Thinning Of Articular Cartilage Research Articles

Disuse atrophy of articular cartilage can be restored by mechanical reloading in mice

BackgroundModerate mechanical stress generated by normal joint loading and movements helps maintain the health of articular cartilage. Despite growing interest in the pathogenesis of cartilage degeneration caused by reduced mechanical stress, its reversibility by mechanical reloading is less understood. This study aimed to investigate the response of articular cartilage exposed to mechanical reloading after unloading in vivo and in vitro.Methods and resultsDisuse atrophy was induced in the knee joint cartilage of adult mice through hindlimb unloading by tail suspension. For in vivo experiments, mice were subjected to reloading with or without daily exercise intervention or surgical destabilization of the knee joint. Microcomputed tomography and histomorphometric analyses were performed on the harvested knee joints. Matrix loss and thinning of articular cartilage due to unloading were fully or partially restored by reloading, and exercise intervention enhanced the restoration. Subchondral bone density decreased by unloading and increased to above-normal levels by reloading. The severity of cartilage damage caused by joint instability was not different even with prior non-weight bearing. For in vitro experiments, articular chondrocytes isolated from the healthy or unloaded joints of the mice were embedded in agarose gel. After dynamic compression loading, the expression levels of anabolic (Sox9, Col2a1, and Acan) and catabolic (Mmp13 and Adamts5) factors of cartilage were analyzed. In chondrocytes isolated from the unloaded joints, similar to those from healthy joints, dynamic compression increased the expression of anabolic factors but suppressed the expression of catabolic factors.ConclusionThe results of this study indicate that the morphological changes in articular cartilage exposed to mechanical unloading may be restored in response to mechanical reloading by shifting extracellular matrix metabolism in chondrocytes to anabolism.

Osseous variations associated with physiological thinning of the glenoid articular cartilage: an osteological study with CT, MRI and arthroscopic correlations

ObjectiveTo investigate the relationship between osseous variations of the glenoid fossa and thinning of the overlaying articular cartilage.Materials and methodsIn total, 360 dry scapulae, comprising adult, children and fetal specimens, were observed for potential presence of osseous variants inside the glenoid fossa. Subsequently, the appearance of the observed variants was evaluated using CT and MRI (each 300 scans), and in-time arthroscopic findings (20 procedures). New terminology of the observed variants was proposed by an expert panel formed by orthopaedic surgeons, anatomists and radiologists.ResultsTubercle of Assaky was observed in 140 (46.7%) adult scapulae, and an innominate osseous depression was identified in 27 (9.0%) adult scapulae. Upon radiological imaging, the tubercle of Assaky was found in 128 (42.7%) CTs and 118 (39.3%) MRIs, while the depression was identified in 12 (4.0%) CTs and 14 (4.7%) MRIs. Articular cartilage above the osseous variations appeared relatively thinner and in several young individuals was found completely absent. Moreover, the tubercle of Assaky featured an increasing prevalence with aging, while the osseous depression develops in the second decade. Macroscopic articular cartilage thinning was identified in 11 (55.0%) arthroscopies. Consequently, four new terms were invented to describe the presented findings.ConclusionPhysiological articular cartilage thinning occurs due to the presence of the intraglenoid tubercle or the glenoid fovea. In teenagers, the cartilage above the glenoid fovea may be naturally absent. Screening for these variations increases the diagnostic accuracy of glenoid defects. In addition, implementing the proposed terminological updates would optimize communication accuracy.

Validation of ultrasonography for measurement of cartilage thickness in the equine carpus.

Articular cartilage thinning is an important hallmark of osteoarthritis (OA), and ultrasonography (US) is a clinically accessible tool potentially suitable for repeated evaluation. The aim of the present prospective methods comparison study was to validate US as a tool for measuring cartilage thickness in the carpus of the horse. Eight Standardbred trotters underwent US examination with 9 and 15 MHz linear transducers. Six anatomical locations in the radiocarpal joint (RCJ) and middle carpal joint (MCJ) were examined. The same joints were assessed by ultrahigh field (9.4 Tesla) magnetic resonance imaging (MRI) and histology. Associations between measurements obtained by the different modalities were assessed by ANOVA, Deming regression, Pearson correlation and Bland–Altman plots. Histologically assessed total cartilage thickness (the noncalcified cartilage (NCC) plus the calcified cartilage zone (CCZ)) overestimated thickness compared to MRI (P < 0.01) and US (P < 0.01). US 15 MHz had substantial agreement with MRI and NCC histology, and repeatability was acceptable (coefficient of variation = 8.6–17.9%) when used for assessment of cartilage thickness in the RCJ. In contrast, 9 MHz US showed poorer agreement with MRI and NCC histology, as it overestimated the thickness of thin cartilage and underestimated the thickness of thicker cartilage in the RCJ and MCJ. Moreover, repeatability was suboptimal (coefficient of variation = 10.4–26.3%). A 15 MHz transducer US is recommended for detecting changes in RCJ cartilage thickness or monitoring development over time, and it has the potential for noninvasive assessment of cartilage health in horses.

