Macrophages In Osteoarthritis Research Articles

The Multifaceted Protective Role of Nuclear Factor Erythroid 2-Related Factor 2 in Osteoarthritis: Regulation of Oxidative Stress and Inflammation.

Osteoarthritis (OA) is a chronic degenerative joint disease characterized by the degradation of joint cartilage, subchondral bone sclerosis, synovitis, and structural changes in the joint. Recent research has highlighted the role of various genes in the pathogenesis and progression of OA, with nuclear factor erythroid 2-related factor 2 (NRF2) emerging as a critical player. NRF2, a vital transcription factor, plays a key role in regulating the OA microenvironment and slowing the disease's progression. It modulates the expression of several antioxidant enzymes, such as Heme oxygenase-1 (HO-1) and NAD(P)H oxidoreductase 1 (NQO1), among others, which help reduce oxidative stress. Furthermore, NRF2 inhibits the nuclear factor kappa-B (NF-κB) signaling pathway, thereby decreasing inflammation, joint pain, and the breakdown of cartilage extracellular matrix, while also mitigating cell aging and death. This review discusses NRF2's impact on oxidative stress, inflammation, cell aging, and various cell death modes (such as apoptosis, necroptosis, and ferroptosis) in OA-affected chondrocytes. The role of NRF2 in OA macrophages, and synovial fibroblasts was also discussed. It also covers NRF2's role in preserving the cartilage extracellular matrix and alleviating joint pain. The purpose of this review is to provide a comprehensive understanding of NRF2's protective mechanisms in OA, highlighting its potential as a therapeutic target and underscoring its significance in the development of novel treatment strategies for OA.

IDN5706 Inhibits Synovial Inflammation via Inducing M2 Polarization of Synovial Macrophages in Osteoarthritis Rats

Introduction: IDN5706 is a tetrahydro derivative of hyperforin. In this study, we aimed to explore the effect of IDN5706 on synovial macrophages in osteoarthritis (OA) rats and the underlying mechanisms. Methods: OA rats were employed for the in vivo experiments, and RAW264.7 cells were employed for the in vitro experiments. Histopathological changes in synovium were examined using hematoxylin-eosin staining. Cell apoptosis in synovium was assessed by TUNEL staining. Macrophage polarization was determined by immunohistochemical analysis and flow cytometry. The mRNA expression and protein level of genes were detected by qRT-PCR and Western blot. The efferocytosis of macrophages was assessed by flow cytometry. Results: IDN5706 reversed the increased CD86-positive cells (M1 macrophages) and decreased CD206-positive cells (M2 macrophages), both in synovium and synovial fluid of OA rats. The in vitro experiments further confirmed the promotion effect of IDN5706 on M2 macrophages, accompanied by the elevated Arg-1 and reduced iNOS. Also, the upregulated p-mTOR in synovium and synovial fluid of OA rats were reversed by IDN5706, and the decreased M1 macrophages and increased M2 macrophages induced by IDN5706 were reversed by the mTOR activator. IDN5706 enhanced the efferocytosis of IL-4-treated RAW264.7 cells, and the animal experiments further revealed the involvement of efferocytosis in the improvement of OA by IDN5706. Conclusions: IDN5706 enhanced the efferocytosis of synovial macrophages by inducing M2 polarization via inhibiting p-mTOR, thus suppressing synovial inflammation and OA development, providing a theoretical basis for IDN5706 as a clinical drug for inflammatory diseases.

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

BackgroundPain 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.MethodsWe 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.ResultsThe 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.ConclusionThese 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.

Targeted knockdown of PGAM5 in synovial macrophages efficiently alleviates osteoarthritis

