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© Author(s) (or their employer(s)) 2023. No commercial re-use. See rights and permissions. Published by BMJ., Background: Postgraduate rheumatology training programmes are already established at a national level in most European countries. However, previous work has highlighted a substantial level of heterogeneity in the organisation and, in part, content of programmes. Objective: To define competences and standards of knowledge, skills and professional behaviours required for the training of rheumatologists. Methods: A European Alliance of Associations for Rheumatology (EULAR) task force (TF) of 23 experts, including two members of the European Union of Medical Specialists (UEMS) section of rheumatology, was convened. The mapping phase consisted of the retrieval of key documents on specialty training in rheumatology and other related specialties across a broad set of international sources. The content of these documents was extracted and represented the foundation for the document draft that underwent several rounds of online discussion within the TF, and afterwards was also distributed to a broad group of stakeholders for collecting feedback. The list of generated competences was voted on during the TF meetings, while the level of agreement (LoA) with each statement was established by anonymous online voting. Results: A total of 132 international training curricula were retrieved and extracted. In addition to the TF members, 253 stakeholders commented and voted on the competences through an online anonymous survey. The TF developed (1) an overarching framework indicating the areas that should be addressed during training, (2) 7 domains defining broad areas that rheumatology trainees should master by the end of the training programme, (3) 8 core themes defining the nuances of each domain and (4) 28 competences that trainees should acquire to cover each of the areas outlined in the overarching framework. A high LoA was achieved for all competences. Conclusion: These points to consider for EULAR-UEMS standards for the training of European rheumatologists are now defined. Their dissemination and use can hopefully contribute to harmonising training across European countries., The project was funded by the European Alliance of Associations for Rheumatology (grant number EDU049).

Synovial fibroblasts and their role in juvenile idiopathic arthritis have received limited attention compared to other immune mediated disease such as rheumatoid arthritis. Furthermore, no review exists regarding synovial fibroblasts, their interaction with immune cells and their potential involvement in juvenile idiopathic arthritis pathogenesis. This scoping review set out to identify and compile the current knowledge of all peer-reviewed studies on synovial fibroblasts from patients with juvenile idiopathic arthritis. The aim was to map the current knowledge and to produce a tool to assist future studies. The entire MEDLINE, EMBASE and Web of Science databases were used to identify all published studies in English regarding synovial fibroblasts from patients with juvenile idiopathic arthritis. We identified 18 eligible studies out of a total of 1778 screened entries. The majority of studies identified synovial fibroblast subsets or functional characteristics that may be involved in disease pathogenesis. We identified mechanisms of cell-cell interaction with leukocytes, pro-inflammatory signaling and unfavorable connective tissue homeostasis that may contribute to cartilage damage or bony overgrowth. All included studies identified mechanisms potentially linking synovial fibroblasts to specific disease traits in juvenile idiopathic arthritis. Most findings were similar to mechanisms also described in synovial fibroblast from adults with arthritis. However, the limited number of studies found identifies an unmet need for additional studies on synovial fibroblasts and their potential role in juvenile idiopathic arthritis.

Fibroblast-like synoviocytes (FLS) play an important pathological role in persistent inflammatory joint diseases such as rheumatoid arthritis (RA). These cells have primarily been characterized in the RA synovial membrane. Here we aim to phenotypically and functionally characterize cultured synovial fluid-derived FLS (sfRA-FLS). Paired peripheral blood mononuclear cells (PBMC) and sfRA-FLS from patients with RA were obtained and monocultures of sfRA-FLS and autologous co-cultures of sfRA-FLS and PBMC were established. The in situ activated sfRA-FLS were CD34-, CD45-, Podoplanin+, Thymocyte differentiation antigen-1+. SfRA-FLS expressed uniform levels of NFкB-related pathway proteins and secreted several pro-inflammatory cytokines dominated by IL-6 and MCP-1. In a co-culture model with autologous PBMC, the ICAM-1 and HLA-DR expression on sfRA-FLS and secretion of IL-1β, IL-6, and MCP-1 increased. In vivo, human sfRA-FLS were cartilage invasive both at ipsilateral and contralateral implantation site. We conclude that, sfRA-FLS closely resemble the pathological sublining layer FLS subset in terms of surface protein expression, cytokine production and leukocyte cross-talk potential. Further, sfRA-FLS are comparable to tissue-derived FLS in their capabilities to invade cartilage at implantation sites but also spread tissue destruction to a distant site. Collectively, sfRA-FLS can serve as a an easy-to-obtain source of pathological sublining FLS in RA.

OBJECTIVE: Immune-mediated inflammatory arthritis (IMIA) is a heterogeneous group of diseases including rheumatoid arthritis (RA), psoriatic arthritis (PsA), and spondyloarthritis (SpA). Disease-modifying antirheumatic drugs (DMARDs) target very different cellular components of the disease processes. Characterization of the pathobiological subtypes of IMIA could provide more specific treatment approaches for each disease. For example, RA has been proposed to consist of at least three synovial pathotypes (lymphoid, myeloid, and fibroid), and only a subgroup of RA patients have erosive disease. The objective of this study was to evaluate the effects of various DMARDs on different synovial cell subsets using human ex vivo models of IMIA.METHODS: Synovial fluid and blood samples were obtained from a study population consisting of patients with RA, PsA, or peripheral SpA with at least one swollen joint (n = 18). The DMARDs used in this study were methotrexate, adalimumab, etanercept, tocilizumab, anakinra, ustekinumab, secukinumab, tofacitinib, and baricitinib. Paired synovial fluid mononuclear cells (SFMCs), peripheral blood mononuclear cells (PBMCs), and fibroblast-like synovial cells (FLSs) were used in three different previously optimized ex vivo models.RESULTS: In SFMCs cultured for 48 hours, all DMARDs except anakinra decreased the production of monocyte chemoattractant protein (MCP)-1. In SFMCs cultured for 21 days, only the two tumor necrosis factor alpha (TNFα) inhibitors adalimumab and etanercept decreased the secretion of tartrate-resistant acid phosphatase (P < 0.01, P < 0.001). In the FLS and PBMC 48-hour co-cultures, only tocilizumab (P < 0.001) and the two Janus kinase inhibitors tofacitinib and baricitinib (both P < 0.05) decreased the production of MCP-1 by around 50%.CONCLUSION: TNFα inhibition was effective in preventing inflammatory osteoclastogenesis, whereas tocilizumab, tofacitinib, and baricitinib had superior efficacy in cultures dominated by FLSs. Taken together, this study reveals that responses to cytokine inhibitors associate with cellular composition in models of IMIA. In particular, this study provides new evidence on the differential effect of DMARDs on leukocytes compared with stromal cells.