RELATIONSHIP BETWEEN THINNING OF ARTICULAR CARTILAGE DUE TO ENDOCHONDRAL OSSIFICATION AND OSTEOARTHRITIS

Purpose: Our purpose is to evaluate the detailed mechanism of articular cartilage degeneration and thinning in osteoarthritis (OA) according to our hypotheses. In non-decalcified specimens, we detected the subchondral bone plate (Subcho.BP) divided into two layers of non-osteocyte and osteocyte region. In addition, several columnar alignments of chondrocytes were present in the non-osteocyte layer. From these new findings, we hypothesized that articular cartilage thinning is due to endochondral ossification and to the resulting thickening of the non-osteocyte region of the Subcho.BP, which are associated with the onset and progression of OA.

Pharmacological Treatment in the Management of Glenohumeral Osteoarthritis.

Glenohumeral osteoarthritis (GH-OA) is a common cause of shoulder pain and is characterized by articular cartilage thinning, glenoid bone loss and deformity, osteophytosis, and other associated changes. The prevalence is estimated to be between 85 and 94% in men and women over the age of 80years. A diagnosis of GH-OA is established based on clinical history, physical examination, and radiographic assessments. Non-pharmacological treatment options may serve as adjuvants to other therapies and should be incorporated for a more holistic approach to management. Pharmacological treatments include oral agents such as acetaminophen, non-steroidal anti-inflammatory drugs (NSAIDs), opioids, corticosteroids and antidepressants. The National Institute for Health and Care Excellence (NICE) UK guidelines recommend NSAIDS as the first-line drugs for OA; topical forms of some of these agents can also be used. However, clinical evidence is largely lacking for its use in GH-OA, although patients with other types of OA, especially the knee and hip, are using these products in efforts to relieve joint pain. Intra-articular injections such as platelet-rich plasma, cortisone, and hyaluronic acid are usually used to control symptoms in moderate to advanced arthritis or in non-surgical candidates. Other non-surgical treatment options include suprascapular nerve block and radiofrequency ablation, and these options have been studied on different levels of evidence. Furthermore, all these treatments have their own indications, contraindications, and adverse effects profiles. Surgical treatment of GH-OA is reserved for patients who do not respond to conservative management or who suffer from debilitating symptoms that severely impair their quality of life.Level of Evidence IV, review article.

Primary Epiphyseal Osteomyelitis (PEO) in 18 Children: A Rare Entity With Atypical Features.

The purpose of this study is to discuss the natural history and management of primary epiphyseal osteomyelitis (PEO), to differentiate clinico-radiologic features of PEO caused by Mycobacterium and other organisms, and to discuss their intermediate-term outcomes. Between 2006 and 2017, 18 patients of PEO were managed at our center. Blood investigations, x-rays, and magnetic resonance imaging of affected part were carried out. Surgical drainage of lesions was done to retrieve infective fluid and tissue for examination. Antibiotics were administered for 1 year in Mycobacterial PEO and for 6 weeks in bacterial PEO. Average follow-up of patients was 5.5 years (range, 2 to 11 y). Boys were more commonly affected (11/18). Distal femur was the most common site involved (12/18). Eleven patients had Mycobacterium tuberculosis as the causative organism, 6 were positive for Staphylococcus aureus, and 1 for Brucella. Swelling and limp were predominant in patients with Tubercular PEO, whereas pain was more common in bacterial PEO. Nine of 11 patients with Tubercular PEO had penetration into the joint, whereas none in bacterial PEO. All patients recovered completely without residual movement restriction or growth alteration. On follow-up magnetic resonance imaging, 4 patients with Tubercular PEO had thinning of articular cartilage. High index of suspicion is required for early diagnosis of PEO. It is important to differentiate Tubercular from other bacterial PEO as it has more subtle symptoms and poor prognosis if left untreated. Aggressive surgical treatment followed by antibiotic therapy of appropriate duration is required to avoid complications related to joint destruction. To our knowledge, this is the largest reported series with longest follow-up.

