Upcoming evidence in clinical practice of two-stage revision arthroplasty for prosthetic joint infection.

Total joint arthroplasty is the recommended treatment for patients with end-stage osteoarthritis, as it reduces disability and pain and restores joint function. However, prosthetic joint infection is a serious complication of this procedure, with the two-stage exchange being the most common treatment method. While there is consensus on diagnosing prosthetic joint infection, there is a lack of agreement on the parameters that can guide the surgeon in performing definitive reimplantation in a two-stage procedure. One approach that has been suggested to improve the accuracy of microbiologic investigations before definitive reimplantation is to observe a holiday period from antibiotic therapy to improve the accuracy of cultures from periprosthetic tissues, but these cultures report some degree of aspecificity. Therefore, several pieces of evidence highlight that performing reimplantation using continuous antibiotic therapy should be considered a safe and effective approach, leading to higher cure rates and a shorter period of disability. Dosage of C-reactive protein (CRP), erythrocyte sedimentation rate (ERS) and D-dimer are helpful in diagnosing prosthetic joint infection, but only D-dimer has shown sufficient accuracy in predicting the risk of infection recurrence after a two-stage procedure. Synovial fluid analysis before reimplantation has been shown to be the most accurate in predicting recurrence, and new cutoff values for leukocyte count and neutrophil percentage have shown a useful predictive rule to identify patients at risk of unfavourable outcome. A new scoring system based on a numerical score calculated from the beta coefficient derived through multivariate analysis of D-dimer levels, synovial fluid leukocytes and relative neutrophils percentage has demonstrated high accuracy when it comes to guiding the second step of two-stage procedure. In conclusion, reimplantation may be a suitable option for patients who are on continuous therapy without local symptoms, and with CRP and ERS within the normal range, with low synovial fluid leukocytes (< 952/mL) and a low relative neutrophil percentage (< 52%) and D-dimer below 1100 µg/mL. A numerical score derived from analysing these three parameters can serve as a valuable tool in determining the feasibility of reimplantation in these patients.

Integration of metabolomics and transcriptomics provides insights into the molecular mechanism of temporomandibular joint osteoarthritis.

The deficiency of clinically specific biomarkers has made it difficult to achieve an accurate diagnosis of temporomandibular joint osteoarthritis (TMJ-OA) and the insufficient comprehension of the pathogenesis of the pathogenesis of TMJ-OA has posed challenges in advancing therapeutic measures. The combined use of metabolomics and transcriptomics technologies presents a highly effective method for identifying vital metabolic pathways and key genes in TMJ-OA patients. In this study, an analysis of synovial fluid untargeted metabolomics of 6 TMJ-OA groups and 6 temporomandibular joint reducible anterior disc displacement (TMJ-DD) groups was conducted using liquid and gas chromatography mass spectrometry (LC/GC-MS). The differential metabolites (DMs) between TMJ-OA and TMJ-DD groups were analyzed through multivariate analysis. Meanwhile, a transcriptomic dataset (GSE205389) was obtained from the GEO database to analyze the differential metabolism-related genes (DE-MTGs) between TMJ-OA and TMJ-DD groups. Finally, an integrated analysis of DMs and DE-MTGs was carried out to investigate the molecular mechanisms associated with TMJ-OA. The analysis revealed significant differences in the levels of 46 DMs between TMJ-OA and TMJ-DD groups, of which 3 metabolites (L-carnitine, taurine, and adenosine) were identified as potential biomarkers for TMJ-OA. Collectively, differential expression analysis identified 20 DE-MTGs. Furthermore, the integration of metabolomics and transcriptomics analysis revealed that the tricarboxylic acid (TCA) cycle, alanine, aspartate and glutamate metabolism, ferroptosis were significantly enriched. This study provides valuable insights into the metabolic abnormalities and associated pathogenic mechanisms, improving our understanding of TMJOA etiopathogenesis and facilitating potential target screening for therapeutic intervention.

Determination of vancomycin and meropenem in serum and synovial fluid of patients with prosthetic joint infections using UPLC-MS/MS.

