Synovial fibroblast gene expression is associated with sensory nerve growth and pain in rheumatoid arthritis.

It has been presumed that rheumatoid arthritis (RA) joint pain is related to inflammation in the synovium; however, recent studies reveal that pain scores in patients do not correlate with synovial inflammation. We developed a machine-learning approach (graph-based gene expression module identification or GbGMI) to identify an 815-gene expression module associated with pain in synovial biopsy samples from patients with established RA who had limited synovial inflammation at arthroplasty. We then validated this finding in an independent cohort of synovial biopsy samples from patients who had early untreated RA with little inflammation. Single-cell RNA sequencing analyses indicated that most of these 815 genes were most robustly expressed by lining layer synovial fibroblasts. Receptor-ligand interaction analysis predicted cross-talk between human lining layer fibroblasts and human dorsal root ganglion neurons expressing calcitonin gene-related peptide (CGRP+). Both RA synovial fibroblast culture supernatant and netrin-4, which is abundantly expressed by lining fibroblasts and was within the GbGMI-identified pain-associated gene module, increased the branching of pain-sensitive murine CGRP+ dorsal root ganglion neurons in vitro. Imaging of solvent-cleared synovial tissue with little inflammation from humans with RA revealed CGRP+ pain-sensing neurons encasing blood vessels growing into synovial hypertrophic papilla. Together, these findings support a model whereby synovial lining fibroblasts express genes associated with pain that enhance the growth of pain-sensing neurons into regions of synovial hypertrophy in RA.

A case of synovial chondromatosis of the knee with 87 free bodies and review of literature.

BACKGROUND: Synovial chondromatosis is a non-malignant synovial disorder characterized by the presence of cartilage formation within the synovial membrane, leading to the emergence of multiple cartilaginous nodules that may be either attached or unattached. The presence of this anatomical feature is frequently observed in articulations such as the knee, hip, elbow, and ankle. CASE REPORT: In this study, we present a case of synovial chondromatosis in the knee joint of a healthy male in his early 60s. Notably, the patient exhibited the simultaneous presence of 87 large loose bodies. The occurrence of a substantial quantity of unattached entities of notable dimensions within the joint is highly uncommon. CONCLUSIONS: The patient had several synovial chondromas, a rare disease. Synovial chondromatosis is a benign disorder; however, growing synovium can cause pyogenic cartilage nodules. Most loose bodies in joints can abrade and degenerate articular cartilage, causing long-term discomfort. Thus, an early-stage procedure to remove loose bodies and carefully excise synovial tissue is necessary to treat this condition.

Synovium microenvironment-responsive injectable hydrogel inducing modulation of macrophages and elimination of synovial fibroblasts for enhanced treatment of rheumatoid arthritis.

Rheumatoid arthritis (RA) is a progressive autoimmune disease accompanied by joint swelling, cartilage erosion and bone damage. Drug therapy for RA has been restricted due to poor therapeutic effect, recurrence and adverse effects. Macrophages and synovial fibroblasts both play important roles in the pathology of RA. Macrophages secrete large amount of pro-inflammatory cytokines, while synovial fibroblasts are tightly correlated with hypoxia synovium microenvironment, cytokine release, recruitment of pro-inflammatory cells, bone and cartilage erosion. Therefore, in this timely research, an injectable and pH-sensitive peptide hydrogel loading methotrexate (MTX) and bismuthene nanosheet/polyethyleneimine (BiNS/PEI) has been developed to reduce the activity of macrophages and eliminate over-proliferated synovial fibroblasts simultaneously. MTX can reduce the cytokine secretion of macrophages/anti-apoptosis property of synovial fibroblasts and BiNS/PEI can eliminate synovial fibroblasts via photodynamic therapy (PDT) and photothermal therapy (PTT) routes. The hydrogel was injected into the acidic inflammatory synovium for precise targeting and served as a drug reservoir for pH responsive and sustained drug release, while improving the bioavailability and reducing the toxicity of MTX. Excellent therapeutic efficacy has been achieved in both in vivo and in vitro studies, and this unique drug delivery system provides a new and robust strategy to eliminate synovial fibroblasts and modulate immune system for RA treatment in clinical.

Macrophage-derived mir-100-5p orchestrates synovial proliferation and inflammation in rheumatoid arthritis through mTOR signaling.