The Role of High Resolution Ultrasound in the Assessment of Knee Osteoarthritis

AbstractBackground: Osteoarthritis (OA) is a common musculoskeletaldisease with high prevalence worldwide. Knee OAis primary diagnosed using conventional radiography, however,changes of articular cartilage or menisci cannot be directlyassessed using conventional radiology. Ultrasound (US) canbe used to assess soft tissue degeneration directly.Aim of Study: The aim of this study is to evaluate theefficacy of High Resolution Ultrasound (HRUS) in the assessmentof structural abnormalities found in knee osteoarthritis.Patients and Methods: The study was conducted upon 20patients with knee OA (12F, 8M; mean age 53 years), eachsubject was evaluated for the presence of medial and lateralfemoral cartilage thinning, medial and lateral femoral andtibial osteophytes, medial and lateral meniscal degeneration,joint effusion and Baker's cyst using ultrasound and MRI,considering MRI as a reference standard.Results: The overall sensitivity of the ultrasound in thedetection of knee OA parameters was good (89.3%), theoverall accuracy was excellent (90%), the overall specificitywas excellent (93.1%). The Positive Predictive Value (PPV)and the Negative Predictive Value (NPV) were 98.3 and 65.9respectively. The p-value was highly significant (<0.01) whencomparing assessment of knee osteoarthritis parameters betweenultrasound and MRI.Conclusion: Ultrasound assessment of the knee joint isan accurate imaging method for detection of knee osteoarthritisparameters including medial and lateral femoral articularcartilage thinning, medial and lateral osteophytes, medial andlateral meniscal degeneration, joint effusion and Baker's cyst.Thus, Knee ultrasound can be used as a complementaryimaging technique to radiography, especially when MRI isnot available, to assess tissue-specific structural OA degenerationnot detected by conventional radiographs.

The Role of Ultrasound in Juvenile Idiopathic Arthritis: A Narrative Review From an Imaging Perspective

Objective: Musculoskeletal sonography (MSKS) is increasingly being used in the setting of juvenile idiopathic arthritis (JIA). The purpose of this narrative review was to describe the role of sonography in JIA and outline strategies for operators of pediatric MSKS. Methods: A literature review through multiple medical databases was conducted by restricting the search to medical subject headings (MeSH). Peer-reviewed English-language articles from 2007 to 2018 were included, which focused the on common sonographic findings of JIA. Results: Twenty-six articles were selected for inclusion in the study after a complete reading. Many studies aimed to address the validity of sonographic techniques in the assessment of JIA, but few studies discussed the specific sonographic appearances of JIA, scanning pitfalls, and appropriate imaging techniques. Conclusion: MSKS reveals subclinical manifestations of arthropathy, but the true value of detecting subclinical disease is not well understood. MSKS is limited in the evaluation of articular cartilage thinning and bony erosions. Responsiveness of MSKS in JIA remains to be formally assessed with higher quality studies.

Disuse histological changes of an unloading environment on joint components in rat knee joints

To clarify the effects of mechanical unloading on joint components, including the articular cartilage by knee compartments, synovial membrane, and the infrapatellar fat pad (IFP), using a hindlimb suspension rat model. Twenty-five male rats were divided into the three following experimental groups: the baseline, control (CON), and hindlimb unloading (HU) groups. Rats in the HU group were subjected to periods of hindlimb unloading by tail suspension. The joint components were evaluated by histopathological and histomorphometric analyses. The unloading condition caused articular cartilage thinning and decreased matrix staining. The tendency of these histological changes in articular cartilage was similar between the knee compartments. In general, the unloading environment resulted in no critical histological changes to the synovial membrane and IFP. At 4 weeks, thickening of the synovial membrane and synovial-like tissue invasion were observed in the HU group, but there was no significant change in inflammation. There was no obvious histological change to the IFP caused by unloading. The unloading environment caused disuse histological changes in the articular cartilage; however, the tendency of the changes was similar between compartments. Furthermore, the unloading condition caused no critical histological changes to the synovial membrane or IFP. Clinically, the unloading environment without joint immobilization over a short period may not have caused critical histological changes to the joint components.

Novel model for age-related OA: ER stress in articular cartilage induces joint degeneration in mice

Purpose: The lifetime risk of osteoarthritis (OA) to an individual is 45% and arthritis is the most common form of disability in the United States, making burden to the individual and society extensive. Age is the most important risk factor for adult-onset OA and ER stress response declines with age. Multiple lines of evidence indicates that pathological ER stress contributes to OA progression. In this work, tested the hypothesis that ER stress in articular chondrocytes alone is sufficient to drive joint degeneration in adult mice. In addition to ER stress, one group of mice was subjected to intense exercise to determine if mechanical stress would hasten joint degeneration. Methods: In the inducible-ER-stress mouse (I-ERS), ER stress was induced by expressing a mis-folded protein, mutant cartilage oligomeric matrix protein, in adult articular chondrocytes (from 16 to 20 weeks). There were three treatment groups: 1) control, 2) ER stress and 3) ER stress with treadmill exercise. Results: Four weeks of ER stress in healthy articular cartilage stimulated joint degeneration including increases matrix metalloproteinase (MMP) activity, inflammation, chondrocyte death, decreases in articular cartilage thickness and proteoglycan content and gait dysfunction, all of which are outcomes associated with OA. Intense exercise increased OARSI score, loss of proteoglycans, articular cartilage thinning and gait dysfunction suggesting that some aspects of OA are exacerbated by mechanical stress. Conclusions: OA is a multifactorial disease involving many risk factors including genetic factors, age, BMI, sex, joint injury and repetitive mechanical stress. Many models recapitulate post-traumatic OA and fewer models address age-related cellular stress. In these studies, physiology relevant ER chondrocyte stress was used to induce joint degeneration in previously healthy joints. This unique model of non-trauma adult-onset OA is useful for 1) defining the role of ER stress in the molecular pathology of OA, 2) for studying age-related OA and 3) testing prevention/treatment strategies and complements other OA models. Defining the role that ER stress plays in OA will lead to a better understand of this complex pathology and may offer new therapeutic strategies.

