Efficiently directing differentiation and homing of mesenchymal stem cells to boost cartilage repair in osteoarthritis via a nanoparticle and peptide dual-engineering strategy.

Mesenchymal stem cells (MSCs) are expected to be useful therapeutics in osteoarthritis (OA), the most common joint disorder characterized by cartilage degradation. However, evidence is limited with regard to cartilage repair in clinical trials because of the uncontrolled differentiation and weak cartilage-targeting ability of MSCs after injection. To overcome these drawbacks, here we synthesized CuO@MSN nanoparticles (NPs) to deliver Sox9 plasmid DNA (favoring chondrogenesis) and recombinant protein Bmp7 (inhibiting hypertrophy). After taking up CuO@MSN/Sox9/Bmp7 (CSB NPs), the expressions of chondrogenic markers were enhanced while hypertrophic markers were decreased in response to these CSB-engineered MSCs. Moreover, a cartilage-targeted peptide (designated as peptide W) was conjugated onto the surface of MSCs via a click chemistry reaction, thereby prolonging the residence time of MSCs in both the knee joint cavity of mice and human-derived cartilage. In a surgery-induced OA mouse model, the NP and peptide dual-modified W-CSB-MSCs showed an enhancing therapeutic effect on cartilage repair in knee joints compared with other engineered MSCs after intra-articular injection. Most importantly, W-CSB-MSCs accelerated cartilage regeneration in damaged cartilage explants derived from OA patients. Thus, this new peptide and NPs dual engineering strategy shows potential for clinical applications to boost cartilage repair in OA using MSC therapy.

The N-terminal fragment of histone deacetylase 4 (1-669aa) promotes chondrocyte apoptosis via the p53-dependent endoplasmic reticulum stress pathway.

Exogenous administration of the histone deacetylation 4 (HDAC4) protein can effectively delay osteoarthritis (OA) progression. However, HDAC4 is unstable and easily degrades into N-terminal (HDAC4-NT) and C-terminal fragments, and the HDAC4-NT can exert biological effects, but little is known about its role in chondrocytes and cartilage. Thus, the roles of HDAC4-NT fragments (1-289aa, 1-326aa and 1-669aa) in chondrocytes and cartilage were evaluated via real-time cell analysis (RTCA), safranin O staining, Sirius Red staining and nanoindentation. Molecular mechanisms were profiled via whole-transcriptome sequencing (RNA-seq) and verified in vitro and in vivo by a live cell real-time monitoring system, flow cytometry, western blotting and immunohistochemistry. The results showed that 1-669aa induced chondrocyte death and cartilage injury significantly, and the differentially expressed genes (DEGs) were enriched mainly in the apoptotic term and p53 signalling pathway. The validation experiments showed that 1-669aa induced chondrocyte apoptosis via the endoplasmic reticulum stress (ERS) pathway, and up-regulated p53 expression was essential for this process. Thus, we concluded that the HDAC4-NT fragment 1-669aa induces chondrocyte apoptosis via the p53-dependent ERS pathway, suggesting that in addition to overexpressing HDAC4, preventing it from degradation may be a new strategy for the treatment of OA.

mTORC2 inhibition by JR-AB2-011 improves IL-1ß-induced inflammation, catabolic response, and apoptosis in human chondrocytes through IκB-α/NF-κB p65.

Osteoarthritis (OA) is a very common chronic joint condition marked by inflammation and cartilage loss. mTOR is a well-known mediator of inflammation, cell survival, and aging; however, its role in OA has not been determined. To explore the role of mTORC2 in OA-and associated pathological changes, we examined the contribution of mTORC2-mediated Akt, rictor and IκB-α/NF-κB p65 pathway in interleukin (IL)-1ß-treated human chondrocytes. We focused on the protein expression of proinflammatory cytokines and catabolic and apoptotic factors, including TNF-α, IL-6, iNOS, MMP13, Bax, and caspase3, which may occur through this signalling pathway in IL-1ß-treated chondrocytes. Chondrocytes were cultured and treated with either 2 ng/mL IL­1ß alone or in combination with increasing concentrations of JR-AB2-011 (50, 100, or 250 µM), a selective mTORC2 inhibitor. The protein levels of phosphorylated (p)­Akt, Akt, rictor, p-NF-κB p65, NF-κB p65, IκB-α, p-IκB-α, iNOS, MMP13, Bax, and caspase3 were evaluated by Western blotting. In IL-1ß-stimulated chondrocytes, mTORC2 activity was increased with increased phosphorylation of Akt and expression of rictor. IL-1ß increased the expression of p-IκBα, p-NF-κB p65, NF-κB p65, IL-6, TNF-α, iNOS, Bax, and caspase3 proteins and decreased the expression of IκB-α. All of these IL-1ß-induced alterations were prevented by JR-AB2-011. The main novel finding in the present study is that selective mTORC2 inhibition by JR-AB2-011 prevents the inflammatory, catabolic, and apoptotic responses induced by IL-1ß via modulation of IκB-α/NF-κB activity. Therefore, we demonstrated a previously unknown function of mTORC2 inhibition that seems to be a potential therapeutic target for OA.