BACKGROUND: Rheumatoid arthritis (RA) is a chronic autoimmune disorder characterized by synovial inflammation, causing substantial disability and reducing life quality. While macrophages are widely appreciated as a master regulator in the inflammatory response of RA, the precise mechanisms underlying the regulation of proliferation and inflammation in RA-derived fibroblast-like synoviocytes (RA-FLS) remain elusive. Here, we provide extensive evidence to demonstrate that macrophage contributes to RA microenvironment remodeling by extracellular vesicles (sEVs) and downstream miR-100-5p/ mammalian target of rapamycin (mTOR) axis. RESULTS: We showed that bone marrow derived macrophage (BMDM) derived-sEVs (BMDM-sEVs) from collagen-induced arthritis (CIA) mice (cBMDM-sEVs) exhibited a notable increase in abundance compared with BMDM-sEVs from normal mice (nBMDM-sEVs). cBMDM-sEVs induced significant RA-FLS proliferation and potent inflammatory responses. Mechanistically, decreased levels of miR-100-5p were detected in cBMDM-sEVs compared with nBMDM-sEVs. miR-100-5p overexpression ameliorated RA-FLS proliferation and inflammation by targeting the mTOR pathway. Partial attenuation of the inflammatory effects induced by cBMDM-sEVs on RA-FLS was achieved through the introduction of an overexpression of miR-100-5p. CONCLUSIONS: Our work reveals the critical role of macrophages in exacerbating RA by facilitating the transfer of miR-100-5p-deficient sEVs to RA-FLS, and sheds light on novel disease mechanisms and provides potential therapeutic targets for RA interventions.

Fibroblast expression of neurotransmitter receptor HTR2A associates with inflammation in rheumatoid arthritis joint.

The study of neuroimmune crosstalk and the involvement of neurotransmitters in inflammation and bone health has illustrated their significance in joint-related conditions. One important mode of cell-to-cell communication in the synovial fluid (SF) is through extracellular vesicles (EVs) carrying microRNAs (miRNAs). The role of neurotransmitter receptors in the pathogenesis of inflammatory joint diseases, and whether there are specific miRNAs regulating differentially expressed HTR2A, contributing to the inflammatory processes and bone metabolism is unclear. Expression of neurotransmitter receptors and their correlated inflammatory molecules were identified in rheumatoid arthritis (RA) and osteoarthritis (OA) synovium from a scRNA-seq dataset. Immunohistochemistry staining of synovial tissue (ST) from RA and OA patients was performed for validation. Expression of miRNAs targeting HTR2A carried by SF EVs was screened in low- and high-grade inflammation RA from a public dataset and validated by qPCR. HTR2A reduction by target miRNAs was verified by miRNAs mimics transfection into RA fibroblasts. HTR2A was found to be highly expressed in fibroblasts derived from RA synovial tissue. Its expression showed a positive correlation with the degree of inflammation observed. 5 miRNAs targeting HTR2A were decreased in RA SF EVs compared to OA, three of which, miR-214-3p, miR-3120-5p and miR-615-3p, mainly derived from monocytes in the SF, were validated as regulators of HTR2A expression. The findings suggest that fibroblast HTR2A may play a contributory role in inflammation and the pathogenesis of RA. Additionally, targeting miRNAs that act upon HTR2A could present novel therapeutic strategies for alleviating inflammation 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.

Targeting STAT6-mediated synovial macrophage activation improves pain in experimental knee osteoarthritis.

