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Arthritis Research & Therapy Jun 2024Kinases are intracellular signalling mediators and key to sustaining the inflammatory process in rheumatoid arthritis (RA). Oral inhibitors of Janus Kinase family (JAKs)...
BACKGROUND
Kinases are intracellular signalling mediators and key to sustaining the inflammatory process in rheumatoid arthritis (RA). Oral inhibitors of Janus Kinase family (JAKs) are widely used in RA, while inhibitors of other kinase families e.g. phosphoinositide 3-kinase (PI3K) are under development. Most current biomarker platforms quantify mRNA/protein levels, but give no direct information on whether proteins are active/inactive. Phosphoproteome analysis has the potential to measure specific enzyme activation status at tissue level.
METHODS
We validated the feasibility of phosphoproteome and total proteome analysis on 8 pre-treatment synovial biopsies from treatment-naive RA patients using label-free mass spectrometry, to identify active cell signalling pathways in synovial tissue which might explain failure to respond to RA therapeutics.
RESULTS
Differential expression analysis and functional enrichment revealed clear separation of phosphoproteome and proteome profiles between lymphoid and myeloid RA pathotypes. Abundance of specific phosphosites was associated with the degree of inflammatory state. The lymphoid pathotype was enriched with lymphoproliferative signalling phosphosites, including Mammalian Target Of Rapamycin (MTOR) signalling, whereas the myeloid pathotype was associated with Mitogen-Activated Protein Kinase (MAPK) and CDK mediated signalling. This analysis also highlighted novel kinases not previously linked to RA, such as Protein Kinase, DNA-Activated, Catalytic Subunit (PRKDC) in the myeloid pathotype. Several phosphosites correlated with clinical features, such as Disease-Activity-Score (DAS)-28, suggesting that phosphosite analysis has potential for identifying novel biomarkers at tissue-level of disease severity and prognosis.
CONCLUSIONS
Specific phosphoproteome/proteome signatures delineate RA pathotypes and may have clinical utility for stratifying patients for personalised medicine in RA.
Topics: Humans; Arthritis, Rheumatoid; Synovial Membrane; Signal Transduction; Proteomics; Female; Phosphoproteins; Middle Aged; Male; Adult; Aged; Proteome
PubMed: 38867295
DOI: 10.1186/s13075-024-03351-4 -
Nature Communications Jun 2024Rheumatoid arthritis (RA) is an autoimmune disease involving antigen-specific T and B cells. Here, we perform single-cell RNA and repertoire sequencing on paired...
Rheumatoid arthritis (RA) is an autoimmune disease involving antigen-specific T and B cells. Here, we perform single-cell RNA and repertoire sequencing on paired synovial tissue and blood samples from 12 seropositive RA patients. We identify clonally expanded CD4 + T cells, including CCL5+ cells and T peripheral helper (Tph) cells, which show a prominent transcriptomic signature of recent activation and effector function. CD8 + T cells show higher oligoclonality than CD4 + T cells, with the largest synovial clones enriched in GZMK+ cells. CD8 + T cells with possibly virus-reactive TCRs are distributed across transcriptomic clusters. In the B cell compartment, NR4A1+ activated B cells, and plasma cells are enriched in the synovium and demonstrate substantial clonal expansion. We identify synovial plasma cells that share BCRs with synovial ABC, memory, and activated B cells. Receptor-ligand analysis predicted IFNG and TNFRSF members as mediators of synovial Tph-B cell interactions. Together, these results reveal clonal relationships between functionally distinct lymphocyte populations that infiltrate the synovium of patients with RA.
Topics: Humans; Arthritis, Rheumatoid; Synovial Membrane; B-Lymphocytes; Female; Male; Middle Aged; CD8-Positive T-Lymphocytes; Lymphocyte Subsets; CD4-Positive T-Lymphocytes; Single-Cell Analysis; Transcriptome; Plasma Cells; Aged; Lymphocyte Activation; Adult
PubMed: 38862501
DOI: 10.1038/s41467-024-49186-0 -
RMD Open Jun 2024To assess the presence and anatomical distribution of activated fibroblasts in the joints and entheses of patients with psoriasis with arthralgia and to test how...