Increased subchondral bone thickness in hips with cam-type femoroacetabular impingement.

Increased thickness of subchondral acetabular bone with associated articular cartilage thinning in hips with femoroacetabular (FAI) cam impingement has been observed on magnetic resonance imaging (MRI). Dynamic attrition by the cam deformity moving into the acetabulum may potentiate trans-articular shear stresses thus causing these subchondral bone changes. We aimed to quantify the hypertrophic changes of subchondral acetabular bone in patients with cam-type FAI. MRI studies were performed on an asymptomatic population of young Swiss army recruits. Subjects underwent clinical examination and completed questionnaires before undergoing an MRI of the hip. Cam deformities were graded and the dimensions of the acetabular subchondral bone quantified. Univariate linear regression was used to determine the association between the presence of cam deformities and the degree of subchondral acetabular sclerosis. There was a strong association between cam deformities and the thickness, area and shape of subchondral sclerosis. The main increase in hypertrophy was observed in the antero-superior acetabulum where impingement typically occurs. The subchondral sclerosis was 0.66 mm thicker in cam-type deformities than in hips without cam-type deformities (95% CI, 0.38-0.93, p value < 0.001). Mechanical stress in the antero-superior acetabular area is elevated in hips with a cam-type deformity. The study supports the concept that cam-type deformity induced stress leads to hypertrophy of subchondral acetabular bone in the area of impingement. This is collocated with the clinically observed cartilage damage caused by the cam mechanism.

Thinning of articular cartilage after joint unloading or immobilization. An experimental investigation of the pathogenesis in mice

Moderate mechanical stress generated by normal joint loading and movement is essential for the maintenance of healthy articular cartilage. However, the effects of reduced loading caused by the absence of weight bearing or joint motion on articular cartilage and subchondral bone is still poorly understood. We aimed to characterize morphological and metabolic responses of articular cartilage and subchondral bone to decreased mechanical stress invivo. Mice were subjected to periods of hindlimb unloading by tail suspension or external fixation of the knee joints. The articular surface was observed with digital microscope and the epiphyseal bone was assessed by micro-CT analysis. Articular cartilage and subchondral bone were further evaluated by histomorphometric, histochemical, and immunohistochemical analyses. The joint surface was intact, but thickness of both the total and uncalcified layer of articular cartilage were decreased both after joint unloading and immobilization. Subchondral bone atrophy with concomitant marrow expansion predisposed osteoclast activity at bone surface to invade into cartilaginous layer. Uncalcified cartilage showed decreased aggrecan content and increased aggrecanase expression. Alkaline phosphatase (ALP) activity was increased at uncalcified cartilage, whereas decreased at calcified cartilage. The distributions of hypertrophic chondrocyte markers remained unchanged. Thinning of articular cartilage induced by mechanical unloading may be mediated by metabolic changes in chondrocytes, including accelerated aggrecan catabolism and exquisitely modulated matrix mineralization, and cartilage matrix degradation and resorption by subchondral osteoclasts. Cartilage degeneration without chondrocyte hypertrophy under unloading condition indicate the possible existence of mechanism which is different from osteoarthritis pathogenesis.

Pterosin B prevents chondrocyte hypertrophy and osteoarthritis in mice by inhibiting Sik3

Osteoarthritis is a common debilitating joint disorder. Risk factors for osteoarthritis include age, which is associated with thinning of articular cartilage. Here we generate chondrocyte-specific salt-inducible kinase 3 (Sik3) conditional knockout mice that are resistant to osteoarthritis with thickened articular cartilage owing to a larger chondrocyte population. We also identify an edible Pteridium aquilinum compound, pterosin B, as a Sik3 pathway inhibitor. We show that either Sik3 deletion or intraarticular injection of mice with pterosin B inhibits chondrocyte hypertrophy and protects cartilage from osteoarthritis. Collectively, our results suggest Sik3 regulates the homeostasis of articular cartilage and is a target for the treatment of osteoarthritis, with pterosin B as a candidate therapeutic.