The US Food and Drug Administration (FDA) has recently added a new ‘black box warning’ on all currently approved Janus kinase (JAK) inhibitors indicated for the treatment of arthritis and other inflammatory conditions based on results from the ORAL Surveillance study of tofacitinib versus tumour necrosis factor alpha inhibitors in rheumatoid arthritis. This is a warning difficult to ignore because the data, being from a randomised controlled trial, are of high fidelity and hard to reproach. It is especially problematic because safety data for all the other JAK inhibitors will be pending for several years. So how might we proceed, without being bound by our stasis? The lack of absolute certainty seems to require a pragmatic approach to the routine care use of JAK inhibitors. The patients who were at greatest risk were older and had other risk factors for the corresponding adverse events, in keeping with effect modification. This highlights the need to focus on risk stratification when tailoring therapy. In this viewpoint, we propose a simple illustration to guide clinical decision-making. First, identify general risk factors for venous thromboembolic event (VTE), major adverse cardiac event (MACE) and cancer (age>65 years and smoking) and whether there is a previous history of VTE, MACE or cancer. Then, evaluate risk based on the number of other risk factors for VTE and the number of other risk factors for MACE. Ultimately, ‘treat-to-target’ will in the end always be ‘treat-to-agreement’. As we have done in the past, and will do in the future, the optimal treatment strategy will have to be tailored based on individual patient risk factors and preferences in a shared-decision process.

Systemic Intermittent Hypoxic Therapy (IHT) relies on the adaptive response to hypoxic stress. We investigated allogenic bone-graft resorption in the lumbar spine in 48 mice. The mice were exposed to IHT for 1 week before surgery or 1 week after surgery and compared with controls after 1 and 4 weeks. Complete graft resorption was observed in 33–36% of the animals in the control group, but none in the preoperative IHT group. Increased bone-graft volume was demonstrated by micro-computed tomography in the preoperative IHT group after 1 week (p = 0.03) while a non-significant difference was observed after 4 weeks (p = 0.12). There were no significant differences in the postoperative IHT group. Increased concentration of immune cells was localized in the graft area, and more positive tartrate-resistant acid phosphatase (TRAP) staining was found in controls compared with IHT allogenic bone grafts. Systemic IHT resulted in a significant increase of the major osteoclast inhibitor osteoprotegerin as well as osteogenic and angiogenic regulators Tgfbr3, Fst3l, Wisp1, and Vegfd. Inflammatory cytokines and receptor activator of nuclear factor kappa-B ligand (RANKL) stimulators IL-6, IL-17a, IL-17f, and IL-23r increased after 1 and 4 weeks, and serum RANKL expression remained constant while Ccl3 and Ccl5 decreased. We conclude that the adaptive response to IHT activates numerous pathways leading to inhibition of osteoclastic activity and inhibition of allogenic bone-graft resorption.

Introduction: Immune checkpoint inhibitor-induced inflammatory arthritis (ICI-IA) is a relatively new disease entity caused by ICI agents during cancer therapy. Reactive arthritis (ReA) is a well-known disease entity caused by urogenital or gastrointestinal bacterial infection or pneumonia. In this sense, ICI-IA and ReA are both defined by a reaction to a well-specified causal event. As a result, comparing these diseases may help to determine therapeutic strategies. Methods: We compared ICI-IA and ReA with special focus on pharmacological management. Specifically regarding treatment, we conducted a literature search of studies published in the PubMed database. Inclusion criteria were studies on treatment with non-steroidal anti-inflammatory drugs (NSAIDs), glucocorticoids (GC), or disease modifying antirheumatic drugs (DMARDs) in ICI-IA or ReA. During systematic selection, 21 studies evaluating ICI-IA and 14 studies evaluating ReA were included. Results: In ICI-IA, prospective and retrospective studies have shown effects of non-steroidal anti-inflammatory drugs (NSAIDs), glucocorticoid (GC), sulfasalazine (SSZ), methotrexate (MTX), hydroxychloroquine (HCQ) and TNFi. In ReA, retrospective studies evaluated NSAIDs and GC. A randomized controlled trial reported the effect of SSZ, and a retrospective study reported the effect of MTX and SSZ in combination with tumor necrosis factor alpha inhibition (TNFi). For both entities, small case reports show treatment effects of interleukin 6 receptor inhibition (IL-6Ri). Discussion: This literature review identified both similarities and differences regarding the pathogenesis and clinical features of ReA and ICI-IA. Studies on treatment reported effectiveness of NSAIDs, GC, MTX, SSZ and TNFi in both diseases. Further, small case reports showed effects of IL-6Ri.

Fibroblasts like synoviocytes (FLS) play several significant roles in rheumatoid arthritis (RA) pathophysiology. This chapter will describe known roles of FLS in disease initiation, joint inflammation, disease persistence and joint destruction. It will describe the newly characterized subsets of FLS based on single cell RNA sequencing studies, and their association to specific aspects of the disease. Finally, we will discuss the future of targeting FLS in the treatment of RA. The FLS in the synovial lining layer are identified by surface complement decay-accelerating factor (CD55) along with lubricin and metallopeptidase expression. Pathological activation of this lining layer subset result in bone and cartilage damage in mice. FLS of the sublining layer are often characterized by THY1 expression, but recent studies have highlighted a heterogeneity where several distinct subsets are identified by additional markers. Sublining FLS expressing human leukocyte antigen-DRA (HLA-DRA) produce C-X-C motif chemokine 12 (CXCL12) and receptor activator of nuclear factor-κB ligand (RANKL) and seems to constitute a pro-inflammatory subset that is associated with inflammation and tertiary lymphoid structures. Another subset of FLS characterized by CD34 expression may discriminate a common progenitor fibroblast subset. Taken together, studies isolating and characterizing gene expression in synovial FLS report both associations of unknown importance and markers that may impose protective or destructive features. This supports evidence of FLS as active players in RA pathology capable of cellular recruitment, local cellular crosstalk and promotion of joint destruction. These discoveries may serve as an atlas for synovial activation in RA and have identified several potential fibroblast markers for the development of targeted treatment.

This letter describes the potential effect of B cell depletion on immune related adverse events associated with immune checkpoint inhibition. B cell depleting agents such as rituximab reduce B cell to plasma cell differentiation and antibody production. This treatment strategy is used in several immune mediated inflammatory diseases such as rheumatoid arthritis and small vessel vasculitis. The immune related adverse events associated with immune checkpoint inhibition resemble immune mediated inflammatory diseases. Here, we report a lower incidence of hypothyroidism in a trial of combined B cell depletion and immune checkpoint inhibitor treatment compared with studies of immune checkpoint inhibitor monotherapy. This letter aims to increase awareness of the immune related adverse events associated with immune checkpoint inhibition in future clinical trials of immune checkpoint inhibition together with B cell depletion (primarily trials of B cell lymphomas). Hopefully, observations from these clinical trials can guide future treatment strategies to treat or prevent immune related adverse events associated with immune checkpoint inhibition.