OBJECTIVES
To assess the presence and anatomical distribution of activated fibroblasts in the joints and entheses of patients with psoriasis with arthralgia and to test how fibroblast activation visualised by gallium-labelled fibroblast activation protein inhibitor-04 (Ga-FAPI-04)-positron emission tomography (PET)/CT correlates with clinical tenderness, musculoskeletal ultrasound findings and progression to psoriatic arthritis (PsA).
METHODS
We conducted a prospective cohort study in patients with psoriasis and arthralgia who underwent clinical and ultrasound evaluation and whole-body PET/CT imaging with Ga-FAPI-04. Ga-FAPI-04 uptake at synovial and entheseal sites was assessed by maximal standardised uptake values (SUVmax) and PET/CT Joint Index (JI); logistic regression models were used to investigate its correlation with clinical and ultrasound findings. Survival analyses were performed on patients with at least 6 months of follow-up.
RESULTS
36 patients with psoriasis were enrolled. Ga-FAPI-04 uptake was found in 318 (7.9%) joints and 369 (7.3%) entheses in 29 (80.6%) participants, with a mean SUVmax (SD) of 3.2 (1.8) for joints and 2.9 (1.6) for entheses. Large joints and the lower limbs were predominantly affected. A significant positive relationship was found between Ga-FAPI-04-PET/CT signal intensity and the 68 tender joint count (SUVmax: p<0.001; PET/CT-JI: p<0.001) and tender entheses count (SUVmax: p<0.001; PET/CT-JI: p=0.002). No correlations were found with ultrasound findings (SUVmax: p=0.969; PET/CT-JI: p=0.720). Patients with relevant synovio-entheseal Ga-FAPI-04 uptake showed a statistically significant higher risk of developing PsA (p=0.02), independent of ultrasound findings.
CONCLUSIONS
Patients with psoriasis presenting with arthralgia show localised signs of resident tissue activation in joints and entheses, which are associated with higher risk of developing PsA.
Topics: Humans; Arthritis, Psoriatic; Male; Female; Middle Aged; Positron Emission Tomography Computed Tomography; Psoriasis; Adult; Prospective Studies; Fibroblasts; Synovial Membrane; Aged; Ultrasonography; Disease Progression
PubMed: 38862244
DOI: 10.1136/rmdopen-2024-004294 -
Frontiers in Immunology 2024Pathological changes in the articular cartilage (AC) and synovium are major manifestations of osteoarthritis (OA) and are strongly associated with pain and functional...
Exosomes derived from miR-146a-overexpressing fibroblast-like synoviocytes in cartilage degradation and macrophage M1 polarization: a novel protective agent for osteoarthritis?
INTRODUCTION
Pathological changes in the articular cartilage (AC) and synovium are major manifestations of osteoarthritis (OA) and are strongly associated with pain and functional limitations. Exosome-derived microRNAs (miRNAs) are crucial regulatory factors in intercellular communication and can influence the progression of OA by participating in the degradation of chondrocytes and the phenotypic transformation in the polarization of synovial macrophages. However, the specific relationships and pathways of action of exosomal miRNAs in the pathological progression of OA in both cartilage and synovium remain unclear.
METHODS
This study evaluates the effects of fibroblast-like synoviocyte (FLS)-derived exosomes (FLS-Exos), influenced by miR-146a, on AC degradation and synovial macrophage polarization. We investigated the targeted relationship between miR-146a and TRAF6, both and , along with the involvement of the NF-κB signaling pathway.
