Osteoarthritis Disease Progression Research Articles

Heme oxygenase 1 linked to inactivation of subchondral osteoclasts in osteoarthritis.

Osteoarthritis (OA) is a chronic progressive osteoarthropathy in the elderly. Osteoclast activation plays a crucial role in the occurrence of subchondral bone loss in early OA. However, the specific mechanism of osteoclast differentiation in OA remains unclear. In our study, gene expression profiles related to OA disease progression and osteoclast activation were screened from the Gene Expression Omnibus (GEO) repository. GEO2R and Funrich analysis tools were employed to find differentially expressed genes (DEGs). Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses demonstrated that chemical carcinogenesis, reactive oxygen species (ROS), and response to oxidative stress were mainly involved in osteoclast differentiation in OA subchondral bone. Furthermore, fourteen DEGs that are associated with oxidative stress were identified. The first ranked differential gene, heme oxygenase 1 (HMOX1), was selected for further validation. Related results showed that osteoclast activation in the pathogenesis of OA subchondral bone is accompanied by the downregulation of HMOX1. Carnosol was revealed to inhibit osteoclastogenesis by targeting HMOX1 and upregulating the expression of antioxidant protein in vitro. Meanwhile, carnosol was found to alleviate the severity of OA by inhibiting the activation of subchondral osteoclasts in vivo. Our research indicated that the activation of osteoclasts due to subchondral bone redox dysplasia may serve as a significant pathway for the advancement of OA. Targeting HMOX1 in subchondral osteoclasts may offer novel insights for the treatment of early OA.

Research progress of Hippo pathway effector molecules in rheumatic immune system diseases.

The core components of the Hippo signaling pathway encompass upstream regulatory molecules, core kinase cascade complexes, and downstream transcriptional regulation complexes. This pathway modulates cellular biological behaviors by influencing the effector molecules of its core components and plays a pivotal role in immune regulation. Effector molecules,such as Yes-associated protein (YAP), transcriptional coactivator with PDZ-binding motif (TAZ), transcriptional enhanced associate domain transcriptional factor (TEAD), monopolar spindle-one binder (MOB1), large tumor suppressor (LATS), can stimulate fibroblast-like synovial cell migration and invasion in rheumatoid arthritis, regulate osteoarthritis disease progression, promote pathological new bone formation in ankylosing spondylitis, sustain submandibular gland development while delaying Sjogren's syndrome progression, mediate alpha-smooth muscle actin in systemic sclerosis, and refine the regulation of target genes associated with pulmonary fibrosis. This article provides an overview of the regulatory mechanisms involving Hippo signaling pathway-related effector molecules in the pathogenesis and progression of rheumatic immune system diseases, to serve as a reference for exploring novel therapeutic targets of rheumatic immune system diseases.

The role of monocyte/macrophage chemokines in pathogenesis of osteoarthritis: A review.

Osteoarthritis (OA) is one of the most common degenerative diseases characterised by joint pain, swelling and decreased mobility, with its main pathological features being articular synovitis, cartilage degeneration and osteophyte formation. Inflammatory cytokines and chemokines secreted by activated immunocytes can trigger various inflammatory and immune responses in articular cartilage and synovium, contributing to the genesis and development of OA. A series of monocyte/macrophage chemokines, including monocyte chemotaxis protein (MCP)-1/CCL2, MCP2/CCL8, macrophage inflammatory protein (MIP)-1α/CCL3, MIP-1β/CCL4, MIP-3α/CCL20, regulated upon activation, normal T-cell expressed and secreted /CCL5, CCL17 and macrophage-derived chemokine/CCL22, was proven to transmit cell signals by binding to G protein-coupled receptors on recipient cell surface, mediating and promoting inflammation in OA joints. However, the underlying mechanism of these chemokines in the pathogenesis of OA remains still elusive. Here, published literature was reviewed, and the function and mechanisms of monocyte/macrophage chemokines in OA pathogenesis were summarised. The symptoms and disease progression of OA were found to be effectively alleviated when the expression of these chemokines is inhibited. Elucidating these mechanisms could contribute to further understand how OA develops and provide potential targets for the early diagnosis of arthritis and drug treatment to delay or even halt OA progression.

