Synovial Fluid Of Patients Research Articles

Correlation of native knee joint septic arthritis and bacteremia: A retrospective study

Objectives: Septic arthritis is an orthopedic emergency, delayed treatment results in rapid, and irreversible joint destruction with significant morbidity and mortality. The correlation between the infectious organism(s) isolated from blood and synovial fluid remains unclear. Native joint septic arthritis (NJSA) is often secondary to bacteremia and other contiguous sources of infection. This investigation examined the correlation in infectious organisms between blood and synovial fluid in patients with NJSA and concurrent bacteremia. Methods: A retrospective chart review was performed on 712 NJSA cases at an urban, level-one trauma center. Patients diagnosed with septic knee arthritis through synovial fluid analysis with white blood cell count >50,000 and/or positive culture from the knee joint on aspiration were included in the study. The organism identified on blood culture in the bacteremic patient was then correlated to the organism found on synovial fluid analysis. Results: We identified 104 patients at our institution with septic knees; 48 with bacteremia and 56 that did not have bacteremia. Of these patients with bacteremia, 34 patients (70.8%) had the same organism isolated in the knee and the blood. This correlation was statistically significant, with P = 0.003. Furthermore, patients with bacteremia underwent a mean of 1.85 ± 0.76 debridement procedures versus 1.21 ± 0.80 procedures in those without (P = 0.001). Conclusion: This study shows a correlation between the organism isolated from the knee and the blood in patients with NJSA in conjunction with bacteremia as well as a correlation between the bacteremic patient and the number of surgical interventions required per patient.

Targeting IL-6 Signaling Pathways for Musculoskeletal Disorders Treatment: Risks and Benefits

Pro-inflammatory mediators like IL-6 effectively mediated the majority of musculoskeletal disorders such rheumatoid arthritis (RA), osteoarthritis (OA), and tendinitis. Increased levels of IL-6 are found in the serum or synovial fluid of patients with these disorders, and these levels are correlated with the incidence and severity of the disease. IL-6 is crucial for the development of cartilage pathology, for example, by inducing a variety of pathways that are involved in the induction and spread of inflammation. The expression of anti-catabolic factors is similarly increased by IL-6, indicating a protective function. The differential impacts of IL-6 classic and trans-signaling may be the reason for this dual role of IL-6, which has so far remained poorly understood. In this article, the experimental and clinical data on the function of inhibiting IL-6 signaling in the development and progression of pathologies of the synovium, cartilage, and bones were thoroughly reviewed. By evaluating the IL-6 targeting approaches that are currently being considered in research and clinical practice, it may provide a glimpse into the future of these illnesses' treatment.

Impaired Efferocytosis by Synovial Macrophages in Patients With Knee Osteoarthritis.

Osteoarthritis (OA) exposes all joint tissues to physiologic stresses, increasing the need to clear apoptotic cells from tissues, including the synovium. We undertook this study to assess the burden of apoptotic cells in synovial tissue in patients with late-stage knee OA and to investigate whether OA impairs the macrophage-mediated clearance of apoptotic cells via efferocytosis. Synovial tissue was collected from individuals with healthy knees and patients with late-stage knee OA during arthroplasty. Synovial apoptotic cell burden was assessed by immunofluorescence for cleaved caspase 3. Efferocytosis of apoptotic Jurkat cells by CD14+ synovial tissue macrophages and peripheral blood-derived macrophages was quantified using immunofluorescence microscopy. Effects of OA on macrophage-mediated efferocytosis were modeled by stimulating blood-derived macrophages with synovial fluid collected from individuals with healthy knees and patients with early- or late-stage knee OA. Patients with late-stage knee OA had more apoptotic synovial cells compared to healthy individuals. There was a marked reduction in the fraction of synovial tissue macrophages engaging in efferocytosis and the quantity of material efferocytosed by individual macrophages in OA patients. Blood-derived macrophages exposed to synovial fluid from patients with knee OA recapitulated the defective efferocytosis, with the greatest effect from patients with early-stage knee OA and higher disease activity (pain and inflammation). Apoptotic cells accumulate in the synovium of patients with late-stage knee OA. Our results suggest that OA impairs critical homeostatic functions of synovial macrophages, leading to accumulation of apoptotic cells.