Numerous studies have suggested that intra-articular administration of antibiotics following primary revision surgery may be one of the methods for treating prosthetic joint infection (PJI). Vancomycin and meropenem are the two most commonly used antibiotics for local application. Determining the concentrations of vancomycin and meropenem in the serum and synovial fluid of patients with PJI plays a significant role in further optimizing local medication schemes and effectively eradicating biofilm infections. This study aimed to establish a rapid, sensitive, and accurate ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method for determining the concentrations of vancomycin and meropenem in human serum and synovial fluid. Serum samples were processed using acetonitrile precipitation of proteins and dichloromethane extraction, while synovial fluid samples were diluted before analysis. Chromatographic separation was achieved in 6 min on a Waters Acquity UPLC BEH C18 column, with the mobile phase consisting of 0.1% formic acid in water (solvent A) and acetonitrile (solvent B). Quantification was carried out using a Waters XEVO TQD triple quadrupole mass spectrometer with an electrospray ionization (ESI) source in positive ion mode. The multiple reaction monitoring (MRM) mode was employed to detect the following quantifier ion transitions: 717.95-99.97 (norvancomycin), 725.90-100.04 (vancomycin), 384.16-67.99 (meropenem). The method validation conformed to the guidelines of the FDA and the Chinese Pharmacopoeia. The method demonstrated good linearity within the range of 0.5-50 µg/ml for serum and 0.5-100 µg/ml for synovial fluid. Selectivity, intra-day and inter-day precision and accuracy, extraction recovery, matrix effect, and stability validation results all met the required standards. This method has been successfully applied in the pharmacokinetic/pharmacodynamic (PK/PD) studies of patients with PJI.

Analysis of the Level of Adiponectin and Selected Cytokines in Patients with Knee Osteoarthritis.

Background and Objectives: Knee osteoarthritis (KOA) is a degenerative disease that is continuously targeting people of different ages, but especially the elderly population, the number of which tends to increase continuously at the global level. Apart from age, excess weight can influence the evolution of the disease, with obesity being associated with a weak inflammation stage and an imbalance between pro-inflammatory and anti-inflammatory cytokines. The present work aimed to analyze specific biomarkers, namely ACRP-30, IL-10, TNF-α, and IL-6, in knee synovial fluid, and correlate them with KOA patients' clinical data, radiographic changes, and functional and pain scores. Materials and Methods: 24 subjects with KOA and over 50 years of age participate in the present study. Synovial fluid was harvested using ultrasound guidance from the target knees of the enrolled KOA patients, and the levels of ACRP-30, IL-10, TNF-α, and IL-6 were measured using enzyme-linked immunosorbent assays (ELISA). All patients underwent a supine X-ray at the target knee and were classified using Kellgren-Lawrence (K-L) grading. The Western Ontario and McMaster University Osteoarthritis Index (WOMAC) was used to assess self-reported physical function, pain, and stiffness. Results: The obtained results highlighted a significant correlation between age and adiponectin level (p = 0.0451, r = -0.412). Also, the IL-10 values are lower in cases where the intensity of the pain is more pronounced (p = 0.0405, r = -0.421). In addition, analyzing the data by gender, it was observed that in the case of males, stiffness is more related to age (p = 0.0079, r = 0.7993), compared to women (p = 0.0203, r = 0.6223). In the case of women, the progression of the disease tends to increase more intensively the WOMAC score's total values (p = 0.00031, r = 0.8342), compared with men (p = 0.0289, r = 7013). Regarding interleukins and BMI, significant correlations were observed only in the case of men. Conclusions: A significant correlation between age and adiponectin, and adiponectin and IL-6, suggests that advanced age may contribute to adiponectin reduction. Comparing men with women, it was observed that men's age is more related to rigidity, and IL-6 and IL-10 are directly correlated to BMI; in addition, women seem to be more sensitive to pain and stiffness.

Advancing the Diagnosis of Periprosthetic Joint Infections: Integrated Microfluidic Platform for Alpha-Defensins-Specific Aptamer Selection and Its Analytical Applications.