ChatGPT-4 and wearable device assisted Intelligent Exercise Therapy for co-existing Sarcopenia and Osteoarthritis (GAISO): a feasibility study and design for a randomized controlled PROBE non-inferiority trial.

BACKGROUND: Sarcopenia and osteoarthritis are prevalent age-related diseases that mutually exacerbate each other, creating a vicious cycle that worsens both conditions. Exercise is key to breaking this detrimental cycle. Facing increasing demand for rehabilitation services within this patient demographic, ChatGPT-4 and wearable device may increase the availability, efficiency and personalization of such health care. AIM: To evaluate the clinical efficacy and cost-effectiveness of a rehabilitation system implemented on mobile platforms, utilizing the integration of ChatGPT-4 and wearable devices. METHODS: The study design is a prospective randomized open blinded end-point (PROBE) non-inferiority trial. 278 patients diagnosed with osteoarthritis and sarcopenia will be recruited and randomly assigned to the intervention group and the control group. In the intervention group patients receive mobile phone-based rehabilitation service where ChatGPT-4 generates personalized exercise therapy, and wearable device guides and monitor the patient to implement the exercise therapy. Traditional clinic based face-to-face exercise therapy will be prescribed and implemented in the control group. All patients will receive three-months exercise therapies following the frequency, intensity, type, time, volume and progression (FITT-VP) principle. The patients will be assessed at baseline, one month, three months, and six months after initiation. Outcome measures will include ROM, gait patterns, Visual Analogue Scale (VAS) for pain assessment, Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC), Knee Injury and Osteoarthritis Outcome Score (KOOS) for functional assessment, Short-Form Health Survey 12 (SF-12) for quality of life, Minimal Clinically Important Difference (MCID), Patient Acceptable Symptom State (PASS), and Substantial Clinical Benefit (SCB) for clinically significant measures. DISCUSSION: A rehabilitation system combining the capabilities of ChatGPT-4 and wearable devices potentially enhance the availability and efficiency of professional rehabilitation services, thus enhancing the therapeutic outcomes for a substantial population concurrently afflicted with sarcopenia and osteoarthritis.

Amentoflavone maintaining extracellular matrix homeostasis and inhibiting subchondral bone loss in osteoarthritis by inhibiting ERK, JNK and NF-κB signaling pathways.

Amentoflavone (AF), a plant biflavone isolated from Selaginella sinensis ethanol extract, is characterized by anti-inflammatory and anti-oxidant properties. According to previous studies, inflammation and oxidative stress are closely related to the pathophysiology of osteoarthritis (OA). However, the effects and mechanisms of AF on OA have not been elucidated.To investigate the inhibitory effects and its molecular mechanism of AF on extracellular matrix (ECM) degradation stimulated by IL-1ß as well as subchondral bone loss induced by RANKL in mice chondrocytes. Quantitative PCR was used to detect the mRNA expression of genes related to inflammation, ECM, and osteoclast differentiation. Protein expression level of iNOS, COX-2, MMP13, ADAMTS5, COL2A1, SOX9, NFATc1, c-fos, JNK, ERK, P65, IκBα was measured by western blotting. The levels of TNF-α and IL-6 in the supernatants were measured by ELISA. The amount of ECM in chondrocytes was measured using toluidine blue staining. The levels of Aggrecan and Col2a1 in chondrocytes were measured using immunofluorescence. Tartrate-resistant acid phosphatase (TRAP) staining, F-actin staining and immunofluorescence were used to detect the effect of AF on osteoclast differentiation and bone resorption. The effect of AF on destabilization of the medial meniscus (DMM)-induced OA mice can be detected in hematoxylin-eosin (H&E) staining, Safranin O green staining and immunohistochemistry.AF might drastically attenuated IL-1ß-stimulated inflammation and reduction of ECM formation by blocking ERK and NF-κB signaling pathways in chondrocytes. Meanwhile, AF suppressed the formation of osteoclasts and the resorption of bone function induced by RANKL. In vivo, AF played a protective role by stabilizing cartilage ECM and inhibiting subchondral bone loss in destabilization of the medial meniscus (DMM)-induced OA mice, further proving its protective effect in the development of OA. Our study show that AF alleviated OA by suppressing ERK, JNK and NF-κB signaling pathways in OA models in vitro and DMM-induced OA mice, suggesting that AF might be a potential therapeutic agent in the treatment of OA.