RESULTS
The expression of miR-146a in the synovial exosomes of OA rats was significantly higher than in healthy rats. , the upregulation of miR-146a reduced chondrocyte apoptosis, whereas its downregulation had the opposite effect. , exosomes derived from miR-146a-overexpressing FLSs (miR-146a-FLS-Exos) reduced AC injury and chondrocyte apoptosis in OA. Furthermore, synovial proliferation was reduced, and the polarization of synovial macrophages shifted from M1 to M2. Mechanistically, the expression of TRAF6 was inhibited by targeting miR-146a, thereby modulating the Toll-like receptor 4/TRAF6/NF-κB pathway in the innate immune response.
DISCUSSION
These findings suggest that miR-146a, mediated through FLS-Exos, may alleviate OA progression by modulating cartilage degradation and macrophage polarization, implicating the NF-κB pathway in the innate immune response. These insights highlight the therapeutic potential of miR-146a as a protective agent in OA, underscoring the importance of exosomal miRNAs in the pathogenesis and potential treatment of the disease.
Topics: MicroRNAs; Animals; Exosomes; Osteoarthritis; Rats; Macrophages; Synoviocytes; Male; TNF Receptor-Associated Factor 6; Cartilage, Articular; Chondrocytes; NF-kappa B; Signal Transduction; Rats, Sprague-Dawley; Fibroblasts; Synovial Membrane; Cells, Cultured; Apoptosis; Toll-Like Receptor 4; Macrophage Activation
PubMed: 38846937
DOI: 10.3389/fimmu.2024.1361606 -
Cureus May 2024Synovial chondromatosis is a rare condition characterized by benign metaplasia of the synovial membrane, leading to nodular growths within the joint space. We present...
Synovial chondromatosis is a rare condition characterized by benign metaplasia of the synovial membrane, leading to nodular growths within the joint space. We present the case of a 58-year-old woman with persistent pain and stiffness in her right elbow, ultimately diagnosed with synovial chondromatosis. Examination revealed joint effusion, tenderness, and restricted range of motion, with palpable loose bodies and ulnar nerve symptomatology. X-ray confirmed the diagnosis. Open synovectomy was performed, with meticulous attention to ulnar nerve protection and decompression. Postoperative care included analgesics, anti-inflammatories, and physiotherapy. Synovial chondromatosis of the elbow requires prompt diagnosis and surgical intervention to alleviate symptoms and prevent complications. Prognosis is favorable with complete removal of the affected tissue. Ulnar nerve palsy should be carefully addressed during surgical management.
PubMed: 38846186
DOI: 10.7759/cureus.59807 -
Journal of Advanced Research Jun 2024Osteoarthritis (OA) is the most common arthritis that is characterized by the progressive synovial inflammation and loss of articular cartilage. Although GYY4137 is a...
INTRODUCTION
Osteoarthritis (OA) is the most common arthritis that is characterized by the progressive synovial inflammation and loss of articular cartilage. Although GYY4137 is a novel and slow-releasing hydrogen sulfide (HS) donor with potent anti-inflammatory properties that may modulate the progression of OA, its underlying mechanism remains unclear.
OBJECTIVES
In this study, we validated the protective role of GYY4137 against OA pathological courses and elucidated its underlying regulatory mechanisms.
METHODS
Cell transfection, immunofluorescence staining, EdU assay, transmission electron microscopy, mitochondrial membrane potential measurement, electrophoretic mobility shift assay, sulfhydration assay, qPCR and western blot assays were performed in the primary mouse chondrocytes or the mouse macrophage cell line raw 264.7 for in vitro study. DMM-induced OA mice model and Macrophage-specific p65 knockout (p65 LysM-CreER) mice on the C57BL/6 background were used for in vivo study.
