiome analysis by 16S rRNA amplicon sequencing was performed on stool samples from mice prior to DMM surgery. At 10 weeks of age, mice were randomly selected for sham (n=6-8) or DMM (n=17-20) surgery, which was performed in sterile Class II cabinets. Knees from operated mice were harvested at 8 or 12wks post DMM. Faecal samples were collected post surgery and at the end of the experiment to test GF status. Joints were sectioned and severity of cartilage degradation was assessed in a blinded method according to OARSI guidelines. No mice were excluded from data analysis. Differences were analysed by ANOVA with Tukey post test correction and P values less than 0.05 were considered statistically significant. Control (specific pathogen free, SPF) male C57BL/6J mice (originally derived from the GF founders) were bred under SPF conditions but housed in an environment to mimic the GF conditions in terms of bedding, food (irradiated) and water, and underwent sham (n=4-6) or DMM (n=11-13) surgery at 10 weeks for analysis after 8 and 12wks. Results: All mice were confirmed GF at the time of surgery but 50% tested positive for microbes at the end of the surgical procedure suggesting contamination at the time of surgery (indicated in red dots). Proteoglycan loss and cartilage erosion were evident in SPF and GF mice 12 wks after surgery. Cartilage damage following DMM was significantly higher than sham in both groups, but was not different between GF and SPF mice. Cartilage damage seen at 8wks was modest and only reached significance over sham-operated in GF mice. No significant differences in scores between the GF and SPF DMM groups were demonstrated. No apparent relationship between those animals that became contaminated at the time of surgery and disease scores was seen. Conclusions: The present study shows that significant cartilage degradation occurs in GF mice undergoing DMM at both timepoints. The disease severity was not altered by the absence of a microbiome, nor apparently when mice lost their GF status during the experimental course. When SPF mice were housed under tightly matched cage conditions to the GF mice, disease appeared indistinguishable. The absence of a gut microbiome does not appear to protect the cartilage from erosion after joint destabilisation.

Subchondral bone microarchitecture and mineral density in human osteoarthritis and osteoporosis: A regional and compartmental analysis.

Osteoarthritis (OA) and osteoporosis (OP) are historically considered to be inversely correlated but there may be an overlap between the pathophysiology of the two diseases. This study aimed to investigate the subchondral bone microarchitecture and matrix mineralization, and the association between them in OA and OP in relation to the degree of cartilage degeneration. Fifty-six osteochondral plugs were collected from 16 OA femoral heads. They were graded on a regional basis according to the stages of cartilage degeneration, as evaluated by a new macroscopic and a modified microscopic grading system. Twenty-one plugs were collected from seven femoral heads with OP. Plugs were scanned by microcomputed tomography and the microarchitectural and mineral properties were obtained for both subchondral plate and trabecular bone. Microarchitecture and material and apparent densities of subchondral bone in OP were similar to regions with early cartilage degeneration but different from regions with advanced cartilage degradation in OA femoral heads. Subchondral trabecular bone was more mineralized than subchondral plate in both OP and OA, and this compartmental difference varied by severity of cartilage degradation. Furthermore, the relationship among trabecular bone volume fraction, tissue mineral density, and apparent bone density was similar in OP and different stages of OA. Subchondral bone microarchitecture and mineral properties in OP are different from OA in a regionalized manner in relation to stages of cartilage degeneration. Both regional and compartmental differences at structural, material, and cellular levels need to be studied to understand the transition of OA subchondral bone from being osteoporotic to sclerotic.