Periprosthetic joint infections (PJIs) pose a significant challenge in orthopedic surgery, particularly total joint arthroplasty (TJA), due to the potential for implant failure and increased patient morbidity. Early and accurate detection of PJIs is crucial for timely intervention and better patient prognosis. Herein, we successfully screened a high-affinity aptamer targeting alpha-defensin complex human neutrophil protein 1-3 (HNP 1-3; potential PJI biomarkers in synovial fluid [SF]) for the first time using systematic evolution of ligands by exponential enrichment (SELEX) on an integrated microfluidic platform. The compact microfluidic device enabled efficient screening, with each round completed within <2 h, comprising five rounds of positive selection, two rounds of negative selection, and one round of competitive selection. A novel one-aptamer-one-antibody assay was further developed from the optimal aptamer screened, and it could accurately quantify HNP 1-3 in SF within 3 h with only ∼50 µL of SF. The assay demonstrated strong binding affinity and specificity for the target protein in SF. Thirteen PJI SF samples were accurately diagnosed and the assay was accurate over a wide dynamic range (0.32-100 mg/L). This study has showcased a rapid and accurate diagnostic tool for PJI detection, which should see widespread use in the clinic, holding promise for potential analytical applications in orthopedic surgery and improving patient care.

Intraoperative calprotectin lateral flow immunoassay can assist decision-making between one- and two-stage revision total hip arthroplasty for patients with suspected periprosthetic joint infection.

Aims: Accurate diagnosis of chronic periprosthetic joint infection (PJI) presents a significant challenge for hip surgeons. Preoperative diagnosis is not always easy to establish, making the intraoperative decision-making process crucial in deciding between one- and two-stage revision total hip arthroplasty (THA). Calprotectin is a promising point-of-care novel biomarker that has displayed high accuracy in detecting PJI. We aimed to evaluate the utility of intraoperative calprotectin lateral flow immunoassay (LFI) in THA patients with suspected chronic PJI. Methods: The study included 48 THAs in 48 patients with a clinical suspicion of PJI, but who did not meet European Bone and Joint Infection Society (EBJIS) PJI criteria preoperatively, out of 105 patients undergoing revision THA at our institution for possible PJI between November 2020 and December 2022. Intraoperatively, synovial fluid calprotectin was measured with LFI. Cases with calprotectin levels ≥ 50 mg/l were considered infected and treated with two-stage revision THA; in negative cases, one-stage revision was performed. At least five tissue cultures were obtained; the implants removed were sent for sonication. Results: Calprotectin was positive (≥ 50 mg/l) in 27 cases; out of these, 25 had positive tissue cultures and/or sonication. Calprotectin was negative in 21 cases. There was one false negative case, which had positive tissue cultures. Calprotectin showed an area under the curve of 0.917, sensitivity of 96.2%, specificity of 90.9%, positive predictive value of 92.6%, negative predictive value of 95.2%, positive likelihood ratio of 10.6, and negative likelihood ratio of 0.04. Overall, 45/48 patients were correctly diagnosed and treated by our algorithm, which included intraoperative calprotectin measurement. This yielded a 93.8% concordance with postoperatively assessed EBJIS criteria. Conclusion: Calprotectin can be a valuable tool in facilitating the intraoperative decision-making process for cases in which chronic PJI is suspected and diagnosis cannot be established preoperatively.

CCR9+CD4+ T cells are associated with disease activity in patients with rheumatoid arthritis.

An increase in CD4+ T cells in the synovium is closely linked to the pathogenesis of rheumatoid arthritis (RA). We aimed to identify the possible causes of the elevated CD4+ T cell levels and to explore the factors influencing disease activity in RA. Fifty-five RA patients, including 28 with active RA (ARA), 27 with inactive RA, and 22 healthy controls, were recruited for this study. The proportion of CCR9+CD4+ T cells and the expression of chemokine receptor 9 (CCR9) on CD4+ T cells were analyzed by flow cytometry. Enzyme-linked immunosorbent assay and chemiluminescent immunoassay were used to evaluate interleukin (IL)-17A and IL-6 levels, respectively. The proportion of CCR9+CD4+ T cells and the expression of CCR9 on CD4+ T cells increased significantly in peripheral blood (PB) and synovial fluid (SF) in ARA compared to those in inactive RA. Furthermore, SF contained more CCR9+CD4+ T cells, IL-6, and IL-17A than PB in RA patients. Moreover, CD4+ T cells in the PB of patients with RA, especially ARA, expressed more CCR9 and secreted more IL-6 and IL-17A after activation. Here, we also demonstrated that both the percentage of CCR9+ cells in CD4+ T cells and the expression of CCR9 on circulating CD4+ T cells were positively correlated with erythrocyte sedimentation rate, hypersensitive C-reactive protein, rheumatoid factor, and anti-cyclic citrullinated peptide antibody. CCR9+CD4+ T cells are elevated in PB and SF, and are associated with disease activity in patients with RA.

