Extracellular vesicles (EVs) are crucial for intercellular communication and are found in various biological fluids. The identification and immunophenotyping of such small particles continue to pose significant challenges. Here, we have developed a workflow for the optimisation of a next-generation panel for in-depth immunophenotyping of circulating plasma EVs using spectral flow cytometry. Our data collection followed a multistep optimisation phase for both instrument setup and 21-colour panel design, thus maximising fluorescent signal recovery. This spectral approach enabled the identification of novel EV subpopulations. Indeed, besides common EVs released by erythrocytes, platelets, leukocytes and endothelial cells, we observed rare and poorly known EV subsets carrying antigens related to cell activation or exhaustion. Notably, the unsupervised data analysis of major EV subsets revealed subpopulations expressing up to five surface antigens simultaneously. However, the majority of EVs expressed only a single surface antigen, suggesting they may not fully represent the phenotype of their parent cells. This is likely due to the small surface area or the biogenesis of EVs rather than antibody steric hindrance. Finally, we tested our workflow by analysing the plasma EV landscape in a cohort of systemic lupus erythematosus (SLE) patients. Interestingly, we observed a significant increase in CD54+ EVs, supporting the notion of elevated circulating ICAM under SLE conditions. To our knowledge, these are the first data highlighting the importance of a spectral flow cytometry approach in deciphering the heterogeneity of plasma EVs paving the way for the routine use of a high-dimensional immunophenotyping in EV research.

Small extracellular vesicles (sEVs) derived from different osteoarthritic (OA) tissues regulate OA-related biological processes through transporting their content (proteins, miRNAs, etc.) to recipient cells. This study aimed to characterize the miRNA profile of synovial fibroblasts-derived small EVs (FS_OA_sEVs) and investigate their role in inflammation in chondrocytes. Chondrocytes were isolated from macroscopically preserved and lesioned OA cartilage (C_OAmin and C_OAmax) and synovial fibroblasts from OA synovium. Synovial fibroblasts-derived small EVs (FS_OA_sEVs) were characterized according to ISEV guidelines and used for miRNA profiling and bioinformatics analysis. miR-21-5p was identified as one of the most abundant, and its target genes, such as KLF6, were enriched in OA-related processes including inflammation. Treatment of C_OAmin chondrocytes with FS_OA_sEVs resulted in decreased expression of COL2A1 and ACAN and an increase in catabolic markers MMP-3 and MMP-13. Moreover, C-OAmin receiving FS_OA_sEVs exhibited increased levels of inflammatory markers and miR-21-5p expression, resembling chondrocytes' phenotype from lesioned OA cartilage, whereas miR-21-5p inhibition reversed their expression of inflammatory markers and miR-21-5p. Compared to C_OA min, C_OAmax chondrocytes exhibited increased miR-21-5p and inflammatory markers expression and decreased KLF6 expression. miR-21-5p inhibition in C_OAmax led to KLF6 upregulation and suppression of inflammatory mediators, whereas co-treatment with siRNA against KLF6 negated this effect, confirming a potential direct regulatory relationship between miR-21-5p and KLF6. Our results provide novel insights into the FS_OA_sEV-mediated inflammation axis, highlighting FS_OA_sEV-derived miR-21-5p as a driver of OA progression via regulating inflammation in chondrocytes.

The aim of this study is to analyze scientific literature to investigate the current research status, focus areas, and developmental trends in nanoparticle systems for rheumatoid arthritis (RA) therapy. To do that, Research articles published from 2003 to 2023 were retrieved from the Web of Science database, and analysis included quantitative output, distribution by country/region, collaborative publishing data, influential authors, high-yield institutions, keywords, hotspots, and development trends. Visual knowledge maps were generated using VOSviewer and Citespace. Findings reveal a steady increase in publications related to nanoparticle systems for RA therapy, indicating growing global interest. China leads with 487 papers (37.433%), followed by the United States (233, 17.909%), India (179, 13.759%), South Korea (89, 6.841%), and Egypt (50, 3.843%). Active collaboration is observed, particularly between the United States and countries such as China, Germany, Saudi Arabia, India, England, and Pakistan. The Chinese Academy of Sciences ranks first in total articles published (55), with Liu Y from China being the most prolific author. The Journal of Controlled Release emerges as a primary outlet in this field. Primary keyword clusters include "Drug delivery systems," "Gold nanoparticles," "Transdermal delivery," "Angiogenesis," "Collagen-induced arthritis," "Rheumatoid arthritis," "Oxidant stress," "Dendritic cells," and "pH sensitive." Research hotspots with great development potential include "Immunopathological Mechanisms," "Novel drugs," and "Smart delivery system." In conclusion, research on nanoparticle systems for RA therapy has significantly expanded over the past two decades, with a focus on elucidating pathogenetic mechanisms and advancing novel drug delivery strategies anticipated to be prominent in the foreseeable future.

Identifying effective druggable targets with disease-specific for diseases is a tremendous challenge in new drug development. However, current studies of druggable targets identification are most based on either druggability or disease-specific, lacking a combination of two factors. To further improve the accuracy of druggable targets discovery, a druggable target discovery strategy for diseases (DTDS) was proposed, which combined druggable targets prediction by machine learning and key targets identification by tissue-level and cellular-level transcriptomics analysis. Rheumatoid arthritis (RA), an autoimmune disease that cannot be treated entirely, was taken as a case. First, the protein-protein interaction network was constructed as the disease background network, and the classification models were established based on the topological parameters of known RA-druggable targets with druggability and non-RA targets without therapeutic effects on RA. 168 potential druggable targets were predicted by the classification models from 264 RA-related targets. Subsequently, 40 RA-specific targets were identified by tissue-level and cellular-level transcriptomics analysis from 168 potential druggable targets. Most of them were RA-druggable targets except PSMB9 and PTPRC. Finally, PSMB9 and PTPRC were further verified by in vitro experiments. The results showed that the inhibitor of PSMB9 or PTPRC could effectively inhibit inflammation and abnormal proliferation of synovial cells, proving that PSMB9 and PTPRC were potential RA-druggable targets, and further indicating that DTDS had high accuracy. In conclusion, the DTDS strategy established in this study is reliable and has been proven in identification of potential RA-druggable targets, which is expected to provide ideas and methods for systematic discovery of potential druggable targets for diseases.