Fatty acid sensing GPCR (GPR84) signaling safeguards cartilage homeostasis and protects against osteoarthritis

It is well known that free fatty acids (FFAs) have beneficial effects on the skeletal system, however, which fatty acid sensing GPCR(s) and how the GPCR(s) regulating cartilage development and osteoarthritis (OA) pathogenesis is largely unknown. In this study, we found Gpr84, a receptor for medium-chain FFAs (MCFA), was the only FFA-sensing GPCR in human and mouse chondrocytes that exhibited elevated expression when stimulated by interleukin (IL)-1β. Gpr84-deficiency upregulated cartilage catabolic regulator expression and downregulated anabolic factor expression in the IL-1β-induced cell model and the destabilization of the medial meniscus (DMM)-induced OA mouse model. Gpr84−/− mice exhibited an aggravated OA phenotype characterized by severe cartilage degradation, osteophyte formation and subchondral bone sclerosis. Moreover, activating Gpr84 directly enhanced cartilage extracellular matrix (ECM) generation while knockout of Gpr84 suppressed ECM-related gene expression. Especially, the agonists of GPR84 protected human OA cartilage explants against degeneration by inducing cartilage anabolic factor expression. At the molecular level, GPR84 activation inhibited IL-1β-induced NF-κB signaling pathway. Furthermore, deletion of Gpr84 had little effect on articular and spine cartilaginous tissues during skeletal growth. Together, all of our results demonstrated that fatty acid sensing GPCR (Gpr84) signaling played a critical role in OA pathogenesis, and activation of GPR84 or MCFA supplementation has potential in preventing the pathogenesis and progression of OA without severe cartilaginous side effect.

TNF alpha-stimulated gene 6 (TSG6) is chondroprotective in vivo but overexpression cannot compensate for loss of FGF2

Purpose: TNF alpha stimulated gene 6 (TSG6) is expressed in response to tissue injury and has purported tissue-protective and anti-inflammatory effects. High levels of TSG6 have been detected in the synovial fluid of patients with OA and following acute knee injury. Our published work shows that TSG6 is regulated after joint destabilisation in mice, in a highly mechanosensitive and fibroblast growth factor 2 (FGF2)-dependent fashion. As FGF2-/- mice develop accelerated OA, and injury-induced FGF2-dependent genes such as Timp1 and inhibin were downregulated in TSG-6-/- mouse joints, we speculated that the chondroprotective role of FGF2 may be mediated through TSG6. We asked whether overexpressing TSG-6 could reduce the severity of OA in wild type or FGF2-/- mice. Methods: OA was induced by cutting the medial menisco-tibial ligament (DMM) in 10-12 week old male and female TSG6-/-, TSG6tg, FGF2-/- mice with their respective controls (C57BL/6 and Balb/c), and male FGF2-/-;TSG6tg+/+ or FGF2-/-;TSG6tg-/- mice. Focal cartilage injury was also performed in male and female TSG6tg mice. 8 and 12 weeks post-surgery, joints were sectioned and the severity of cartilage degradation was assessed according to a modified OARSI score. FGF2 levels were measured in cartilage injury conditioned medium from FGF2-/-, Balb/c, TSG-6tg+/+, TSG-6tg-/+, TSG-6tg-/- hips by V-PLEX bFGF kit from Meso Scale Discovery (MSD, Rockville, MD). Results: Accelerated OA was seen in TSG6-/- male mice at 12 but not 8 weeks post DMM compared with wild type (C57BL/6). A reciprocal improvement in OA scores was seen in male TSG6Tg mice 12 weeks after DMM. FGF2−/− mice developed accelerated OA post DMM as shown previously, but overexpression of TSG6 (FGF2-/-;TSG6tg+/+) was unable to overcome this. There was no significant difference observed in cartilage repair scores 8 weeks after focal cartilage injury in male or female TSG-6tg mice compared with wild type (Balb/c). Secreted FGF2 in cartilage injury conditioned medium was not influenced by genotype indicating that the bioavailability of FGF2 is not determined by TSG-6. Conclusions: TSG-6 is weakly chondroprotective in murine OA. Despite being strongly FGF2 dependent in vivo, TSG6 overexpression does not protect FGF2-/- mice from accelerated OA. The chondroprotective role of FGF2 cannot be explained by FGF2-dependent regulation of TSG6.

SOST Deficiency Aggravates Osteoarthritis in Mice by Promoting Sclerosis of Subchondral Bone.