The alterations in nerve growth factor concentration in plasma and synovial fluid before and after total knee arthroplasty.

Total knee arthroplasty (TKA) is an effective procedure for pain relief; however, the emergence of postsurgical pain remains a concern. In this study, we investigated the production of nerve growth factor (NGF) and mediators that affect NGF production and their function in the synovial fluid and plasma after TKA. This study included 19 patients (20 knees) who had rheumatoid arthritis (RA), systemic lupus erythematosus (SLE), and knee osteoarthritis (OA) who underwent TKA, categorized into OA and non-OA groups. The levels of NGF, inflammatory cytokines, and lipid mediators were analyzed before and after surgery. The intraoperative synovial fluid NGF concentration was more than seven times higher in the non-OA group than in the OA group. The intra-articular NGF levels increased significantly by more than threefold postoperatively in the OA group but not in the non-OA group. Moreover, the levels of inflammatory cytokines and lipid mediators were increased in the synovial fluid of both groups. The intra-articular cytokines or NGF concentrations positively correlated with postoperative pain. Targeted NGF control has the potential to alleviate postsurgical pain in TKA, especially in patients with OA, emphasizing the importance of understanding NGF dynamics under different knee conditions.

Associations with unplanned repeat irrigation and debridement of native septic arthritis.

PURPOSE: To identify associations with unplanned repeat irrigation and debridement (I&D) after arthrotomy for native septic arthritis. METHODS: A retrospective review identified patients with native septic arthritis treated with open arthrotomies. The primary outcome was unplanned repeat I&D within 90 days. Associations evaluated for included comorbidities, ability to bear weight, fever, immunosuppressed status, purulence, C-reactive protein, erythrocyte sedimentation rate, white blood cell count (synovial fluid and serum levels), and synovial fluid polymorphonuclear cell percentage (PMN%). RESULTS: There were 59 arthrotomies in 53 patients involving the knee (n = 32), shoulder (n = 10), elbow (n = 8), ankle (n = 6), and hip (n = 3). The median patient age was 52, and a 71.2% were male. An unplanned repeat I&D was required in 40.7% (n = 24). The median time to the second I&D was 4 days (interquartile range 3 to 9). On univariate analysis, unplanned repeat I&Ds were associated with fever (p = 0.03), purulence (p = 0.01), bacteria growth on cultures (p = 0.02), and the use of deep drains (p = 0.05). On multivariate analysis, the only variables that remained associated with unplanned repeat I&Ds were fever (odds ratio (OR) 5.5, 95% confidence interval (CI) 1.3, 23.6, p = 0.02) and purulence (OR 5.3, CI 1.1, 24.4, p = 0.03). CONCLUSIONS: An unplanned repeat I&D was required in 40.7% of patients and was associated with fever and purulence. These findings highlight the difficulty of controlling these infections and support the need for future research into better methods of management. LEVEL OF EVIDENCE: Diagnostic, Level III.

Evaluation of predictive factors of septic wrist to avoid overdiagnosis.