The aging population has led to a global issue of osteoarthritis (OA), which not only impacts the quality of life for patients but also poses a significant economic burden on society. While biotherapy offers hope for OA treatment, currently available treatments are unable to delay or prevent the onset or progression of OA. Recent studies have shown that as nanoscale bioactive substances that mediate cell communication, exosomes from stem cell sources have led to some breakthroughs in the treatment of OA and have important clinical significance. This paper summarizes the mechanism and function of stem cell exosomes in delaying OA and looks forward to the development prospects and challenges of exosomes.

Rheumatoid arthritis (RA) is defined as a chronic autoimmune disease that severely influences a patient's quality of life. Extracellular vesicles (EVs) have gained much attention in recent years as one of the most potent therapeutic agents for the treatment of RA. A systematic review was performed with the purpose of assessing the current evidence relating to the therapeutic applications of EVs in RA. The systematic search was performed in the databases of PubMed, Scopus, and Web of Science, from inception times to September 2024. All studies investigating the use of EVs for the treatment of RA were included. The quality appraisal of selected articles and data extraction regarding EV characteristics, therapeutic applications, and associated outcomes were performed. Of the 1418 initially identified articles, 59 studies met inclusion criteria. Regarding their cellular origins, most EVs were derived from mesenchymal stem cells, followed by immune cells. The main therapeutic mechanisms included modulation of the immune response, reduction of inflammation, and repair of tissues. Recent trends are toward increasing interest in engineered EVs and combination therapies. Indeed, most studies reported positive outcomes with regard to lowered inflammation and improved joint function. On the other hand, standardization of the metrics of evaluation considerably varied between different studies. EVs are promising therapeutic agents in the treatment of RA by modulating immune responses. Standardization, delivery systems, and clinical translation are challenges yet to be overcome. Future studies will be directed to optimize EV engineering, targeted delivery systems, and large-scale clinical trials.

Exosomes are integral to the pathophysiology of osteoarthritis (OA) due to their roles in mediating intercellular communication and regulating inflammatory processes. Exosomes are integral to the transport of bioactive molecules, such as proteins, lipids and nucleic acids, which can influence chondrocyte behavior and joint homeostasis. Given their properties of regeneration and ability to target damaged tissues, exosomes represent a promising therapeutic avenue for OA treatment. Exosomes have potential in promoting cartilage repair, reducing inflammation and improving overall joint function. However, several challenges remain, including the need for standardized isolation and characterization methods, variability in exosomal content, and regulatory hurdles. The present review aims to provide a comprehensive overview of the current understanding of exosome mechanisms in OA and their therapeutic potential, while also addressing the ongoing challenges faced in translating these findings into clinical practice. By consolidating existing research, the present review aims to pave the way for future studies aimed at optimizing exosome‑based therapies for effective OA management.

Chronic osteomyelitis is a chronic bone infection characterized by progressive osteonecrosis and dead bone formation, which is closely related to persistent infection and chronic inflammation. Exosomes derived from bone marrow-derived mesenchymal stem cells (BMSC) play an important role in bone tissue regeneration and the modulation of inflammatory processes. However, their role and mechanism of action in osteomyelitis have not been reported so far. This paper explores the potential effect of BMSC-derived exosomes on osteomyelitis in vitro model with the aim of providing a theoretical basis for the treatment of osteomyelitis in the future. In this study, exosomes were isolated and extracted from BMSCs and identified. MC3T3-E1 cells were treated with Staphylococcal protein A (SPA) to establish an in vitro model of osteomyelitis. Next, the effects of BMSC-derived exosomes on cell proliferation, apoptosis, angiogenesis, and autophagy in MC3T3-E1 cells treated with SPA were evaluated. Results showed that the proliferation ability of MC3T3-E1 cells increased after co-culture with BMSC-derived exosomes. Moreover, exosomes induced autophagy and osteogenic differentiation in MC3T3-E1 cells. The mRNA and protein levels of factors related to proliferation, differentiation, apoptosis, autophagy, and angiogenesis including β-Catenin, Runx2, Bcl-2, VEGFA, and Beclin-1 upregulated in SPA-treated MC3T3-E1 cells, whereas the levels of inflammatory cytokines including TNF-α, IL-1β, and IL-6 decreased in the supernatant. The results showed that exosomes derived from BMSCs may participate in the attenuation of osteomyelitis by inducing proliferation and osteogenic differentiation and regulating the inflammatory state in bone cells.

Musculoskeletal conditions impact 1.71 billion individuals, posing significant challenges due to their complexity, varying clinical courses, and unclear molecular mechanisms. Conventional spectrum treatments often prove inadequate, underscoring the importance of targeted therapies. Recently, RNA-based technologies have emerged as a groundbreaking approach in therapeutics, showing applications in joint related ailments. This perspective aims to examine endeavors exploring the use of RNA-based treatments in both experimental and clinical contexts for addressing joint issues like osteoarthritis, rheumatoid arthritis, and cartilage injuries. The cited studies demonstrate how mRNA can stimulate the production of proteins that aid in controlling inflammation, fostering tissue regeneration and repairing cartilage damage. In summary, this perspective offers an overview of the progress made in mRNA-based technologies for treating related conditions by highlighting favorable findings from preclinical research and encouraging results from clinical trials. With advancements in the field, mRNA therapeutics have the potential to revolutionize treatment approaches for musculoskeletal disorders, bringing renewed hope to the future of musculoskeletal conditions.

To develop a protocol for largescale analysis of synovial fluid proteins, for the identification of biological networks associated with subtypes of osteoarthritis.