Dithiotreitol pre-treatment of synovial fluid samples improves microbiological counts in peri-prosthetic joint infection

PurposeSynovial fluid cultures of periprosthetic joint infections (PJI) may be limited by bacteria living in the fluids as biofilm-aggregates. The antibiofilm pre-treatment of synovial fluids with dithiotreitol (DTT) could improve bacterial counts and microbiological early stage diagnosis in patients with suspected PJI.MethodsSynovial fluids collected from 57 subjects, affected by painful total hip or knee replacement, were divided into two aliquots, one pre-treated with DTT and one with normal saline. All samples were plated for microbial counts. Sensitivity of cultural examination and bacterial counts of pre-treated and control samples were then calculated and statistically compared.ResultsDithiothreitol pre-treatment led to a higher number of positive samples, compared to controls (27 vs 19), leading to a statistically significant increase in the sensitivity of the microbiological count examination from 54.3 to 77.1% and in colony-forming units count from 1884 ± 2.129 CFU/mL with saline pre-treatment to 20.442 ± 19.270 with DTT pre-treatment (P = 0.02).ConclusionsTo our knowledge, this is the first report showing the ability of a chemical antibiofilm pre-treatment to increase the sensitivity of microbiological examination in the synovial fluid of patients with peri-prosthetic joint infection. If confirmed by larger studies, this finding may have a significant impact on routine microbiological procedures applied to synovial fluids and brings further support to the key role of bacteria living in biofilm-formed aggregates in joint infections.

Comprehensive analysis of microRNA expression in lumbar facet joint capsules and synovium of patients with osteoarthritis: Comparison between early-stage and late-stage osteoarthritis samples from a single individual

BackgroundMicroRNA is attracting attention as a therapeutic target for osteoarthritis. We focused on joint capsules and synovium in lumbar facet joint osteoarthritis. The purpose of this study was to identify microRNAs that are upregulated in lumbar facet joint capsules and synovium with osteoarthritis. MethodsWe included patients who underwent spinal fusion for degenerative lumbar spine diseases. We selected patients who had both early-stage and late-stage facet joint osteoarthritis in a single individual. We extracted joint capsule and synovium samples from these patients and isolated microRNAs. During the screening phase, we compared early-stage and late-stage osteoarthritis samples from the same individual. We identified microRNAs with >2-fold change in expression in 75% or more of patients with late-stage osteoarthritis using next generation sequencing. During the technical validation phase, the same samples were used for real-time polymerase chain reaction. We identified microRNAs with >2-fold change in expression in 62.5% or more of patients with late-stage osteoarthritis. ResultsOf 40 patients who underwent spinal fusion, we selected eight patients with both early-stage and late-stage facet joint osteoarthritis. During the screening phase, we identified eight upregulated microRNAs out of 2274 microRNAs in late-stage OA. In late-stage OA, two microRNAs (miR-133a-5p and miR-144-3p) were upregulated in seven patients and six microRNAs (miR-133a-3p, miR-133b, miR-206, miR-20a-5p, miR-301a-3p, and miR-32-5p) were upregulated in six patients. During the technical validation phase, we found significant upregulation of miR-144-3p expression in late-stage osteoarthritis compared with early-stage osteoarthritis. Expression of the other microRNAs was not significantly different according to the paired-t test. However, miR-133a-3p, miR-133b, and miR-206 were upregulated >2-fold in 62.5% or more of patients with late-stage osteoarthritis. ConclusionsSome of the microRNAs identified in this study might be involved in joint capsule degeneration or synovitis.

Metabolite asymmetric dimethylarginine (ADMA) functions as a destabilization enhancer of SOX9 mediated by DDAH1 in osteoarthritis.

Osteoarthritis (OA) is a degenerative disease with a series of metabolic changes accompanied by many altered enzymes. Here, we report that the down-regulated dimethylarginine dimethylaminohydrolase-1 (DDAH1) is accompanied by increased asymmetric dimethylarginine (ADMA) in degenerated chondrocytes and in OA samples. Global or chondrocyte-conditional knockout of ADMA hydrolase DDAH1 accelerated OA development in mice. ADMA induces the degeneration and senescence of chondrocytes and reduces the extracellular matrix deposition, thereby accelerating OA progression. ADMA simultaneously binds to SOX9 and its deubiquitinating enzyme USP7, blocking the deubiquitination effects of USP7 on SOX9 and therefore leads to SOX9 degradation. The ADMA level in synovial fluids of patients with OA is increased and has predictive value for OA diagnosis with good sensitivity and specificity. Therefore, activating DDAH1 to reduce ADMA level might be a potential therapeutic strategy for OA treatment.