Tranexamic Acid Attenuates the Progression of Posttraumatic Osteoarthritis in Mice

Background: Posttraumatic osteoarthritis (OA) is a common disorder associated with a high socioeconomic burden, particularly in young, physically active, and working patients. Tranexamic acid (TXA) is commonly used in orthopaedic trauma surgery as an antifibrinolytic agent to control excessive bleeding. Previous studies have reported that TXA modulates inflammation and bone cell function, both of which are dysregulated during posttraumatic OA disease progression. Purpose: To evaluate the therapeutic effects of systemic and topical TXA treatment on the progression of posttraumatic OA in the knee of mice. Study Design: Controlled laboratory study. Methods: OA was induced via anterior cruciate ligament (ACL) transection on the right knee of female mice. Mice were treated with TXA or vehicle intraperitoneally daily or intra-articularly weekly for 4 weeks, starting on the day of surgery. Articular cartilage degeneration, synovitis, bone erosion, and osteophyte formation were scored histologically. Micro−computed tomography evaluation was conducted to measure the subchondral bone microstructure and osteophyte volume. Cartilage thickness and bone remodeling were assessed histomorphometrically. Results: Both systemic and topical TXA treatment significantly reduced cartilage degeneration, synovitis, and bone erosion scores and increased the ratio of hyaline to calcified cartilage thickness in posttraumatic OA. Systemic TXA reversed ACL transection-induced subchondral bone loss and osteophyte formation, whereas topical treatment had no effect. Systemic TXA decreased the number and surface area of osteoclasts, whereas those of osteoblasts were not affected. No effect of topical TXA on osteoblast or osteoclast parameters was observed. Conclusion: Both systemic and topical TXA exerted protective effects on the progression of posttraumatic OA. Drug repurposing of TXA may, therefore, be useful for the prevention or treatment of posttraumatic OA, particularly after ACL surgery. Clinical Relevance: TXA might be beneficial in patients with posttraumatic OA of the knee.

Endoplasmic reticulum stress-related protein GRP78 and CHOP levels in synovial fluid correlate with disease progression of primary knee osteoarthritis: A cross-sectional study.

Endoplasmic reticulum (ER) stress has been shown to play an important role in osteoarthritis (OA). This study was aimed at assessing the relationship of endoplasmic reticulum (ER) stress-related glucose-regulated protein 78 (GRP78) and CCAAT/enhancer-binding protein homologous protein (CHOP) concentrations in the serum/synovial fluid (SF) with disease severity of primary knee osteoarthritis (pkOA). Patients with pkOA together with healthy individuals were consecutively recruited from our hospital. The levels of GRP78 and CHOP in serum / SF were detected using enzyme-linked immunosorbent assay. The levels of IL-6 and MMP-3 were also examined. Radiographic progression of pkOA was evaluated based on Kellgren-Lawrence (K-L) grades. Receiver Operating Characteristic (ROC) curves were used to assess the diagnostic value of GRP78/CHOP levels with regard to K-L grades. The assessment of clinical severity was conducted using the visual analogue scale (VAS), Oxford knee score (OKS), and Lequesne algofunctional index (LAI). A total of 140 pkOA patients and 140 healthy individuals were included. Serum GRP78 and CHOP levels in pkOA patients were not significantly different from those in healthy individuals. The SF GRP78 and CHOP levels in healthy controls were not detected due to ethical reasons. Compared to those with K-L grade 2 and 3, the pkOA patients with K-L grade 4 had higher GRP78 and CHOP levels in the SF with statistical significance. In addition, the pkOA patients with K-L grade 3 exhibited drastically upregulated GRP78 and CHOP concentrations in the SF compared to those with K-L grade 2. Positive correlations of GRP78 and CHOP levels with K-L grades, IL-6, and MMP-3 levels in the SF were observed. ROC curve analysis indicated that both GRP78 and CHOP levels may act as decent indicators with regard to OA. GRP78 and CHOP concentrations in the SF were positively correlated with VAS/LAI score and negatively associated with OKS score. The study indicated that GRP78 and CHOP levels in the SF but not the serum were positively correlated with disease severity of pkOA.

Evaluation of an Unloading Concept for Knee Osteoarthritis: A Pilot Study in a Small Patient Group.

Joint compressive forces have been identified as a risk factor for osteoarthritis disease progression. Therefore, unloader braces are a common treatment with the aim of relieving pain, but their effects are not clearly documented in the literature. A knee brace concept was tested with the aim of reducing joint loads and pain in knee osteoarthritis patients by applying an extension moment exclusively during the stance phase. The ideal effects were evaluated during gait based on musculoskeletal modeling of six patients, and experimental tests with a prototype brace were conducted on one patient. The effects were evaluated using electromyography measurements and musculoskeletal models to evaluate the muscle activation and knee compressive forces, respectively. The ideal brace simulations revealed a varying reduction of the first peak knee force between 3.5% and 33.8% across six patients whereas the second peak was unaffected. The prototype reduced the peak vasti muscle activation with 7.9% and musculoskeletal models showed a reduction of the first peak knee compressive force of up to 26.3%. However, the prototype brace increased the knee joint force impulse of up to 17.1% and no immediate pain reduction was observed. The reduction of the first peak knee compressive force, using a prototype on a single patient, indicates a promising effect from an applied knee extension moment for reducing knee joint loads during normal gait. However, further clinical experiments with this brace method are required to evaluate the long-term effects on both pain and disease progression in knee osteoarthritis patients.