Inhibitory Effects of Reynoutria japonica Houtt. on Pain and Cartilage Breakdown in Osteoarthritis Based on Its Multifaceted Anti-Inflammatory Activity: An In Vivo and In Vitro Approach.

In the past 30 years, the number of years lived with disability due to osteoarthritis (OA) has doubled, making it an increasing global health burden. To address this issue, interventions that inhibit the progressive pathology driven by age-related low-grade inflammation, the primary mechanism of OA, are being actively pursued. Recent investigations have focused on modulating the age-related low-grade inflammatory pathology of this disease as a therapeutic target. However, no agent has successfully halted the disease's progression or reversed its irreversible course. Reynoutria japonica Houtt. (RJ), a promising East Asian herbal medicine, has been utilized for several diseases due to its potent anti-inflammatory activity. This study aims to determine RJ's capacity to inhibit OA symptoms and associated inflammation, exploring its potential for further development. In vivo and in vitro experiments demonstrated RJ's anti-OA activity and modulation of multifaceted inflammatory targets. RJ significantly inhibited pain, gait deterioration, and cartilage destruction in a monosodium iodoacetate-induced OA rat model, with its analgesic effect further confirmed in an acetic acid-induced writhing model. RJ exhibited consistent anti-inflammatory activity against multiple targets in serum and cartilage of the OA rat model and lipopolysaccharide-induced RAW 264.7 cells. The inhibition of inflammatory cytokines, including interleukin-1ß, interleukin-6, matrix metalloproteinase-13, tumor necrosis factor-α, and nitric oxide synthase 2, suggests that RJ's alleviation of OA manifestations relates to its multifaceted anti-inflammatory activity. These results indicate that RJ merits further investigation as a disease-modifying drug candidate targeting OA's inflammatory pathology. To further characterize the pharmacological properties of RJ, future studies with expanded designs are warranted.

The Role of Adipokines between Genders in the Pathogenesis of Osteoarthritis.

Osteoarthritis (OA) is a chronic, progressive, degenerative joint disease characterized by joint pain, stiffness, and limited movement. It presents significant intra- and inter-individual variability-in particular, between genders. Recent research has increasingly focused on the role of adipokines-especially leptin, adiponectin, and resistin-in the development of OA. Adipokines, peptide hormones primarily secreted by adipose tissue, are involved in crucial physiological processes related to metabolism and immunity. They can also impact bone and cartilage turnover by interacting with joint cells such as osteoblasts, osteoclasts, chondrocytes, and mesenchymal stem cells, thereby linking inflammation with bone cartilage homeostasis. This review aims to elucidate the structure and functions of various adipokines, their serum and synovial levels, and their association with clinical presentation and radiographic progression in OA patients, with a focus on differences between sexes. A narrative literature review was conducted using three databases specifically analyzing sex differences. OA patients generally show elevated serum and synovial levels of leptin, chemerin, and visfatin, as well as high plasma levels of resistin and visfatin. In contrast, synovial levels of adiponectin and omentin are reduced in OA patients compared to healthy individuals, with an inverse relationship to disease severity, suggesting a potential protective role. Resistin and leptin were positively correlated with pain severity and radiographic progression, while adiponectin's role in OA remains controversial. Regarding sex differences, male OA patients exhibited higher serum levels of leptin, chemerin, and omentin compared to healthy controls, with a positive correlation to the BMI and estrogen levels, potentially explaining the sexual dimorphism observed in this condition. Studies on visfatin and lipocalin did not reveal significant differences in synovial or serum levels between the sexes. The role of resistin remains controversial. Adipokines influence the joint microenvironment and contribute to the progression of osteoarthritis (OA). However, the precise biological mechanisms are not yet fully understood due to the complex interactions between the metabolic, mechanical, and immune systems. Further research is needed to clarify their roles in OA and to identify targeted therapies for managing this degenerative disease.

Injections of the Hand and Wrist: Part I. Trigger Finger, First Carpometacarpal Joint Osteoarthritis, and Palmar Fibromatosis.