Mammalian Glycosylation Patterns Protect Citrullinated Chemokine MCP-1/CCL2 from Partial Degradation.

Monocyte chemoattractant protein-1 (MCP-1/CCL2) is a potent chemotactic agent for monocytes, primarily produced by macrophages and endothelial cells. Significantly elevated levels of MCP-1/CCL2 were found in synovial fluids of patients with rheumatoid arthritis (RA), compared to osteoarthritis or other arthritis patients. Several studies suggested an important role for MCP-1 in the massive inflammation at the damaged joint, in part due to its chemotactic and angiogenic effects. It is a known fact that the post-translational modifications (PTMs) of proteins have a significant impact on their properties. In mammals, arginine residues within proteins can be converted into citrulline by peptidylarginine deiminase (PAD) enzymes. Anti-citrullinated protein antibodies (ACPA), recognizing these PTMs, have become a hallmark for rheumatoid arthritis (RA) and other autoimmune diseases and are important in diagnostics and prognosis. In previous studies, we found that citrullination converts the neutrophil attracting chemokine neutrophil-activating peptide 78 (ENA-78) into a potent macrophage chemoattractant. Here we report that both commercially available and recombinant bacterially produced MCP-1/CCL2 are rapidly (partially) degraded upon in vitro citrullination. However, properly glycosylated MCP-1/CCL2 produced by mammalian cells is protected against degradation during efficient citrullination. Site-directed mutagenesis of the potential glycosylation site at the asparagine-14 residue within human MCP-1 revealed lower expression levels in mammalian expression systems. The glycosylation-mediated recombinant chemokine stabilization allows the production of citrullinated MCP-1/CCL2, which can be effectively used to calibrate crucial assays, such as modified ELISAs.

Matrix Metalloproteinase-9 Level in Synovial Fluid-Association with Joint Destruction in Early Rheumatoid Arthritis.

Background and objective: Matrix metalloproteinases (MMPs) are the key enzymes in the pathogenesis of cartilage and joint damage and potentially a new biomarker of the early erosive form of rheumatoid arthritis (RA). Firstly, the study aimed to compare the level of MMP-9 in plasma (PL) and synovial fluid (SF) of patients with RA and osteoarthritis (OA). Secondly, the goal was to examine the association of MMP-9 level in PL and SF with early erosive changes in RA, and finally, to determine the association of MMP-9 level with serological parameters of the disease (rheumatoid factor-RF and anti-citrulline protein antibodies-ACPA). Materials and Methods: A total of 156 subjects were involved in this study (84 patients with RA and 72 patients with OA, who were involved as a control group). MMP-9 level was measured in PL and SF of all subjects by the sandwich enzyme-linked immunosorbent assay (ELISA) method. Standard radiographs of the hands and feet were used to detect joint damage and classification into erosive or non-erosive RA. The Larsen score (LS) was used for the quantitative assessment of joint damage, and its annual change (∆ LS) was used to assess the radiographic progression of the disease. Results: MMP-9 level in PL and SF was significantly higher in RA compared to controls (PL: 19.26 ± 7.54 vs. 14.57 ± 3.11 ng/mL, p< 0.01; SF: 16.17 ± 12.25 vs. 0.75 ± 0.53 ng/mL, p < 0.001) as well as in SF of patients with erosive compared to non-erosive RA (18.43 ± 12.87 vs. 9.36 ± 7.72; p < 0.05). Faster radiographic progression was recorded in erosive compared to non-erosive early RA (11.14 ± 4.75 vs. 6.13 ± 2.72; p < 0.01). MMP-9 level in SF, but not in PL, significantly correlates with the radiographic progression in both erosive and non-erosive RA (ρ = 0.38 and ρ = 0.27). We did not find a significant association between RF and MMP-9 level in early RA, but the ACPA level significantly correlates with MMP-9 level in SF (r = 0.48). Conclusion: The level of MMP-9 in plasma and synovial fluid of patients with RA is significantly higher compared to patients with osteoarthritis. The level of MMP-9 in synovial fluid is significantly higher in erosive than non-erosive early RA. It is significantly associated with the radiographic progression of the disease and the level of anti-citrulline protein antibodies.