THE MECHANICAL REGULATION OF SEMA3A IN PATIENT SYNOVIAL FLUID WITH CHRONIC KNEE VARUS MALALIGNMENT

AbstractOBJECTIVEKnee varus malalignment increases medial knee compartment loading and is associated with knee osteoarthritis (OA) progression and severity1. Altered biomechanical loading and dysregulation of joint tissue biology drive OA progression, but mechanistic links between these factors are lacking. Subchondral bone structural changes are biomechanically driven, involve bone resorption, immune cell influx, angiogenesis, and sensory nerve invasion, and contribute to joint destruction and pain2. We have investigated mechanisms underlying this involving RANKL and alkaline phosphatase (ALP), which reflect bone resorption and mineralisation respectively3 and the axonal guidance factor Sema3A. Sema3A is osteotropic, expressed by mechanically sensitive osteocytes, and an inhibitor of sensory nerve, blood vessel and immune cell invasion4. Sema3A is also differentially expressed in human OA bone5.HYPOTHESIS: Medial knee compartment overloading in varus knee malalignment patients causes dysregulation of bone derived Sema3A signalling directly linking joint biomechanics to pathology and pain.METHODSSynovial fluid obtained from 30 subjects with medial knee OA (KL grade II-IV) undergoing high tibial osteotomy surgery (HTO) was analysed by mesoscale discovery and ELISA analysis for inflammatory, neural and bone turnover markers. 11 of these patients had been previously analysed in a published patient-specific musculoskeletal model6 of gait estimating joint contact location, pressure, forces, and medial-lateral condyle load distribution in a published data set included in analyses. Data analysis was performed using Pearson's correlation matrices and principal component analyses. Principal Components (PCs) with eigenvalues greater than 1 were analysed.RESULTSPC1 (32.94% of variation) and PC2 (25.79% of variation) from PCA analysis and correlation matrices separated patients according to correlated clusters of established inflammatory markers of OA pain and progression (IL6/IL8, r=0.754, p<0.001) and anti-inflammatory mediators (IL4/IL10, r=0.469, p=0.005). Bone turnover marker ALP was positively associated with KL grade (r=0.815, p=0.002) and negatively associated with IL10 (r=−0.402, p=0.018) and first peak knee loading pressures (r=−0.688, p=0.019). RANKL was positively associated with IL4 (r=0.489, p=0.003). Synovial fluid Sema3A concentrations showed separate clustering from all OA progression markers and was inversely correlated with TNF-α (r=−0.423, p=0.022) in HTO patients. Sema3A was significantly inversely correlated with total predicted force in the medial joint compartment (r=−0.621, p=0.041), mean (r=−0.63, p=0.038) and maximum (r=−0.613, p=0.045) calculated medial compartment joint pressures during the first phase and mean (r=−0.618, p=0.043) and maximum (r=−0.641, p=0.034) medial compartment joint pressures during midstance outputs of patient-specific musculoskeletal model.CONCLUSIONSThis study shows joint inflammatory status and mechanical overloading influence subchondral bone-remodelling. Synovial Sema3A concentrations are inversely correlated to patient-specific musculoskeletal model estimations of pathological medial overloading. This study reveals Sema3A as a biological mediator with capacity to induce OA pain and disease progression that is directly regulated by gait mechanical loading.Declaration of Interest(b) declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research reported:I declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research project.

Targeted delivery of miR125a-5p and human Factor VIII attenuates molecular mediators of hemophilic arthropathy

Hemophilic arthropathy (HA) due to repeated bleeding into the joint cavity is a major cause of morbidity in patients with hemophilia. The molecular mechanisms contributing to this condition are not well characterized. MicroRNAs (miRs) are known to modulate the phenotype of multiple joint diseases such as osteoarthritis (OA) and rheumatoid arthritis (RA). Since miR125a is known to modulate disease progression in OA and RA, we performed a targeted screen of miR125a-5p and its target genes in a murine model of chronic HA. A digital PCR analysis demonstrated significant downregulation of miR125a-5p (2-fold vs control joint). Further molecular evaluation revealed elevated expression of the immunological markers STAT1 (7.6-fold vs control joint) and TRAF6 (10.6 fold vs control joint), which are direct targets of miR125a-5p. We then studied the impact of targeted overexpression of miR125a-5p using an Adeno-associated virus (AAV) vector in modulating the molecular mediators of HA. AAV5-miR125a vectors were administered intra-articularly either alone or in combination with a low dose of AAV8-based human factor 8 (F8) gene in a murine model of HA. We observed significantly increased expression of miR125a-5p in AAV5-miR125a administered mice (~12 fold vs injured joint) or in combination with AAV8-F8 vectors (~44 fold vs injured joint). The activity assay revealed ~17 %–20 % FVIII levels in mice that received low dose liver-directed F8 gene therapy. Further immunohistochemical analysis, demonstrated a decrease in inflammatory markers (STAT1 and TRAF6) and cartilage-degrading matrix metalloproteinases (MMPs) 3, 9, 13 in the joints of treated animals. These data highlight the crucial role of miR125a-5p in the development of HA.