Elisabeth Busk Rasmussen,1 Lotte Lindgreen Eriksen,2 Stinne Ravn Greisen,1 Anne Louise Hansen,1 Mikkel Carstensen,1 Thomas Damgaard Sandahl,2 Sidsel Støy,2 Tue Wenzel Kragstrup1,3 1Department of Biomedicine, Aarhus University, Aarhus, Denmark; 2Department of Hepatology and Gastroenterology, Aarhus University Hospital, Aarhus, Denmark; 3Diagnostic Center, Silkeborg Regional Hospital, Silkeborg, DenmarkCorrespondence: Tue Wenzel KragstrupDepartment of Biomedicine, Aarhus University, Wilhelm Meyers Allé 4, Aarhus C, DK-8000, DenmarkTel +4587167265Email kragstrup@biomed.au.dkObjective: Alcoholic hepatitis (AH) holds a high mortality, and vast macrophage infiltration of the liver is involved in the progressive liver injury. No efficient medical treatment exists, and macrophages may be a future treatment target. Here, we examine associations between non-classical monocyte subsets and cell surface markers of migration with disease activity in patients with severe AH.Methods: We analyzed samples from two cohorts of patients with AH. Cohort 1 included 15 AH patients, followed for 30 days, and 8 healthy controls (HCs). Cohort 2 included 23 AH patients, followed for 90 days, and 9 HCs. Peripheral blood mononuclear cells (PBMCs) from both cohorts were analyzed by flow cytometry. Liver biopsies from cohort 2 were analyzed by RNA sequencing.Results: Circulating non-classical monocytes in all but absent in patients with AH compared to HC in both cohorts (both p< 0.0001). The frequency of non-classical monocytes was significantly associated with Maddrey’s discriminant function (mDF) (r=− 0.79, p=0.0008, cohort 1), Child–Pugh score (CP) (r=− 0.56, p=0.03, cohort 1), Model for End-Stage Liver Disease (MELD) (r=− 0.54, p=0.02, cohort 2) and C-reactive protein (CRP) (r=− 0.51, p=0.027, cohort 2). The surface expression of CD11b was increased on non-classical monocytes in patients with AH compared to HC (p< 0.0001) (cohort 1). The mRNA expression of CD11b was increased in liver biopsies in patients with AH compared to HC (cohort 2) (p< 0.0001).Conclusion: In this study, we describe an almost complete depletion of circulating non-classical monocytes in the blood in two independent cohorts of patients with AH, which may be associated with a possible harmful recruitment of these cells to the liver. These results contribute to a better understanding of the disease, which hopefully can lead to therapies that target the acute inflammatory response leading to severe AH.Keywords: monocytes, non-classical, alcoholic hepatitis, CD11b, CCR2, CX3CR1

Aims Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) binds to angiotensin converting enzyme 2 (ACE2) enabling entrance of the virus into cells and causing the infection termed coronavirus disease of 2019 (COVID-19). Here, we investigate associations between plasma ACE2 and outcome of COVID-19. Methods and results This analysis used data from a large longitudinal study of 306 COVID-19 positive patients and 78 COVID-19 negative patients (MGH Emergency Department COVID-19 Cohort). Comprehensive clinical data were collected on this cohort, including 28-day outcomes. The samples were run on the Olink® Explore 1536 platform which includes measurement of the ACE2 protein. High admission plasma ACE2 in COVID-19 patients was associated with increased maximal illness severity within 28 days with OR = 1.8, 95%-CI: 1.4–2.3 (P < 0.0001). Plasma ACE2 was significantly higher in COVID-19 patients with hypertension compared with patients without hypertension (P = 0.0045). Circulating ACE2 was also significantly higher in COVID-19 patients with pre-existing heart conditions and kidney disease compared with patients without these pre-existing conditions (P = 0.0363 and P = 0.0303, respectively). Conclusion This study suggests that measuring plasma ACE2 is potentially valuable in predicting COVID-19 outcomes. Further, ACE2 could be a link between COVID-19 illness severity and its established risk factors hypertension, pre-existing heart disease and pre-existing kidney disease.

Background Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) binds to angiotensin converting enzyme 2 (ACE2) enabling entrance of the virus into cells and causing the infection termed coronavirus disease of 2019 (COVID-19). COVID-19 is a disease with a very broad spectrum of clinical manifestations, ranging from asymptomatic and subclinical infection to severe hyperinflammatory syndrome and death. Methods This study used data from a large longitudinal study of 306 COVID-19 positive patients and 78 COVID-19 negative patients (MGH Emergency Department COVID-19 Cohort with Olink Proteomics). Comprehensive clinical data were collected on this cohort, including 28-day outcomes classified according to the World Health Organization (WHO) COVID-19 outcomes scale. The samples were run on the Olink® Explore 1536 platform which includes measurement of the ACE2 protein. Findings High baseline levels of ACE2 in plasma from COVID-19 patients were associated with worse WHOmax category at 28 days with OR=0.56, 95%-CI: 0.44-0.71 (P < 0.0001). This association was significant in regression models with correction for baseline characteristics, pre-existing medical conditions, and laboratory test results. High levels of ACE2 in plasma from COVID-19 patients were also significantly associated with worse WHO category at the time of blood sampling at both day 0, day 3, and day 7 (P = 0.0004, P < 0.0001, and P < 0.0001, respectively). The levels of ACE2 in plasma from COVID-19 patients with hypertension were significantly higher compared to patients without hypertension (P = 0.0045). The plasma ACE2 levels were also significantly higher in COVID-19 patients with pre-existing heart conditions and kidney disease compared with patients without these pre-existing conditions (P = 0.0363 and P = 0.0303, respectively). There was no difference in plasma ACE2 levels comparing patients with or without pre-existing lung disease, diabetes, or immunosuppressive conditions (P = 0.953, P = 0.291, and P = 0.237, respectively). The associations between high plasma levels of ACE2 and worse WHOmax category during 28 days were more pronounced in COVID-19 positive patients compared with COVID-19 negative patients but the difference was not significant in the two-way ANOVA analysis. Interpretation This study suggests that measuring ACE2 is potentially valuable in predicting COVID-19 outcomes. Further, ACE2 levels could be a link between severe COVID-19 disease and its risk factors, namely hypertension, pre-existing heart disease and pre-existing kidney disease. The design of the data analysis using the Olink platform does not allow assessment of quantitative differences. However, previous studies have described a positive correlation between plasma ACE2 and ACE1 activity. This is interesting because ACE1 (serum ACE) analysis is a standardized test in most hospital laboratories. Therefore, our study encourages quantitative investigations of both plasma ACE 1 and 2 in COVID-19. Key Points Question Can plasma levels of the receptor for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), angiotensin converting enzyme 2 (ACE2), predict outcome of coronavirus disease of 2019 (COVID-19). Findings In this study of 306 COVID-19 positive patients, high baseline levels of ACE2 in plasma from COVID-19 patients were associated with worse outcome measured by the World Health Organization (WHO) COVID-19 outcomes scale. Meaning Measuring ACE2 is potentially valuable in predicting COVID-19 outcomes and link COVID-19 disease and the risk factors hypertension, pre-existing heart disease and pre-existing kidney disease.