Unilateral anterior crossbite induces aberrant mineral deposition in degenerative temporomandibular cartilage in rats

Purpose: Rrecently, we have developed a unilateral anterior crossbite (UAC) procedure that could induce osteoarthritic (OA)-like changes in the temporomandibular joints (TMJs) cartilage of rats and mice. Herein, we employed imaging, histological, and molecular biology techniques, and also in vitra assay, to investigate whether there is mineral deposition in the UAC induced OA cartilage, what the composition the mineral deposition has, and what a role of the minerals may take in OA cartilage. Methods: Spraguee-Dawley rats were subjected to a unilateral anterior crossbite (UAC) procedure. Histology, electron microscopy, and energy dispersive spectroscopy (EDS) were used to examine cartilage matrix structures and compositions of mineral deposit in TMJs. Protein and/or RNA expression of modulators of phenotypic markers, mineralization and matrix degradation was analyzed by immunohistochemistry and/or real-time PCR. Synthetic BCP and CPPD crystals were used to stimulate chondrogenic ATDC5 cells for their impact on activities of cellular differentiation and secretion. Results: The proteoglycan content in UAC cartilage began to decrease as early as 2 weeks post-operation and nearly absent at 20 weeks post-UAC without significant inflammatory response. The collagen arrangement was obviously interrupted and the cartialge thickness reduced from 8 weeks but the thickness of the calcified zone was increased. The expression of Col-II, Col-X, and aggrecan was decreased. The control group chondrocytesunder transmission electron microscopy (TEM) were found in well-delimited lacunae that were surrounded by the tightly-organized collagen fibers. In UAC cartilage, however, collagen fibers were disorganized and fragmented with increasing inter-fibers spaces and were worsened with time. Mineral deposition was observed around the hypertrophic chondrocytes, beginning 2 weeks after the UAC procedure. Those peri-cellular minerals were contained in abundant matrix vesicles (with diameters ranging from 50-200 nm) that were located in close proximity to the cell member of hypertrophic chondrocytes, supporting the cellular origin of the vesicles. Immunohistochemistry and/or real-time PCR analyses demonstrated biphasic changes in the expression of MGP mRNA in TMJ joints of UAC rat, which increased at 2 and 4 weeks post-operation and decreased after 8 weeks post-operation when compared to age-matched controls. The expression of CD73 and Ank was significantly reduced in TMJ cartilage of UAC group at all time points. The expression of Npp1 was significantly reduced in UAC cartilage at 2 and 4 weeks post-operation. In contrast to the expression of anti-mineralization molecules, the expression of Tnap, which hydrolyzes PPi into Pi to promote the formation of BCP crystals, was up-regulated in the UAC groups at 4, 8, and 12 weeks post-operation, and this effect waned at 20 weeks post-operation. The expression of matrix metalloproteinase, Mmp13, which cleaves type II collagen, was also up-regulated in the UAC groups in the similar time-dependent manner as seen in the expression of Tnap. Using EDS technology we confirmed an increase in calcium (Ca) and phosphorus (P) content in UAC cartilage vs their age-matched controls, beginning at 2 weeks post-operation. Only one of four joints tested in 12- and 20-week UAC group was noticed an isolated mineral which was located closing to the deep zonal mineral deposition. They were rich in calcium and phosphorus, but had a low carbon content. Their averaged Ca/P ratio was 1.42 (ranging from 1.38 to 1.48) and 1.44 (range from 1.39 to 1.49) at 12- and 20-weeks UAC group, respectively, closely resemble to those of octacalcium phosphate (Ca/P=1.33) and tricalcium phosphate (Ca/P=1.50), two subtypes of BCP crystals. We then exposed ATDC5 cells to the synthetic BCP crystals. We found that BCP crystals (50μg/ml) reduced Col-II, Col-X and aggrecan expression, but upregulated the expression of Adamts5, Mmp3 and Mmp9, however, only transiently stimulated Tnf-α and Il-1β expression. Conclusions: The present data indicated that mechanical stimuli by UAC enhance mineral accumulation of BCP-like minerals in the cartilage matrix of TMJ condyle at deep layer. The UAC induced aberrant mineralization deposition of BCP-like minerals can stimulate matrix degradation via promoting cartilage-degrading enzymes, and may contribute, at least in part, to the thinning of articular cartilage by expansion of the calcified cartilage, facilitate osteoarthritis progression.

Suppressive Activity of Glucosamine Hydrochloride on Nitric Oxide Production from Synoviocytes In Vitro

Background: Knee osteoarthritis (OA) is well known to be one of the most common joint disease in elderly people and characterized by a slowly progression of articular cartilage thinning and remodeling of the joints. Glucosamine hydrochloride (GH) and chondroitin sulfate, either alone or combined, are used for the treatment of OA and reported that these substances are favorably modify the clinical symptoms, especially pain, however, the underlying therapeutic mechanisms of these substances are not completely clear. The present study was undertaken to examine the influence of GH on the production of nitric oxide (NO), which is one of the important final effector molecules associated with OA, from synoviocytes from OA patients by an in vitro cell culture technique. Methods: Synoviocytes (1x105 cell/ml) were stimulated with 100.0 ng/ml periostin in the presence of various concentrations of GH. After 24 h, NO content in culture supernatants was examined by the Griess method. We also examined the influence of GH on transcription factor, NF-κB, activation and iNOS mRNA expression in synoviocytes 4 and 12 h after periostin stimulation, respectively. Results: Addition of GH into cell cultures caused the suppression of NO production from synoviocytes induced by periostin stimulation through the suppression of NF-κB activation and iNOS mRNA expression. The minimum concentration of GH that caused significant suppression of NO production was 1.5 mg/ml. Conclusion: These results strongly suggest that the ability of GH to suppress NO production from synoviocytes may account, at least in part, for the clinical efficacy of GH on OA.