Signatures of disease progression in knee osteoarthritis: insights from an integrated multi-scale modeling approach, a proof of concept.

Introduction: Knee osteoarthritis (KOA) is characterized by articular cartilage degeneration. It has been widely accepted that the mechanical joint environment plays a significant role in the onset and progression of this disease. In silico models have been used to study the interplay between mechanical loading and cartilage degeneration, hereby relying mainly on two key mechanoregulatory factors indicative of collagen degradation and proteoglycans depletion. These factors are the strain in collagen fibril direction (SFD) and maximum shear strain (MSS) respectively. Methods: In this study, a multi-scale in silico modeling approach was used based on a synergy between musculoskeletal and finite element modeling to evaluate the SFD and MSS. These strains were evaluated during gait based on subject-specific gait analysis data collected at baseline (before a 2-year follow-up) for a healthy and progressive early-stage KOA subject with similar demographics. Results: The results show that both SFD and MSS factors allowed distinguishing between a healthy subject and a KOA subject, showing progression at 2 years follow-up, at the instance of peak contact force as well as during the stance phase of the gait cycle. At the peak of the stance phase, the SFD were found to be more elevated in the KOA patient with the median being 0.82% higher in the lateral and 0.4% higher in the medial compartment of the tibial cartilage compared to the healthy subject. Similarly, for the MSS, the median strains were found to be 3.6% higher in the lateral and 0.7% higher in the medial tibial compartment of the KOA patient compared to the healthy subject. Based on these intersubject SFD and MSS differences, we were additionally able to identify that the tibial compartment of the KOA subject at risk of progression. Conclusion/discussion: We confirmed the mechanoregulatory factors as potential biomarkers to discriminate patients at risk of disease progression. Future studies should evaluate the sensitivity of the mechanoregulatory factors calculated based on this multi-scale modeling workflow in larger patient and control cohorts.

Three-dimensional ultrasound to investigate synovitis in first carpometacarpal osteoarthritis: A feasibility study.

Synovitis is one of the defining characteristics of osteoarthritis (OA) in the carpometacarpal (CMC1) joint of the thumb. Quantitative characterization of synovial volume is important for furthering our understanding of CMC1 OA disease progression, treatment response, and monitoring strategies. In previous studies, three-dimensional ultrasound (3-D US) has demonstrated the feasibility of being a point-of-care system for monitoring knee OA. However, 3-D US has not been tested on the smaller joints of the hand, which presents unique physiological and imaging challenges. To develop and validate a novel application of 3-D US to monitor soft-tissue characteristics of OA in a CMC1 OA patient population compared to the current gold standard, magnetic resonance imaging (MRI). A motorized submerged transducer moving assembly was designed for this device specifically for imaging the joints of the hands and wrist. The device used a linear 3-D scanning approach, where a 14L5 2-D transducer was translated over the region of interest. Two imaging phantoms were used to test the linear and volumetric measurement accuracy of the 3-D US device. To evaluate the accuracy of the reconstructed 3-D US geometry, a multilayer monofilament string-grid phantom (10mm square grid) was scanned. To validate the volumetric measurement capabilities of the system, a simulated synovial tissue phantom with an embedded synovial effusion was fabricated and imaged. Ten CMC1 OA patients were imaged by our 3-D US and a 3.0 T MRI system to compare synovial volumes. The synovial volumes were manually segmented by two raters on the 2D slices of the 3D US reconstruction and MR images, to assess the accuracy and precision of the device for determining synovial tissue volumes. The Standard Error of Measurement and Minimal Detectable Change was used to assess the precision and sensitivity of the volume measurements. Paired sample t-tests were used to assess statistical significance. Additionally, rater reliability was assessed using Intra-Class Correlation (ICC) coefficients. The largest percent difference observed between the known physical volume of synovial extrusion in the phantom and the volume measured by our 3D US was 1.1% (p-value=0.03). The mean volume difference between the 3-D US and the gold standard MRI was 1.78% (p-value=0.48). The 3-D US synovial tissue volume measurements had a Standard Error Measurement (SEm ) of 11.21mm3 and a Minimal Detectible Change (MDC) of 31.06mm3 , while the MRI synovial tissue volume measurements had an SEM of 16.82mm3 and an MDC of 46.63mm3 . Excellent inter- and intra-rater reliability (ICCs=0.94-0.99) observed across all imaging modalities and raters. Our results indicate the feasibility of applying 3-D US technology to provide accurate and precise CMC1 synovial tissue volume measurements, similar to MRI volume measurements. Lower MDC and SEm values for 3-D US volume measurements indicate that it is a precise measurement tool to assess synovial volume and that it is sensitive to variation between volume segmentations. The application of this imaging technique to monitor OA pathogenesis and treatment response over time at the patient's bedside should be thoroughly investigated in future studies.