The primary vasculitides constitute a heterogeneous group of immune mediated diseases of incompletely understood pathogenesis currently classified by the size of blood vessels affected (Chapel Hill classification). In recent years, several drugs with well-characterized immunological targets have been tested in clinical trials in large vessel vasculitis and small vessel vasculitis. Such trials provide "reverse translational" or bedside to bench information about underlying pathogenic mechanisms. Therefore, the aim of this systematic literature review was to examine the evidence base for a more refined mechanistic immunological classification of vasculitis. A total of 40 studies (20 randomized controlled trials (RCTs), 16 prospective studies, 1 retrospective cohort study and 3 case series) were included for full qualitative assessment. RCTs concerning biologic therapy for large vessel vasculitis mainly supports interleukin 6 receptor inhibition (tocilizumab). RCTs concerning biologic therapy for granulomatosis with polyangiitis and microscopic polyangiitis mainly support anti-CD20 treatment (rituximab) and complement inhibition with a small molecule C5a receptor antagonist (avacopan) is an emerging treatment option. The biologic treatment of eosinophilic granulomatosis with polyangiitis is centered around interleukin 5 inhibition (mepolizumab). Studies on tumor necrosis factor alpha inhibition (adalimumab, infliximab, and etanercept) showed negative results in giant cell arteritis but some effect in Takayasu arteritis. Taken together, clinical studies with cytokine and cell specific drugs are dissecting the heterogeneous immunopathogenic mechanisms of vasculitis and support a mechanistic immunological classification. Especially, cytokine antagonism is pointing towards immunological distinctions between eosinophilic granulomatosis with polyangiitis and granulomatosis with polyangiitis/microscopic polyangiitis and differences between giant cell arteritis and Takayasu arteritis.

Axial spondyloarthritis (axSpA), psoriatic arthritis (PsA), psoriasis, inflammatory bowel disease (IBD), and noninfectious uveitis form a distinct group among the immune mediated inflammatory diseases. Thus, many patients suffer from more than one of these disease manifestations. Here, we will use the term spondylitis-psoriasis-enthesitis-enterocolitis-dactylitis-uveitis-peripheral synovitis (SPEED-UP) spectrum disease. The aim is to review the new targeted pharmacological treatment options for all these diseases. All biological or targeted synthetic drugs with U.S. Food and Drug Administration (FDA) or European Medicines Agency (EMA) approval for any of the diagnoses axSpA, PsA, psoriasis, IBD, or non-infectious uveitis were included. Some of the drugs have documented efficacy in more than one of the diseases, e.g. tumor necrosis factor (TNF) inhibitors. However, other drugs are particularly effective for a specific inflamed tissue and approved in only one or two of the disease entities, e.g. abatacept for peripheral arthritis and vedolizumab for inflammatory bowel disease. This contributes with bedside to bench understanding of the immunology underlying this disease spectrum and provides clinicians with an overview that can assist stratified treatment decisions. We hope that this review will help guide clinicians to speed up treatment of patients with this disease spectrum. ispartof: AUTOIMMUNITY REVIEWS vol:20 issue:2 ispartof: location:Netherlands status: published

Background The aetiologies and pathogeneses of the joint diseases rheumatoid arthritis (RA) and spondyloarthritis (SpA) are still not fully elucidated. To increase our understanding of the molecular pathogenesis, we analysed the protein composition of synovial fluid (SF) from rheumatoid arthritis (RA) and spondyloarthritis (SpA) patients. Methods Fifty-six synovial fluid samples (RA, n = 32; SpA, n = 24) were digested with trypsin, and the resulting peptides were separated by liquid chromatography and analysed by tandem mass spectrometry. Additionally, the concentration of cell-free DNA (cfDNA) in the synovial fluid was measured, and plasma C-reactive protein (CRP) was determined. Results Three hundred thirty five proteins were identified within the SF. The more abundant proteins seen in RA SF were inflammatory proteins, including proteins originating from neutrophil granulocytes, while SpA SF had less inflammatory proteins and a higher concentration of haptoglobin. The concentration of cell-free DNA in the SF increased together with proteins that may have originated from neutrophils. Plasma CRP levels in both RA and SpA, correlated to other acute phase reactants. Conclusions The proteomic results underline that neutrophils are central in the RA pathology but not in SpA, and even though inhibitors of neutrophils (migration, proteinase inhibitors) were present in the SF it was not sufficient to interrupt the disease process.

ObjectiveTo update the European League Against Rheumatism (EULAR) recommendations for the pharmacological treatment of psoriatic arthritis (PsA).MethodsAccording to the EULAR standardised operating procedures, a systematic literature review was followed by a consensus meeting to develop this update involving 28 international taskforce members in May 2019. Levels of evidence and strengths of recommendations were determined.ResultsThe updated recommendations comprise 6 overarching principles and 12 recommendations. The overarching principles address the nature of PsA and diversity of both musculoskeletal and non-musculoskeletal manifestations; the need for collaborative management and shared decision-making is highlighted. The recommendations provide a treatment strategy for pharmacological therapies. Non-steroidal anti-inflammatory drugs and local glucocorticoid injections are proposed as initial therapy; for patients with arthritis and poor prognostic factors, such as polyarthritis or monoarthritis/oligoarthritis accompanied by factors such as dactylitis or joint damage, rapid initiation of conventional synthetic disease-modifying antirheumatic drugs is recommended. If the treatment target is not achieved with this strategy, a biological disease-modifying antirheumatic drugs (bDMARDs) targeting tumour necrosis factor (TNF), interleukin (IL)-17A or IL-12/23 should be initiated, taking into account skin involvement if relevant. If axial disease predominates, a TNF inhibitor or IL-17A inhibitor should be started as first-line disease-modifying antirheumatic drug. Use of Janus kinase inhibitors is addressed primarily after bDMARD failure. Phosphodiesterase-4 inhibition is proposed for patients in whom these other drugs are inappropriate, generally in the context of mild disease. Drug switches and tapering in sustained remission are addressed.ConclusionThese recommendations provide stakeholders with an updated consensus on the pharmacological management of PsA, based on a combination of evidence and expert opinion.

INTRODUCTION: The development of companion diagnostics (CDx) will increase efficacy and cost-benefit markedly, compared to the currently prevailing trial-and-error approach for treatment. Recent improvements in high-throughput protein technology have resulted in large amounts of predictive biomarkers that are potentially useful components of future CDx assays. Current high multiplex protein arrays are suitable for discovery-based approaches, while low-density and more simple arrays are suitable for use in point-of-care facilities.AREA COVERED: This review discusses the technical platforms available for protein array focused CDx, explains the technical details of the platforms and provide examples of clinical use, ranging from multiplex arrays to low-density clinically applicable arrays. We thereafter highlight recent predictive biomarkers within different disease areas, such as oncology and autoimmune diseases. Lastly, we discuss some of the challenges connected to the implementation of CDx assays as point-of-care tests.EXPERT OPINION: Recent advances in the field of protein arrays have enabled high-density arrays permitting large biomarker discovery studies, which are beneficial for future CDx assays. The density of protein arrays range from a single protein to proteome-wide arrays, allowing the discovery of protein signatures that may correlate with drug response. Protein arrays will undoubtedly play a key role in future CDx assays.