Family physicians are well-positioned to provide injections for patients who have pain due to hand and finger conditions, especially when initial treatments such as splinting and nonsteroidal anti-inflammatory drugs are ineffective. Corticosteroid injections can offer pain relief; however, potential risks such as infection, cartilage damage, and skin depigmentation should be discussed. Techniques and procedures for injections vary. Corticosteroid injections for ste-nosing flexor tenosynovitis (trigger finger) can be performed with or without ultrasound guidance. To maximize benefits of corticosteroid injection for carpometacarpal joint osteoarthritis, topical nonsteroidal anti-inflammatory drugs and other conservative treatment modalities should be used concurrently. Because of the risks of disease recurrence and adverse effects, corticosteroid injections for palmar fibromatosis should be approached with caution in the context of shared decision-making.

Intra-articular corticosteroid injections for osteoarthritis: A qualitative study of patients' and clinicians' experiences.

BACKGROUND: Osteoarthritis is a leading cause of joint pain and disability. Intra-articular corticosteroid injections (IACs) are often used in primary care once other recommended treatments have failed. Evidence shows that IACs provide short-term relief of osteoarthritis symptoms, yet little is known about patients' and primary care clinicians' experiences and beliefs about their use. We explored patients' and primary care clinicians' views about IACs, including the benefits, disadvantages, perceived risks of treatment, when they are used, and factors that affect decision-making. METHODS: We conducted individual interviews with patients and primary care clinicians and used inductive thematic analysis to investigate their views and experiences of intra-articular corticosteroid injections for osteoarthritis (IACs). FINDINGS: We interviewed 38 patients and 19 primary care clinicians. We identified 6 patient themes: variation in access; awareness of IACs; views of risk and trust; effectiveness of IACs; variation in onset and effect duration; and an alternative to undesirable treatments. In the interviews with clinicians, we identified an overarching theme of caution and competence, which included eight subthemes: confidence and (dis)comfort with practical procedures; risk of adverse outcomes; training; uncertainty about evidence and guidelines; technical uncertainties; IACs use on the osteoarthritis pathway; perceived benefits and impacts of IACs; and the possibility of placebo. CONCLUSION: Patients and clinicians valued IACs' potential to relieve symptoms and improve quality of life. Variability in patients' access to treatment appears related to clinicians' confidence in delivering injections and their concerns about the evidence base. Variation in dose frequency and timing reflect clinicians' uncertainty about current guidance. Despite variation in effectiveness patients preferred IACs to other forms of pain medication and to delay or avoid surgery. IACs were mostly used as an adjunct treatment before surgery was offered. These findings can inform further research into the effectiveness of IACs and improvements in information and guidance.

Anti-inflammatory effects of polydeoxyribonucleotide and adipose tissue-derived mesenchymal stem cells in a canine cell model of osteoarthritis.

IMPORTANCE: A relatively new therapeutic agent for osteoarthritis (OA), polydeoxyribonucleotide (PDRN), shows potential in treating human OA due to its regenerative and anti-inflammatory effects. However, studies on PDRN for canine OA are limited, and no study has investigated their use with mesenchymal stem cells (MSCs) conventionally used for OA treatment. OBJECTIVE: This study aimed to evaluate the potential of PDRN and explore its combined effect with adipose tissue-derived MSCs (AdMSCs) in treating canine OA. METHODS: To study the impact of PDRN, canine chondrocytes, synoviocytes, and AdMSCs were exposed to various PDRN concentrations, and viability was assessed using cell counting kit-8. The OA model was created by treating chondrocytes and synoviocytes with lipopolysaccharide, followed by treatment under three different conditions: PDRN alone, AdMSCs alone, and a combination of PDRN and AdMSCs. Using real-time quantitative polymerase chain reaction, the anti-inflammatory effects and mechanisms were investigated by quantitatively assessing pro-inflammatory cytokines, collagen degradation markers, adenosine A2a receptor (ADORA2A), and nuclear factor-kappa B. RESULTS: PDRN alone and combined with AdMSCs significantly reduced the expression of pro-inflammatory cytokines and collagen degradation markers in an OA model. PDRN promoted AdMSC proliferation and upregulated ADORA2A expression. AdMSCs exhibited comprehensive anti-inflammatory effects through paracrine effects, and both substances reduced inflammatory gene expression through different mechanisms, potentially enhancing therapeutic effects. CONCLUSIONS AND RELEVANCE: The results indicate that PDRN is a safe and effective anti-inflammatory material that can be used independently or as an adjuvant for AdMSCs. Although additional research is necessary, this study is significant because it provides a foundation for future research at the cellular level.