Senescence-Driven Inflammatory and Trophic Microenvironment Imprints Mesenchymal Stromal/Stem Cells in Osteoarthritic Patients.

Senescent cells promote progressive tissue degeneration through the establishment of a combined inflammatory and trophic microenvironment. The cellular senescence state has therefore emerged as a central driving mechanism of numerous age-related diseases, including osteoarthritis (OA), the most common rheumatic disease. Senescence hallmarks are detectable in chondrocytes, synoviocytes and sub-chondral bone cells. This study investigates how the senescence-driven microenvironment could impact the cell fate of resident osteoarticular mesenchymal stromal/stem cells (MSCs) that are hence contributing to OA disease progression. For that purpose, we performed a comparative gene expression analysis of MSCs isolated from healthy donors that were in vitro chronically exposed either to interferon-gamma (IFN-γ) or Transforming Growth Factor beta 1 (TGFβ1), two archetypical factors produced by senescent cells. Both treatments reduced MSC self-renewal capacities by upregulating different senescence-driven cycle-dependent kinase inhibitors. Furthermore, a common set of differentially expressed genes was identified in both treated MSCs that was also found enriched in MSCs isolated from OA patients. These findings highlight an imprinting of OA MSCs by the senescent joint microenvironment that changes their matrisome gene expression. Altogether, this research gives new insights into OA etiology and points to new innovative therapeutic opportunities to treat OA patients.

Markers of Pain in 3 Different Murine Models of Posttraumatic Osteoarthritis: A Comparison Using Gait Analysis, Imaging, Histology, and Gene Expression

Background: Animal models play an important role in studying posttraumatic osteoarthritis (PTOA) disease progression. Different models exist, such as destabilization of the medial meniscus (DMM), anterior cruciate ligament (ACL) surgical transection (ACLs), and noninvasive ACL rupture. Purpose: To study the effects of PTOA on nociception in 3 different murine models and to relate these findings to macroscopic and microscopic changes in joint tissues. Study Design: Controlled laboratory study. Methods: A total of 42 male C57BL/6 mice, 12 weeks old, were randomly assigned to 4 groups: intact control (n = 10), DMM (n = 10), ACLs (n = 11), and closed ACL rupture (ACLc; n = 11) groups. Gait analysis was performed on 5 mice from the DMM group and 6 mice from ACLs and ACLc groups at 0, 1, 2, 4, 8, and 12 weeks after injury. At the 12-week time point, all mice underwent radiographs and then either micro–computed tomography imaging followed by histology and immunohistochemistry or gene expression analysis of the dorsal root ganglion and tibialis anterior muscle. Results: The peripheral and central pain markers were expressed at significantly higher levels in the synovium of both ACL injury groups when compared with the DMM group. Muscle atrophy genes were significantly upregulated in the ACL injury groups. Pain-related gait behavior started at 4 weeks for the ACL rupture groups and at 12 weeks for the DMM group. High-resolution radiographic imaging and histology demonstrated divergent changes in bone microstructure between the ACLs and DMM groups, suggesting different mechanical loading environments in these models. Conclusion: The principal finding of this study is the presence of markers of nociception at both the gene and the protein levels, with earlier expression in the ACL injury groups when compared with the DMM group. The second finding of this study is that the noninvasive ACL rupture model demonstrated changes comparable with those of the commonly used surgical ACL transection model, supporting use of this clinically realistic model in future studies of PTOA. Clinical Relevance: Quantitative clinical outcomes (imaging, pain scale, gait changes) related to osteoarthritis severity in an animal study, allowing for better understanding of clinical outcomes of osteoarthritis progression after ACL injuries in humans.