Synovial Fluid To detect molecular Endotypes by Unbiased Proteomics in Osteoarthritis (STEpUP OA) is an international consortium utilising clinical data (capturing pain, radiographic severity and demographic features) and knee synovial fluid from 17 participating cohorts. 1746 samples from 1650 individuals comprising OA, joint injury, healthy and inflammatory arthritis controls, divided into discovery (n = 1045) and replication (n = 701) datasets, were analysed by SomaScan Discovery Plex V4.1 (>7000 SOMAmers/proteins). An optimised approach to standardisation was developed. Technical confounders and batch-effects were identified and adjusted for. Poorly performing SOMAmers and samples were excluded. Variance in the data was determined by principal component (PC) analysis.

A synovial fluid standardised protocol was optimised that had good reliability (<20% co-efficient of variation for >80% of SOMAmers in pooled samples) and overall good correlation with immunoassay. 1720 samples and >6290 SOMAmers met inclusion criteria. 48% of data variance (PC1) was strongly correlated with individual SOMAmer signal intensities, particularly with low abundance proteins (median correlation coefficient 0.70), and was enriched for nuclear and non-secreted proteins. We concluded that this component was predominantly intracellular proteins, and could be adjusted for using an 'intracellular protein score' (IPS). PC2 (7% variance) was attributable to processing batch and was batch-corrected by ComBat. Lesser effects were attributed to other technical confounders. Data visualisation revealed clustering of injury and OA cases in overlapping but distinguishable areas of high-dimensional proteomic space.

We have developed a robust method for analysing synovial fluid protein, creating a molecular and clinical dataset of unprecedented scale to explore potential patient subtypes and the molecular pathogenesis of OA. Such methodology underpins the development of new approaches to tackle this disease which remains a huge societal challenge.

Osteoarthritis (OA) is a chronic progressive degenerative joint disease that affects a significant portion of the equine population and humans worldwide. Current treatment options for equine OA are limited and incompletely curative. Horses provide an excellent large-animal model for studying human OA. Recent advances in the field of regenerative medicine have led to the exploration of extracellular vesicles (EVs)-cargoes of microRNA, proteins, lipids, and nucleic acids-to evaluate their diagnostic value in terms of disease progression and severity, as well as a potential cell-free therapeutic approach for equine OA. EVs transmit molecular signals that influence various biological processes, including the inflammatory response, apoptosis, proliferation, and cell communication. In the present review, we summarize recent advances in the isolation and identification of EVs, the use of their biologically active components as biomarkers, and the distribution of the gap junction protein connexin 43. Moreover, we highlight the role of mesenchymal stem cell-derived EVs as a potential therapeutic tool for equine musculoskeletal disorders. This review aims to provide a comprehensive overview of the current understanding of the pathogenesis, diagnosis, and treatment strategies for OA. In particular, the roles of EVs as biomarkers in synovial fluid, chondrocytes, and plasma for the early detection of equine OA are discussed.

Osteoporosis is a widespread chronic condition. Although standard treatments are generally effective, they are frequently constrained by side effects and the risk of developing drug resistance. A promising area of research is the investigation of extracellular vesicles (EVs), including exosomes, microvesicles, and apoptotic bodies, which play a crucial role in bone metabolism. Exosomes, in particular, have shown significant potential in both the diagnosis and treatment of osteoporosis. EVs derived from osteoclasts, osteoblasts, mesenchymal stem cells, and other sources can influence bone metabolism, while exosomes from inflammatory and tumor cells may exacerbate bone loss, highlighting their dual role in osteoporosis pathology. This review offers a comprehensive overview of EV biogenesis, composition, and function in osteoporosis, focusing on their diagnostic and therapeutic potential. We examine the roles of various types of EVs and their cargo-proteins, RNAs, and lipids-in bone metabolism. Additionally, we explore the emerging applications of EVs as biomarkers and therapeutic agents, emphasizing the need for further research to address current challenges and enhance EV-based strategies for managing osteoporosis.

Background/Objectives: Osteoarthritis (OA) is the most common joint disease in the adult population. OA is the result of multiple mechanisms leading to inflammation and the degradation of the cartilage. A complex series of etiological actors have been identified so far, including extracellular vesicles (EVs). The EV content of the synovial fluid (SF) can release inflammatory mediators that enhance OA progression. An intra-articular viscosupplementation of high-MW hyaluronic acid (HyA) constitutes the first-line conservative treatment for OA. Although attractive for the potential pharmacological implications, the possibility that HyA may interact with EVs in the context of OA has not yet been specifically investigated; therefore, the present study aimed to fill this gap. Methods: We studied the effect of a HyA preparation (a blend of crosslinked and linear polymers, CLHyA) on the relevant inflammatory markers in chondrocytes (HC cells or primary chondrocytes isolated from patients with advanced OA) exposed to the EVs collected from IL-1β-stimulated THP-1 human monocytes (EVs+). Results: EVs+ caused specific inflammatory responses in chondrocytes that could be prevented by coincubation with CLHyA. This anti-inflammatory activity is likely dependent on the direct binding of CLHyA to CD44 receptors highly expressed in EVs+ and on the subsequent hindrance to EVs+ diffusion and docking to target cells. Conclusions: On the whole, the tight interactions identified herein between HMW HyA and EVs+ represent a novel, pharmacologically exploitable mechanism potentially relevant in the context of OA treatment.