BACKGROUND: Pain from osteoarthritis (OA) is one of the top causes of disability worldwide, but effective treatment is lacking. Nociceptive factors are released by activated synovial macrophages in OA, but depletion of synovial macrophages paradoxically worsens inflammation and tissue damage in previous studies. Rather than depleting macrophages, we hypothesized that inhibiting macrophage activation may improve pain without increasing tissue damage. We aimed to identify key mechanisms mediating synovial macrophage activation and test the role of STAT signaling in macrophages on pain outcomes in experimental knee OA. METHODS: We induced experimental knee OA in rats via knee destabilization surgery, and performed RNA sequencing analysis on sorted synovial tissue macrophages to identify macrophage activation mechanisms. Liposomes laden with STAT1 or STAT6 inhibitors, vehicle (control), or clodronate (depletion control) were delivered selectively to synovial macrophages via serial intra-articular injections up to 12 weeks after OA induction. Treatment effects on knee and hindpaw mechanical pain sensitivity were measured during OA development, along with synovitis, cartilage damage, and synovial macrophage infiltration using histopathology and immunofluorescence. Lastly, crosstalk between drug-treated synovial tissue and articular chondrocytes was assessed in co-culture. RESULTS: The majority of pathways identified by transcriptomic analyses in OA synovial macrophages involve STAT signaling. As expected, macrophage depletion reduced pain, but increased synovial tissue fibrosis and vascularization. In contrast, STAT6 inhibition in macrophages led to marked, sustained improvements in mechanical pain sensitivity and synovial inflammation without worsening synovial or cartilage pathology. During co-culture, STAT6 inhibitor-treated synovial tissue had minimal effects on healthy chondrocyte gene expression, whereas STAT1 inhibitor-treated synovium induced changes in numerous cartilage turnover-related genes. CONCLUSION: These results suggest that STAT signaling is a major mediator of synovial macrophage activation in experimental knee OA. STAT6 may be a key mechanism mediating the release of nociceptive factors from macrophages and the development of mechanical pain sensitivity. Whereas therapeutic depletion of macrophages paradoxically increases inflammation and fibrosis, blocking STAT6-mediated synovial macrophage activation may be a novel strategy for OA-pain management without accelerating tissue damage.

The FTO-CMPK2 Pathway in Fibroblast-like Synoviocytes Modulates Rheumatoid Arthritis Synovial Inflammation and Cartilage Homeostasis via mtDNA Regulation.

In rheumatoid arthritis (RA), a debilitating autoimmune disorder marked by chronic synovial inflammation and progressive cartilage degradation, fibroblast-like synoviocytes (FLS) are key pathogenic players. Current treatments targeting these cells are limited. Our study focused on the Fat Mass and Obesity-associated protein (FTO), known for its roles in cell proliferation and inflammatory response modulation, and its involvement in RA. We specifically examined the inflammatory regulatory roles of FTO and CMPK2, a mitochondrial DNA synthesis protein, in FLS. Utilizing a combination of in vitro and in vivo methods, including FTO inhibition and gene knockdown, we aimed to understand FTO's influence on RA progression and chondrocyte functionality. Our findings showed that increased FTO expression in RA synovial cells enhanced their proliferation and migration and decreased senescence and apoptosis. Inhibiting FTO significantly slowed the disease progression in our models. Our research also highlighted that the FTO-CMPK2 pathway plays a crucial role in regulating synovial inflammation through the mtDNA-mediated cGAS/STING pathway, affecting chondrocyte homeostasis. This study indicates that targeting the FTO-CMPK2 axis could be a promising new therapeutic strategy for managing RA.

Isorhamnetin Downregulates MMP2 and MMP9 to Inhibit Development of Rheumatoid Arthritis through SRC/ERK/CREB Pathway.