Osteoarthritis (OA) is a common degenerative disease worldwide and new therapeutics that target inflammation and the crosstalk between immunocytes and chondrocytes are being developed to prevent and treat OA. These attempts involve repolarizing pro-inflammatory M1 macrophages into the anti-inflammatory M2 phenotype in synovium. In this study, we found that phosphoglycerate mutase 5 (PGAM5) significantly increased in macrophages in OA synovium compared to controls based on histology of human samples and single-cell RNA sequencing results of mice models. To address the role of PGAM5 in macrophages in OA, we found conditional knockout of PGAM5 in macrophages greatly alleviated OA symptoms and promoted anabolic metabolism of chondrocytes in vitro and in vivo. Mechanistically, we found that PGAM5 enhanced M1 polarization via AKT-mTOR/p38/ERK pathways, whereas inhibited M2 polarization via STAT6-PPARγ pathway in murine bone marrow-derived macrophages. Furthermore, we found that PGAM5 directly dephosphorylated Dishevelled Segment Polarity Protein 2 (DVL2) which resulted in the inhibition of β-catenin and repolarization of M2 macrophages into M1 macrophages. Conditional knockout of both PGAM5 and β-catenin in macrophages significantly exacerbated osteoarthritis compared to PGAM5-deficient mice. Motivated by these findings, we successfully designed mannose modified fluoropolymers combined with siPGAM5 to inhibit PGAM5 specifically in synovial macrophages via intra-articular injection, which possessed desired targeting abilities of synovial macrophages and greatly attenuated murine osteoarthritis. Collectively, these findings defined a key role for PGAM5 in orchestrating macrophage polarization and provides insights into novel macrophage-targeted strategy for treating OA.

Research progress and hotspots on macrophages in osteoarthritis: A bibliometric analysis from 2009 to 2022.

Macrophages in the synovium, as immune cells, can be polarized into different phenotypes to play an anti-inflammatory role in the treatment of osteoarthritis. In this study, bibliometric methods were used to search the relevant literature to find valuable research directions for researchers and provide new targets for osteoarthritis prevention and early treatment. Studies about the application of macrophages in the treatment of osteoarthritis were searched through the Web of Science core database from 2009 to 2022. Microsoft Excel 2019, VOSviewer, CiteSpace, R software, and 2 online websites were used to analyze the research status and predict the future development of the trend in research on macrophages in osteoarthritis. The number of publications identified with the search strategy was 1304. China and the United States ranked first in the number of publications. Shanghai Jiao Tong University ranked first in the world with 37 papers. Osteoarthritis and Cartilage was the journal with the most publications, and "exosomes," "stem cells," "macrophage polarization," "regeneration," and "innate immunity" may remain the research hotspots and frontiers in the future. The findings from the global trend analysis indicate that research on macrophages in the treatment of osteoarthritis is gradually deepening, and the number of studies is increasing. Exosomes may become a research trend and hotspot in the future.

Expression pattern analysis of m6A regulators reveals IGF2BP3 as a key modulator in osteoarthritis synovial macrophages

BackgroundDisruption of N6 methyl adenosine (m6A) modulation hampers gene expression and cellular functions, leading to various illnesses. However, the role of m6A modification in osteoarthritis (OA) synovitis remains unclear. This study aimed to explore the expression patterns of m6A regulators in OA synovial cell clusters and identify key m6A regulators that mediate synovial macrophage phenotypes.MethodsThe expression patterns of m6A regulators in the OA synovium were illustrated by analyzing bulk RNA-seq data. Next, we built an OA LASSO-Cox regression prediction model to identify the core m6A regulators. Potential target genes of these m6A regulators were identified by analyzing data from the RM2target database. A molecular functional network based on core m6A regulators and their target genes was constructed using the STRING database. Single-cell RNA-seq data were collected to verify the effects of m6A regulators on synovial cell clusters. Conjoint analyses of bulk and single-cell RNA-seq data were performed to validate the correlation between m6A regulators, synovial clusters, and disease conditions. After IGF2BP3 was screened as a potential modulator in OA macrophages, the IGF2BP3 expression level was tested in OA synovium and macrophages, and its functions were further tested by overexpression and knockdown in vitro.ResultsOA synovium showed aberrant expression patterns of m6A regulators. Based on these regulators, we constructed a well-fitting OA prediction model comprising six factors (FTO, YTHDC1, METTL5, IGF2BP3, ZC3H13, and HNRNPC). The functional network indicated that these factors were closely associated with OA synovial phenotypic alterations. Among these regulators, the m6A reader IGF2BP3 was identified as a potential macrophage mediator. Finally, IGF2BP3 upregulation was verified in the OA synovium, which promoted macrophage M1 polarization and inflammation.ConclusionsOur findings revealed the functions of m6A regulators in OA synovium and highlighted the association between IGF2BP3 and enhanced M1 polarization and inflammation in OA macrophages, providing novel molecular targets for OA diagnosis and treatment.