Exosomes are nanosized, lipid membrane vesicles secreted by cells, facilitating intercellular communication by transferring cargo from parent to recipient cells. This capability enables biological crosstalk across multiple tissues and cells. Extensive research has been conducted on their role in the pathogenesis of degenerative musculoskeletal diseases such as osteoarthritis (OA), a chronic and painful joint disease that particularly affects cartilage. Currently, no effective treatment exists for OA. Given that exosomes naturally modulate synovial joint inflammation and facilitate cartilage matrix synthesis, they are promising candidates as next generation nanocarriers for OA therapy. Recent advancements have focused on engineering exosomes through endogenous and exogenous approaches to enhance their joint retention, cartilage and chondrocyte targeting properties, and therapeutic content enrichment, further increasing their potential for OA drug delivery. Notably, charge-reversed exosomes that utilize electrostatic binding interactions with cartilage anionic aggrecan glycosaminoglycans have demonstrated the ability to penetrate the full thickness of early-stage arthritic cartilage tissue following intra-articular administration, maximizing their therapeutic potential. These exosomes offer a non-viral, naturally derived, cell-free carrier for OA drug and gene delivery applications. Efforts to standardize exosome harvest, engineering, and property characterization methods, along with scaling up production, will facilitate more efficient and rapid clinical translation. This article reviews the current state-of-the-art, explores opportunities for exosomes as OA therapeutics, and identifies potential challenges in their clinical translation.

Patients with Rheumatoid arthritis (RA) have an elevated risk of lung cancer compared to the healthy population. However, there are few studies on the relationship between RA and lung adenocarcinoma (LUAD), especially the mechanisms at the genetic level. In this study, we investigated the link between RA and LUAD regarding Ferroptosis-Related Genes. The RNA-seq data of RA (GSE77298 and GSE 82107) and LUAD(GSE75037) in the Gene Expression Omnibus (GEO) database were obtained. 259 ferroptosis-related genes were obtained from the website ( http://www.zhounan.org/ferrdb/ ).The differential genes obtained from the RA and LUAD datasets were intersected with ferroptosis-related genes to obtain the ferroptosis-related differentially expressed genes (FRDEGs). Next, the mRNA-miRNA network was constructed, then Gene Set Enrichment Analysis (GSEA) for target genes were performed. The CIBERSORT algorithm was used to analyze the immune infiltration. Finally, the results were validated using external datasets (GSE89408 and GSE48780) and The Cancer Genome Atlas (TCGA) dataset. We obtained FRDEGs common to LUAD and RA: FANCD2, HELLS, RRM2, G6PD, VLDLR. These five genes play important roles in the progression of RA and LUAD. They also hold great diagnostic value for both diseases. Also, we found that LUAD and RA share common signaling pathways and similar immune mechanisms.

Rheumatoid arthritis (RA) is a systemic inflammatory disease whose precise pathogenesis remains mysterious. The involvement of epigenetic regulation in the pathogenesis of RA is one of the most anticipated findings, among which non-coding RNAs (ncRNAs) hold great application promise as diagnostic and therapeutic biomarkers for RA. Extracellular vesicles (EVs) are a heterogeneous group of nano-sized, membrane-enclosed vesicles that mediate intercellular communication and substance exchange, especially the transfer of ncRNAs from donor cells, thereby regulating the functional activities and biological processes of recipient cells. In light of the significant correlation between EVs, ncRNAs, and RA, we first documented expression levels of EVs and their-encapsulated ncRNAs in RA individuals, and methodically discussed their-implicated signaling pathways and phenotypic changes. The last but not least, we paied special attention to the therapeutic benefits of gene therapy reagents specifically imitating or silencing candidate ncRNAs with exosomes as carriers on RA animal models, and briefly highlighted their clinical application advantage and foreground. In conclusion, the present review may be conducive to a deeper comprehension of the diagnostic and therapeutic roles of EVs-enwrapped ncRNAs in RA, with special emphasis on exosomal ncRNAs, which may offer hints for the monitoring and treatment of RA.

Kashin-Beck disease (KBD) is an endemic, chronic degenerative joint disease in China. Exosomes miRNAs, as signaling molecules in intercellular communication, can transfer specific biological martials into target cell to regulate their function and might participate in the pathogenesis of KBD. We isolated serum and chondrocytes-derived exosomes, miRNA sequencing revealed exosomes miRNA profiles and differentially expressed miRNAs (DE-miRNAs) were identified. The target genes were predicted of known and novel DE-miRNAs with TargetScan 5.0 and miRanda 3.3a database. Single-cell RNA sequencing (scRNA-seq) was performed to identify chondrocyte clusters and their gene signatures in KBD. And we performed comparative analysis between the serum and chondrocytes-derived exosomes DE-miRNA target genes and differentially expressed genes of each cell clusters. A total of 20 DE-miRNAs were identified in serum-derived exosomes. In the miRNA expression of chondrocytes-derived exosomes, 53 DE-miRNAs were identified. 16,063 predicted targets were identified as the target genes in the serum-derived exosomes, 57,316 predicted targets were identified as the target genes in the chondrocytes-derived exosomes. Seven clusters were labeled by cell type according to the expression of previously described markers. Three hundred fifteen common genes were found among serum/chondrocytes-derived exosomes DE-miRNA target genes and DEGs identified by scRNA-seq analysis. We firstly integratly analyzed the serum and chondrocytes exosomes miRNA with single-cell RNA sequencing (scRNA-seq) data of KBD chondrocyte, the results showed that DE-miRNAs in exosomes might play a potential role in regulating genes expression in different KBD chondrocytes clusters by exosomes mediating cell-cell communications functions, which could improve the new diagnosis and treatment methods for KBD.

Arthritis, a diverse group of inflammatory joint disorders, poses great challenges in early diagnosis and targeted treatment. Timely intervention is imperative, yet conventional diagnostic methods are not able to detect subtle early symptoms. Hence, there is an urgent need for specific biomarkers that discriminate between different arthritis forms and for early diagnosis. The pursuit of such precise diagnostic tools has prompted a growing interest in extracellular vesicles (EVs). EVs, released by cells in a regulated fashion, are detectable in body fluids, including synovial fluid (SF), which fills the joint space. They provide insights into the intricate molecular landscapes of arthritis, and this has stimulated the search for minimally invasive EV-based diagnostics. As such, the analysis of EVs in SF has become a focus for identifying EV-based biomarkers for joint disease endotyping, prognosis, and progression. EVs are composed of a lipid bilayer and a wide variety of different cargo types, of which proteins and RNAs are widely investigated. In contrast, membrane lipids of EVs, especially the abundance, presence, or absence of specific lipids and their contribution to the biological activity of EVs, are largely overlooked in EV research. Furthermore, the identification of specific combinations of different EV components acting in concert in EVs can fuel the definition of composite biomarkers. We here provide a state-of-the-art overview of the knowledge on SF-derived EVs with emphasis on lipid analysis and we give an example of the added value of integrated proteomics and lipidomics analysis in the search for composite EV-associated biomarkers.