OBJECTIVE: To investigate the effect of isorhamnetin on the pathology of rheumatoid arthritis (RA). METHODS: Tumor necrosis factor (TNF)- α -induced fibroblast-like synoviocytes (FLS) was exposed to additional isorhamnetin (10, 20 and 40 µ mol/L). Overexpression vectors for matrix metalloproteinase-2 (MMP2) or MMP9 or SRC were transfected to explore their roles in isorhamnetin-mediated RA-FLS function. RA-FLS viability, migration, and invasion were evaluated. Moreover, a collagen-induced arthritis (CIA) rat model was established. Rats were randomly divided to sham, CIA, low-, medium-, and high-dosage groups using a random number table (n=5 in each group) and administed with normal saline or additional isorhamnetin [2, 10, and 20 mg/(kg·day)] for 4 weeks, respectively. Arthritis index was calculated and synovial tissue inflammation was determined in CIA rats. The levels of MMP2, MMP9, TNF-α, interleukin-6 (IL-6), and IL-1 ß, as well as the phosphorylation levels of SRC, extracellular regulated kinase (ERK), and cyclic adenosine monophosphate response element-binding (CREB), were detected in RA-FLS and synovial tissue. Molecular docking was also used to analyze the binding of isorhamnetin to SRC. RESULTS: In in vitro studies, isorhamnetin inhibited RA-FLS viability, migration and invasion (P<0.05). Isorhamnetin downregulated the levels of MMP2, MMP9, TNF-α, IL-6, and IL-1 ß in RA-FLS (P<0.05). The overexpression of either MMP2 or MMP9 reversed isorhamnetin-inhibited RA-FLS migration and invasion, as well as the levels of TNF-α, IL-6, and IL-1 ß (P<0.05). Furthermore, isorhamnetin bound to SRC and reduced the phosphorylation of SRC, ERK, and CREB (P<0.05). SRC overexpression reversed the inhibitory effect of isorhamnetin on RA-FLS viability, migration and invasion, as well as the negative regulation of MMP2 and MMP9 (P<0.05). In in vivo studies, isorhamnetin decreased arthritis index scores (P<0.05) and alleviated synovial inflammation. Isorhamnetin reduced the levels of MMP2, MMP9, TNF-α, IL-6, and IL-1 ß, as well as the phosphorylation of SRC, ERK, and CREB in synovial tissue (P<0.05). Notably, the inhibitory effect of isorhamnetin was more pronounced at higher concentrations (P<0.05). CONCLUSION: Isorhamnetin exhibited anti-RA effects through modulating SRC/ERK/CREB and MMP2/MMP9 signaling pathways, suggesting that isorhamnetin may be a potential therapeutic agent for RA.

Periplogenin inhibits pathologic synovial proliferation and infiltration in rheumatoid arthritis by regulating the JAK2/3-STAT3 pathway.

Rheumatoid arthritis (RA) is an autoimmune disease that affects joints, causing inflammation, synovitis, and erosion of cartilage and bone. Periplogenin is an active ingredient in the anti-rheumatic and anti-inflammatory herb, cortex periplocae. We conducted a study using a CIA model and an in vitro model of fibroblast-like synoviocytes (FLS) induced by Tumor Necrosis Factor-alpha (TNF-α) stimulation. We evaluated cell activity, proliferation, and migration using the CCK8 test, EDU kit, and transwell assays, as well as network pharmacokinetic analysis of periplogenin targets and RA-related effects. Furthermore, we measured inflammatory factors and matrix metalloproteinases (MMPs) expression using ELISA and qRT-PCR assays. We also evaluated joint destruction using HE and Safranin O-Fast Green Staining and examined the changes in the JAK2/3-STAT3 pathway using western blot. The results indicated that periplogenin can effectively inhibit the secretion of inflammatory factors, suppress the JAK2/3-STAT3 pathway, and impede the proliferation and migration of RA FLS. Thus, periplogenin alleviated the Synovial inflammatory infiltration of RA.

Coptisine inhibits aggressive and proliferative actions of fibroblast like synoviocytes and exerts a therapeutic potential for rheumatoid arthritis.

OBJECTIVE: Coptisine, a natural bioactive small molecular compound extracted from traditional Chinese herb Coptis chinensis, has been shown to exhibit anti-tumor effect. However, its contribution to autoimmune diseases such as rheumatoid arthritis (RA) is unknown. Here, we evaluate the effect of coptisine in controlling fibroblast-like synoviocytes (FLS)-mediated synovial proliferation and aggression in RA and further explore its underlying mechanism(s). METHODS: FLS were separated from synovial tissues obtained from patients with RA. Protein expression was measured by Western blot or immunohistochemistry. Gene expression was detected by quantitative RT-PCR. The EdU incorporation was used to measure cell proliferation. Migration and invasion were determined by Boyden chamber assay. RNA sequencing analysis was used to seek for the target of coptisine. The in vivo effect of coptisine was evaluated in collagen-induced arthritis (CIA) model. RESULTS: Treatment with coptisine reduced the proliferation, migration, and invasion, but not apoptosis of RA FLS. Mechanistically, we identified PSAT1, an enzyme that catalyzes serine/one-carbon/glycine biosynthesis, as a novel targeting gene of coptisine in RA FLS. PSAT1 expression was increased in FLS and synovial tissues from patients with RA compared to healthy control subjects. Coptisine treatment or PSAT1 knockdown reduced the TNF-α-induced phosphorylation of p38, ERK1/2, and JNK MAPK pathway. Interestingly, coptisine administration improved the severity of arthritis and reduced synovial PSAT1 expression in mice with CIA. CONCLUSIONS: Our data demonstrate that coptisine treatment suppresses aggressive and proliferative actions of RA FLS by targeting PSAT1 and sequential inhibition of phosphorylated p38, ERK1/2, and JNK MAPK pathway. Our findings suggest that coptisine might control FLS-mediated rheumatoid synovial proliferation and aggression, and be a novel potential agent for RA treatment.