Metformin Attenuates the Inflammatory Response via the Regulation of Synovial M1 Macrophage in Osteoarthritis

Osteoarthritis (OA), the most common chronic inflammatory joint disease, is characterized by progressive cartilage degeneration, subchondral bone sclerosis, synovitis, and osteophyte formation. Metformin, a hypoglycemic agent used in the treatment of type 2 diabetes, has been evidenced to have anti-inflammatory properties to treat OA. It hampers the M1 polarization of synovial sublining macrophages, which promotes synovitis and exacerbates OA, thus lessening cartilage loss. In this study, metformin prevented the pro-inflammatory cytokines secreted by M1 macrophages, suppressed the inflammatory response of chondrocytes cultured with conditional medium (CM) from M1 macrophages, and mitigated the migration of M1 macrophages induced by interleukin-1ß (IL-1ß)-treated chondrocytes in vitro. In the meantime, metformin reduced the invasion of M1 macrophages in synovial regions brought about by the destabilization of medial meniscus (DMM) surgery in mice, and alleviated cartilage degeneration. Mechanistically, metformin regulated PI3K/AKT and downstream pathways in M1 macrophages. Overall, we demonstrated the therapeutic potential of metformin targeting synovial M1 macrophages in OA.

Exosomes derived from umbilical cord mesenchymal stem cells protect cartilage and regulate the polarization of macrophages in osteoarthritis

BackgroundOsteoarthritis (OA) is one of the most common joint diseases and a major global public health concern. Mesenchymal stem cells (MSCs) have been widely used for the treatment of OA owing to their paracrine secretion of trophic factors, a phenomenon in which exosomes may play a major role. Here, we investigate the potential of exosomes from human umbilical cord-derived MSCs (hUC-MSCs-Exos) in alleviating OA.MethodsThe hUC-MSCs-Exos were harvested from hUC-MSC-conditioned medium using ultracentrifugation. Rats with surgically-induced OA were intra-articularly injected with hUC-MSCs-Exos. The effect of hUC-MSCs-Exos in repairing osteoarticular cartilage was assessed using hematoxylin and eosin (HE) staining, safranin-O and fast green staining and immunohistochemistry. The in vitro experiments were further carried out to verify the therapeutic effect. The effects of hUC-MSCs-Exos on the proliferation and migration of human chondrocytes were evaluated using the cell counting kit-8, EdU-555 cell proliferation kit, and transwell assays. Annexin V-FITC/PI staining were used to evaluate the effect of exosomes on chondrocyte apoptosis. An in vitro model of human articular chondrocytes treated with interleukin 1 beta (IL-1β) was used to evaluate the effects of exosomes, analyses involved using quantitative real-time polymerase chain reaction (qRT-PCR), immunofluorescence, and western blotting. The role of hUC-MSCs-Exos in macrophage polarization was examined in the monocyte cell line, Tohoku Hospital Pediatrics-1 (THP-1) by qRT-PCR and immunofluorescence.ResultsThe results showed that hUC-MSCs-Exos prevented severe damage to the knee articular cartilage in the rat OA model. We confirmed the high efficacy of hUC-MSCs-Exos in promoting chondrocyte proliferation and migration and inhibiting chondrocyte apoptosis. Additionally, hUC-MSCs-Exos could reverse IL-1β-induced injury of chondrocytes and regulate the polarization of macrophages in vitro.ConclusionsThere is potential for hUC-MSCs-Exos to be used as a treatment strategy for OA.

Nrf2-mediated anti-inflammatory polarization of macrophages as therapeutic targets for osteoarthritis.