As the initial part in the development of osteoarthritis (OA), subchondral bone sclerosis has been considered to be initiated by excess mechanical loading and proven to be correlated to other pathological changes. Sclerostin, which is an essential mechanical stress response protein, is encoded by the SOST gene. It is expressed in osteocytes and mature chondrocytes and has been proven to be closely correlated to OA. However, the relationship and mechanism between the SOST gene and the development of OA remain unclear. The aim of the present study was to investigate the role of the SOST gene in OA pathogenesis in the subchondral bone. A knee anterior cruciate ligament transection (ACLT) mouse osteoarthritis (OA) model on SOST-knockout (SOST KO) and wild-type (WT) mice was established. The pathogenic and phenotypic changes in the subchondral bone were investigated by histology, micro-CT, immunohistochemistry, TRAP staining, Masson staining, and Toluidine blue staining. It was found that sclerostin expression decreased in both the calcified cartilage and mineralized subchondral structures during the development of OA. Joint instability induced a severe cartilage degradation phenotype, with higher OARSI scores in SOST KO mice, when compared to WT mice. SOST KO mice with OA exhibited a higher BMD and BV/TV ratio, as well as a higher rate of bone remodeling and TRAP-positive cell number, when compared to the WT counterparts, but the difference was not significant between the sham-operation groups. It was concluded that loss of sclerostin aggravates knee OA in mice by promoting subchondral bone sclerosis and increasing catabolic activity of cartilage.

Suppression of ABCD2 dysregulates lipid metabolism via dysregulation of miR-141:ACSL4 in human osteoarthritis.

Even though increasing evidence indicates the importance of peroxisomal lipid metabolism in regulating biological and pathological events, its involvement in cartilage development has not been well studied. Here, we identified the importance of peroxisomal function, particularly the functional integrity of ABCD2, in the pathogenesis of osteoarthritis (OA). Knockdown of ABCD2 in OA chondrocytes induced the accumulation of very long chain fatty acids (VLCFAs) and apoptotic cell death. Moreover, knockdown of ABCD2 altered profiles of miRNAs that affect the expression level of ACSL4, a known direct regulator of lipid metabolism. Suppression of ACSL4 in human chondrocytes-induced VLCFA accumulation, MMP-13 expression, and apoptotic cell death. In vivo morph-down of the ACSL4 homologue in zebrafish resulted in significant defects in cartilage development and in vivo knockdown of ACSL4 in cartilage tissue of an OA model mice promoted severe cartilage degradation. In summary, to the best of our knowledge, this is the first report suggesting that the regulatory network among peroxisomal ABCD2:ACSL4:VLCFA serves as a novel regulator of cartilage homeostasis, and these data may provide novel insights into the role of peroxisomal fatty acid metabolism in pathogenesis of human OA. SIGNIFICANCE OF THE STUDY: Our study indicates that peroxisomal dysfunction is closely related to OA pathogenesis. Particularly, the functional integrity of ABCD2 may play an important role in OA pathogenesis via the accumulation of VLCFAs and stimulation of apoptotic death through altering profiles of miRNAs that target ACSL4. Our findings suggest that targeting the regulatory network among the peroxisomal ABCD2:ACSL4:VLCFA axis may provide a new potential therapeutic strategy for OA pathogenesis.