Reduction of Edema and Pain in Transcutaneous Electrical Nerve Stimulation Treated-Arthritic Rat.

Transcutaneous electrical nerve stimulation (TENS) has been used to reduce pain or improve motor function in musculoskeletal and neurological disorders in the clinic. Although some studies have suggested electrotherapy as an intervention for edema, the effects and mechanisms of TENS on inflammation-induced edema remain unclear. Thus, we aimed to investigate the effects of TENS on arthritic pain with edema. 1% carrageenan was injected into the right tibiofemoral joint of 69 male Sprague-Dawley rats (200-250 g). After the development of arthritic pain, low-frequency (4-Hz, Low-TENS, n = 25) and high-frequency (100-Hz, High-TENS, n = 25) TENS with sub-motor threshold or placebo-TENS (n = 19) was applied for 20-min to medio-lateral part of the ipsilateral side. Weight bearing and knee-bend tests were used to assess pain-like behaviors. Also, we examined the size of edema and measured tumor necrosis factor-alpha (TNF-α) and interleukin-1 beta (IL-1ß) levels in the synovium by western blot. Eight rats in each of the two TENS groups were injected with Naloxone. Edema was reduced in the low- and high-frequency TENS groups at 6-h. TENS-treated rats showed reduced pain in the knee-bend test at 6-h. We observed decreased weight load shifts on the ipsilateral side in TENS groups. Naloxone reduced these effects. TNF-α and IL-1ß expression decreased in the synovial membrane at 6-h. These results suggest that low- and high-frequency TENS have acutely positive effects on inflammatory edema, with the management of arthritic pain and reduction in pro-inflammatory mediators. Therefore, Low-TENS and High-TENS may be useful in treating acute inflammatory pain and edema.

Emerging Roles of Macrophage Polarization in Osteoarthritis: Mechanisms and Therapeutic Strategies.

Osteoarthritis (OA) is the most common chronic degenerative joint disease in middle-aged and elderly people, characterized by joint pain and dysfunction. Macrophages are key players in OA pathology, and their activation state has been studied extensively. Various studies have suggested that macrophages might respond to stimuli in their microenvironment by changing their phenotypes to pro-inflammatory or anti-inflammatory phenotypes, which is called macrophage polarization. Macrophages accumulate and become polarized (M1 or M2) in many tissues, such as synovium, adipose tissue, bone marrow, and bone mesenchymal tissues in joints, while resident macrophages as well as other stromal cells, including fibroblasts, chondrocytes, and osteoblasts, form the joint and function as an integrated unit. In this study, we focus exclusively on synovial macrophages, adipose tissue macrophages, and osteoclasts, to investigate their roles in the development of OA. We review recent key findings related to macrophage polarization and OA, including pathogenesis, molecular pathways, and therapeutics. We summarize several signaling pathways in macrophage reprogramming related to OA, including NF-κB, MAPK, TGF-ß, JAK/STAT, PI3K/Akt/mTOR, and NLRP3. Of note, despite the increasing availability of treatments for osteoarthritis, like intra-articular injections, surgery, and cellular therapy, the demand for more effective clinical therapies has remained steady. Therefore, we also describe the current prospective therapeutic methods that deem macrophage polarization to be a therapeutic target, including physical stimulus, chemical compounds, and biological molecules, to enhance cartilage repair and alleviate the progression of OA.

EPCR deficiency ameliorates inflammatory arthritis in mice by suppressing the activation and migration of T cells and dendritic cells.