Human extracellular matrix (hECM) proteins aids the initial attachment and initiation of an infection, by specific binding to bacterial cell surface proteins. However, the importance of hECM proteins in structure, integrity and antibiotic resilience of a biofilm is unknown. This study aims to determine how specific hECM proteins affect S. aureus USA300 JE2 biofilms.Biofilms were grown in the presence of synovial fluid from rheumatoid arteritis patients to mimic in vivo conditions, where bacteria incorporate hECM proteins into the biofilm matrix. Difference in biofilm structure, with and without addition of hECM to growth media, was visualized by confocal laser scanning microscopy. Two enzymatic degradation experiments were used to study biofilm matrix composition and importance of hECM proteins: enzymatic removal of specific hECM proteins from growth media, before biofilm formation, and enzymatic treatment of 24-hour-old biofilms.hECM addition changed the overall biofilm structure, with larger dispersion of cells within the biofilm matrix. Fibrin, elastin, and collagen were important in forming and maintaining the biofilm structure. Their absence, from growth media, reduced biofilm formation 5-fold, indicating that they are important for biofilm initiation. Their enzymatic degradation, in an established biofilm, caused dispersal, showing that these proteins are critical for structural integrity. A combination of antibiotics with hECM degrading enzymes did not improve the treatment outcome.We conclude that while hECM proteins are an integral part of the biofilm matrix, we find no evidence that these matrix components are directly responsible for the biofilm’s unique antibiotic resilience. The hECM proteins are however highly important in determining biofilm structure and initiation. When utilizing in vitro biofilm models, we therefore recommend addition of hECM proteins to standard growth media, in order to mimic biofilm properties and structure seen in vivo.

Objective: A reduction in haemoglobin level is a frequent complication among rheumatoid arthritis (RA) patients. Hepcidin has been linked to disturbed erythropoiesis. The objective of this study was to investigate the longitudinal changes in hepcidin in patients with early RA. Method: Hepcidin plasma concentrations were measured by enzyme-linked immunosorbent assay in patients with early RA (n = 80) and healthy volunteers (HV, n = 40). Haemoglobin and other iron-related proteins were also measured. At baseline, all patients had active disease and were treatment naïve. Patients were treated with disease-modifying anti-rheumatic drugs (DMARDs) and with additional adalimumab (ADA, n = 42) or placebo (PLA, n = 38) during 52 weeks, using a treat-to-target strategy, aiming for a 28-joint Disease Activity Score (DAS28)

The aim was to examine anti-tumor necrosis factor α (anti-TNFα) therapy influence changes on Th17 and Th22 cells in patients with spondyloarthritis (SpA), and its correlation with changes in clinical and magnetic resonance imaging (MRI) activity and chronicity scores. The Th17 and Th22 cells were assessed at baseline, after 12 and 52 weeks of anti-TNFα therapy by flow cytometry (ClinicalTrials.gov NCT4682724). The percentages of both Th17 and Th22 cells were increased by 70% at baseline compared with healthy controls (both p

Background: Fibroblasts-like synoviocytes (FLS) present in the stromal environment are key effector cells in the persistence of synovial inflammation and joint damage in rheumatoid arthritis (RA). A particular subset of FLS characterized as Thy-1+Podoplanin+CD34- is significantly expanded in RA patients and has been described to be disease-associated1. Currently, no treatment targeting the stromal environment in RA is available2. Objectives: To identify a novel treatment option targeting the stromal environment in RA to modify the disease-associated subset of fibroblasts. Methods: An in vitro model was established with FLS derived from synovial fluid mononuclear cells (SFMC) from RA and osteoarthritis (OA) patients (n=6). FLS between passage 2-5 were used for further analyses. Untreated FLS in cultures were analyzed by flow cytometry for expression of the surface proteins CD34, CD45, Thy-1 and Podoplanin. The potential effect of cell detachment solutions on the expression of the surface proteins was examined by applying either trypsin or lidocaine. To evaluate on the inflammation status of the FLS, MCP-1 levels in supernatants from FLS in mono-cultures stimulated with either antibodies targeting galectin-9 (Gal-9), an anti-Gal-9 antibody-matched isotype control, LPS or steroid were analyzed by ELISA and compared to culture medium alone. Cell viability were examined using an MTT assay. Data are expressed as mean (95% CI) and analysed by a paired t-test. P-values Results: FLS derived from SFMC were characterized as CD34-CD45- and the majority co-expressed Thy-1 and Podoplanin [80.5%, (66.3-94.7%)] confirming the pathogenic phenotype of these cells. This phenotype was not altered by using different methods to detach the cells from the cell-culture plates. FLS receiving anti-Gal-9 antibody treatment showed a significant decreased fold change in MCP-1 secretion (0.84, [0.70;0.98]) compared with unstimulated cells (p=0.036). Treatment with an isotype control did not result in a significant decrease in MCP-1 secretion. This tendency was specific to FLS derived from RA patients as FLS derived from OA patients showed no significant decrease in MCP-1 secretion upon anti-Gal-9 antibody treatment. FLS derived from both RA or OA patients showed a significant increased fold change in secretion of MCP-1 upon LPS stimulation and significantly decreased levels of MCP-1 upon steroid treatment, consistent with the pathogenic phenotype of these cells. None of the different stimulations resulted in morphological changes of the FLS examined by light microscopy. Further, no significant changes in cell viability were detected after anti-Gal-9 antibody treatment. Conclusion: FLS cultures derived from RA patients at passage 2-5 consist mainly of disease-associated fibroblasts and secrete significantly lower amounts of MCP-1 when treated with an anti-Gal-9 antibody whitout affecting cell viability. Thus suggesting that Gal-9 neutralization may represent a novel treatment option targeting the stromal environment and inflammation in RA. References: [1] Mizoguchi, F. et al. Functionally distinct disease-associated fibroblast subsets in rheumatoid arthritis. Nat. Commun. 9, 789 (2018). [2] Dakin, S. G. et al. Pathogenic stromal cells as therapeutic targets in joint inflammation. Nat. Rev. Rheumatol. 14, 714–726 (2018). Disclosure of Interests: Ditte Koster: None declared, Malene Hvid: None declared, Aida Solhoj Hansen: None declared, Tue Wenzel Kragstrup Consultant for: Bristol-Myers Squibb, Speakers bureau: Pfizer, Bristol-Myers Squibb, Eli-Lilly, Novartis, UCB, Pierre Busson: None declared, Bent Deleuran: None declared, Morten Aagaard Nielsen: None declared