Estrogen deficiency accelerates lumbar facet joints arthritis

Dramatic increase in the prevalence of lumbar facet joint (LFJ) arthritis in women around the age of menopause indicates a protective role for estrogen in LFJ arthritis. To date, there is no evidence for this indication and the mechanism of such an effect remains poorly understood. In this study, ovariectomized (OVX) mice were used to mimic the estrogen-deficient status of post-menopausal women. Micro-CT and immunohistochemistry was employed to assess the morphological and molecular changes in ovariectomy-induced LFJ arthritis. The results show that the LFJ subchondral bone mass was significantly decreased in OVX mice, with increased cavities on the interface of the subchondral bone. Severe cartilage degradation was observed in ovariectomy-induced LFJ arthritis. Increased blood vessels and innervations were also found in degenerated LFJ, particularly in the subchondral bone area. 17β-Estradiol treatment efficiently suppressed LFJ subchondral bone turnover, markedly inhibited cartilage degradation, and increased blood vessel and nerve ending growth in degenerated LFJ in OVX mice. Our study reveals that estrogen is a key factor in regulating LFJ metabolism. Severe LFJ degeneration occurs when estrogen is absent in vivo. Collapsed subchondral bone may be the initiation of this process, and estrogen replacement therapy can effectively prevent degeneration of LFJ under estrogen-deficient conditions.

Effectiveness of Radiofrequency Hyperthermia for Treating Cartilage in Guinea Pigs with Primary Osteoarthritis.

Objective Autophagy was reported to be essential for maintaining chondrocyte function, and reduced autophagy leads to osteoarthritis (OA). Previous studies showed involvement of heat shock stress in the control of autophagy in cells. This study sought to investigate the effect of hyperthermia on the expression of autophagy-related proteins in articular cartilage and the progression of naturally occurring OA in Hartley guinea pigs. Design Radiofrequency pulses of 13.56 MHz were applied to the animals' knees for 20 minutes to induce hyperthermia. The knee joints were resected at 8 hours, 24 hours, 72 hours, 7 days, and 6 months after hyperthermia. Serial sections of knees were examined for histopathological changes. The expression levels of Unc-51-like kinase 1 (ULK1) and Beclin1 were analyzed by immunohistochemistry. Results Analysis of the distribution of positive cells showed that, in cases of moderate OA, ULK1 and Beclin1 expression levels were significantly decreased in the superficial zone (SZ) and middle zone (MZ) ( P < 0.01) compared with normal cartilage. Seven days after exposure to radiofrequency waves, expression levels of ULK1 and Beclin1 were augmented in the SZ in animals with mild OA. The severity of cartilage degradation was significantly reduced ( P < 0.01) in the radiofrequency-treated knees versus the untreated knees. Conclusions This study showed that heat stimulation enhanced autophagy in healthy knee chondrocytes and chondrocytes in knees with mild OA. The study also showed that long-term periodic application of hyperthermia suppresses aging-related progression of OA. The activation of autophagy by radiofrequency hyperthermia may be an effective therapeutic approach for osteoarthritis.

Different responses of articular cartilage to strenuous running and joint immobilization

ABSTRACTObjective: To compare the pathological changes in cartilage derived from rats that developed osteoarthritis either by joint immobilization or by strenuous treadmill running in order to better understand their respective pathomechanism.Method: A total of 24 male Wistar rats were randomly assigned to three groups: sedentary control (CON), immobilization (IM), and strenuous running (SR). For rats in the IM group, unilateral knee joint was immobilized in flexion. Rats in the SR group underwent treadmill running with high intensity. Eight weeks later, all animals were sacrificed. Femoral condyles were collected to take histological observation for cartilage characteristic and immunohistochemistry for collagen type II. In addition, cartilage samples were obtained to assess gene expression of aggrecan, collagen type II, biglycan, and fibromodulin by quantitative RT-PCR.Results: Gross and histological observation showed osteoarthritic changes in groups SR and IM; however, more severe cartilage degradation was revealed in the latter. Proteoglycan and collagen II content decreased in groups SR and IM in comparison to group CON, with more loss in group IM. In group SR, mRNA levels in femoral cartilage were found to be unaltered for all the molecules measured. On the contrary, these molecules were significantly downregulated in group IM.Conclusion: Differences in gross observation, histological characteristics, and gene expression of proteoglycans and collagen II suggest that both knee immobilization and strenuous running would lead to degenerative change of cartilage, but at different stages of the degenerative process.