OBJECTIVES: Endothelial protein C receptor (EPCR) is highly expressed in synovial tissues of patients with RA, but the function of this receptor remains unknown in RA. This study investigated the effect of EPCR on the onset and development of inflammatory arthritis and its underlying mechanisms. METHODS: CIA was induced in EPCR gene knockout (KO) and matched wild-type (WT) mice. The onset and development of arthritis was monitored clinically and histologically. T cells, dendritic cells (DCs), EPCR and cytokines from EPCR KO and WT mice, RA patients and healthy controls (HCs) were detected by flow cytometry and ELISA. RESULTS: EPCR KO mice displayed >40% lower arthritis incidence and 50% less disease severity than WT mice. EPCR KO mice also had significantly fewer Th1/Th17 cells in synovial tissues with more DCs in circulation. Lymph nodes and synovial CD4 T cells from EPCR KO mice expressed fewer chemokine receptors CXCR3, CXCR5 and CCR6 than WT mice. In vitro, EPCR KO spleen cells contained fewer Th1 and more Th2 and Th17 cells than WT and, in concordance, blocking EPCR in WT cells stimulated Th2 and Th17 cells. DCs generated from EPCR KO bone marrow were less mature and produced less MMP-9. Circulating T cells from RA patients expressed higher levels of EPCR than HC cells; blocking EPCR stimulated Th2 and Treg cells in vitro. CONCLUSION: Deficiency of EPCR ameliorates arthritis in CIA via inhibition of the activation and migration of pathogenic Th cells and DCs. Targeting EPCR may constitute a novel strategy for future RA treatment.

Iron overload causes macrophages to produce a pro-inflammatory phenotype in the synovium of hemophiliac arthritis via the acetyl-p53 pathway.

AIM: Haemophiliac arthritis (HA) is caused by spontaneous intra-articular hemorrhage and repeated intra-articular hematomas, leading to iron overload, which, in turn, induces M1 macrophage polarisation and inflammatory cytokine secretion, resulting in synovitis. Here, we explored the mechanism by which iron overload in HA induces the polarisation of M1 macrophages, providing a new approach for the treatment of HA synovitis. METHODS: The synovium from the knee joints of normal amputees and patients with HA was collected. Pathological changes in the synovial tissues were analysed using hematoxylin and eosin staining. Iron tissue deposition was evaluated using the iron assay kit and Prussia Blue staining, while macrophage phenotype was determined using immunofluorescence. The levels of pro-inflammatory cytokines and p53 acetylation were determine using western blotting. An in vitro iron overload model was established by inducing THP-1 macrophages with ferric ammonium citrate, and the involvement of acetylated p53 in M1 macrophage polarisation was investigated. RESULTS: Compared to control samples, the iron content in the synovium of patients with HA was significantly increased. The protein levels of M1 macrophage markers, pro-inflammatory cytokines, and acetylated p53, were also significantly elevated in the synovial tissues of patients with HA. Similar results were observed in the in vitro iron overload model. Furthermore, the inhibition of p53 acetylation in vitro reversed these iron overload-induced effects. CONCLUSION: In patients with HA, iron overload induced synovial p53 acetylation, leading to macrophage polarisation toward the M1 phenotype and increased inflammatory cytokine secretion, resulting in synovitis. HIGHLIGHTS: Synovial iron overload is associated with changes in P53 acetylation in hemophiliac arthritis (HA). Acetylated p53, a known regulator of macrophage polarization, is highly expressed in HA synovium, suggesting a potential role in M1 polarization. HA synovial macrophages predominantly polarize into the pro-inflammatory M1 phenotype, secreting elevated levels of pro-inflammatory cytokines.

Profiling joint tissues at single-cell resolution: advances and insights.

Advances in the profiling of human joint tissues at single-cell resolution have provided unique insights into the organization and function of these tissues in health and disease. Data generated by various single-cell technologies, including single-cell RNA sequencing and cytometry by time-of-flight, have identified the distinct subpopulations that constitute these tissues. These timely studies have provided the building blocks for the construction of single-cell atlases of joint tissues including cartilage, bone and synovium, leading to the identification of developmental trajectories, deciphering of crosstalk between cells and discovery of rare populations such as stem and progenitor cells. In addition, these studies have revealed unique pathogenetic populations that are potential therapeutic targets. The use of these approaches in synovial tissues has helped to identify how distinct cell subpopulations can orchestrate disease initiation and progression and be responsible for distinct pathological outcomes. Additionally, repair of tissues such as cartilage and meniscus remains an unmet medical need, and single-cell methodologies can be invaluable in providing a blueprint for both effective tissue-engineering strategies and therapeutic interventions for chronic joint diseases such as osteoarthritis and rheumatoid arthritis.