BACKGROUND: The existing diagnostic criteria for septic wrist are nonspecific, exposing patients with noninfectious etiologies to surgical morbidity. This study aimed to identify predictors differentiating septic wrist from other etiologies. METHODS: An institutional review board-approved retrospective review was conducted on patients with a presumed diagnosis of septic wrist (2003-2022). Bivariate and multiple regression analyses were performed to identify correlation between confirmed septic wrist and comorbidities (autoimmune diseases, immunosuppression, crystalline arthropathy, intravenous [IV] drug use, smoking), penetrating trauma, fever, multi-joint involvement, inflammatory markers (erythrocyte sedimentation rate [ESR]/C-reactive protein [CRP]/white blood cells [WBC]), serum uric acid level, blood cultures, imaging findings, and synovial fluid analysis. Categorical data were reported as median [interquartile range]. RESULTS: Hundred and sixty-eight (58 females and 110 males) patients were included. The median length of hospitalization and follow-up were 6[7] days and 1[3] months. Eighty-nine (53%) patients had septic wrist confirmed with Gram stain/culture, 48 (29%) patients received alternative diagnoses, and 31 (18%) patients had undetermined diagnoses. Concomitant septic wrist and crystalline arthropathy were identified in 9 patients (6.6% of total patients). Out of the 48 patients who received alternative diagnoses, 12 (25%) underwent open drainage. Elevated synovial WBC count (95,409.4 ± 85,926.2) showed a trend of association with septic wrist (p = 0.08). Negative synovial crystals (p = 0.01), positive blood culture (p = 0.04), negative history of crystalline arthropathy (p = 0.08), and multi-joint involvement (p = 0.05) were identified as predictors of septic wrist with a combined sensitivity of 87.5%, specificity of 86.2%, and area under the curve 0.93. CONCLUSIONS: Current diagnostic criteria for septic wrist have low specificity. Negative history of crystalline arthropathy, multi-joint involvement, absence of synovial crystals, and positive blood culture are helpful indicators for predicting septic wrist in patients presenting with a painful, erythematous, and swollen wrist.

Extracellular microRNAs induce dendritic cell-dependent joint inflammation and potentiate osteoclast differentiation via TLR7/8 engagement.

OBJECTIVES: Monocyte-derived dendritic cells (DCs) are key players in the induction of inflammation, autoreactive T cell activation and loss of tolerance in rheumatoid arthritis (RA), but the precise mechanisms underlying their activation remain elusive. Here, we hypothesized that extracellular microRNAs released in RA synovial fluids may represent a novel, physiological stimulus triggering unwanted immune response via TLR8-expressing DC stimulation. METHODS: Human monocyte-derived DCs were stimulated with a mixture of GU-rich miRNAs upregulated in RA tissues and released in synovial fluids (Ex-miRNAs). Activation of DCs was assessed in terms of NF-κB activation by Western blot, cytokine production by ELISA, T cell proliferation and polarization by allogeneic mixed lymphocyte reaction. DC differentiation into osteoclasts was evaluated in terms of tartrate-resistant acid phosphatase production and formation of resorption pits in dentine slices. Induction of joint inflammation in vivo was evaluated using a murine model of DC-induced arthritis. TLR7/8 involvement was assessed by specific inhibitors. RESULTS: Ex-miRNAs activate DCs to secrete TNFα, induce joint inflammation, start an early autoimmune response and potentiate the differentiation of DCs into aggressive osteoclasts. CONCLUSIONS: This work represents a proof of concept that the pool of extracellular miRNAs overexpressed in RA joints can act as a physiological activator of inflammation via the stimulation of TLR8 expressed by human DCs, which in turn exert arthritogenic functions. In this scenario, pharmacological inhibition of TLR8 might offer a new therapeutic option to reduce inflammation and osteoclast-mediated bone destruction in RA.

Peptidoglycan in osteoarthritis synovial tissue is associated with joint inflammation.

OBJECTIVES: Peptidoglycan (PG) is an arthritogenic bacterial cell wall component whose role in human osteoarthritis is poorly understood. The purpose of this study was to determine if PG is present in synovial tissue of osteoarthritis patients at the time of primary total knee arthroplasty (TKA), and if its presence is associated with inflammation and patient reported outcomes. METHODS: Intraoperative synovial tissue and synovial fluid samples were obtained from 56 patients undergoing primary TKA, none of whom had history of infection. PG in synovial tissue was detected by immunohistochemistry (IHC) and immunofluorescence microscopy (IFM). Synovial tissue inflammation and fibrosis were assessed by histopathology and synovial fluid cytokine quantification. Primary human fibroblasts isolated from arthritis synovial tissue were stimulated with PG to determine inflammatory cytokine response. RESULTS: A total of 33/56 (59%) of primary TKA synovial tissue samples were positive for PG by IHC, and PG staining colocalized with markers of synovial macrophages and fibroblasts by IFM. Synovial tissue inflammation and elevated IL-6 in synovial fluid positively correlated with PG positivity. Primary human fibroblasts stimulated with PG secreted high levels of IL-6, consistent with ex vivo findings. Interestingly, we observed a significant inverse correlation between PG and age at time of TKA, indicating younger age at time of TKA was associated with higher PG levels. CONCLUSION: Peptidoglycan is commonly found in synovial tissue from patients undergoing TKA. Our data indicate that PG may play an important role in inflammatory synovitis, particularly in patients who undergo TKA at a relatively younger age.