The objective of this study was to characterize extracellular vesicles (EVs) in plasma and synovial fluid obtained from horses with and without naturally occurring post-traumatic osteoarthritis (PTOA).

EVs were isolated from plasma and synovial fluid from horses with (n = 6) and without (n = 6) PTOA.

Plasma and synovial fluid EVs were characterized with respect to quantity, size, and surface markers. Small RNA sequencing was performed, and differentially expressed microRNAs (miRNAs) underwent bioinformatic analysis to identify putative targets and to explore potential associations with specific biological processes.

Plasma and synovial fluid samples from horses with PTOA had a significantly higher proportion of exosomes and a lower proportion of microvesicles compared to horses without PTOA. Small RNA sequencing revealed several differentially expressed miRNAs, including miR-144, miR-219-3p, and miR-199a-3l in plasma and miR-199a-3p, miR-214, and miR-9094 in synovial fluid EVs. Bioinformatics analysis of the differentially expressed miRNAs highlighted their potential role in fibrosis, differentiation of chondrocytes, apoptosis, and inflammation pathways in PTOA.

We have identified dynamic molecular changes in the small noncoding signatures of plasma and synovial fluid EVs in horses with naturally occurring PTOA. These findings could serve to identify promising biomarkers in the pathogenesis of PTOA, to facilitate the development of targeted therapies, and to aid in establishing appropriate translational models of PTOA.

Rheumatoid arthritis (RA) is a systemic autoimmune disease characterized by chronic inflammation of the joints and bone destruction. Because of systemic administration and poor targeting, traditional anti-rheumatic drugs have unsatisfactory treatment efficacy and strong side effects, including myelosuppression, liver or kidney function damage, and malignant tumors. Consequently, mesenchymal stem cells (MSCs)-involved therapy is proposed for RA therapy as a benefit of their immunosuppressive and tissue-repairing effects. This review summarizes the progress of MSCs-involved RA therapy through suppressing inflammation and promoting tissue regeneration and predicts their potential clinical application.

Inflammatory bone disease is a general term for a series of diseases caused by chronic inflammation, which leads to the destruction of bone homeostasis, that is, the osteolytic activity of osteoclasts increases, and the osteogenic activity of osteoblasts decreases, leading to osteolysis. Macrophages are innate immune cell with plasticity, and their polarization is related to inflammatory bone diseases. The dynamic balance of macrophages between the M1 phenotype and the M2 phenotype affects the occurrence and development of diseases. In recent years, an increasing number of studies have shown that extracellular vesicles existing in the extracellular environment can act on macrophages, affecting the progress of inflammatory diseases. This process is realized by influencing the physiological activity or functional activity of macrophages, inducing macrophages to secrete cytokines, and playing an anti-inflammatory or pro-inflammatory role. In addition, by modifying and editing extracellular vesicles, the potential of targeting macrophages can be used to provide new ideas for developing new drug carriers for inflammatory bone diseases.

Mechanical overloading can promote cartilage senescence and osteoarthritis (OA) development, but its impact on synovial macrophages and the interaction between macrophages and chondrocytes remain unknown. Here, we found that macrophages exhibited M1 polarization under mechanical overloading and secreted ectosomes that induced cartilage degradation and senescence. By performing miRNA sequencing on ectosomes, we identified highly expressed miR-350-3p as a key factor mediating the homeostatic imbalance of chondrocytes caused by M1-polarized macrophages, this result being confirmed by altering the miR-350-3p level in chondrocytes with mimics and inhibitor. In experimental OA mice, miR-350-3p was increased in synovium and cartilage, while intra-articular injection of antagomir-350-3p inhibited the increase of miR-350-3p and alleviated cartilage degeneration and senescence. Further studies showed that macrophage-derived ectosomal miR-350-3p promoted OA progression by inhibiting nuclear receptor binding SET domain protein 1(NSD1) in chondrocytes and regulating histone H3 lysine 36(H3K36) methylation. This study demonstrated that the targeting of macrophage-derived ectosomal miRNAs was a potential therapeutic method for mechanical overload-induced OA.

Spinal cord injury (SCI) is a devastating condition with a complex pathology that affects a significant portion of the population and causes long-term consequences. After primary injury, an inflammatory cascade of secondary injury occurs, followed by neuronal cell death and glial scar formation. Together with the limited regenerative capacity of the central nervous system, these are the main reasons for the poor prognosis after SCI. Despite recent advances, there is still no effective treatment. Promising therapeutic approaches include stem cells transplantation, which has demonstrated neuroprotective and immunomodulatory effects in SCI. This positive effect is thought to be mediated by small extracellular vesicles (sEVs); membrane-bound nanovesicles involved in intercellular communication through transport of functional proteins and RNA molecules. In this review, we summarize the current knowledge about sEVs and microRNA as their cargo as one of the most promising therapeutic approaches for the treatment of SCI. We provide a comprehensive overview of their role in SCI pathophysiology, neuroprotective potential and therapeutic effect.