Macrophages are the most abundant immune cells within the synovial joints, and also the main innate immune effector cells triggering the initial inflammatory responses in the pathological process of osteoarthritis (OA). The transition of synovial macrophages between pro-inflammatory and anti-inflammatory phenotypes can play a key role in building the intra-articular microenvironment. The pro-inflammatory cascade induced by TNF-α, IL-1β, and IL-6 is closely related to M1 macrophages, resulting in the production of pro-chondrolytic mediators. However, IL-10, IL1RA, CCL-18, IGF, and TGF are closely related to M2 macrophages, leading to the protection of cartilage and the promoted regeneration. The inhibition of NF-κB signaling pathway is central in OA treatment via controlling inflammatory responses in macrophages, while the nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway appears not to attract widespread attention in the field. Nrf2 is a transcription factor encoding a large number of antioxidant enzymes. The activation of Nrf2 can have antioxidant and anti-inflammatory effects, which can also have complex crosstalk with NF-κB signaling pathway. The activation of Nrf2 can inhibit the M1 polarization and promote the M2 polarization through potential signaling transductions including TGF-β/SMAD, TLR/NF-κB, and JAK/STAT signaling pathways, with the regulation or cooperation of Notch, NLRP3, PI3K/Akt, and MAPK signaling. And the expression of heme oxygenase-1 (HO-1) and the negative regulation of Nrf2 for NF-κB can be the main mechanisms for promotion. Furthermore, the candidates of OA treatment by activating Nrf2 to promote M2 phenotype macrophages in OA are also reviewed in this work, such as itaconate and fumarate derivatives, curcumin, quercetin, melatonin, mesenchymal stem cells, and low-intensity pulsed ultrasound.

Phenotype Diversity of Macrophages in Osteoarthritis: Implications for Development of Macrophage Modulating Therapies.

Chronic inflammation is implicated in numerous human pathologies. In particular, low-grade inflammation is currently recognized as an important mechanism of osteoarthritis (OA), at least in some patients. Among the signs of the inflammatory process are elevated macrophage numbers detected in the OA synovium compared to healthy controls. High macrophage counts also correlate with clinical symptoms of the disease. Macrophages are central players in the development of chronic inflammation, pain, cartilage destruction, and bone remodeling. However, macrophages are also involved in tissue repair and remodeling, including cartilage. Therefore, reduction of macrophage content in the joints correlates with deleterious effects in OA models. Macrophage population is heterogeneous and dynamic, with phenotype transitions being induced by a variety of stimuli. In order to effectively use the macrophage inflammatory circuit for treatment of OA, it is important to understand macrophage heterogeneity and interactions with surrounding cells and tissues in the joint. In this review, we discuss functional phenotypes of macrophages and specific targeting approaches relevant for OA treatment development.

Integrated comparison of the mRNAome in cartilage, synovium, and macrophages in osteoarthritis.

The precise molecular mechanisms associated with osteoarthritis (OA), the most common musculoskeletal disorder, are poorly understood. There are currently no effective treatments to prevent the initiation and progression of the disease. In recent years, the development of mRNAome has made it possible to identify new mechanisms and therapeutic targets. However, the differentially expressed genes screened by different microarrays are not completely the same. In order to avoid this shortcoming, we integrate the different genes from different tissues and data sets, and select the commonly expressed genes for further studies.

Research progress of cellular senescence in the pathogenesis of osteoarthritis

To review the pathological effects of cellular senescence in the occurrence and development of osteoarthritis (OA) and potential therapeutic targets. The role of chondrocyte senescence, synovial cell senescence, mesenchymal stem cells senescence in OA, and the biological mechanism and progress of chondrocyte senescence were summarized by consulting relevant domestic and abroad literature. The existing evidence has basically made clear that chondrocyte senescence, mesenchymal stem cells senescence, and cartilage repair abnormalities, and the occurrence and development of OA have a certain causal relationship, and the role of the senescence of synovial cells, especially synovial macrophages in OA is still unclear. Transcription factors and epigenetics are the main mechanisms that regulate the upstream pathways of cellular senescence. Signal communication between cells can promote the appearance of senescent phenotypes in healthy cells. Targeted elimination of senescent cells and promotion of mesenchymal stem cells rejuvenation can effectively delay the progress of OA. Cellular senescence is an important biological phenomenon and potential therapeutic target in the occurrence and development of OA. In-depth study of its biological mechanism is helpful to the early prevention and treatment of OA.

An Emerging Target in the Battle against Osteoarthritis: Macrophage Polarization.