Spatial distribution of elements during osteoarthritis disease progression using synchrotron X-ray fluorescence microscopy

The osteochondral interface is a thin layer that connects hyaline cartilage to subchondral bone. Subcellular elemental distribution can be visualised using synchrotron X-ray fluorescence microscopy (SR-XFM) (1 μm). This study aims to determine the relationship between elemental distribution and osteoarthritis (OA) progression based on disease severity. Using modified Mankin scores, we collected tibia plates from 9 knee OA patients who underwent knee replacement surgery and graded them as intact cartilage (non-OA) or degraded cartilage (OA). We used a tape-assisted system with a silicon nitride sandwich structure to collect fresh-frozen osteochondral sections, and changes in the osteochondral unit were defined using quantified SR-XFM elemental mapping at the Australian synchrotron's XFM beamline. Non-OA osteochondral samples were found to have significantly different zinc (Zn) and calcium (Ca) compositions than OA samples. The tidemark separating noncalcified and calcified cartilage was rich in zinc. Zn levels in OA samples were lower than in non-OA samples (P = 0.0072). In OA samples, the tidemark had less Ca than the calcified cartilage zone and subchondral bone plate (P < 0.0001). The Zn–strontium (Sr) colocalisation index was higher in OA samples than in non-OA samples. The lead, potassium, phosphate, sulphur, and chloride distributions were not significantly different (P > 0.05). In conclusion, SR-XFM analysis revealed spatial elemental distribution at the subcellular level during OA development.

Trimethylamine-N-oxide sensitizes chondrocytes to mechanical loading through the upregulation of Piezo1.

Mechanical strain plays a crucial role in chondrocyte apoptosis and osteoarthritis (OA) disease progression through Piezo1. Trimethylamine-N-oxide (TMAO) is a diet-derived metabolite that correlates positively with multiple chronic diseases. Herein, we explored the potential role of TMAO in sensitizing chondrocytes to Piezo1-mediated mechanotransduction. The cytotoxicity of TMAO on chondrocytes was assayed. Piezo1 expression was measured after TMAO intervention. Pathological mechanical loading or Yoda1 (a specific Piezo1 channel activator) was administered in chondrocytes. The calcium levels and cytoskeleton in chondrocytes were observed by fluorescence microscopy. Flow cytometry, western blotting, and mitochondrial membrane potential assays were utilized to evaluate apoptosis. A rat OA model was constructed by anterior cruciate ligament transection. Hematoxylin-eosin staining, Safranin-O/Fast Green staining, immunochemistry, and TUNEL were applied to estimate OA severity. TMAO intervention alone did not affect chondrocyte viability up to 600μM. TMAO significantly increased Piezo1 expression and up-regulated intracellular calcium levels, further leading to cytoskeletal damage. Mechanical strain or Yoda1 treatment significantly induced chondrocyte apoptosis. Notably, TMAO intervention further aggravated chondrocyte apoptosis and cartilage destruction under pathological mechanical loading. TMAO significantly up-regulated Piezo1 expression and sensitized chondrocytes to mechanical loading, which may be closely related to the pathogenesis of OA.

Inhibition of Leukotriene A4 Hydrolase Suppressed Cartilage Degradation and Synovial Inflammation in a Mouse Model of Experimental Osteoarthritis.

Chronic inflammation plays an important role in the osteoarthritis (OA) pathology but how this influence OA disease progression is unclear. Leukotriene B4 (LTB4) is a potent proinflammatory lipid mediator generated from arachidonic acid through the sequential activities of 5-lipoxygenase, 5-lipoxygenase-activating protein, Leukotriene A4 hydrolase (LTA4H) and its downstream product LTB4. The aim of this study is to investigate the involvement and the potential therapeutic target of the LTB4 pathway in OA disease progression. Both clinical human cartilage samples (n = 7) and mice experimental OA models (n = 6) were used. The levels of LTA4H and leukotriene B4 receptor 1 were first examined using immunostaining in human OA/non-OA cartilage and mice experimental OA models. We also determined whether the LTA4H pathway was associated with cartilage degeneration and synovitis inflammation in OA mice models and human articular chondrocytes. We found that both LTA4H and LTB4 receptor (BLT1) were highly expressed in human and mice OA cartilage. Inhibition of LTA4H suppressed cartilage degeneration and synovitis in OA mice model. Furthermore, inhibition of LTA4H promoted cartilage regeneration by upregulating chondrogenic genes expression such as aggrecan (ACAN), collagen 2A1 (COL2A1), and SRY-Box transcription factor 9 (SOX9). Our results indicate that the LTA4H pathway is a crucial regulator of OA pathogenesis and suggest that LTA4H could be a therapeutic target in combat OA.