Rheumatoid arthritis is a chronic systemic autoimmune disease that involves mainly synovitis and joint injury and is one of the main causes of disability. The pathogenesis of rheumatoid arthritis is complicated, and the treatment cycle is long. The traditional methods of inhibiting inflammation and immunosuppression are no longer sufficient for treatment of the disease, so there is an urgent need to seek new treatments. The exocrine microenvironment is a kind of microvesicle with a lipid bilayer membrane structure that can be secreted by most cells in the body. This structure contains cell-specific proteins, lipids and nucleic acids that can transmit this information from one cell to another. To achieve cell-to-cell communication. Exocrine microRNAs can be contained in exocrine cells and can be selectively transferred to target receptor cells via exocrine signaling, thus regulating the physiological function of target cells. This article focuses on the pathological changes that occur during the development of rheumatoid arthritis and the biological regulation of exocrine and exocrine microRNAs in rheumatoid joints. Research on the roles of exocrine and exocrine microRNAs in regulating the inflammatory response, cell proliferation/apoptosis, autophagy, effects on fibroblast-like synoviocytes and immune regulation in rheumatoid arthritis was reviewed. In addition, the challenges faced by this new treatment are discussed.

Rheumatoid arthritis (RA) is a prevalent inflammatory autoimmune disease characterised by persistent inflammation and joint damage with elevated levels of reactive oxygen species (ROS). Current treatment modalities for RA have significant limitations, including poor bioavailability, severe side effects, and inadequate targeting of inflamed joints. Herein, we synthesised cerium/manganese oxide nanoparticles (NPs) as efficient drug carriers with antioxidant and catalytic-like functions that can eliminate ROS to facilitate the polarization of macrophages phenotype from M1 to M2 and alleviate inflammation. Methotrexate (MTX), a first-line RA medication, was loaded into the NPs, which were further modified with bovine serum albumin (BSA) and integrated into dissolving hyaluronic acid-based microneedles (MNs) for transdermal delivery.

This innovative approach significantly enhanced drug delivery efficiency, reduced RA inflammation, and successfully modulated macrophage polarization toward an anti-inflammatory phenotype.

This research not only presents a promising drug delivery strategy for RA but also contributes broadly to the field of immune disease treatment by offering an advanced approach for macrophage phenotypic reprogramming.

During the 2023 Padua Days on Muscle and Mobility Medicine the 2024 meeting was scheduled from 28 February to 2 March 2024 (2024Pdm3). During autumn 2023 the program was expanded with Scientific Sessions which will take place over five days (in 2024 this includes February 29), starting from the afternoon of 27 February 2024 in the Conference Rooms of the Hotel Petrarca, Thermae of Euganean Hills (Padua), Italy. As per consolidated tradition, the second day will take place in Padua, for the occasion in the Sala San Luca of the Monastery of Santa Giustina in Prato della Valle, Padua, Italy. Confirming the attractiveness of the Padua Days on Muscle and Mobility Medicine, over 100 titles were accepted until 15 December 2023 (many more than expected), forcing the organization of parallel sessions on both 1 and 2 March 2024. The five days will include lectures and oral presentations of scientists and clinicians from Argentina, Austria, Belgium, Brazil, Bulgaria, Canada, Denmark, Egypt, France, Germany, Iceland, Ireland, Italy, Romania, Russia, Slovenia, Switzerland, UK and USA. Only Australia, China, India and Japan are missing from this edition. But we are confident that authors from those countries who publish articles in the PAGEpress: European Journal of Translational Myology (EJTM: 2022 ESCI Clarivate's Impact Factor: 2.2; SCOPUS Cite Score: 3.2) will decide to join us in the coming years. Together with the program established by 31 January 2024, the abstracts will circulate during the meeting only in the electronic version of the EJTM Issue 34 (1) 2024. See you soon in person at the Hotel Petrarca in Montegrotto Terme, Padua, for the inauguration scheduled the afternoon of 27 February 2024 or on-line for free via Zoom. Send us your email address if you are not traditional participants listed in Pdm3 and EJTM address books.

The pathogenesis of rheumatoid arthritis (RA) is heavily impacted by the inflammation and activation of fibroblast-like synoviocytes (FLS). The objective of this investigation is to clarify the involvement of exosomes derived from FLS stimulated by tumour necrosis factor α (TNF-α) in angiogenesis and the underlying mechanisms. FLS cells were obtained from synovial fluid of RA patients and exosomes were obtained from FLS cell supernatant with TNF-α stimulation by ultracentrifugation. Exosomes were subsequently analysed using transmission electron microscopy, nanoparticle tracking analysis, and western blotting. The functional effects of exosomes with TNF-α stimulation on human umbilical vein endothelial cells (HUVEC) migration, invasion, and angiogenesis was evaluated using wound scratch healing test, transwell invasion assay, and tube formation assay. DNA nanoball-seq (DNBSEQ) sequencing platform was utilised to analysis different expression miRNA from exosomes, miRNA and mRNA from HUVEC. The expression level of miR-200a-3p was determined through quantitative real-time polymerase chain reaction (qRT-PCR). The quantification of KLF6 and VEGFA expression levels were performed by qRT-PCR and western blot analysis. The validation of the association between miR-200a-3p and KLF6 was established through a fluorescence enzyme reporting assay. In comparison to exosome induced by PBS, exosome induced by TNF-α exhibited a substantial exacerbation of invasion, migration, and angiogenesis in HUVEC. 4 miRNAs in exosomes and HUVEC cells, namely miR-1246, miR-200a-3p, miR-30a-3p, and miR-99b-3p was obtained. MiR-200a-3p maintained high consistency with the sequencing results. We obtained 5 gene symbols, and KLF6 was chose for further investigation. The expression of miR-200a-3p in exosomes induced by TNF-α and in HUVEC treated with these exosomes demonstrated a significantly increase. Additionally, HUVEC cells displayed a notable decrease in KLF6 expression and a significant elevation in VEGFA expression. This was further confirmed by the fluorescence enzyme report assay, which provided evidence of the direct targeting of KLF6 by miR-200a-3p. Exosomes induced by TNF-α have the ability to enhance the migration, invasion, and angiogenesis of HUVEC cells via the miR-200a-3p/KLF6/VEGFA axis.