Osteoarthritis (OA) is one of the most prevalent chronic joint diseases worldwide, which causes a series of problems, such as joint pain, muscle atrophy, and joint deformities. Benefiting from some advances in the clinical treatment of OA, the quality of life of OA patients has been improved. However, the clinical need for more effective treatments for OA is still very urgent. Increasing findings show that macrophages are a critical breakthrough in OA therapy. Stimulated by different factors, macrophages are differentiated into two phenotypes: the pro-inflammatory M1 type and anti-inflammatory M2 type. In this study, various therapeutic reagents for macrophage-dependent OA treatment are summarized, including physical stimuli, chemical compounds, and biological molecules. Subsequently, the mechanisms of action of various approaches to modulating macrophages are discussed, and the signaling pathways underlying these treatments are interpreted. The NF-κB signaling pathway plays a vital role in the occurrence and development of macrophage-mediated OA, as NF-κB signaling pathway agonists promote the occurrence of OA, whereas NF-κB inhibitors ameliorate OA. Besides, several signaling pathways are also involved in the process of OA, including the JNK, Akt, MAPK, STAT6, Wnt/β-catenin, and mTOR pathways. In summary, macrophage polarization is a critical node in regulating the inflammatory response of OA. Reagents targeting the polarization of macrophages can effectively inhibit inflammation in the joints, which finally relieves OA symptoms. Our work lays the foundation for the development of macrophage-targeted therapeutic molecules and helps to elucidate the role of macrophages in OA.

Analysis of distribution, phenotype andclinical correlation of CD163 positive macrophages in osteoarthritis

Purpose: The aim of this study was to analyze the distribution characteristics of CD163+ macrophages in peripheral blood and synovium of patients with osteoarthritis (OA),and investigate the correlation of CD163+ macrophages, intracellular TNF-a, IL-10 expression with the disease duration , inflammation level and body mass index (BMI). Methods: A total of 45 patients with OA and 5 patients with fracture tibial plateau were enrolled in this research. The percentages of CD163+ macrophages in synovium and peripheral blood and their intracellular levels of TNF-α and IL-10 were detected by flow cytometry. Meanwhile, immunofluorescence was used to observe the distribution of CD163+ macrophages in synovial tissue of patients with OA or fracture. Statistical comparisons were performed by independent sample t test. Results: ①There were significant differences in the percentage of CD163+ macrophages in synovial and peripheral blood between OA and fracture patients. (p<0.05,respectively). ②The expression of TNF-α in CD163+ macrophages of synovium and blood of OA patients with disease duration below 10 years was significantly higher than that of patients with OA disease duration above 10 years ( p<0.05,respectively ) .while the expression of IL-10 was significantly lower than that of patients with OA disease duration above 10 years ( p < 0.05,respectively). ③CRP was positively correlated with BMI in patients with OA disease duration below 10 years (p<0.05), while showed no correlation with BMI in patients with OA duration above 10 years (P>0.05). ④Immunofluorescence confocal microscopy showed a large number of CD163+ macrophages invaded around the synovial vessel of patients with OA. Conclusions: Our results suggest that there were significant differences in the distribution of CD163+ macrophages in synovium and peripheral blood of OA patients at different stages of disease, and they showed different macrophage subtypes. The correlation between inflammatory markers and BMI was also significant varied in different stages of disease.

Dual role of IL6 mediated by mesenchymal stromal cell signaling to joint macrophages in osteoarthritis