Targeting PAR2-mediated inflammation in osteoarthritis: a comprehensive in vitro evaluation of oleocanthal's potential as a functional food intervention for chondrocyte protection and anti-inflammatory effects.

BACKGROUND: Osteoarthritis (OA) is a prevalent degenerative joint disease characterized by chronic inflammation and progressive cartilage degradation, ultimately leading to joint dysfunction and disability. Oleocanthal (OC), a bioactive phenolic compound derived from extra virgin olive oil, has garnered significant attention due to its potent anti-inflammatory properties, which are comparable to those of non-steroidal anti-inflammatory drugs (NSAIDs). This study pioneers the investigation into the effects of OC on the Protease-Activated Receptor-2 (PAR-2) mediated inflammatory pathway in OA, aiming to validate its efficacy as a functional food-based therapeutic intervention. METHODS: To simulate cartilage tissue in vitro, human bone marrow-derived mesenchymal stem cells (BMSCs) were differentiated into chondrocytes. An inflammatory OA-like environment was induced in these chondrocytes using lipopolysaccharide (LPS) to mimic the pathological conditions of OA. The therapeutic effects of OC were evaluated by treating these inflamed chondrocytes with various concentrations of OC. The study focused on assessing key inflammatory markers, catabolic enzymes, and mitochondrial function to elucidate the protective mechanisms of OC. Mitochondrial function, specifically mitochondrial membrane potential (ΔΨm), was assessed using Rhodamine 123 staining, a fluorescent dye that selectively accumulates in active mitochondria. The integrity of ΔΨm serves as an indicator of mitochondrial and bioenergetic function. Additionally, Western blotting was employed to analyze protein expression levels, while real-time polymerase chain reaction (RT-PCR) was used to quantify gene expression of inflammatory cytokines and catabolic enzymes. Flow cytometry was utilized to measure cell viability and apoptosis, providing a comprehensive evaluation of OC's therapeutic effects on chondrocytes. RESULTS: The results demonstrated that OC significantly downregulated PAR-2 expression in a dose-dependent manner, leading to a substantial reduction in pro-inflammatory cytokines, including TNF-α, IL-1ß, and MCP-1. Furthermore, OC attenuated the expression of catabolic markers such as SOX4 and ADAMTS5, which are critically involved in cartilage matrix degradation. Importantly, OC was found to preserve mitochondrial membrane potential (ΔΨm) in chondrocytes subjected to inflammatory stress, as evidenced by Rhodamine 123 staining, indicating a protective effect on cellular bioenergetics. Additionally, OC modulated the Receptor Activator of Nuclear Factor Kappa-Β Ligand (RANKL)/Receptor Activator of Nuclear Factor Kappa-Β (RANK) pathway, suggesting a broader therapeutic action against the multifactorial pathogenesis of OA. CONCLUSIONS: This study is the first to elucidate the modulatory effects of OC on the PAR-2 mediated inflammatory pathway in OA, revealing its potential as a multifaceted therapeutic agent that not only mitigates inflammation but also protects cartilage integrity. The preservation of mitochondrial function and modulation of the RANKL/RANK pathway further underscores OC's comprehensive therapeutic potential in counteracting the complex pathogenesis of OA. These findings position OC as a promising candidate for integration into nutritional interventions aimed at managing OA. However, further research is warranted to fully explore OC's therapeutic potential across different stages of OA and its long-term effects in musculoskeletal disorders.

A RANKL-UCHL1-sCD13 negative feedback loop limits osteoclastogenesis in subchondral bone to prevent osteoarthritis progression.

Abnormal subchondral bone remodeling plays a pivotal role in the progression of osteoarthritis (OA). Here, we analyzed subchondral bone samples from OA patients and observed a significant upregulation of ubiquitin carboxy-terminal hydrolase L1 (UCHL1) specifically in subchondral bone osteoclasts. Notably, we found a strong correlation between UCHL1 expression and osteoclast activity in the subchondral bone during OA progression in both human and murine models. Conditional UCHL1 deletion in osteoclast precursors exacerbated OA progression, while its overexpression, mediated by adeno-associated virus 9, alleviated this process in male mice. Mechanistically, RANKL stimulates UCHL1 expression in osteoclast precursors, subsequently stabilizing CD13, augmenting soluble CD13 (sCD13) release, and triggering an autocrine inhibitory effect on the MAPK pathway, thereby suppressing osteoclast formation. These findings unveil a previously unidentified negative feedback loop, RANKL-UCHL1-sCD13, that modulates osteoclast formation and presents a potential therapeutic target for OA.

Evaluation of the protective dose-dependent effect of metformin for induced osteoarthritis in rats.