The Diagnostic Value of miR-124a Expression in Peripheral Blood and Synovial Fluid of Patients with Rheumatoid Arthritis

Introduction: Rheumatoid arthritis (RA), a chronic autoimmune disorder, is currently a severe health threat. Previous studies have documented the altered expression of various miRNAs in RA patients. This study determined the expression of miR-124a in RA patients and estimated its diagnostic value for RA. Methods: A total of 80 RA patients were enrolled as the study subjects, and 36 patients with osteoarthritis were included, with another 36 healthy people as the controls. miR-124a expression levels in peripheral blood plasma, peripheral blood mononuclear cells (PBMCs), and synovial fluid were measured using reverse transcription quantitative polymerase chain reaction, followed by Pearson correlation analysis. Additionally, the association between miR-124a and major clinical indicators was assessed, such as rheumatoid factor (RF), erythrocyte sedimentation rate (ESR), and disease activity score of 28 joints (DAS28). The diagnostic efficacy of miR-124a expression in plasma, PBMCs, and synovial fluid for RA was evaluated by the receiver operating characteristic curve, and the difference in the area under the curve (AUC) was analyzed. Results: miR-124a was downregulated in RA patients, and the expression levels of miR-124a in plasma, PBMCs, and synovial fluid showed a certain degree of positive correlation. miR-124a was inversely linked with RF, ESR, and DAS28. For the diagnosis of RA patients, the AUC of plasma miR-124a was 0.899 and the cut-off value was 0.800, with 68.75% sensitivity and 94.44% specificity; the AUC of miR-124a in PBMCs was 0.937 and the cut-off value was 0.805, with 82.50% sensitivity and 91.67% specificity; the AUC of miR-124a in plasma combined with PBMCs was 0.961, with a higher diagnostic value than independent plasma or PBMCs; the AUC of miR-124a in synovial fluid was 0.929 and the cut-off value was 0.835, with 80.00% sensitivity and 88.89% specificity. Conclusion: miR-124a expression is downregulated in the plasma, PBMCs, and synovial fluid of RA patients and has a high diagnostic value for RA.

ОКИСНА МОДИФІКАЦІЯ БІЛКІВ У СИНОВІАЛЬНІЙ РІДИНІ ХВОРИХ НА ОСТЕОАРТРИТ ПІСЛЯ SARS-CoV2-ІНФЕКЦІЇ

The pandemic of coronavirus disease 2019 (COVID-19) caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has made a serious public health threat worldwide. It has caused many complications and deaths, so millions of people are at risk in more and more countries. A special group consists of patients with chronic diseases, particularly osteoarthritis. It is known that the development of osteoarthritis is associated with oxidative stress and excessive production of free radicals. Proteins are highly sensitive to free radical oxidation, so their oxidation level reflects the oxidant-antioxidant balance in the body. The aim of the work was to determine the concentration of products of oxidative modification of proteins and sulfhydryl groups in the synovial fluid of patients with osteoarthritis after SARS-CoV2 infection. All study participants were divided into two groups. The first group (n=22) is patients with osteoarthritis of the knee joints II-III degree. The second group (n=14) is patients with osteoarthritis of the knee joints II-III degree, who suffered a mild and moderate form of COVID-19 6-9 months ago. In the conducted studies, the products of protein oxidation in the synovial fluid of patients of all experimental groups were evaluated. The concentration of products of oxidative modification of proteins was determined by the level of carbonyl derivatives, which are detected in the reaction with 2,4-dinitrophenylhydrazine. The concentration of total, protein-bound and non-protein sulfhydryl (SH) groups was measured by the Ellman method. Processing of research results was carried out using generally accepted methods of variational statistics. It was established that in the synovial fluid of patients with osteoarthritis of the knee joints who contracted COVID-19, the concentration of products of oxidative modification of proteins increased. Under the same experimental conditions, the concentration of sulfhydryl groups decreased. The detected changes indicate a violation of the oxidant-antioxidant balance in the synovial fluid of patients with osteoarthritis after SARS-CoV-2 infection. This indicates the prevalence of free radical processes and the development of oxidative stress directly in the joint. Our results indicate that patients with osteoarthritis may develop more severe joint damage after infection with COVID-19.