RESULTS
We found that GYY4137 can alleviate OA progress by suppressing synovium pyroptosis in vivo. Moreover, our in vitro data revealed that GYY4137 attenuates inflammation-induced NLRP3 and caspase-1 activation and results in a decrease of IL-1β production in macrophages. Mechanistically, GYY4137 increased persulfidation of NF-kB p65 in response to inflammatory stimuli that results in a decrease of cellular reactive oxygen species (ROS) accumulation and ameliorates mitochondrial dysfunctions. Using site-directed mutagenesis, we showed that HS persulfidates cysteine38 in p65 protein and hampers p65 transcriptional activity, and p65 mutant impaired macrophage responses to GYY4137.
CONCLUSION
These findings suggest a mechanism by which GYY4137 through redox modification of p65 participates in inhibiting NLRP3 activation by OA to regulate inflammatory responses. Thus, we propose that GYY4137 represents a promising novel therapeutic strategy for the treatment of OA.
PubMed: 38844123
DOI: 10.1016/j.jare.2024.05.033 -
Frontiers in Veterinary Science 2024Here, we describe a case of a 5-year-old show-jumping stallion presented with severe lameness, swelling, and pain on palpation of the left metacarpophalangeal joint...
Case report: Equine metacarpophalangeal joint partial and full thickness defects treated with allogenic equine synovial membrane mesenchymal stem/stromal cell combined with umbilical cord mesenchymal stem/stromal cell conditioned medium.
Here, we describe a case of a 5-year-old show-jumping stallion presented with severe lameness, swelling, and pain on palpation of the left metacarpophalangeal joint (MCj). Diagnostic imaging revealed full and partial-thickness articular defects over the lateral condyle of the third metacarpus (MC3) and the dorsolateral aspect of the first phalanx (P1). After the lesion's arthroscopic curettage, the patient was subjected to an innovative regenerative treatment consisting of two intra-articular injections of equine synovial membrane mesenchymal stem/stromal cells (eSM-MSCs) combined with umbilical cord mesenchymal stem/stromal cells conditioned medium (UC-MSC CM), 15 days apart. A 12-week rehabilitation program was accomplished, and lameness, pain, and joint effusion were remarkably reduced; however, magnetic resonance imaging (MRI) and computed tomography (CT) scan presented incomplete healing of the MC3's lesion, prompting a second round of treatment. Subsequently, the horse achieved clinical soundness and returned to a higher level of athletic performance, and imaging exams revealed the absence of lesions at P1, fulfillment of the osteochondral lesion, and cartilage-like tissue formation at MC3's lesion site. The positive outcomes suggest the effectiveness of this combination for treating full and partial cartilage defects in horses. Multipotent mesenchymal stem/stromal cells (MSCs) and their bioactive factors compose a novel therapeutic approach for tissue regeneration and organ function restoration with anti-inflammatory and pro-regenerative impact through paracrine mechanisms.
PubMed: 38840629
DOI: 10.3389/fvets.2024.1403174 -
Veterinaria Italiana Dec 2023Cells obtained from chicken embryos are often preferred for in vitro studies. These cells, which easily adapt to rapid and continuous growth in the appropriate cell...
Cells obtained from chicken embryos are often preferred for in vitro studies. These cells, which easily adapt to rapid and continuous growth in the appropriate cell culture environment, are thought to be one of the effective methods in the investigation of leg diseases that are frequently observed in poultry. Leg diseases, especially affecting the joints in chickens, cause locomotor problems and adversely affect animal welfare. In addition, they cause significant health problems and increase mortality. It is known that synovial fibroblasts play an important role in joint diseases. In this study, chicken embryonic synovial fibroblasts were isolated from tissue explants taken from the tibio-metatarsal joint region of brown layer chicken embryos. Characterization of cells was evaluated by immunocytochemistry and hemacolor staining. chicken embryonic synovial fibroblasts showed a strong positive reaction to the vimentin antibody. As a result of hemacolor staining, it was noted that the cell morphology was spindle-shaped. The absence of macrophages in chicken embryonic synovial fibroblast culture was confirmed by the carbon powder uptake. In this present study, we aim to present a useful cell culture protocol such as primary culture, passage, and characterization suitable for chicken embryonic synovial fibroblast to be used in the new scientific research.