Performance of adenosine deaminase in synovial fluid for the diagnosis of tuberculous arthritis: A systematic review and meta-analysis.

OBJECTIVES: Adenosine deaminase (ADA) activity has shown good performance in diagnosing pleural, peritoneal, and meningeal tuberculosis. This meta-analysis aimed to evaluate the performance of measuring ADA activity in synovial fluid for the early diagnosis of joint tuberculosis. METHODS: We searched published information in MEDLINE, Embase, Cochrane Library, Web of Science, and MedRxiv databases, as well as unpublished information in the American College of Rheumatology and European League Against Rheumatism for conference abstracts (2012-2021). We also scanned the reference lists of articles. Two reviewers independently applied the criteria for selection, assessed quality, and extracted data (PROSPERO number CRD42021284472). RESULTS: Seven independent studies (N=305 subjects) that compared ADA activity in synovial fluid with a composite reference diagnostic method for tuberculosis were included. Overall, the risk of bias was judged low. Studies were classified as high quality (n=3; 148 subjects) and low quality (n=4; 157 subjects). Pooled sensitivity and specificity of ADA activity was 94% (95% confidence interval [CI], 0.89-98; I2=23%) and 88% (95% CI, 83-92; I2=83%), respectively. The random-effects model for pooled diagnostic Odds ratio was 67.1 (95%CI, 20.3-222.2; I2=30%). The receiver operating characteristic curve area was 0.96 (95% CI, 0.92-0.99). Meta-regression did not identify the quality of the study, country of publication, or the type of assay as a source of heterogeneity. CONCLUSIONS: Measuring ADA activity in synovial fluid demonstrates good performance for the early diagnosis of joint tuberculosis.

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

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

Single-cell profiling identifies IL1Bhi macrophages associated with inflammation in PD-1 inhibitor-induced inflammatory arthritis.

Inflammatory arthritis (IA) is a common rheumatic adverse event following immune checkpoint inhibitors treatment. The clinical disparities between IA and rheumatoid arthritis (RA) imply disease heterogeneity and distinct mechanisms, which remain elusive. Here, we profile CD45+ cells from the peripheral blood or synovial fluid (SF) of patients with PD-1-induced IA (PD-1-IA) or RA using single-cell RNA sequencing. We report the predominant expansion of IL1Bhi myeloid cells with enhanced NLRP3 inflammasome activity, in both the SF and peripheral blood of PD-1-IA, but not RA. IL1Bhi macrophages in the SF of PD-1-IA shared similar inflammatory signatures and might originate from peripheral IL1Bhi monocytes. Exhausted CD8+ T cells (Texs) significantly accumulated in the SF of patients with PD-1-IA. IL1Bhi myeloid cells communicated with CD8+ Texs possibly via the CCR1-CCL5/CCL3 and CXCL10-CXCR3 axes. Collectively, these results demonstrate different cellular and molecular pathways in PD-1-IA and RA and highlight IL1Bhi macrophages as a possible therapeutic target in PD-1-IA.

The acute phase reactant orosomucoid-2 directly promotes rheumatoid inflammation.

Acute phase proteins involved in chronic inflammatory diseases have not been systematically analyzed. Here, global proteome profiling of serum and urine revealed that orosomucoid-2 (ORM2), an acute phase reactant, was differentially expressed in rheumatoid arthritis (RA) patients and showed the highest fold change. Therefore, we questioned the extent to which ORM2, which is produced mainly in the liver, actively participates in rheumatoid inflammation. Surprisingly, ORM2 expression was upregulated in the synovial fluids and synovial membranes of RA patients. The major cell types producing ORM2 were synovial macrophages and fibroblast-like synoviocytes (FLSs) from RA patients. Recombinant ORM2 robustly increased IL-6, TNF-α, CXCL8 (IL-8), and CCL2 production by RA macrophages and FLSs via the NF-κB and p38 MAPK pathways. Interestingly, glycophorin C, a membrane protein for determining erythrocyte shape, was the receptor for ORM2. Intra-articular injection of ORM2 increased the severity of arthritis in mice and accelerated the infiltration of macrophages into the affected joints. Moreover, circulating ORM2 levels correlated with RA activity and radiographic progression. In conclusion, the acute phase protein ORM2 can directly increase the production of proinflammatory mediators and promote chronic arthritis in mice, suggesting that ORM2 could be a new therapeutic target for RA.