The chicken combs extract alleviates pain and cartilage degradation in rat model osteoarthritis

Osteoarthritis (OA) is a degenerative and inflammatory disorder on particular joint inducing annihilation of articular cartilage. This study is aimed to investigate the therapeutic effect of extract of chicken combs (CCE) on pain severity and cartilage degeneration in an experimental model of rat OA. OA was induced in rats by intra-articular injection of monosodium iodoacetate (MIA) to the right knee. CCE, hyaluronic acid and celecoxib were administrated orally every day after MIA injection. Pain severity was estimated by evaluation of secondary tactile allodynia using the von Frey assessment test. The severity of cartilage degradation was examined by histological analysis and Mankin scoring system. Protein expression was observed by immunohistochemistry. Real-time polymerase chain reaction was used to measure mRNA level. CCE decreased secondary tactile allodynia revealed by a promoted pain withdrawal latency and pain withdrawal threshold. Cartilage destruction in the osteoarthritic joints was improved by CCE treatment. CCE also suppressed the expression of metalloproteinase (MMP)-3, -13, interleukin-1β, inducible nitric oxide synthase and nitrotyrosine increased in osteoarthritic joints. The mRNA level of MMP-1, 3, and -13 was down-regulated by CCE treatment. On the other hand, CCE treatment induced the gene expression of tissue inhibitor of metalloproteinase-1 and -3. CCE treatment demonstrates the therapeutic effect of pain relief and attenuates cartilage degeneration through the suppression of inflammatory mediators and metalloproteinases performing a pivotal function in OA pathogenesis.

Section of the anterior cruciate ligament in the rabbit as animal model for osteoarthritis progression.

The purpose of this study was to determine whether instability of knee is a risk factor in the progression of osteoarthritis (OA). Twenty-four mature New Zealand White rabbits were randomly divided into four groups. The control group received 0.3ml saline in the first, fourth and seventh days in the right knee, while the other three groups received the same dosage 4% papain and its activator 0.03ML-cystein. The P3w group knees were harvested at three weeks after the last papain injection, the P6w group knees received a sham surgery at three weeks and were harvested at six weeks after the last papain injection, while the P+ACLT group knees received ACL transection at three weeks and were harvested at six weeks after the last papain injection. Cartilage degradation of femoral condyles and tibial plateaus were evaluated by X-rays, macroscopy, light microscopic and transmission electron microscopy (TEM). According to X-rays grade scale, macroscopic grade scale, light microscopic modified Mankin scale and TEM, in the P3w knees, cartilage degeneration of femoral condyles and tibial plateaus were significantly severe compared to those of the control group (P < 0.05), but the differences were not apparent in comparison with the P6w knees (P > 0.05). However, in P+ACLT knees, cartilage degeneration of femoral condyles and tibial plateaus appeared more severe in comparison with P6w knees, and the difference was significant (P < 0.05). Instability of knee plays a significant role in increasing the severity of cartilage degradation in rabbit knees and should be considered as a risk factor in OA knee progression. Our data may suggest that reconstruction of knee stability may prevent or delay the progression of OA.

Lithium Protects Against Cartilage Degradation in Osteoarthritis

To determine the actions of lithium chloride (LiCl) on catabolic events in human articular chondrocytes, and the effects of LiCl on the progression and severity of cartilage degradation in interleukin-1β (IL-1β)-treated mouse knee joints and after surgical induction of osteoarthritis (OA) in a mouse model. Human articular chondrocytes were treated with LiCl followed by IL-1β, and the expression levels of catabolic genes were determined by real-time polymerase chain reaction. To understand the mechanism by which LiCl affects catabolic events in articular chondrocytes after IL-1β treatment, the activation of NF-κB was determined using luciferase reporter assays, and the activities of MAPKs and the STAT-3 signaling pathway were determined by immunoblot analysis of total cell lysates. Cultures of mouse femoral head explants treated with IL-1β and a mouse model of surgically induced OA were used to determine the effects of LiCl on proteoglycan loss and cartilage degradation. LiCl treatment resulted in decreased catabolic marker messenger RNA levels and activation of NF-κB, p38 MAPK, and STAT-3 signaling in IL-1β-treated articular chondrocytes. Furthermore, LiCl directly inhibited IL-6-stimulated activation of STAT-3 signaling. Consequently, the loss of proteoglycan and severity of cartilage destruction in LiCl-treated mouse knee joints 8 weeks after OA induction surgery or in LiCl-treated mouse femoral head explants after IL-1β treatment were markedly reduced compared to that in vehicle-treated joints or explants. LiCl reduced catabolic events in IL-1β-treated human articular chondrocytes and attenuated the severity of cartilage destruction in IL-1β-treated mouse femoral head explants and in the knee joints of mice with surgically induced OA, acting via inhibition of the activities of the NF-κB, p38, and STAT-3 signaling pathways.