Objective: To evaluate the potential protective effect of metformin against osteoarthritis development in rats. METHODS: The experimental study was conducted at the Iraqi Centre for Cancer Research and Medical Genetics, Mustansiriyah University, Baghdad, Iraq, from December 2021 to February 2022, and comprised male Sprague- Dawley mice who were divided into 5 equal groups: negative control group, osteoarthritis group subjected to monoiodoacetate induction, positive control group treated with celecoxib 30mg/kg, metformin 100mg/kg group, and metformin 200mg/kg group. Body mass index, inflammatory biomarkers, and serum C-terminal cross-linked telopeptide of type II collagen levels were noted for all the animals. Data was analysed using SPSS 24. RESULTS: Of the 35 mice, 7(20%) were in each of the 5 groups. Compared to the osteoarthritis group, metformintreated mice showed significantly reduced body mass index, inflammatory biomarker levels, and blood levels of Cterminal cross-linked telopeptide of type II collagen (p=0.05). Metformin 200mg/kg treatment had more beneficial effects than 100mg/kg dose on inflammatory biomarkers and serum C-terminal cross-linked telopeptide of type II collagen (p=0.05). CONCLUSIONS: A beneficial protective effect against the onset of osteoarthritis was produced by metformin in a dosedependent way. Additionally, metformin could lessen cartilage damage as demonstrated by a decrease in the serum levels of C-terminal cross-linked telopeptide of type II collagen in the osteoarthritis group.

[The diabetic hand - a forgotten complication]. / Den diabetiska handen ­ ett i dag bortglömt syndrom.

The term ¼the diabetic hand« traditionally denotes complications affecting the hand in individuals with diabetes mellitus, such as restricted finger movement, numbness, and pain. Trigger finger, Dupuytren's disease, carpal tunnel syndrome, ulnar nerve entrapment, and osteoarthritis of the first carpometacarpal joint are all conditions that are more prevalent among individuals with both type 1 and type 2 diabetes. This overview aims to shed light on a somewhat neglected area in diabetes complications, i.e. the diabetic hand, to increase the knowledge among physicians and surgeons as well as nurses, physiotherapists, and occupational therapists treating patients with diabetes.

SIRT2 Alleviates Inflammatory Response, Apoptosis, and ECM Degradation in Osteoarthritic Chondrocytes by Stabilizing PCK1.

BACKGROUND: It has been reported that Sirtuin 2 (SIRT2) prevents phosphoenolpyruvate carboxykinase 1 (PCK1) degradation, which can be involved in aging-induced osteoarthritis (OA), but the molecular mechanism of SIRT2/PCK1 in chondrocytes has not been clarified. Therefore, this study aims to explore the mechanism of SIRT2/PCK1 in chondrocyte inflammation. METHOD: To establish the OA model in vitro, chondrocytes cultured with interleukin-1ß (IL-1ß, 10 ng/mL) and manipulation of SIRT2 and PCK1 expression in the constructed cells to elucidate the interaction between the two genes. 1,9-Dimethyl-Methylene Blue (DMMB) was used to detect cellular glycosaminoglycan (GAG) content. Inflammatory factor levels were assessed using Enzyme-linked Immunosorbent Assay (ELISA). Apoptosis was detected by osmotic dye. The expression of B-cell lymphoma-2 (Bcl-2), Bcl-2 Associated X (Bax), Wnt Family Member 1 (Wnt1), catenin Beta 1 (ß-catenin), Aggrecan, Collagen II, matrix metallopeptidase 13 (MMP-13) proteins in cells were analyzed using Western blot. RESULTS: PCK1 gained lower expressions in OA cell models. Overexpression of PCK1 or SIRT2 in the IL-1ß chondrocyte model of inflammation promoted GAG content, inhibited apoptosis and Wnt/ß-catenin protein expression, and lowered the levels of inflammatory factors. PCK1 silencing was proved to have the opposite effect. SIRT2 overexpression rescued the increased inflammation, MMP-13 expression, and apoptosis and the decreased Aggrecan and Collagen II expression caused by PCK1 silencing. PCK1 silencing also reversed the positive effects of SIRT2 overexpression on chondrocytes. CONCLUSION: SIRT2 inhibits articular chondrocyte extracellular matrix (ECM) degradation, inflammatory factor expression, and apoptosis via PCK1.

Histone demethylase PHF8 protected against chondrocyte injury and alleviated posttraumatic osteoarthritis by epigenetically enhancing WWP2 expression.