Comparative study of 1H-NMR metabolomic profile of canine synovial fluid in patients affected by four progressive stages of spontaneous osteoarthritis.

The study aimed to assess the metabolomic profile of the synovial fluid (SF) of dogs affected by spontaneous osteoarthritis (OA) and compare any differences based on disease progression. Sixty client-owned dogs affected by spontaneous OA underwent clinical, radiographic, and cytologic evaluations to confirm the diagnosis. The affected joints were divided into four study groups based on the Kallgreen-Lawrence classification: OA1 (mild), OA2 (moderate), OA3 (severe), and OA4 (extremely severe/deforming). The osteoarthritic joint's SF was subjected to cytologic examination and 1H-NMR analysis. The metabolomic profiles of the study groups' SF samples were statistically compared using one-way ANOVA. Sixty osteoarthritic joints (45 stifles, 10 shoulders and 5 elbows) were included in the study. Fourteen, 28, and 18 joints were included in the OA1, OA2, and OA3 groups, respectively (0 joints in the OA4 group). Metabolomic analysis identified 48 metabolites, five of which were significantly different between study groups: Mannose and betaine were elevated in the OA1 group compared with the OA2 group, and the 2-hydroxyisobutyrate concentration decreased with OA progression; in contrast, isoleucine was less concentrated in mild vs. moderate OA, and lactate increased in severe OA. This study identified different 1H-NMR metabolomic profiles of canine SF in patients with progressive degrees of spontaneous OA, suggesting 1H-NMR metabolomic analysis as a potential alternative method for monitoring OA progression. In addition, the results suggest the therapeutic potentials of the metabolomic pathways that involve mannose, betaine, 2-hydroxyisobutyrate, isoleucine, and lactate.

Adipocytes in synovial fluid cytology: An approach for diagnosing synovial lipomatosis.

A 2-year-old neutered male bullmastiff dog was presented with chronic left hind limb lameness. Physical examination revealed left stifle effusion and medial buttress without cranial tibial thrust. Radiographs showed joint effusion and new bone formation at the patella apex. Magnetic resonance imaging showed increased synovial fluid, widening of the joint space, abnormal infrapatellar fat body and thinning of the cranial cruciate ligament. Synoviocentesis and cytologic evaluation of synovial fluid revealed marked mononuclear inflammation with abundant fatty tissue, suggesting synovial lipomatosis in conjunction with the imaging findings. The disease was confirmed histologically after sampling the lesion during arthrotomy. Synovial lipomatosis, characterized by extensive synovial adipose tissue proliferation of the synovial membrane, is a rare "tumor-like" disorder that usually affects the stifle. Although the etiology remains unclear, joint trauma, inflammation, instability, and lipid abnormalities have been proposed as causes. Inflammatory factors may promote synoviocyte and adipocyte hyperplasia that perpetuate the process. Surgical removal may be suggested to eliminate triggers and prevent future recurrences. The report provides the first cytological description of adipocytes in synovial fluid associated with the diagnosis of synovial lipomatosis in dogs. This case report underscores the potential effectiveness of cytologic analysis of synovial fluid smears, in combination with magnetic resonance imaging (MRI), for diagnosing this condition and reducing complications associated with arthrotomy for sampling purposes. Additionally, the case highlights that synovial lipomatosis should be considered as a potential differential diagnosis for synovial masses in dogs. Further cases are needed to validate these observations in veterinary medicine.

Anti-inflammatory effects of infliximab and methotrexate on peripheral blood and synovial fluid mononuclear cells: ex vivo study.