Inflammatory markers associated with osteoarthritis after destabilization surgery in young mice with and without Receptor for Advanced Glycation End-products (RAGE).

HtrA1, Ddr-2, and Mmp-13 are reliable biomarkers for osteoarthritis (OA), yet the exact mechanism for the upregulation of HtrA-1 is unknown. Some have shown that chondrocyte hypertrophy is associated with early indicators of inflammation including TGF-β and the Receptor for Advanced Glycation End-products (RAGE). To examine the correlation of inflammation with the expression of biomarkers in OA, we performed right knee destabilization surgery on 4-week-old-wild type and RAGE knock-out (KO) mice. We assayed for HtrA-1, TGF-β1, Mmp-13, and Ddr-2 in articular cartilage at 3, 7, 14, and 28 days post-surgery by immunohistochemistry on left and right knee joints. RAGE KO and wild type mice both showed staining for key OA biomarkers. However, RAGE KO mice were significantly protected against OA compared to controls. We observed a difference in the total number of chondrocytes and percentage of chondrocytes staining positive for OA biomarkers between RAGE KO and control mice. The percentage of cells staining for OA biomarkers correlated with severity of cartilage degradation. Our results indicate that the absence of RAGE did protect against the development of advanced OA. We conclude that HtrA-1 plays a role in lowering TGF-β1 expression in the process of making articular cartilage vulnerable to damage associated with OA progression.