Synovial microflora of large joints in patients of a multidisciplinary hospital

Relevance. There is no doubt that microorganisms participate in occurrence and development of septic process in joints. However, the issue of etiological significance of each agent remains controversial, because, in spite of the general trends, indicating the participation of numerous microorganisms in the development of articular pathology, each result of microbiological analysis concerns only a specific case in the territorial and clinical aspects. The aim of the study - microbiological research of synovial fluid obtained from knee joint during synovitis after its aspiration in patients of various departments of the Stavropol Regional Clinical Hospital. Materials and Methods . There were studied 198 samples of synovial fluid. Primary inoculation of puncture was performed with subsequent isolation, identification of the cultures by mass spectrometry and assessment of their antibiotic sensitivity by discodiffusion method. Results and Discussion . 11 cultures of bacterial pathogens were isolated. Gram-positive cocci - 82 %, of which 77.8 % - microorganisms of Staphylococcus genus (44.4 % S.aureus , 33.4 % S.epidermidis ), 22.2 % - other gram-positive cocci: one strain of each, Enterococcus faecium and Streptococcus mitis . Gram-negative pathogens are represented by K.neumoniae and P.aeruginosa with a total content of 18 %. Highly virulent microorganisms S.aureus , K.neumoniae and P.aeruginosa are isolated from the synovial fluid of patients of the surgical departments (orthopedotraumatological No. 1, No. 2) and the rheumatological department. Microorganisms with low virulence E.faecium , S.mitis and S.epidermidis are isolated from synovial fluid of patients of various departments. No obvious resistance of isolated pathogens to antimicrobial drugs has been registered. Conclusion . The presence and species affiliation of the microorganisms identified in synovial fluid allows predicting their etiological significance in development of septic process in joints. Their role as causative agents of nosocomial infections typical for a medical institution is not excluded. The presence of articular pathology in each of the examined departments dictates the need for a clear understanding of the importance of timely and high-quality joint aspiration followed by microbiological examination in almost all patients with damage of large joints, including patients without clinical signs of septic arthritis. Such an approach that makes it possible to identify a greater number of causative agents of septic arthritis and quickly evaluate the dynamics of their antimicrobial resistance should become an obligatory part of a comprehensive research and treatment of a patient with arthritis in multi-fi eld hospitals.

Altered lipid metabolism in synovial fibroblasts of individuals at risk of developing rheumatoid arthritis

ObjectiveFibroblast-like synoviocytes (FLS) can augment the inflammatory process observed in synovium of patients with rheumatoid arthritis (RA). A recent transcriptomic study in synovial biopsies revealed changes in metabolic pathways before disease onset in absence of synovial tissue inflammation. This raises the question whether alterations in cellular metabolism in tissue resident FLS underlie disease pathogenesis. Materials and methodsTo study this, we compared the metabolic profile of FLS isolated from synovial biopsies from individuals with arthralgia who were autoantibody positive but without any evidence of arthritis (RA-risk individuals, n = 6) with FLS from patients with RA (n = 6), osteoarthritis (OA, n = 6) and seronegative controls (n = 6). After synovial digestion, FLS were cultured in vitro and cellular metabolism was assessed using quantitative PCR, flow cytometry, XFe96 Seahorse Analyzer and tritium-labelled oleate oxidation assays. ResultsReal-time metabolic profiling revealed that basal (p < 0.0001) and maximum mitochondrial respiration (p = 0.0024) were significantly lower in RA FLS compared with control FLS. In all donors, basal respiration was largely dependent on fatty acid oxidation while glucose was only highly used by FLS from RA patients. Moreover, we showed that RA-risk and RA FLS are less metabolically flexible. Strikingly, mitochondrial fatty acid β-oxidation was significantly impaired in RA-risk (p = 0.001) and RA FLS (p < 0.0001) compared with control FLS. ConclusionOverall, this study showed several metabolic alterations in FLS even in absence of synovial inflammation, suggesting that these alterations already start before clinical manifestation of disease and may drive disease pathogenesis.