Background The many different disease modifying anti-rheumatic drugs used for the treatment of immune mediated inflammatory arthritis have very distinctive modes of action. However, the downstream effects of the different drugs are not completely understood. As more treatment options become available detailed knowledge about the different drugs will be important in the future when planning treatment strategies. Adalimumab is a fully humanized monoclonal antibody against tumor necrosis factor-alpha (TNFα). Earlier studies have found that TNFα inhibitors can decrease the production of interleukin 1 (IL-1), IL-6, IL-8 (aka CXC motif chemokine 8 (CXCL8)), monocyte chemotactic protein 1 (MCP-1) and granulocyte-macrophage colony-stimulating factor (GM-CSF).[1] However, these studies have been limited to the measurement of a subset of predefined downstream signaling molecules. Objectives The aim of this study was to make an unbiased investigation of downstream effects of adalimumab on production of a wide range of cytokines, chemokines and growth factors by synovial fluid mononuclear cells (SFMCs). Methods SFMCs were obtained from a study population consisting of patients with active RA or peripheral SpA with at least one swollen joint (for obtaining synovial fluid) (n=14). SFMCs were cultured for 48 hours with and without addition of adalimumab 5 μg/ml measuring the secretion of 96 cytokines, chemokines and growth factors by an Olink proseek multiplex panel. Results In SFMCs cultured for 48 hours, adalimumab decreased the production of IL-8 (CXCL8) (P=0.0001), CXC motif chemokine 5 (CXCL5) (P=0.0003), CXCL9 (P=0.03) and CXCL10 (P=0.03), and monocyte chemotactic protein 2 (MCP-2) (P=0.04) and increased the production of hepatocyte growth factor (HGF) (P=0.008) and tumor necrosis factor-like weak inducer of apoptosis (TWEAK) (P=0.03) after Bonferroni correction (all corrected P values). In this unbiased analysis, adalimumab did not change the production of several other cytokines and chemokines including IL-12B (P=0.07), IL-6 (P=0.07), IL-10 (P=0.12), IL-18 (P=0.19), and IL-4 (P=0.63). Conclusion This study illuminates the downstream effects of neutralizing TNFα not previously investigated. Adalimumab had a pronounced effect on downregulation of the inflammatory CXCL subfamily chemokines IL-8, CXCL5, CXCL9, and CXCL10. This helps explain findings of diminished inflammatory cell migration into joints seen in the first trials with TNFα inhibitors and could be an important mechanism of action of TNFα inhibitors.[2] Further characterization of downstream effects of the multiple DMARDs used for the treatment of immune mediated inflammatory arthritis will help guide treatment strategies for these patients. References [1] Brennan FM, Chantry D, Jackson A, Maini R, Feldmann M. Inhibitory effect of TNF alpha antibodies on synovial cell interleukin-1 production in rheumatoid arthritis. Lancet1989;2:244–7. [2] Taylor PC, Peters AM, Paleolog E, Chapman PT, Elliott MJ, McCloskey R, et al. Reduction of chemokine levels and leukocyte traffic to joints by tumor necrosis factor alpha blockade in patients with rheumatoid arthritis. Arthritis Rheum. 2000;43:38–47. Disclosure of Interests Anne Faurschou Boisen: None declared, Elisabeth Busk Rasmussen: None declared, Tue Wenzel Kragstrup Consultant for: Bristol-Myers Squibb, Speakers bureau: Pfizer, Bristol-Myers Squibb, Eli-Lilly, Novartis, UCB

Background The pathophysiology of osteoarthritis (OA) involves wear and tear, and a state of low-grade inflammation. Tissue repair responses include transforming growth factor beta (TGFβ)-induced myofibroblast production of extracellular matrix. Fibronectins are an essential part of the extracellular matrix, and injection of fibronectin fragments into rabbit joints is a previously established animal model of OA. Fibronectin containing the ED-A domain is currently being used as drug delivery target in the development of anti-inflammatory drugs (e.g. Dekavil). Methods In this study, samples of synovial membrane were obtained from patients with knee OA undergoing joint replacement surgery. Immunostaining for ED-A fibronectin and the myofibroblast marker alpha smooth muscle actin (αSMA) was performed on fibroblast-like synovial cells (FLS) and synovial membranes. RAW 264.7 macrophages were incubated with recombinant ED-A fibronectin. Results The staining of ED-A fibronectin in OA FLS was increased by TGFβ but not by TNFα, lipopolysaccharide, or IL-6 (n = 3). ED-A fibronectin co-stained with the myofibroblast marker αSMA in both the OA FLS (n = 3) and in the OA synovial membranes (n = 8). ED-A fibronectin staining was associated with both number of lining layer cells (rho = 0.85 and p = 0.011) and sublining cells (rho = 0.88 and p = 0.007) in the OA synovium (n = 8), and co-distributed with TNFα (n = 5). Recombinant ED-A fibronectin increased the production of TNFα by RAW 264.7 macrophages (n = 3). Conclusions The disease process in OA shares features with the chronic wound healing response. Our findings support utilizing ED-A fibronectin for drug delivery or therapeutic targeting to reduce pro-inflammatory responses in OA.

Background: Apremilast (Otezla®) is a phosphodiesterase 4 (PDE4) inhibitor approved for the treatment of psoriasis and psoriatic arthritis (PsA), but the reason why apremilast shows clinical effect is not fully understood. The objective of this study was to study the downstream effects of apremilast on cells of inflamed joints in immune-mediated inflammatory arthritis. Methods: Synovial fluid was obtained from patients with active rheumatoid arthritis (RA), PsA or peripheral spondyloarthritis (SpA; n = 18). The in vitro models consisted of synovial fluid mononuclear cells (SFMCs) or fibroblast-like synovial cells (FLSs) cultured for 48 h, SFMCs cultured for 21 days, an osteoclast pit formation assay, and a mineralization assay. Results: In SFMCs cultured for 48 h, apremilast decreased the production of interleukin (IL)-12/IL-23p40 (the shared subunit of IL-12 and IL-23), colony-stimulating factor 1, CD6, and CD40 and increased the production of C-X-C motif chemokine 5 dose-dependently. Apremilast had a very different response signature compared with the tumor necrosis factor alpha inhibitor adalimumab with a substantially greater inhibition of IL-12/IL-23p40. In SFMCs cultured for 21 days, apremilast increased the secretion of IL-10. In FLS cultures, apremilast decreased matrix metalloproteinase-3 production. Apremilast decreased osteoclastogenesis but did not affect mineralization by human osteoblasts. Conclusion: This study reveals the downstream effects of apremilast in ex vivo models of arthritis with a strong inhibition of IL-12/IL-23p40 by SFMCs. Our findings could explain some of the efficacy of apremilast seen in IL-12/IL-23-driven immune-mediated inflammatory diseases such as psoriasis and PsA.