AXONAL GUIDANCE SIGNALLING IS REGULATED BY MECHANICAL LOADING AND INFLAMMATION IN A 3D MODEL OF HUMAN STEM CELL-DERIVED OSTEOCYTES

Osteoarthritis (OA) is a common cause of chronic pain. Subchondral bone is highly innervated, and bone structural changes directly correlate with pain in OA. Mechanisms underlying skeletal–neural interactions are under-investigated. Bone derived axon guidance molecules are known to regulate bone remodelling. Such signals in the nervous system regulate neural plasticity, branching and neural inflammation. Perturbation of these signals during OA disease progression may disrupt sensory afferents activity, affecting tissue integrity, nociception, and proprioception.Osteocyte mechanical loading and IL-6 stimulation alters axon guidance signalling influencing innervation, proprioception, and nociception.Human Y201 MSC cells, embedded in 3D type I collagen gels (0.05 × 106 cell/gel) in 48 well plastic or silicone (load) plates, were differentiated to osteocytes for 7 days before stimulation with IL-6 (5ng/ml) with soluble IL-6 receptor (sIL-6r (40ng/ml) or unstimulated (n=5/group), or mechanically loaded (5000 μstrain, 10Hz, 3000 cycles) or not loaded (n=5/group). RNA extracted 1hr and 24hrs post load was quantified by RNAseq whole transcriptome analysis (NovaSeq S1 flow cell 2 × 100bp PE reads and differentially expressed neurotransmitters identified (&gt;2-fold change in DEseq2 analysis on normalised count data with FDR p&lt;0.05). After 24 hours, extracted IL-6 stimulated RNA was quantified by RT-qPCR for neurotrophic factors using 2–∆∆Ct method (efficiency=94-106%) normalised to reference gene GAPDH (stability = 1.12 REfinder). Normally distributed data with homogenous variances was analysed by two-tailed t test.All detected axonal guidance genes were regulated by mechanical load. Axonal guidance genes were both down-regulated (Netrin1 0.16-fold, p=0.001; Sema3A 0.4-fold, p&lt;0.001; SEMA3C (0.4-fold, p&lt;0.001), and up-regulated (SLIT2 2.3-fold, p&lt;0.001; CXCL12 5-fold, p&lt;0.001; SEMA3B 13-fold, p&lt;0.001; SEMA4F 2-fold, p&lt;0.001) by mechanical load. IL6 and IL6sR stimulation upregulated SEMA3A (7-fold, p=0.01), its receptor Plexin1 (3-fold, p=0.03). Neutrophins analysed in IL6 stimulated RNA did not show regulation.Here we show osteocytes regulate multiple factors which may influence innervation, nociception, and proprioception upon inflammatory or mechanical insult. Future studies will establish how these factors may combine and affect nerve activity during OA disease progression.

Survivorship, clinical outcomes and indications for revision in uncemented unicompartmental knee arthroplasty: systematic review.

Modern uncemented unicompartmental knee arthroplasty (UKA) relies on the mechanics of the implant design and a biological bond at the bone-implant interface to create a secure fixation of its components. The aim of this systematic review was to determine implant survivorship, clinical outcomes and indications for revision in uncemented UKAs. A search strategy was employed using keywords related to UKAs and uncemented fixation to identify suitable studies. Both prospective and retrospective studies with a minimum of two year mean follow-up were included. Data was gathered on study design, implant type, patient demographics, survivorship, clinical outcome scores and the indications for revision. Methodological quality was assessed using a ten-point risk of bias scoring tool. Eighteen studies were included in the final review. The mean follow-up of studies ranged between 2-11 years. The primary outcome of survival demonstrated 5 year survivorship ranged between 91.7-100.0% and 10-year survivorship between 91.0-97.5%. Clinical and functional outcome scores were found to be excellent in the majority of studies with the remaining reporting good results. Revisions represented 2.7% of the total operations performed. There were 145 revisions with an overall revision rate of 0.8 per 100 observed component years. Osteoarthritis disease progression (30.2%) and bearing dislocations (23.8%) were the most common causes of implant failure. This review finds uncemented UKAs demonstrate comparable survivorship, clinical outcomes and safety profile to cemented UKAs to consider this fixation a suitable alternative in clinical use.

T Helper 17–Like Regulatory T Cells in Equine Synovial Fluid Are Associated With Disease Severity of Naturally Occurring Posttraumatic Osteoarthritis