Deficiency of Thrombospondin-4 in Mice Does Not Affect Skeletal Growth or Bone Mass Acquisition, but Causes a Transient Reduction of Articular Cartilage Thickness.

Although articular cartilage degeneration represents a major public health problem, the underlying molecular mechanisms are still poorly characterized. We have previously utilized genome-wide expression analysis to identify specific markers of porcine articular cartilage, one of them being Thrombospondin-4 (Thbs4). In the present study we analyzed Thbs4 expression in mice, thereby confirming its predominant expression in articular cartilage, but also identifying expression in other tissues, including bone. To study the role of Thbs4 in skeletal development and integrity we took advantage of a Thbs4-deficient mouse model that was analyzed by undecalcified bone histology. We found that Thbs4-deficient mice do not display phenotypic differences towards wildtype littermates in terms of skeletal growth or bone mass acquisition. Since Thbs4 has previously been found over-expressed in bones of Phex-deficient Hyp mice, we additionally generated Thbs4-deficient Hyp mice, but failed to detect phenotypic differences towards Hyp littermates. With respect to articular cartilage we found that Thbs4-deficient mice display transient thinning of articular cartilage, suggesting a protective role of Thbs4 for joint integrity. Gene expression analysis using porcine primary cells revealed that Thbs4 is not expressed by synovial fibroblasts and that it represents the only member of the Thbs gene family with specific expression in articular, but not in growth plate chondrocytes. In an attempt to identify specific molecular effects of Thbs4 we treated porcine articular chondrocytes with human THBS4 in the absence or presence of conditioned medium from porcine synovial fibroblasts. Here we did not observe a significant influence of THBS4 on proliferation, metabolic activity, apoptosis or gene expression, suggesting that it does not act as a signaling molecule. Taken together, our data demonstrate that Thbs4 is highly expressed in articular chondrocytes, where its presence in the extracellular matrix is required for articular cartilage integrity.

Pentosan Polysulfate: a Novel Glycosaminoglycan-Like Molecule for Effective Treatment of Alphavirus-Induced Cartilage Destruction and Inflammatory Disease.

Arthritogenic alphaviruses such as Ross River virus (RRV) and chikungunya virus (CHIKV) cause large-scale epidemics of severe musculoskeletal disease and have been progressively expanding their global distribution. Since its introduction in July 2014, CHIKV now circulates in the United States. The hallmark of alphavirus disease is crippling pain and inflammation of the joints, a similar immunopathology to rheumatoid arthritis. The use of glycans as novel therapeutics is an area of research that has increased in recent years. Here, we describe the promising therapeutic potential of the glycosaminoglycan (GAG)-like molecule pentosan polysulfate (PPS) to alleviate virus-induced arthritis. Mouse models of RRV and CHIKV disease were used to characterize the extent of cartilage damage in infection and investigate the potential of PPS to treat disease. This was assessed using histological analysis, real-time PCR, and fluorescence-activated cell sorting (FACS). Alphaviral infection resulted in cartilage destruction, the severity of which was alleviated by PPS therapy during RRV and CHIKV clinical disease. The reduction in cartilage damage corresponded with a significant reduction in immune infiltrates. Using multiplex bead arrays, PPS treatment was found to have significantly increased the anti-inflammatory cytokine interleukin-10 and reduced proinflammatory cytokines, typically correlated with disease severity. Furthermore, we reveal that the severe RRV-induced joint pathology, including thinning of articular cartilage and loss of proteoglycans in the cartilage matrix, was diminished with treatment. PPS is a promising new therapy for alphavirus-induced arthritis, acting to preserve the cartilage matrix, which is damaged during alphavirus infection. Overall, the data demonstrate the potential of glycotherapeutics as a new class of treatment for infectious arthritis. The hallmark of alphavirus disease is crippling pain and joint arthritis, which often has an extended duration. In the past year, CHIKV has expanded into the Americas, with approximately 1 million cases reported to date, whereas RRV continues to circulate in the South Pacific. Currently, there is no licensed specific treatment for alphavirus disease, and the increasing spread of infection highlights an urgent need for therapeutic intervention strategies. Pentosan polysulfate (PPS) is a glycan derivative that is orally bioavailable, has few toxic side effects, and is currently licensed under the name Elmiron for the treatment of cystitis in the United States. Our findings show that RRV infection damages the articular cartilage, including a loss of proteoglycans within the joint. Furthermore, treatment with PPS reduced the severity of both RRV- and CHIKV-induced musculoskeletal disease, including a reduction in inflammation and joint swelling, suggesting that PPS is a promising candidate for drug repurposing for the treatment of alphavirus-induced arthritis.