Topics: Animals; Fibroblasts; Chick Embryo; Synovial Membrane; Cell Culture Techniques; Chickens; Cells, Cultured
PubMed: 38828859
DOI: 10.12834/VetIt.3030.21925.2 -
Osteoarthritis and Cartilage Open Sep 2024Total hip arthroplasty is a successful procedure for treating advanced osteoarthritis (OA). Metal bearing surfaces remain one of the most widely implanted prosthesis,...
Cobalt ions induce a cellular senescence secretory phenotype in human synovial fibroblast-like cells that may be an early event in the development of adverse local tissue reactions to hip implants.
OBJECTIVES
Total hip arthroplasty is a successful procedure for treating advanced osteoarthritis (OA). Metal bearing surfaces remain one of the most widely implanted prosthesis, however approximately 10% of patients develop adverse local tissue reactions (ALTRs), namely lymphocytic predominant soft tissue reaction with or without necrosis and osteolysis resulting in high revision rates. The mechanism(s) for these reactions remains unclear although T lymphocyte mediated type IV hypersensitivity to cobalt (Co) and chromium (Cr) ions have been described. The purpose of this study was to determine the prolonged effects of Co and Cr metal ions on synovial fibroblasts to better understand the impact of the synovial membrane in the development of ALTRs.
METHODS
Human synovial fibroblast-like cells were isolated from donors undergoing arthroplasty. DNA content and Alamar blue assay were used to determine cellular viability against exposure to Co and Cr. A beta-galactosidase assay was used to determine the development of cellular senescence. Western blotting and RT-qPCR were employed to determine changes in senescent associated secretory factors, signaling and anti-oxidant enzyme expression. A fluorescent assay was used to measure accumulation of hydrogen peroxide.
RESULTS
We demonstrate that prolonged cobalt exposure results in a downregulation of the enzyme catalase resulting in cytosolic accumulation of hydrogen peroxide, decreased Akt activity and cellular senescence. Senescent fibroblasts demonstrated upregulation of proinflammatory cytokines IL-1β and TNFα in addition to the neurotrophic factor NGF.
CONCLUSION
Our results provide evidence that metal ions induce a senescent associated secretory phenotype in synovial fibroblasts that could contribute to the development of adverse local tissue reactions.
PubMed: 38828014
DOI: 10.1016/j.ocarto.2024.100490 -
Nature Communications May 2024Synovial tissue inflammation is a hallmark of rheumatoid arthritis (RA). Recent work has identified prominent pathogenic cell states in inflamed RA synovial tissue, such...
Synovial tissue inflammation is a hallmark of rheumatoid arthritis (RA). Recent work has identified prominent pathogenic cell states in inflamed RA synovial tissue, such as T peripheral helper cells; however, the epigenetic regulation of these states has yet to be defined. Here, we examine genome-wide open chromatin at single-cell resolution in 30 synovial tissue samples, including 12 samples with transcriptional data in multimodal experiments. We identify 24 chromatin classes and predict their associated transcription factors, including a CD8 + GZMK+ class associated with EOMES and a lining fibroblast class associated with AP-1. By integrating with an RA tissue transcriptional atlas, we propose that these chromatin classes represent 'superstates' corresponding to multiple transcriptional cell states. Finally, we demonstrate the utility of this RA tissue chromatin atlas through the associations between disease phenotypes and chromatin class abundance, as well as the nomination of classes mediating the effects of putatively causal RA genetic variants.
Topics: Arthritis, Rheumatoid; Humans; Chromatin; Synovial Membrane; T-Box Domain Proteins; Epigenesis, Genetic; Single-Cell Analysis; Transcription Factors; Fibroblasts; Transcription Factor AP-1; Transcription, Genetic; CD8-Positive T-Lymphocytes
PubMed: 38821936
DOI: 10.1038/s41467-024-48620-7