Purpose: Osteoarthritis (OA) is a joint disease affecting > 5 million Canadians and patients have limited palliative and joint-replacement surgical options. Stromal cell therapy is emerging as a compelling treatment for OA. Our group led the first-in-Canada Ph1/2 trial with bone marrow mesenchymal stromal cells (BM-MSCs) in OA patients successfully; and significant improvements in patient outcomes were observed. This was accompanied with reduction in pro-inflammatory monocytes/macrophages (MΦs) in the synovial fluid (SF), suggestive of a clinical MSC anti-inflammatory effect. Understanding mechanism of action of MSCs in OA will enable effective donor MSC screening, and generation of more efficient engineered MSCs. While IL6 is a known pro-inflammatory mediator in the joint, and anti-IL6 therapeutics have been effective in treatment of Rheumatoid Arthritis, blocking single pro-inflammatory cytokines has been less effective in OA. This leads us to believe that IL6 might have complex roles. Indeed, MSCs-induced polarization of macrophages to inflammation resolving subtypes via IL6 signaling. This effect is not well studied in an OA joint where high levels of IL6 may have pro-inflammatory effects. We hypothesize that IL-6, which is known as pro-inflammatory mediator in OA is mediating anti-inflammatory function of MSCs through signaling via membrane-bound, but not soluble receptors. Methods: We co-cultured peripheral blood-derived MΦs with OA BM-MSCs in the presence of early and late SF from OA patients; or in the presence of IL6 antibody. Polarization of MΦs were assessed by analysis of surface markers and IL10 secretion. Levels of IL6 soluble IL6 receptor (sIL6R) were measured by ELISA. Results: Preliminary quantification of BM- MSC from OA patients revealed high IL6 secretion is associated with positive patient-reported outcomes. IL6 antibody abrogated MSCs-induced anti-inflammatory polarization of MΦs as shown by reduction of CD163 and CD206, suggesting a key role for IL6 in MSC anti-inflammatory effects on MΦs. Paradoxically, IL6 levels were increased in SF from late stage OA, whereas levels of sIL6R were comparable in early and late OA-SF. Further, the effect of MSCs on anti-inflammatory polarization of MΦs was attenuated in the presence of late OA-SF compared to early OA-SF. This could be due to enhanced IL6 trans-signaling (mediated by sIL6R) in late-stage OA SF expected to occur at the expense of reduced IL6 classical pathway (mediated by membrane bound IL6 receptor, mIL6R). To discriminate between classical vs. trans-signalling, we are using soluble gp130, a non-signaling decoy receptor which complexes IL6-sIL6R. Our preliminary result showed that sgp130 treatment enhanced the expression of CD163 in MΦs exposed to pooled OA SF. The classical IL6 pathway will be studied specifically by silencing mIL6R in MΦs using siRNA Conclusions: MSCs exert their anti-inflammatory action in OA, at least partly through polarization of MΦs to anti-inflammatory phenotype mediated by IL6. The ratio of IL6 signaling through trans-vs. classical signaling pathways is an important arbiter of MSC-modulation of joint inflammation.

Dual role of IL6 mediated by mesenchymal stromal cell signaling to joint macrophages in osteoarthritis

Background & Aim Osteoarthritis (OA) is a joint disease affecting> 5 million Canadians and patients have limited palliative and joint-replacement surgical options. Stromal cell therapy is emerging as a compelling treatment for OA. Our first-in-Canada Ph1/2 trial with bone marrow mesenchymal stromal cells (BM-MSCs) in OA patients showed significant improvements in patient outcomes. Levels of pro-inflammatory monocytes/macrophages (MΦs) were reduced in the synovial fluid (SF), suggestive of a clinical MSC anti-inflammatory action. Understanding mechanism of action of MSCs in OA will enable effective donor MSC screening, and generation of more efficient engineered MSCs. While IL6 is a known pro-inflammatory mediator in the joint, blocking IL6 has not been so effective in OA suggesting complex roles for IL6. Indeed, MSCs induced homeostatic polarization of macrophages via IL6. This effect is not well studied in an OA joint where high levels of IL6 may have pro-inflammatory roles. We hypothesize that IL-6 mediates anti-inflammatory function of MSCs via IL6 membrane-bound receptor (mIL6R), but not soluble IL6 receptors (sIL6R). Methods, Results & Conclusion We co-cultured blood-derived MΦs with OA-MSCs in the presence of SF from OA patients; or in the presence of IL6 antibody. Polarization of MΦs were assessed by analysis of surface markers. Levels of IL10, IL6 and sIL6R were measured by ELISA. Preliminary analysis of IL6 from MSC from OA patients revealed high IL6 secretion is associated with positive patient-reported outcomes. IL6 antibody abrogated MSCs-induced homeostatic polarization of MΦs, suggesting a key role for IL6 in MSC anti-inflammatory effects. Paradoxically, IL6 levels were increased in SF from late stage OA, whereas sIL6R levels were comparable in early and late OA-SF. Further, the effect of MSCs on MΦs polarization was attenuated in the presence of late OA-SF compared to early OA-SF. This could be due to enhanced IL6 trans-signaling (via sIL6R) in late-OA SF expected to occur at the expense of reduced IL6 classical pathway (via mIL6R). To discriminate between classical vs. trans-signalling, we are using soluble gp130, a non-signaling decoy receptor which complexes IL6-sIL6R without affecting the classical pathway. In conclusion, MSCs exert their anti-inflammatory action in OA, at least partly via polarization of MΦs to homeostatic phenotype mediated by IL6. The ratio of IL6 signaling through trans-vs. classical pathways is an important arbiter of MSC-modulation of joint inflammation.