Membrane-type 1 matrix metalloproteinase controls osteo- and chondrogenesis by a proteolysis-independent mechanism mediated by its cytoplasmic tail

s / Osteoarthritis and Cartilage 23 (2015) A26eA81 A64 deregulated genes included Has1, ADAMTS4, Tnf, IL6, IL18, Inhba, Cd68, Ngf, Ccr2, Wnt 16, Tnfaip6 and Il1r. Conclusions: Deletion of JNK2 retarded the development of experimental murine OA, showing the importance of the kinase for cartilage homeostasis in vivo and implicating an intracellular signalling pathway in OA for the first time. 68 MEMBRANE-TYPE 1 MATRIX METALLOPROTEINASE CONTROLS OSTEOAND CHONDROGENESIS BY A PROTEOLYSIS-INDEPENDENT MECHANISM MEDIATED BY ITS CYTOPLASMIC TAIL Q. Yang y, M. Attur y, T. Kirsch z, Y.J. Lee z, S. Yakar x, Z. Liu x, S.B. Abramson y, P. Mignatti yk. yNYU Sch. of Med. Dept. of Med., New York, NY, USA; zNYU Sch. of Med. Dept. of Orthopedic Surgery, New York, NY, USA; xNYU Coll. of Dentistry Dept. of Basic Sci. and Craniofacial Biology, New York, NY, USA; kNYU Sch. of Med. Dept. of Cell Biology, New York, NY, USA Purpose: We aimed to understand the mechanism by which membrane-type 1 matrix metalloproteinase (MT1-MMP, MMP-14) controls bone and cartilage homeostasis. MT1-MMP, a cell-membrane-bound proteinase with an extracellular catalytic site and a 20-amino acid cytoplasmic tail, degrades the extracellular matrix and plays a key role in postnatal bone formation. MT1-MMP is overexpressed in osteoarthritis, and mutation of MT1-MMP in humans causes the multicentric osteolysis and arthritis disease, Winchester syndrome. The genetic deficiency of MT1-MMP (MT1-MMP-/-) in the mouse causes dwarfism, osteopenia and severe arthritis. Conditional deletion of MT1-MMP in bone marrow-derived mesenchymal progenitor cells (BM-MSC) fully recapitulates this phenotype, showing that MT1-MMP controls osteogenic differentiation inMSC. It has been proposed that the phenotype of MT1-MMP-/mice results from the lack of MT1-MMP proteolytic activity. However, mounting evidence shows a variety of proteolysisindependent functions of MT1-MMP. The unique tyrosine (Y573) in the MT1-MMP cytoplasmic tail is fundamental for the control of intracellular signaling. We have shown that Y573 mediates Ras-ERK1/2 signaling, and that Y573 mutation (Y573D) blocks this functionwithout affecting MT1-MMP proteolytic activity. Methods: We generated a mouse with the Y573D mutation in MT1MMP (MT1-MMP Y573D) and characterized its skeletal phenotype by histological and microCT analyses. We isolated BM-MSC and induced them to differentiate into osteoblasts, chondrocytes and adipocytes, using qRT-PCR to analyze gene expression. Mouse C3H10T1/2MSCwere transfected with MT1-MMP cDNA and analyzed for Wnt signaling and Runx2 activation by luciferase reporter assays. Results: Analysis of the long bones of both homozygote and heterozygote MT1-MMP Y573D mice showed increased trabecular bone relative to wt littermates (BV/TV %; wt: 10 4±8; homo: 23.2±2.0; Tb.Th (mm) wt: 20.5±3.8; homo: 40.1±7.0; Tb.N./mm wt: 10.5±4.2; homo: 19.5 4.2; Tb.Sp (mm): 325.2±52.1; 190.6 49.1; p 0.01), a phenotypeopposite to that of MT1-MMP-/mice. MT1-MMP Y573D mice also showed marked thinning of articular cartilage, with disorganized tissue architecture, clustering and cloning of chondrocytes, as well as pronounced decrease in bone marrow-associated and total body fat (fat weight/body weight wt: 0.270±0.0105; homo: 0.0135±0.006; p1⁄40.0029). These findings indicated a defect in MSC differentiation. Therefore, we isolated BM-MSC from wt and homozygous mutant littermates, and induced them to differentiate into osteoblast and chondrocytes, and myeloid precursors were induced to differentiate into osteoclasts. The Y573D mutation dramatically increased MSC expression of osteoblast markers (ALP, COL1, BSP, OCN, RUNX2) and strongly downregulated chondrocyte (ADA4, ADA5, SOX9, MMP13) and osteoclast markers (TRACP, CALCR, CTSK, DC-STAMP, MMP9, iNOS). These findings indicated thatWnt signaling is upregulated inMT1MMP Y573D-expressing MSC. The canonical Wnt pathway is indeed unique, as it exerts opposite effects on osteoblast and chondrocyte differentiation fromMSC. Therefore, we analyzedWnt signaling and Runx2 activation. We transiently transfected C3H10T1/2 MSC cells in osteoblast medium with the cDNAs for wt MT1-MMP and MT1-MMP Y573D. As controls the cellswere transfectedwith the empty vector (pcDNA) orwith MT1-MMP E240A, a mutant devoid of proteolytic activity. MT1-MMP Y573D dramatically upregulated both Wnt signaling (2.8-4.6-fold; p1⁄40.007) and Runx2 activity (8.7-46.5-fold; 1⁄40.009) relative to wt MT1MMP and MT1-MMP E240A. Conclusions: MT1-MMP controls Wnt signaling by a mechanism independent of extracellular proteolysis and mediated by its cytoplasmic tail. MT1-MMP is a bifunctional protein, with an extracellular proteolytic activity that promotes bone formation through ECM remodeling and a cytoplasmic tail that controls osteogenesis by interacting with a key pro-osteogenic signaling pathway. 69 A DUAL ROLE FOR NOTCH SIGNALING IN JOINT CARTILAGE MAINTENANCE AND OSTEOARTHRITIS Z. Liu y, J. Chen z, A.J. Mirando z, C. Wang y, M.J. Zuscik y, R.J. O'Keefe x, M.J. Hilton z. yUniv. of Rochester Med. Ctr., Rochester, NY, USA; zDuke Univ. Sch. of Med., Durham, NC, USA; xWashington Univ. Sch. of Med.,