Recently, there is a growing interest in the research based on extracellular vesicles (EVs) which represent paracrine factors secreted by almost all cell types. Both, normal and pathological cells are able to release various types of EVs with different physiological properties, functions and compositions. EVs play an important role in intercellular communication, mechanism and tissue repair. Moreover, EVs could help not only in the treatment of diseases but also in their diagnostics. This work focused on the evaluation of the potential of EVs being used as biomarkers for the diagnosis of osteoarthritis (OA) based on a comparison of the composition of EVs separated from platelet-poor plasma (PPP) of healthy donors and OA patients at different stages of OA. OA is established as a complex syndrome with extensive impact on multiple tissues within the synovial joint. It is a chronic disease of musculoskeletal system that mainly affects the elderly. Depending on the use of the Kellgren-Lawrence classification system, there are four grades of OA which have a negative impact on patients' quality of life. It is very difficult to detect OA in its early stages, so it is necessary to find a new diagnostic method for its timely detection. PPP samples were prepared from whole blood. PPP-EVs were separated from 3 groups of donors-healthy control, early stage OA, end-stage OA, and their content was compared and correlated. EVs from PPP were separated by size exclusion chromatography and characterized in terms of their size, yield and purity by NTA, western blotting, ELISA and flow cytometry. Detection of surface markers expression in EVs was performed using MACSPlex approach. Inflammatory and growth factors in EVs were analysed using MAGPix technology. Our study confirmed significant differences between EVs surface markers of patients and healthy controls correlating with the age of donor (CD63, CD31 and ROR1) and stage of OA (CD45, CD326 and CD56), respectively. Circulating EVs have been under extensive investigation for their capability to predict OA pathology diagnosis as potential targets for biomarker discovery. Taken together, obtained results indicated that PPP-EVs surface markers could be used as potential biomarkers in the early diagnosis of OA.

With the increasingly prominent problem of population aging, osteoarthritis (OA), which is closely related to aging, has become a serious illness affecting the lives and health of elderly individuals. However, effective treatments are still lacking. OA is typically considered a low-grade inflammatory state. The inflammatory infiltration of macrophages, neutrophils, T cells, and other cells is common in diseased joints. These cells create the inflammatory environment of OA and are involved in the onset and progression of the disease. Exosomes, a type of complex vesicle containing abundant RNA molecules and proteins, play a crucial role in the physiological and pathological processes of an organism. In comparison to other therapeutic methods such as stem cells, exosomes have distinct advantages of precise targeting and low immunogenicity. Moreover, research and techniques related to exosomes are more mature, indicating a promising future in disease treatment. Many studies have shown that the impact of exosomes on the inflammatory microenvironment directly or indirectly leads to the occurrence of various diseases. Furthermore, exosomes can be helpful in the management of illnesses. This article provides a comprehensive review and update on the research of exosomes, a type of extracellular vesicle, in the treatment of OA by modulating the inflammatory microenvironment. It also combines innovative studies on the modification of exosomes. In general, the application of exosomes in the treatment of OA has been validated, and the introduction of modified exosome technology holds potential for enhancing its therapeutic efficacy.

Synovial fibroblasts are critical for maintaining homeostasis in major autoimmune diseases involving joint inflammation, including osteoarthritis and rheumatoid arthritis. However, little is known about the interactions among different cell subtypes and the specific sets of signaling pathways and activities that they trigger.

Using social network analysis, pattern recognition, and manifold learning approaches, we identified patterns of single-cell communication in OA (osteoarthritis) and RA (rheumatoid arthritis).

Our results suggest that OA and RA have distinct cellular communication patterns and signaling pathways. The LAMININ (Laminin) and COLLAGEN (Collagen) pathways predominate in osteoarthritis, while the EGF (Epidermal growth factor), NT (Neurotrophin) and CDH5 (Cadherin 5) pathways predominate in rheumatoid arthritis, with a central role for THY1 (Thy-1 cell surface antigen) +CDH11 (Cadherin 11) + cells. The OA opens the PDGF (Platelet-derived growth factors) pathway (driver of bone angiogenesis), the RA opens the EGF pathway (bone formation) and the SEMA3 (Semaphorin 3A) pathway (involved in immune regulation). Interestingly, we found that OA no longer has cell types involved in the MHC complex (Major histocompatibility complex) and their activity, whereas the MHC complex functions primarily in RA in the presentation of inflammatory antigens, and that the complement system in OA has the potential to displace the function of the MHC complex. The specific signaling patterns of THY1+CDH11+ cells and their secreted ligand receptors are more conducive to cell migration and lay the foundation for promoting osteoclastogenesis. This subpopulation may also be involved in the accumulation of lymphocytes, affecting the recruitment of immune cells. Members of the collagen family (COL1A1 (Collagen Type I Alpha 1 Chain), COL6A2 (Collagen Type VI Alpha 2 Chain) and COL6A1 (Collagen Type VI Alpha 1 Chain)) and transforming growth factor (TGFB3) maintain the extracellular matrix in osteoarthritis and mediate cell migration and adhesion in rheumatoid arthritis, including the PTN (Pleiotrophin) / THBS1 (Thrombospondin 1) interaction.

Increased understanding of the interaction networks between synovial fibroblast subtypes, particularly the shared and unique cellular communication features between osteoarthritis and rheumatoid arthritis and their hub cells, should help inform the design of therapeutic agents for inflammatory joint disease.