Understanding the dual role of IL6 signaling mediated by mesenchymal stromal cell signaling to joint macrophages in osteoarthritis

Understanding the dual role of IL6 signaling mediated by mesenchymal stromal cell signaling to joint macrophages in osteoarthritis

Inhomogeneity of immune cell composition in the synovial sublining: linear mixed modelling indicates differences in distribution and spatial decline of CD68+ macrophages in osteoarthritis and rheumatoid arthritis.

BackgroundInhomogeneity of immune cell distribution in the synovial sublining layer was analyzed in order to improve our mechanistic understanding of synovial inflammation and explore potential refinements for histological biomarkers in rheumatoid arthritis (RA) and osteoarthritis (OA).MethodsSynovial tissue of 20 patients (11 RA, 9 OA) was immunohistochemically stained for macrophages (CD68), synovial fibroblasts (CD55), T cells (CD3), plasma cells (CD38), endothelial cells (vWF) and mast cells (MCT). The synovial sublining layer was divided into predefined adjacent zones and fractions of the stained area (SA) were determined by digital image analysis for each cell marker.ResultsDistribution of CD68, CD55, CD38 and MCT staining of the sublining area was heterogeneous (Friedman ANOVA p < 0.05). The highest expression for all markers was observed in the upper layer close to the lining layer with a decrease in the middle and lower sublining. The SA of CD68, CD55 and CD38 was significantly higher in all layers of RA tissue compared to OA (p < 0.05), except the CD38 fraction of the lower sublining. Based on receiver operating characteristics analysis, CD68 SA of the total sublining resulted in the highest area under the curve (AUC 0.944, CI 95 % 0.844–1.0, p = 0.001) followed by CD68 in the upper and middle layer respectively (both AUC 0.933, CI 95 % 0.816–1.0, p = 0.001) in both RA and OA. Linear mixed modelling confirmed significant differences in the SA of sublining CD68 between OA and RA (p = 0.0042) with a higher concentration of CD68+ towards the lining layer and more rapid decline towards the periphery of the sublining in RA compared to OA (p = 0.0022).ConclusionsImmune cells are inhomogeneously distributed within the sublining layer. RA and OA tissue display differences in the number of CD68 macrophages and differences in CD68 decline within the synovial sublining.

Folate receptor positive macrophages in osteoarthritis and effects of triamcinolone

Folate receptor positive macrophages in osteoarthritis and effects of triamcinolone

Functional folate receptor beta-expressing macrophages in osteoarthritis synovium and their M1/M2 expression profiles

Objective: The distribution of folate receptor (FR)-β+ macrophages and their M1/M2 expression profiles were examined in osteoarthritis (OA) synovial tissues, and compared to those in rheumatoid arthritis (RA) synovial tissues and CD163+ macrophages in both OA and RA synovial tissues.Method: The phenotypes and fluorescein isothiocyanate (FITC)–folate uptake of FR-β+ synovial macrophages were analysed by flow cytometry. The distribution of FR-β+ macrophages in OA and RA synovial tissues was examined by immunofluorescent microscopy. Tumour necrosis factor (TNF)-α, inducible nitric oxide synthase (iNOS), interleukin (IL)-10, and transforming growth factor (TGF)-β expression in FR-β+ macrophages was detected by double-immunostaining in both OA and RA synovial tissues.Results: FR-β+ macrophages were predominantly present in the synovial lining layer in OA patients. The proportion of CD163–FR-β+ cells in synovial mononuclear cells (MNCs) was increased in OA compared to RA synovial tissues. FR-βhigh macrophages from OA synovial tissues represented the majority of folic acid-binding cells. Although FR-β+ or CD163+ macrophages in the synovial tissues of OA and RA patients expressed a mixed pattern of M1 and M2 macrophage markers, there were more M2 markers expressing synovial macrophages in OA than in RA patients.Conclusions: The distribution and M1/M2 expression profiles of FR-β+ synovial macrophages were different between OA and RA synovial tissues. Thus, the findings underscore that the M1/M2 paradigm using surface markers FR-β and CD163 is an oversimplification of macrophage subsets. Functional FR-β present on OA synovial macrophages provides a potential tool for the diagnosis and treatment of OA.