Aberrant mechanical forces were considered as an important factor for osteoarthritis (OA) pathogenesis. Plant homeodomain finger-containing protein 8 (PHF8) participated in osteogenic differentiation and inflammatory progression. However, the role of PHF8 in aberrant force-related OA remains to be elucidated. In this study, a fluid shear stress (FSS) model in ATDC5 cells and an anterior cruciate ligament transection (ACLT) animal model were constructed. The results revealed the decrease of PHF8 in aberrant force-induced cartilage damage in vitro and in vivo. PHF8 overexpression alleviated the aberrant force-induced cell apoptosis, extracellular matrix degradation, and inflammation. Chromatin immunoprecipitation (ChIP) assays demonstrated that PHF8 epigenetically regulated WWP2 expression through demethylating H3K9me2 at WWP2 promoter, which was influenced by FSS treatment. C-X-C chemokine receptor type 4 (CXCR4) was identified as a potential substrate of WWP2. Co-immunoprecipitation (Co-IP) and ubiquitination experiments further demonstrated WWP2 decreased the stability of CXCR4 via the ubiquitination pathway. Subsequently, rescue experiments validated reintroduction of WWP2 significantly attenuated the effects of PHF8 deletion on FSS-induced chondrocyte injury, and CXCR4 overexpression reversed the protective effects of WWP2 overexpression on chondrocyte injury in FSS-treated ATDC5 cells. Moreover, delivery of a PHF8 adeno-associated virus (AAV) into articular cartilage remarkably ameliorated the breakdown of cartilage matrix by ACLT in mice. In conclusion, our findings highlighted the importance of PHF8/WWP2/CXCR4 signaling pathway in aberrant force-induced cartilage injury, which might provide a novel insight on future epigenetic-based treatment of posttraumatic OA.

[Arthroscopic removal of the fragmented medial coronoid process in dogs - Evaluation of treatment results with special reference to postoperative computed tomography and residual particles]. / Arthroskopische Entfernung des fragmentierten Processus coronoideus medialis bei Hunden.

OBJECTIVE: The aim of this study was to assess the treatment success of dogs with medial coronoid disease (here: Fragmented Coronoid Process [FCP]) following arthroscopically performed subtotal coronoidectomy by the use of subjective and objective gait analysis and the evaluation of osteoarthritis progression in addition to their correlation with the findings from postoperative computed tomography. In particular, the clinical relevance of residual particles remaining postoperatively was to be evaluated. MATERIAL AND METHODS: Thirty elbows from 24 dogs of different breeds were examined clinically, orthopedically, using gait analysis as well as radiographically at least 6 months after unilateral or bilateral subtotal coronoidectomy. The results were compared with the preoperative findings of the corresponding examinations and postoperative computed tomography. RESULTS: 79.2% (19/24) of the patients improved by at least one degree of lameness in the orthopedic examination, 66.7% (16/24) were free of lameness. 60% (18/30 joints) showed an increase in the degree of arthrosis. Postoperative computed tomography revealed small residual bone particles (<1 mm) in the joint space or soft tissue in 56.7% (17/30) of the joints. No significant association was evident between residual lameness and residual particles remaining within the joint postoperatively. An association between persistent lameness and progression of arthrosis was also not be demonstrable. CONCLUSION: No evidence was apparent supporting a link between residues of small bone particles after arthroscopic subtotal coronoidectomy and postoperative lameness. The progression of osteoarthritis could not be interrupted despite the surgical intervention. CLINICAL RELEVANCE: When small residual particles (<1 mm) remain after subtotal coronoidectomy, these do not appear to have any influence on the clinical outcome.

Gene therapy for fat-1 prevents obesity-induced metabolic dysfunction, cellular senescence, and osteoarthritis.

Obesity is one of the primary risk factors for osteoarthritis (OA), acting through cross talk among altered biomechanics, metabolism, adipokines, and dietary free fatty acid (FA) composition. Obesity and aging have been linked to cellular senescence in various tissues, resulting in increased local and systemic inflammation and immune dysfunction. We hypothesized that obesity and joint injury lead to cellular senescence that is typically associated with increased OA severity or with aging and that the ratio of omega-6 (ω-6) to omega-3 (ω-3) FAs regulates these pathologic effects. Mice were placed on an ω-6-rich high-fat diet or a lean control diet and underwent destabilization of the medial meniscus to induce OA. Obesity and joint injury significantly increased cellular senescence in subcutaneous and visceral fat as well as joint tissues such as synovium and cartilage. Using adeno-associated virus (AAV) gene therapy for fat-1, a fatty acid desaturase that converts ω-6 to ω-3 FAs, decreasing the serum ω-6:ω-3 FA ratio had a strong senomorphic and therapeutic effect, mitigating metabolic dysfunction, cellular senescence, and joint degeneration. In vitro coculture of bone marrow-derived macrophages and chondrocytes from control and AAV8-fat1-treated mice were used to examine the roles of various FA mediators in regulating chondrocyte senescence. Our results suggest that obesity and joint injury result in a premature "aging" of the joint as measured by senescence markers, and these changes can be ameliorated by altering FA composition using fat-1 gene therapy. These findings support the potential for fat-1 gene therapy to treat obesity- and/or injury-induced OA clinically.