Validation of Levels of Decorin as A Reliable Biomarker of Osteoarthritis: Comparison of Serum and Synovial Fluid Levels

Objective: The proteoglycan decorin, which plays an important part in the aetiology of osteoarthritis as well as a role in the binding of collagen, was the focus of this study's background information and objectives (OA). This investigation was carried out with the objectives of determining the levels of decorin in the blood and synovial fluid of patients who suffered from knee OA and determining whether or not these levels had a correlation with OA and the Western Ontario and McMaster Universities Osteoarthritis (WOMAC) score. Place of Study: Islam medical college Sialkot Duration: December 2021 to May 2022 Methods: In this prospective study, a total of 60 persons were enrolled in the study and control groups: 30 people with knee osteoarthritis and 30 people with other knee sjoint problems. For the purpose of determining how well the knee functions, the WOMAC score was utilised.Both the levels of decorin in the blood and the levels of decorin in the synovial fluid were analysed by means of an enzyme-linked immunosorbent assay (ELISA). We used a technique known as binary logistic regression, which contained both a single and a multi-categorical predictor, in order to evaluate the potential risk factors for OA. This allowed us to determine which characteristics would increase the likelihood of developing OA. Results: It was discovered that the levels of serum decorin in the group that was diagnosed with OA were statistically and considerably higher than the levels found in the group that acted as the control (P 0.001). There was not a significant difference in the amounts of decorin found in synovial fluid between those who had OA and the control group. WOMAC score (OR)=1.073, 95% confidence interval (CI): 1.032-1.116, P&lt;0 .001 This association between OA and high serum decorin levels was shown to be statistically significant (OR=1.114, 95%CI: 1.030–1.205, P=0.007). OA was demonstrated to be strongly linked with high serum decorin levels. There was shown to be a positive association between the levels of serum decorin and the WOMAC score in patients who had OA. Conclusions: An increase in serum decorin levels may be indicative of changes in the structure of the extracellular matrix.The fact that there is a positive connection between the serum decorin level and the WOMAC score lends credence to the previous assertion that this conclusion is correct. It was found that having a higher WOMAC score and having higher serum decorin levels are both risk factors associated with osteoarthritis (OA). On the other hand, there was no link found between osteoarthritis and the amount of decorin found in the joint fluid.

NLRC4-mediated activation of CD1c+ DC contributes to perpetuation of synovitis in rheumatoid arthritis.

The individual contribution of specific myeloid subsets such as CD1c+ conventional DC (cDC) to perpetuation of rheumatoid arthritis (RA) pathology remains unclear. In addition, the specific innate sensors driving pathogenic activation of CD1c+ cDC in patients with RA and their functional implications have not been characterized. Here, we assessed phenotypical, transcriptional, and functional characteristics of CD1c+ and CD141+ cDC and monocytes from the blood and synovial fluid of patients with RA. Increased levels of CCR2 and the IgG receptor CD64 on circulating CD1c+ cDC was associated with the presence of this DC subset in the synovial membrane in patients with RA. Moreover, synovial CD1c+ cDC are characterized by increased expression of proinflammatory cytokines and high abilities to induce pathogenic IFN-γ+IL-17+CD4+ T cells in vitro. Finally, we identified the crosstalk between Fcγ receptors and NLRC4 as a potential molecular mechanism mediating pathogenic activation, CD64 upregulation, and functional specialization of CD1c+ cDC in response to dsDNA-IgG in patients with RA.

Contribution of MicroRNA-27b-3p to Synovial Fibrotic Responses in Knee Osteoarthritis.