Introduction: Renal osteodystrophy (ROD) is a type of bone metabolic disorder in patients with chronic kidney disease (CKD). Inflammation is associated with bone loss in ROD. However, its precise mechanism has not yet been elucidated. The present study was conducted to investigate whether exosomes (Exos) in bone marrow (BM) are involved in the pathogenesis of high-turnover ROD.Methods: Bone mass, osteoclast number, and pro-inflammatory cytokines levels of BM supernatant were detected in adenine-induced ROD rats. The effect of Exos derived from BM (BM-Exos) of ROD (ROD-Exos) on inflammatory genes and osteoclast differentiation of BM-derived macrophages (BMMs) were further examined. Then, exosomal miRNA sequencing was performed and an miRNA-mRNA-pathway network was constructed.Results: we found increased osteoclasts and decreased bone mass in ROD rats, as well as inflammatory activation in the BM niche. Furthermore, BMMs from ROD rats displayed overproduction of proinflammatory cytokines and increased osteoclast differentiation, accompanied by nuclear factor κB (NF-κB) signaling activation. Mechanistically, we found that ROD-Exos activates NF-κB signaling to promote the release of proinflammatory cytokines and increase osteoclast differentiation of BMMs. Meanwhile, a total of 24 differentially expressed miRNAs were identified between BM-Exos from ROD and normal control (NC). The miRNA-mRNA-pathway network suggests that rno-miR-9a-5p, rno-miR-133a-3p, rno-miR-30c-5p, rno-miR-206-3p, and rno-miR-17-5p might play pivotal roles in inflammation and osteoclast differentiation. Additionally, we validated that the expression of miR-9a-5p is upregulated in ROD-Exos.Conclusion: The BM niche of ROD alters the miRNA cargo of BM-Exos to promote inflammation and osteoclast differentiation of BMMs, at least partially contributing to the pathogenesis of high-turnover ROD.

Extracellular vesicular miRNAs (EV-miRNAs) play essential roles as intercellular communication molecules in knee Osteoarthritis (OA). We isolated cartilage-derived extracellular vesicles (EVs), to perform miRNA sequencing, which revealed EV-miRNA profiles and identified differentially expressed miRNAs (DE-miRNAs) between cartilage injury and cartilage non-injury groups. The target genes of known and novel DE-miRNAs were predicted with multiMiR package in 14 miRNA-target interaction databases. Meanwhile, single-cell RNA sequencing (scRNA-seq) was performed to identify chondrocyte clusters and their gene signatures in knee OA. Then we performed comparative analysis between target genes of the cartilage-derived EV-DE-miRNAs target genes and cluster-specific maker genes of characteristic chondrocyte clusters. Finally, the functional analysis of the cartilage-derived EVs DE-miRNA target genes and cluster-specific marker genes of each cell population were performed. The EV-miRNA profile analysis identified 13 DE-miRNAs and 7638 target genes. ScRNA-seq labelled seven clusters by cell type according to the expression of multiple characteristic markers. The results identified 735, 184, 303 and 879 common genes between EV-DE-miRNA target genes and cluster-specific marker genes in regulatory chondrocytes (RegCs), fibrocartilage chondrocytes (FC), prehypertrophic chondrocytes (PreHTCs) and mitochondrial chondrocytes (MTC), respectively. We firstly integrated the association between the cartilage-derived EV-DE-miRNA target genes and distinguished cluster-specific marker genes of each chondrocyte clusters. KEGG pathway analysis further identified that the DE-miRNAs target genes were significantly enriched in MAPK signaling pathway, Focal adhesion and FoxO signaling pathway. Our results provided some new insights into cartilage injury and knee OA pathogenesis which could improve the new diagnosis and treatment methods for OA.

Extracellular vesicles (EVs) contribute to osteoarthritis pathogenesis through their release into joint tissues and synovial fluid. Synovial fluid-derived EVs have the potential to be direct biomarkers in the causal pathway of disease but also enable understanding of their role in disease progression. Utilizing a temporal model of osteoarthritis, we defined the changes in matched synovial fluid and plasma-derived EV small non-coding RNA and protein cargo using sequencing and mass spectrometry. Data exploration included time series clustering, factor analysis and gene enrichment interrogation. Chondrocyte signalling was analysed using luciferase-based transcription factor activity assays. EV protein cargo appears to be more important during osteoarthritis progression than small non-coding RNAs. Cluster analysis revealed plasma-EVs represented a time-dependent response to osteoarthritis induction associated with supramolecular complexes. Clusters for synovial fluid-derived EVs were associated with initial osteoarthritis response and represented immune/inflammatory pathways. Factor analysis for plasma-derived EVs correlated with day post-induction and were primarily composed of proteins modulating lipid metabolism. Synovial fluid-derived EVs factors represented intermediate filament and supramolecular complexes reflecting tissue repair. There was a significant interaction between time and osteoarthritis for CRE, NFkB, SRE, SRF with a trend for osteoarthritis synovial fluid-derived EVs at later time points to have a more pronounced effect.

Extracellular vesicles (EVs) are implicated in the pathogenesis of rheumatoid arthritis (RA) but little is known about the composition of specific small EV (sEV) subpopulations. This study aimed to characterize the CD63, CD81 and CD9 tetraspanin profile in the membrane of single EVs in plasma from treatment naïve RA patients and assess potential discrepancies between methotrexate (MTX) responder groups. EVs isolated from plasma were characterized using transmission electron microscopy, and detection of surface markers (CD63, CD81 and CD9) on single EVs was performed on the ExoView platform. All RA patients (N = 8) were newly diagnosed, treatment naïve, females, ACPA positive and former smokers. The controls (N = 5) were matched for age and gender. After three months of MTX treatment, responders (N = 4) were defined as those with ΔDAS28 > 1.2 and DAS28 ≤ 3.2 post-treatment. The isolated EVs were 50-200 nm in size. The RA patients had a higher proportion of both CD9 and CD81 single positive sEVs compared to healthy controls, while there was a decrease in CD81/CD9 double positive sEVs in patients. Stratification of RA patients into MTX responders and non-responders revealed a distinctly higher proportion of CD81 single positive sEVs in the responder group. The proportion of CD81/CD9 double positive sEVs (anti-CD9 captured) was lower in the non-responders, but increased upon 3 months of MTX treatment. Our exploratory study revealed distinct tetraspanin profiles in RA patients suggesting their implication in RA pathophysiology and MTX treatment response.