Effect of psychological stress on the structure of the temporomandibular joint and the expression of MMP-3 and TIMP-3 in the cartilage in rats

Our aim was to observe the effects of psychological stress on the structure of the temporomandibular joint (TMJ), and to evaluate the expression of matrix metallopeptidase-3 (MMP-3) and tissue inhibitor of metalloproteinase-3 (TIMP-3) in condylar chondrocytes in rats. The rats were divided into 3 groups of 12 according to the duration of psychological stress: 3 weeks or 6 weeks, and 6 weeks of recovery. A fourth group of 12 rats was used as controls. Each rat was evaluated by the open-field test and the weight measured. The results confirmed psychological stress in 24 of the 36 rats (67%). The tissues of the TMJ were stained with haematoxylin and eosin and pathological changes were studied under a light microscope. MMP-3 and TIMP-3 expression was investigated using the SP kit. The experimental groups showed thinning of articular cartilage, shedding of collagen fibres, cracks in the articular discs, and other structural changes that were aggravated with time, from three weeks to six weeks. The 6-week recovery group showed an improvement in these changes, which indicated the initiation of joint repair. The MMP-3 expression rate correlated with the degree of joint lesion, while the TIMP-3 rate showed an opposite trend and was highest in the 6-week recovery group. Our findings clearly indicate that psychological stress may play an important part in the development of TMJ diseases in rats; further studies should be made to extrapolate the results to other models before clinical use.

Posterior horn medial meniscal root tear: the prequel

To determine whether subarticular marrow changes deep to the posterior horn medial meniscal root anchor might predict subsequent medial meniscal root tear. Fifteen patients with MR-diagnosed posterior horn medial meniscal root (PHMMR) tear and a knee MRI antecedent to the tear were identified at three imaging centers over a 7-year period. The pre- and post-tear MR images were evaluated for marrow signal changes deep to the root anchor, meniscal root signal intensity, medial compartment articular cartilage thinning, and meniscal body extrusion. Images of 29 age- and gender-matched individuals with two MRIs of the same knee were reviewed as a control group. MRI in 11 of 15 (73%) cases with subsequent PHMMR tear demonstrated linear subcortical marrow edema deep to the meniscal root anchor on the antecedent MRI compared to only 1 of 29 (3%) non-tear controls (p < 0.0001). The abnormal signal resolved on post-tear MRI in all but two patients. Cyst-like changes deep to the PHMMR were present on initial MRI in three of 15 (23%) cases and three of 29 (10%) controls, persisting in all but one case on follow-up imaging. The PHMMR was gray on the initial MRI in seven of 15 (47%) of cases that developed tears compared to four of 29 (14%) controls (p < 0.0001). There was medial meniscal extrusion (MME) prior to tear in two of 15 (13%) patients and in ten of 15 (67%) patients after PHMMR failure. In the control group, MME was present in one (3%) and three (10%) of 29 subjects on the initial and follow-up MRIs, respectively. Articular cartilage loss was noted in two of 15 (15%) cases before tear and nine of 15 (69%) on follow-up imaging, as compared to one (3%) and four (14%) of 29 subjects in the control group. Subcortical marrow edema deep to the PHMMR may result from abnormal stresses and thus be a harbinger of meniscal root failure. This hypothesis is supported by resolution of these marrow signal changes after root tear. Following tear, extrusion of the meniscal body results in increased stress on the medial weight-bearing surfaces often leading to articular cartilage loss; we observed this sequence in six of our 15 patients with PHMMR tears.