Synovial fibrosis contributes to osteoarthritis (OA) pathology, but the underlying mechanisms remain unknown. We have observed increased microRNA-27b-3p (miR-27b-3p) levels in synovial fluid of patients with late-stage radiographic knee OA. Here, we investigated the contribution of miR-27b-3p to synovial fibrosis in patients with severe knee OA and in a mouse model of knee OA. We stained synovium sections obtained from patients with radiographic knee OA scored according to the Kellgren/Lawrence scale and mice that underwent destabilization of the medial meniscus (DMM) for miR-27b-3p using in situ hybridization. We examined the effects of intraarticular injection of miR-27b-3p mimic into naive mouse knee joints and intraarticular injection of a miR-27b-3p inhibitor into mouse knee joints after DMM. We performed transfection with miR-27b-3p mimic and miR-27b-3p inhibitor in human OA fibroblast-like synoviocytes (FLS) using reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR) array, RNA sequencing, RT-qPCR, Western blotting, immunofluorescence, and migration assays. We observed increased miR-27b-3p expression in the synovium from patients with knee OA and in mice with DMM-induced arthritis. Injection of the miR-27b-3p mimic in mouse knee joints induced a synovial fibrosis-like phenotype, increased synovitis scores, and increased COL1A1 and α-smooth muscle actin (α-SMA) expression. In the mouse model of DMM-induced arthritis, injection of the miR-27b-3p inhibitor decreased α-SMA but did not change COL1A1 expression levels or synovitis scores. Transfection with the miR-27b-3p mimic in human OA FLS induced profibrotic responses, including increased migration and expression of key extracellular matrix (ECM) genes, but transfection with the miR-27b-3p inhibitor had the opposite effects. RNA sequencing identified a PPARG/ADAMTS8 signaling axis regulated by miR-27b-3p in OA FLS. Human OA FLS transfected with miR-27b-3p mimic and then treated with the PPARG agonist rosiglitazone or with ADAMTS8 small interfering RNA exhibited altered expression of select ECM genes. Our findings demonstrate that miR-27b-3p has a key role in ECM regulation associated with synovial fibrosis during OA.

Study of the Element Concentrations in Synovial Fluids of Patients with Arthritis and Rheumatic Diseases

Study of the Element Concentrations in Synovial Fluids of Patients with Arthritis and Rheumatic Diseases

CCL24, CXCL9 and CXCL10 are increased in synovial fluid in patients with juvenile idiopathic arthritis requiring advanced treatment.

Oligoarticular JIA disease progression and outcomes are variable. Our objective is to detect protein markers that would allow for earlier intervention to potentially halt disease progression. In this retrospective study of serial SF samples, elevated expression of CCL24, CXCL9 and CXCL10 was linked to the eventual need for advanced medications. Serial SF samples were selected from patients with persistent and extended oligoarticular JIA. The samples were separated into two groups: those who did and did not receive advanced medications throughout their disease course. Protein antibody arrays and Luminex assays were performed to determine changes in protein expression. CCL24, CXCL9 and CXCL10 expression levels were significantly higher in patients who eventually required advanced treatment than in those who did not. The expression levels of CCL24 and CXCL9 were consistently elevated in paired samples of those who later received advanced medications. In the persistent oligoarticular JIA group, CXCL10 levels remained elevated over time in those who required advanced treatment. Conversely, CCL24 levels decreased in patients who did not require advanced treatment. In the extended samples, the levels of CCL24 and CXCL10 expression increased significantly over time in the patients who ultimately required advanced treatment. In patients with oligoarticular JIA, regardless of disease onset and progression, the consistent elevation of any or all three markers, the CCL24, CXCL9 and CXCL10 in SFs was associated with the future use of advanced therapy, which could be reflective of disease severity.

Association of synovial expression of growth and differentiation factor 5 (GDF5) with radiographic severity of knee osteoarthritis

ObjectivesThe growth and differentiation factor 5 (GDF5) gene plays a significant role in facilitating chondrogenesis, and GDF5 polymorphism is a genetic factor contributing to osteoarthritis (OA). However, the role of GDF5 expression in the synovial membrane remains unclear. The aim of this study was to determine the expression of GDF5 in the synovium in patients with primary knee OA. Materials and methodsThirty patients scheduled for total knee arthroplasty were enrolled. Patients were grouped according to the Kellgren and Lawrence classification (KL) as grade 3 (15 patients) and grade 4 (15 patients). Synovial tissue was collected, and the GDF5 expression level was determined by real-time PCR. Ten synovial samples were randomly selected to evaluate the degree of synovitis. ResultsBaseline characteristics did not differ between the two groups. The expression of GDF5 was significantly higher in the KL4 group (median expression 3.50, range 1.45–13.62) than in the KL3 group (median expression 1.81, range 0–9.46) (p value = 0.02). Histological staining of the synovium indicated low-grade synovitis in both groups. ConclusionsGDF5 expression in the synovium was positively associated with the radiographic severity of knee OA. The difference in GDF5 expression between the KL3 and KL4 groups supports the hypothesis that, through GDF5, the synovium may have important roles in cartilage maintenance and homeostasis in primary knee OA.