OBJECTIVE: To investigate the effects of methotrexate (MTX) and the tumour necrosis factor inhibitor infliximab (IFX) on immune cells derived from peripheral blood mononuclear cells (PBMCs) and synovial fluid mononuclear cells (SFMCs) of inflammatory arthritis patients. METHOD: Phytohaemagglutinin (PHA)-induced proliferation of healthy donors' PBMCs and synovial intermediate monocytes (CD14+CD16+ cells) in SFMCs derived from psoriatic arthritis (PsA) and rheumatoid arthritis (RA) patients was determined by flow cytometry following co-culture with IFX and MTX. PHA-induced interferon-γ (IFN-γ) production in PBMCs was measured by enzyme-linked immunosorbent assay. The drugs' effect on mRNA expression in SFMCs was determined by quantitative polymerase chain reaction. RESULTS: The combination of IFX 10 µg/mL + MTX 0.1 µg/mL had the strongest inhibitory effect on PBMC proliferation (91%), followed by MTX 0.1 µg/mL (86%) and IFX 10 µg/mL (49%). In PHA-stimulated PBMCs, IFN-γ production was reduced by IFX 10 µg/mL, MTX 0.1 µg/mL, and IFX 10 µg/mL + MTX 0.1 µg/mL by 68%, 90%, and 85%, respectively. In SFMCs, IFX 10 µg/mL significantly reduced CD14+CD16+ cells compared to medium (PsA 54%, p < 0.01; RA 46%, p < 0.05), while MTX had no effect on this population. IFX + MTX led to a similar suppression of CD14+CD16+ cells as achieved by IFX alone. The drugs had different impacts on SFMC gene expression. CONCLUSION: Both IFX and MTX effectively inhibited PBMC proliferation and IFN-γ production, but only IFX reduced synovial monocytes and pro-inflammatory gene expression in SFMCs, suggesting a differential impact of IFX and MTX on critical inflammatory cell populations ex vivo.

Patients with higher numbers of allergies have lower levels of anti-inflammatory cytokines and increased pain at the time of ACL reconstruction.

BACKGROUND: Synovial fluid biomarkers are well studied indicators of inflammation and healing in the setting of orthopedic injuries. However, it has not been studied if patients with one or more allergies have a difference in the concentrations of synovial fluid inflammatory cytokines compared to patients without allergies. The purpose of the current study is to analyze the concentration of 10 pro- and anti-inflammatory cytokines in the synovial fluid of isolated ACL injury patients with and without at least one allergy. STUDY DESIGN: Retrospective Case-Control. METHODS: A database of patients who underwent surgery for isolated ACL injury between September 2011 and July 2023 was analyzed. All patients had SF aspirated from the operative knee prior to the surgical incision and the concentrations of pre- and anti-inflammatory biomarkers were quantified. From this cohort, 24 patients were identified to have allergies by chart review. These patients were matched 1:1 to 24 patients without allergies based on age and sex. RESULTS: There were no significant differences between the allergy and no allergy cohorts with respect to age (28.5 ± 10.3 vs. 29.5 ± 8.9, p = 0.76) and sex (70.8 % female vs. 70.8 % female, p = 1.00). The allergy cohort had a decreased concentration of TIMP-1 (492.41 ± 616.20 ng/mL vs. 1041.48 ± 942.04 ng/mL, p = 0.03) and IL-1Ra (101.70 ± 93.37 pg/mL vs. 359.94 ± 399.77 pg/mL, p = 0.01) compared to patients without allergies. A linear regression analysis found a significant association between increasing number of patients reported allergies and decreasing concentration of TIMP-1 (ß = -231.59, p = 0.03) and IL-1Ra (ß = -71.69p = 0.03) concentrations when controlling for age and sex. Finally, the allergy cohort was found to have a significantly higher value for the VAS pain scale at the time of surgery (26.84 ± 24.73 vs. 7.37 ± 10.98, p < 0.01) compared to those without an allergy. CONCLUSION: Patients undergoing ACL reconstruction with at least one allergy were found to have decreased concentrations of the anti-inflammatory cytokines TIMP-1 and IL-1Ra in their synovial fluid compared to those without allergies on the day of surgery. Furthermore, an increase in total number of allergies was found to be an associated with a decrease in TIMP-1 and IL-1Ra levels. Finally, the allergy cohort also had a higher value for the VAS pain scale at the time of surgery, implicating the role of a patient's innate immune system to their biologic and symptomatic response to injury.