Nonspecific binding of monoclonal antibodies (mAbs) to Fc-receptors on leukocytes is an important cause of background fluorescence in flow cytometry, and failing to block such nonspecific binding can lead to erroneous results. A major part of previous studies on blocking reagents for flow cytometry have been done in mice, and published results are not completely in agreement. In humans, Fc-receptors are found on most leukocytes, with highest abundance on monocytes/macrophages. Therefore, in the present study our aim was to thoroughly investigate the efficiency of different commonly used blocking reagents regarding inhibition of nonspecific binding of mouse mAbs to human peripheral blood mononuclear cells (MNCs) and monocyte-derived macrophages (MDMs). Monocytes and MDMs showed strong nonspecific binding of IgG1 and IgG2a isotypes, but not IgG2b. In contrast, B-cells, T-cells, and NK-cells did not substantially bind any of the mouse isotype control antibodies evaluated (IgG1, IgG2a, and IgG2b). Importantly, we show that binding of IgG1 and IgG2a to monocytes and MDMs can be eliminated by blocking, either with a commercial Fc-blocking reagent, with mouse or human serum, or with mouse or human IgG in high concentration. Previously, isotype controls have been widely used in flow cytometry assays. However, we show that such controls may be highly unreliable, and we believe they should not be used as gating controls, or to determine background signal. Based on these results, as well as considerations of price and applicability, our recommendation is not to use isotypes as gating controls in flow cytometry, but instead to use 100 μg/mL of purified human IgG as blocking reagent for elimination of nonspecific binding of mouse mAbs to human MNCs and MDMs. © 2016 International Society for Advancement of Cytometry

Background:Rheumatoid arthritis (RA) is an immune mediated inflammatory disease with autoimmune features, including antibodies to citrullinated proteins and peptides (ACPAs). Several in vitro studies have suggested a pathogenic role of ACPAs in RA. However, in vivo proof of this concept has been hampered by the lack of therapeutic strategies to reduce or deplete ACPA in serum and synovial fluid. Previously, we constructed a chitosan-hyaluronic acid nanoparticle formulation with the ability to use neutrophil recruitment as a delivery mechanism to inflamed joints. Specifically, nanoparticles got phagocytosed and then released to synovial fluid upon death of the short-lived neutrophilsObjectives:We hypothesized that reducing ACPA levels would have a therapeutic effect by blocking cytokine production. In this study, we prepared and tested a series of therapeutic nanoparticles for specific targeting of ACPA in synovial fluid.Methods:Nanoparticles were prepared by the microdroplet method and then decorated with synthetic cyclic citrullinated peptide aptamer PEP2, PEG/hexanoic acid and fluorophore (Cy5.5). Nanoparticles were characterized by dynamic light scattering (DLS), scanning electron microscopy (SEM) and high-performance liquid chromatography (HPLC). Nanoparticles were then used in a series of in vitro assays, including cell uptake with flow cytometry (FACS) detection, and in vivo studies including disease activity scores, cytokine measurements and near-infrared imaging.Results:We screened a series of citrullinated peptide epitopes and identified a fibrinogen-derived 21-amino-acid-long citrullinated peptide showing high selectivity toward autoantibodies in RA samples. We incorporated this aptamer in the chitosan-hyaluronic acid nanoparticle formulation previously described. Average nanoparticle size was 230 nm ± 10 nm by DLS and SEM; z potential was -0.0012. Purity by HPLC was over 95%. Attachment efficiency of the aptamer was 92% by HPLC. FACS study showed selective uptake of Cy5.5 labelled aptamer-nanoparticle conjugates by neutrophils in the concentration range 0.5-4 nM. Similar to previous studies,1there was no apparent immunogenicity for this nanoparticle formulation measured by cytokine secretion from human peripheral blood leukocytes. In vivo, over 50% reduction of disease activity was achieved in three weeks treatment using as little as 1 nM drug candidate (dosed every 48 hours) in the collagen-induced (CIA) mouse model of RA (N=30; pConclusion:Overall, we have designed a first-in-class therapeutic nanoparticle drug for specific targeting of anti-citrullinated protein antibodies. The marked effect of this nanoparticle observed in vivo holds promise for targeting ACPAs as a therapeutic option in RA.References:[1]Khatri S, Hansen J, Mendes AC, Chronakis IS, Hung S-C, Mellins ED, Astakhova K. Bioconjug Chem. 2019 Oct 16;30(10):2584–259Disclosure of Interests:Sangita Khatri: None declared, Jonas Hansen: None declared, Mads Hartvig Clausen Shareholder of: iBio Tech ApS, Tue Wenzel Kragstrup Shareholder of: iBio Tech ApS, Consultant of: Bristol-Myers Squibb, Speakers bureau: TWK has engaged in educational activities talking about immunology in rheumatic diseases receiving speaking fees from Pfizer, Bristol-Myers Squibb, Eli Lilly, Novartis, and UCB., Shu-Chen Hung: None declared, Elisabeth Mellins: None declared, Kira Astakhova: None declared

Background:During cancer treatment with immune checkpoint inhibitors (ICI) such as the anti-PD-1 antibody pembrolizumab, 2-4% of patients develop inflammatory arthritis as an immune related adverse event and half of patients with pre-existing inflammatory arthritis have disease flares. This type of adverse events shows striking similarities with traditional immune mediated inflammatory arthritis. However, the underlying immunological mechanisms of inflammatory arthritis associated with ICI are not fully understood.Objectives:We aimed to develop an in vitro model of inflammatory arthritis associated with ICI, and to use this model to investigate monocyte differentiation and activation following treatment with pembrolizumab.Methods:First, synovial fluid mononuclear cells (SFMCs) and peripheral blood mononuclear cells (PBMCs) from patients with immune mediated inflammatory arthritis (rheumatoid arthritis and peripheral spondyloarthritis, n=22) and PBMCs from healthy controls were incubated with pembrolizumab and assessed for monocyte chemoattractant protein 1 (MCP-1) secretion by ELISA. Then, cytokine production in SFMCs was studied in more detail by the multiplex V-PLEX proinflammatory panel and by intracellular flow cytometry. Finally, pembrolizumab treated SFMCs were incubated with the different disease modifying anti-rheumatic drugs adalimumab, tocilizumab, tofacitinib, and baricitinib.Results:Pembrolizumab significantly increased MCP-1 production in the SFMCcultures (P=0.0031). In contrast, pembrolizumab did not change MCP-1 production by PBMCs from neither patients nor healthy controls (P=0.77 and P=0.43). Pembrolizumab also increased the production of TNFα (P=0.049), IFNγ (P=0.047), and IL-12p70 (P=0.031), but did not change the production of IL-6 (P=0.98). Among SFMCs treated with pembrolizumab there was an increased frequency of intermediate monocytes (P=0.044). Interestingly, pembrolizumab also increased the MCP-1 production within the intermediate monocytes only (P=0.028). In contrast, among SFMCs treated with LPS, only the classical monocyte subset was increased (P=0.0045) and MCP-1 production increased in both intermediate and classical monocyte subsets. Lastly, the TNFα inhibitor adalimumab and the Janus kinase inhibitors baricitinib and tofacitinib attenuated the pembrolizumab-induced MCP-1 production (P=0.0004, P=0.033, and P=0.025, respectively) while this was not seen with the IL-6 inhibitor tocilizumab (P=0.75).Conclusion:We have developed a very simple in vitro model of inflammatory arthritis associated with ICI. Using this model, we found that pembrolizumab specifically activated intermediate monocytes and induced TNFα, IFNγ, and IL-12p70 production, whereas IL-6 was unchanged. These findings were supported by effectful reduction of MCP-1 secretion with TNFα inhibition but not with IL-6 inhibition. This model could potentially be used to further study the effects of ICIs and the underlying immunological mechanisms of inflammatory arthritis associated with ICI.References:None.Disclosure of Interests:Anne Sofie Sørensen: None declared, Morten Nørgaard Andersen: None declared, Kristian Juul-Madsen: None declared, Cæcilie Skejø: None declared, Henrik Schmidt: None declared, Thomas Vorup-Jensen: None declared, Tue Wenzel Kragstrup Shareholder of: iBio Tech ApS, Consultant of: Bristol-Myers Squibb, Speakers bureau: TWK has engaged in educational activities talking about immunology in rheumatic diseases receiving speaking fees from Pfizer, Bristol-Myers Squibb, Eli Lilly, Novartis, and UCB.