Exosomal microRNA-363 mediates the destructive effect of M1 macrophages on chondrocytes by repressing G3BP2.

M1 polarization of synovial macrophages contributes to cartilage degeneration and osteoarthritis (OA) development. However, limited knowledge is available about how M1 macrophages affect the biological properties of chondrocytes. This study aimed to explore the role of exosomal microRNAs (miRs) released from M1 macrophages in modulating the proliferation and survival of chondrocytes. Through bioinformatic analysis and experimental validation, we indicated that miR-363 was selectively induced in M1 macrophages (CD68+CD80+) but not M2 macrophages (CD68+CD206+). The upregulation of miR-363 in M1 macrophages depended on the activation of STAT1 signaling. Clinically, OA patients had a significantly higher miR-363 level in synovial fluid than control individuals without OA. Functional studies revealed that inhibition of miR-363 blocked the M1 macrophage polarization induced by lipopolysaccharide and IFN-γ. Moreover, exosomal miR-363 released from M1 macrophages significantly suppressed the proliferation and survival and induced inflammatory gene expression in chondrocytes. G3BP2 was identified as a target gene for miR-363 and could be negatively regulated by miR-363. Knockdown of G3BP2 recapitulated the effect of miR-363 overexpression on chondrocytes. Most importantly, enforced expression of G3BP2 attenuated miR-363-induced apoptosis and inflammatory response in chondrocytes. In conclusion, miR-363 plays an indispensable role in M1 macrophage polarization and can be released from M1 macrophages via exosomes to cause chondrocyte injury and inflammation. The miR-363/G3BP2 axis may represent a promising target for the prevention of OA development.

Identification of hub genes through integrated single-cell and microarray transcriptome analysis in osteoarthritic meniscus.

BACKGROUND: Osteoarthritis (OA) is marked by the progressive degradation of joint cartilage and subchondral bone. The precise molecular mechanisms driving meniscus deterioration in OA, especially at the single-cell level, remain poorly understood. METHOD: We analyzed two datasets from the GEO database, GSE220243 and GSE98918, focusing on meniscus tissue sequencing data from OA and non-OA patients. The standard Seurat procedure was employed to process single-cell data and identify differentially expressed genes (DEGs). Immune cell infiltration was assessed using the Microenvironment Cell Populations (MCP) counter and CIBERSORT algorithms. For the microarray data, DEGs were identified with the limma package, and Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were performed using ClusterProfiler. The overlapping DEGs from both datasets were imported into Cytoscape to generate protein-protein interaction (PPI) networks and identify hub genes. Transcription factor (TF) and miRNA interaction networks were analyzed using NetworkAnalyst, and gene-related predictive drugs were enriched through the DSigDB platform. RESULT: After quality control, 34,763 cells from the OA patients and 34,145 cells from the healthy controls were analyzed. UMAP identified and SingleR annotated 14 cell clusters. The 10 largest cell clusters were selected for further analysis. The OA group exhibited a notable increase in macrophages and a reduction in cytotoxic lymphocytes and endothelial cells in the meniscus. In GSE98918, 220 DEGs were identified, and the MCODE plug-in in Cytoscape pinpointed a key module containing 12 candidate genes. The MCC methodfiltered the top 20 DEGs in each GSE220243 cluster. Overlapping DEGs from GSE220243 and GSE98918 identified COL1A1, COL3A1, COL5A2, COL6A3, LOX, and VEGFA as significantly decreased in OA, with COL3A1, COL5A2, LOX, and VEGFA upregulated in meniscal chondrocytes. The interaction network highlighted 3 key miRNAs and 13 shared TFs. Ten gene-related predictive drug molecules were identified. CONCLUSION: This research highlights crucial genes in the OA meniscus and uncovers their differing regulatory patterns between chondrocytes and non-chondrocytes. These findings enhance our understanding of the molecular mechanisms driving OA pathogenesis and aid in identifying potential drug targets.