Autophagy: A New Therapeutic Target in Cartilage Injury and Osteoarthritis

Autophagy is a cellular homeostasis mechanism that plays an essential role in energy and nutrient regulation as well as in the removal of damaged and dysfunctional macromolecules and organelles.1 Failure of autophagy results in the increased production of reactive oxygen species and to abnormal gene expression, and it can lead to cell death. Autophagy failure as observed in aging tissues and cells leads to neurodegeneration, cardiomyopathies, abnormal skeletal development, and premature death.2 In human osteoarthritis (OA) and in a mouse model, autophagy proteins were expressed at reduced levels; this measure was associated with increased cell death and degradation in articular cartilage.3 An important suppressor of autophagy is the protein mammalian target of rapamycin (mTOR) as part of the complex mTORC1. Rapamycin is a lipophilic macrolide antibiotic that is used as an immunosuppressive drug in solidorgan transplantation for the management of renal cell carcinoma, and it is under evaluation for the treatment of certain types of cancer. It can induce autophagy in a variety of cell types.4 Rapamycin was tested for chondroprotective activities in an in vitro model of cartilage injury and in a mouse model of OA. Cartilage injury was induced by subjecting bovine tissue explants to singleimpact loading, which results in cell death and extracellular matrix damage; single-impact loading was also associated with a suppression of autophagy regulators. Treatment of the explants with rapamycin improved cell viability and attenuated tissue damage.5 In the mouse model, OA was induced by transection of the medial meniscotibial ligament and the medial collateral ligament. Intraperitoneal administration of rapamycin affected the mTOR signaling pathway, as indicated by inhibition of phosphorylation of ribosomal protein S6, a target of mTORC1, and by activation of light chain 3 (LC3), a main marker of autophagy.6 The severity of cartilage degradation was reduced in the rapamycin-treated group compared with the control group, and this reduction was associated with a decrease in synovitis.6 Rapamycin treatment also maintained cartilage cellularity and decreased the expression of ADAMTS-5, an important enzyme in cartilage extracellular matrix degradation,7 and of interleukin-1β, a potent inflammatory and catabolic stimulus in OA.8 These results suggest that rapamycin reduces the severity of traumatic cartilage injury and of experimental OA at least in part by autophagy activation. Drugs that inhibit one of a large number of extracellular matrix-degrading enzymes may not provide sufficient impact to change the overall course of the disease and failed in previous clinical trials.9,10 By contrast, inhibiting the failure of a critical cellular homeostasis mechanism, such as autophagy, leads to global changes in gene expression and can ultimately cause cell death and extracellular matrix destruction,2 two main features of OA-affected cartilage.11 The in vitro and in vivo studies of rapamycin establish proof of principle for chondroprotective effects of rapamycin. The two models are associated with the rapid development of cartilage lesions; it remains to be determined whether rapamycin is also effective in models of spontaneous or aging-associated OA. However, short-term use of rapamycin may be effective and feasible in clinical conditions associated with joint injury.12 One important limitation of rapamycin for long-term administration in humans is its immunosuppressive effect. This will certainly be a concern if the drug were to be used to slow progression of established disease. However, the clinical feasibility of using mTOR inhibitors for the management of OA is supported by recent advances in the development of new and safer rapamycin analogs. It can also be anticipated that, with more precise resolution of the different effector mechanisms that are regulated by mTOR, there will be new drugs with greater specificity.13 Recent observations on suppression of autophagy in aging and injured cartilage and chondroprotective activities of rapamycin suggest that pharmacologic enhancement of autophagy may be a promising new approach to prevent or manage OA.

Autophagy activation by rapamycin reduces severity of experimental osteoarthritis

ObjectivesOsteoarthritis is associated with cell death and extracellular matrix degradation in articular cartilage. Autophagy is an essential cellular homeostasis mechanism that was found to be deficient in ageing and osteoarthritic...

Anatomical significance of a posterior horn of medial meniscus: the relationship between its radial tear and cartilage degradation of joint surface

BackgroundTraumatic injury and surgical meniscectomy of a medial meniscus are known to cause subsequent knee osteoarthritis. However, the difference in the prevalence of osteoarthritis caused by the individual type of the medial meniscal tear has not been elucidated. The aim of this study was to investigate what type of tear is predominantly responsible for the degradation of articular cartilage in the medial compartment of knee joints.MethodsFive hundred and forty eight cadaveric knees (290 male and 258 female) were registered in this study. The average age of cadavers at death was 78.8 years old (range: 52-103 years). The knees were macroscopically examined and their medial menisci were classified into four groups according to types of tears: "no tear", "radial tear of posterior horn", "other types of tear" and "worn-out meniscus" groups. The severity of cartilage degradation in their medial compartment of knee joints was evaluated using the international cartilage repair society (ICRS) grading system. We statistically compared the ICRS grades among the groups using Mann-Whitney U test.ResultsThe knees were assigned into the four groups: 416 "no tear" knees, 51 "radial tear of posterior horn" knees, 71 "other types of tear" knees, and 10 "worn-out meniscus" knees. The knees with substantial meniscal tears showed the severer ICRS grades of cartilage degradation than those without meniscal tears. In addition, the ICRS grades were significantly severer in the "radial tear of posterior horn" group than in the "other types of tear" group, suggesting that the radial tear of posterior horn in the medial meniscus is one of the risk factors for cartilage degradation of joint surface.ConclusionsWe have clarified the relationship between the radial tear of posterior horn in the medial meniscus and the severer grade of cartilage degradation. This study indicates that the efforts should be made to restore the anatomical role of the posterior horn in keeping the hoop strain, when patients' physical activity levels are high and the tear pattern is simple enough to be securely sutured.