Background: Infiltration of cluster of differentiation (CD) 3+ (CD3+) T cells into the synovium and synovial fluid occurs in most patients with posttraumatic osteoarthritis. During disease progression, proinflammatory T helper 17 cells and anti-inflammatory regulatory T cells infiltrate the joint in response to inflammation. This study aimed to characterize the dynamics of regulatory T and T helper 17 cell populations in synovial fluid from equine clinical patients with posttraumatic osteoarthritis to determine whether phenotype and function are associated with potential immunotherapeutic targets. Hypothesis: An imbalance of the ratio of regulatory T cells and T helper 17 cells would be associated with disease progression in posttraumatic osteoarthritis, suggesting opportunities for immunomodulatory therapy. Study Design: Descriptive laboratory study. Methods: Synovial fluid was aspirated from the joints of equine clinical patients undergoing arthroscopic surgery for posttraumatic osteoarthritis resulting from intra-articular fragmentation. Joints were classified as having mild or moderate posttraumatic osteoarthritis. Synovial fluid was also obtained from nonoperated horses with normal cartilage. Peripheral blood was obtained from horses with normal cartilage and those with mild and moderate posttraumatic osteoarthritis. Synovial fluid and peripheral blood cells were analyzed by flow cytometry, and native synovial fluid was analyzed by enzyme-linked immunosorbent assay. Results: CD3+ T cells represented 81% of lymphocytes in synovial fluid, which increased in animals with moderate posttraumatic osteoarthritis to 88.3% (P = .02). CD14+ macrophages were doubled in those with moderate posttraumatic osteoarthritis compared with mild posttraumatic osteoarthritis and controls (P < .001). Less than 5% of CD3+ T cells found within the joint were forkhead box P3 protein+ (Foxp3+) regulatory T cells, but a 4- to 8-times higher percentage of nonoperated and mild posttraumatic osteoarthritis joint regulatory T cells secreted interleukin (IL)–10 than peripheral blood Tregs (P < .005). T regulatory-1 cells that secreted IL-10 but did not express Foxp3 accounted for approximately 5% of CD3+ T cells in all joints. T helper 17 cells and Th17-like regulatory T cells were increased in those with moderate posttraumatic osteoarthritis (P < .0001) compared with mild and nonoperated patients. IL-10, IL-17A, IL-6, chemokine (C-C motif) ligand (CCL) 2 (CCL2), and CCL5 concentrations detected by enzyme-linked immunosorbent assay in synovial fluid were not different between groups. Conclusions: An imbalance of the ratio of regulatory T cells and T helper 17 cells and an increase in T helper 17 cell–like regulatory T cells in synovial fluid from joints with more severe disease provide novel insights into immunological mechanisms that are associated with posttraumatic osteoarthritis progression and pathogenesis. Clinical Relevance: Early and targeted use of immunotherapeutics in the mitigation of posttraumatic osteoarthritis may improve patient clinical outcomes.

Noise correction of atomic force microscopy-based integrin frequency analysis

As the most common joint disorder, Osteoarthritis (OA) is problematic for countries with an aging population. In OA, the cartilage degrades which causes bone on bone contact which results in constant pain for those affected. Treatments for OA are centered around symptom relief since there is no drug approved that will slow/stop OA disease progression. The cells that make up the cartilage are called chondrocytes. These chondrocytes must regulate themselves to build and break down the cartilage when necessary through cell signaling. When this signaling isn’t done properly, the chondrocytes may break down too much cartilage and result in OA. To better understand the signaling within the chondrocytes, an Atomic Force Microscope (AFM) was used to measure the movement and forces of mechanical receptor molecules (integrins) on the surface of the cell. However, because the measurements were near the sensitivity limit of the AFM, there was not enough certainty that noise (i.e., thermal, instrument and building) was not a dominant factor within the data. The focus of this project was to subtract model-based noise out of the displacement data to correct the data for further analysis. Instrument noise and building noise was found from the negative control, bare silica. Random thermal noise curves were generated based on theoretical thermal noise RMS values. A Fast Fourier Transform (FFT) was applied to each individual curve and each noise FFT was subtracted from the chondrocyte displacement curve in Python code. Inverse FFTs were applied to the resulting curves, resulting in noise-corrected time domain displacement curves.

Strategic application of imaging in DMOAD clinical trials: focus on eligibility, drug delivery, and semiquantitative assessment of structural progression.

Despite decades of research efforts and multiple clinical trials aimed at discovering efficacious disease-modifying osteoarthritis (OA) drugs (DMOAD), we still do not have a drug that shows convincing scientific evidence to be approved as an effective DMOAD. It has been suggested these DMOAD clinical trials were in part unsuccessful since eligibility criteria and imaging-based outcome evaluation were solely based on conventional radiography. The OA research community has been aware of the limitations of conventional radiography being used as a primary imaging modality for eligibility and efficacy assessment in DMOAD trials. An imaging modality for DMOAD trials should be able to depict soft tissue and osseous pathologies that are relevant to OA disease progression and clinical manifestations of OA. Magnetic resonance imaging (MRI) fulfills these criteria and advances in technology and increasing knowledge regarding imaging outcomes likely should play a more prominent role in DMOAD clinical trials. In this perspective article, we will describe MRI-based tools and analytic methods that can be applied to DMOAD clinical trials with a particular emphasis on knee OA. MRI should be the modality of choice for eligibility screening and outcome assessment. Optimal MRI pulse sequences must be chosen to visualize specific features of OA.