Three-dimensional ultrasound to investigate synovitis in first carpometacarpal osteoarthritis: A feasibility study.

BACKGROUND: Synovitis is one of the defining characteristics of osteoarthritis (OA) in the carpometacarpal (CMC1) joint of the thumb. Quantitative characterization of synovial volume is important for furthering our understanding of CMC1 OA disease progression, treatment response, and monitoring strategies. In previous studies, three-dimensional ultrasound (3-D US) has demonstrated the feasibility of being a point-of-care system for monitoring knee OA. However, 3-D US has not been tested on the smaller joints of the hand, which presents unique physiological and imaging challenges. PURPOSE: To develop and validate a novel application of 3-D US to monitor soft-tissue characteristics of OA in a CMC1 OA patient population compared to the current gold standard, magnetic resonance imaging (MRI). METHODS: A motorized submerged transducer moving assembly was designed for this device specifically for imaging the joints of the hands and wrist. The device used a linear 3-D scanning approach, where a 14L5 2-D transducer was translated over the region of interest. Two imaging phantoms were used to test the linear and volumetric measurement accuracy of the 3-D US device. To evaluate the accuracy of the reconstructed 3-D US geometry, a multilayer monofilament string-grid phantom (10 mm square grid) was scanned. To validate the volumetric measurement capabilities of the system, a simulated synovial tissue phantom with an embedded synovial effusion was fabricated and imaged. Ten CMC1 OA patients were imaged by our 3-D US and a 3.0 T MRI system to compare synovial volumes. The synovial volumes were manually segmented by two raters on the 2D slices of the 3D US reconstruction and MR images, to assess the accuracy and precision of the device for determining synovial tissue volumes. The Standard Error of Measurement and Minimal Detectable Change was used to assess the precision and sensitivity of the volume measurements. Paired sample t-tests were used to assess statistical significance. Additionally, rater reliability was assessed using Intra-Class Correlation (ICC) coefficients. RESULTS: The largest percent difference observed between the known physical volume of synovial extrusion in the phantom and the volume measured by our 3D US was 1.1% (p-value = 0.03). The mean volume difference between the 3-D US and the gold standard MRI was 1.78% (p-value = 0.48). The 3-D US synovial tissue volume measurements had a Standard Error Measurement (SEm ) of 11.21 mm3 and a Minimal Detectible Change (MDC) of 31.06 mm3 , while the MRI synovial tissue volume measurements had an SEM of 16.82 mm3 and an MDC of 46.63 mm3 . Excellent inter- and intra-rater reliability (ICCs = 0.94-0.99) observed across all imaging modalities and raters. CONCLUSION: Our results indicate the feasibility of applying 3-D US technology to provide accurate and precise CMC1 synovial tissue volume measurements, similar to MRI volume measurements. Lower MDC and SEm values for 3-D US volume measurements indicate that it is a precise measurement tool to assess synovial volume and that it is sensitive to variation between volume segmentations. The application of this imaging technique to monitor OA pathogenesis and treatment response over time at the patient's bedside should be thoroughly investigated in future studies.

Knee Osteotomy Decreases Joint Inflammation Based on Synovial Histology and Synovial Fluid Analysis.