Comprehensive Analysis and Functional Characteristics of Differential Expression of N6-Methyladenosine Methylation Modification in the Whole Transcriptome of Rheumatoid Arthritis.

N6-methyladenosine (m6A) modification is the most prevalent chemical modification in eukaryotic mRNA and is associated with the development of various immune diseases. However, the role of m6A methylation in rheumatoid arthritis (RA) development is unclear. We preliminarily explored the role of m6A methylation-related mRNAs in RA for its clinical application. The discovery of m6A methylation-modifying genes in this study may provide a fresh perspective on the development of drugs for RA treatment. High-throughput sequencing combined with methylated RNA immunoprecipitation (MeRIP-seq) and RNA sequencing were used to assess whole-transcriptome m6A modifications in the synovium of patients with RA. The relationship between m6A-modified target genes and RA inflammation and macrophages was determined. The expression of the m6A-modified significant transcript-enriched inflammatory signaling pathway was assessed through animal experiments. Differentially expressed m6A genes were correlated with macrophage activation involved in immune response, vascular endothelium, MAPK signaling pathway, PI3K - Akt signaling pathway, and other inflammatory processes. Furthermore, combined analysis with m6A-seq and RNA-seq revealed 120 genes with significant changes in both m6A modification and mRNA expression. We selected the top 3 candidate mRNAs that were upregulated and downregulated simultaneously. The expression of phosphatase and tensin homolog deleted on chromosome ten (PTEN) mRNA and protein in RA patients was lower than that in healthy control (HC). SHC-binding protein 1 (SHCBP1) and neurexophilin-3 (NXPH3) mRNA expressions were increased in RA patients. The expression of M1 macrophages was increased in RA patients. RA markers are such as rheumatoid factor (RF) and peptide containing citrulline (CCP). Further animal experiments showed that the expression of synovial MAPK, PI3K, and Akt1 proteins in the RA model was increased, and the PTEN, p-PTEN protein expression was decreased. PI3K, Akt1, PTEN, and p-PTEN were correlated to RA joint inflammation. This study revealed a unique pattern of differential m6A methylation modifications in RA and concluded that m6A modification is related to the occurrence of RA synovial inflammation.

Acid sensor ASIC1a induces synovial fibroblast proliferation via Wnt/β-catenin/c-Myc pathway in rheumatoid arthritis

Rheumatoid arthritis (RA) is a chronic autoimmune disease characterized by synovial hyperplasia and progressive joint destruction in the middle and late stages. Notably, activated rheumatoid arthritis synovial fibroblasts (RASFs) exhibit tumor-like features, including an increased proliferation rate that largely contributes to pannus formation and joint destruction. Our previous studies have demonstrated that acid-sensing ion channel 1a (ASIC1a) was highly expressed in RASFs, and acidic microenvironment of synovial fluid in patients with RA can activate ASIC1a to promote synovial inflammation, leading to the progression of RA. However, the role and possible mechanism of ASIC1a in RASF proliferation remains unclear. The present study aimed to investigate the effect of ASIC1a activation upon acidosis on RASF proliferation and its molecular mechanism in vivo and in vitro. The results of in vitro experiments showed that activation of ASIC1a upon acidosis promoted the proliferation of RASFs, which could be attenuated by the specific ASIC1a inhibitor Psalmotoxin-1 (PcTx-1) or specific siRNA for ASIC1a. Mechanistically, Wnt/β-catenin/c-Myc signaling pathway was involved in ASIC1a-induced RASF proliferation. The results of in vivo experiments indicated that intra-articular injection of PcTx-1 reduced synovial hyperplasia and ameliorated cartilage degradation in rats with adjuvant arthritis (AA). Collectively, these results suggest that activation of ASIC1a upon acidosis promotes RASF proliferation, and the mechanism may be related to Wnt/β-catenin/c-Myc pathway.