PURPOSE: To examine the biological changes in the joints of patients with knee osteoarthritis (OA) before and after around-knee osteotomy (AKO), focusing on synovial fluid (SF) and synovial pathological changes. METHODS: Patients who underwent AKO for medial compartment knee OA between 2019 and 2021 were examined. SF and synovium were obtained at the time of AKO and plate removal after bone union (mean, 16.8 months [range: 11-38 months] postoperatively). SF volume and interleukin (IL)-6 concentrations in SF were assayed using enzyme-linked immunosorbent assay. Synovitis was assessed histologically using a semiquantitative scoring system. Macrophage infiltration was assessed by immunohistochemistry using a semiquantitative score for F4/80 expression. The M1/M2 ratio was calculated using percentage of cells positive for CD80 and CD163. The expression of proinflammatory cytokines was assessed by the percentage of IL-1ß- and IL-6-positive cells. The number of vascular endothelial growth factor-positive luminal structures was counted to assess angiogenesis. The change in each parameter was compared before and after AKO using the Wilcoxon matched-pairs signed-rank test. RESULTS: Twenty-four knees of 21 patients were included. SF volume and IL-6 concentration significantly decreased postoperatively (12.6 ± 2.1 mL vs 4.2 ± 0.6 mL; P < .0001 and 50.5 ± 8.6 pg/mL vs 20.7 ± 3.8 pg/mL; P = .0001, respectively). A significant reduction in synovitis score (P = .0001), macrophage infiltration (P < .0003), M1/M2 ratio (P < .0007), angiogenesis (P < .0001), and the percentage of IL-1ß- and IL-6-positive cells in the intima (P < .008 and P < .002, respectively) was found after AKO. CONCLUSIONS: SF volume and IL-6 concentrations in the SF decreased and inflammatory synovium pathology improved after AKO. In addition to biomechanical changes, the biological environment of the joint can be improved after AKO. LEVEL OF EVIDENCE: Level IV, retrospective therapeutic case series.

B7-H3 promotes angiogenesis in rheumatoid arthritis.

OBJECTIVE: The primary pathological changes of rheumatoid arthritis (RA) include chronic synovial inflammation, bone destruction, and aggressive pannus formation on cartilage, in which angiogenesis plays a critical role. B7-H3, an important immune checkpoint molecule, represents a novel target in tumor therapy and plays a significant role in the pathogenesis of autoimmune diseases. However, its biological mechanism in RA remains unclear. METHODS: Hematoxylin-eosin (HE) staining and immunohistochemistry were used to explore the histological characteristics and expression of B7-H3, CD34, and vascular endothelial growth factor (VEGF) in patients with RA and collagen-induced arthritis (CIA) mice. ELISA was used to detect VEGF, soluble B7-H3, and disease markers in the peripheral blood of patients. A monoclonal anti-B7-H3 antibody was used to treat CIA mice by blocking B7-H3-mediated signaling. RESULTS: The ELISA and HE staining results showed a positive correlation between the expression of B7-H3 and the degree of joint cavity destruction and pannus formation. B7-H3 expression also correlated with increased expression of the vessel biomarkers CD34 and VEGF. Anti-B7-H3 effectively reduced pannus formation in CIA mice. CONCLUSION: B7-H3 modulates angiogenic activity in the joint synovium, demonstrating its therapeutic value in the context of RA.

BMP4 up-regulated by 630 nm LED irradiation is associated with the amelioration of rheumatoid arthritis.

Rheumatoid arthritis (RA) is caused by inflammatory response of joints with cartilage and damage of synovium and bone erosion. In our previous studies, it has showed that irradiation of 630 nm LED reduce inflammation of synovial fibroblasts and cartilage and bone destruction in RA. However, the key genes and mechanism in ameliorating RA by irradiation of 630 nm LED remains unknown. In this study, human fibroblast-like synoviocytes (FLS) cell line MH7A and primary human RA-FLSs were treated with TNF-α and 630 nm LED irradiation with the different energy density. The mRNA sequencing was performed to screen the differentially expressed genes (DEGs). In all datasets, 10 DEGs were identified through screening. The protein interaction network analysis showed that 8 out of the 10 DEGs interacted with each other including IL-6, CXCL2, CXCL3, MAF, PGF, IL-1RL1, RRAD and BMP4. This study focused on BMP4, which is identified as important morphogens in regulating the development and homeostasis. CCK-8 assay results showed that 630 nm LED irradiation did not affect the cell viability. The qPCR and ELISA results showed that TNF-α stimulation inhibited BMP4 mRNA and protein level and irradiation of 630 nm LED increased the BMP4 mRNA and protein level in MH7A cells. In CIA and transgenic hTNF-α mice models, H&E staining showed that irradiation of 630 nm LED decreased the histological scores assessed from inflammation and bone erosion, while BMP4 expression level was up-regulated after 630 nm LED irradiation. Pearson correlation analysis shown that BMP4 protein expression was negatively correlated with the histological score of CIA mice and transgenic hTNF-α mice. These results indicated that BMP4 increased by irradiation of 630 nm LED was associated with the amelioration of RA, which suggested that BMP4 may be a potential targeting gene for photobiomodulation.