Background:Fibroblast-like synoviocytes (FLS) are central cellular components in persistent inflammatory joint diseases such as rheumatoid arthritis (RA). Pathological subsets of FLS have been identified from synovial tissue. However, the synovial tissue obtained from arthroplasty procedures is acquired at late disease stages and the cellular yield obtained from synovial tissue biopsies is fairly low. Collectively, challenging the robustness of human RAin vivoandin vitromodels. FLS obtained from the synovial fluid (SF-FLS) are proposed as an alternative source of FLS, but a detailed phenotypical and functional characterization of FLS subsets from the synovial fluid has not been performed.Objectives:The aim of this study was to determine the phenotypical and functional characteristics of synovial fluid-derived fibroblast-like synoviocytes in rheumatoid arthritis.Methods:In the present study, paired peripheral blood mononuclear cells (PBMC) and SF-FLS from patients with RA were obtained (n=7). FLS were isolated from the synovial fluid by a strict trypsinization protocol1and their cellular characteristics and functionality were evaluated at passage 4. Monocultures (SF-FLS) and autologous co-cultures (SF-FLS and PBMC) were established from five patients with RA and subsequently evaluated by flow cytometry, Western blotting and multiplex immunoassays. Human cartilage-sponges (n=3) with SF-FLS and without SF-FLS (n=3) were co-implanted subcutaneously in SCID mice (n=15), mice with only cell-free human cartilage-sponges were used as controls (n=12). After 45 days, the implants were evaluated using stained sections to determine the SF-FLS invasion score based on perichondrocytic cartilage degradation. Data are expressed as median (25-75 percentile). P-values Results:The homogeneous subpopulations of FLS, isolated from the synovial fluid, were negative for CD34 and CD45 [98.9%, (97.5-99.7]) and positive for Thy-1 and PDPN [94.6%, (79.9-97.4]). Without stimulation, RA SF-FLS showed high and comparable levels of NFκB related pathway proteins and secreted multiple pro-inflammatory cytokines and chemokines dominated by IL-6 [2648 pg/mL, (1327-6116)] and MCP-1 [2458 pg/mL, (692-8719)]. SF-FLS increased their ICAM-1 and HLA-DR expression after encountering autologous PBMCs (pin vivowas at primary site, [1.6, (1.3-1.7)] and contralateral implantation site [1.5, (1.1-2.2)]. The invasion score of the human SF-FLS-containing implants both at primary and contralateral site were significantly higher compared with cartilage-sponges evaluated from SF-FLS-free control mice (pConclusion:This phenotypical and functional characterization of SF-FLS, acquired and activated at the site of pathology, lays a foundation for establishingin vivoandin vitroFLS models. These FLS models will be beneficial in our understanding of the role of this cellular subset in arthritis and for characterization of drugs specifically targeting this pathological RA FLS subset.References:[1]Nielsen M. A. et al. Responses to Cytokine Inhibitors Associated with Cellular Composition in Models of Immune-Mediated Inflammatory Arthritis. ACR Open Rheumatology, 2(1):3-10.http://doi.org/10.1002/acr2.11094Disclosure of Interests:Ditte Køster: None declared, Johanne Hovgaard Egedal: None declared, Malene Hvid: None declared, Martin Roelsgaard Jakobsen: None declared, Ulf Müller-Ladner Speakers bureau: Biogen, Bent Deleuran: None declared, Tue Wenzel Kragstrup Shareholder of: iBio Tech ApS, Consultant of: Bristol-Myers Squibb, Speakers bureau: TWK has engaged in educational activities talking about immunology in rheumatic diseases receiving speaking fees from Pfizer, Bristol-Myers Squibb, Eli Lilly, Novartis, and UCB., Elena Neumann: None declared, Morten Aagaard Nielsen: None declared

Objective Active rheumatoid arthritis (RA) is accompanied by increased appendicular and axial bone loss, closely associated to the degree of inflammation. The programmed death-1 (PD-1) pathway is important for maintaining peripheral tolerance, and its ligand PD-L2 has recently been associated with bone morphogenetic protein activity. Here, we report that PD-L2 plays a central role in RA osteoimmunology. Methods Femoral bone mineral density (BMD) and trabecular bone microstructure were evaluated by micro-CT in wild type (WT) and PD-L2−/− mice. Osteoclasts were generated from RA synovial fluid mononuclear cells and peripheral blood monocytes. The effects of recombinant PD-L2, was evaluated by tartrate-resistant acid phosphatase (TRAP) activity and the development of bone erosions in the presence of anti-citrullinated protein antibodies (ACPA). Plasma soluble (s)PD-L2 levels were measured in patients with early (e)RA (n ​= ​103) treated with methotrexate alone or in combination with the TNF inhibitor Adalimumab. Results PD-L2−/− mice had a decreased BMD and deteriorated trabecular bone microstructure that was not related to the RANKL/OPG pathway. PD-L2 decreased TRAP activity in osteoclasts and decreased ACPA-induced erosions. In the RA synovial membrane PD-L2 was highly expressed especially in the lining layer and plasma sPD-L2 levels were increased in eRA patients and decreased with treatment. One-year sPD-L2 correlated inversely with erosive progression two years after treatment initiation with methotrexate and placebo. Conclusion PD-L2 regulates bone homeostasis in RA. Our findings provide new insight into the relationship between the immune system and bone homeostasis, and suggest a potential therapeutic target for limiting inflammatory bone loss in RA., Highlights • PD-L2 is closely related to bone homeostasis in a mouse model. • PD-L2 inhibits osteoclastogenesis and osteoclast activation in vitro. • PD-L2 is highly expressed by cells in the synovial membrane of rheumatoid arthritis. • PD-L2 is associated with less radiographic progression in patients with early rheumatoid arthritis.

Background The pathogenesis of spondyloarthritis (SpA) involves both inflammation and new bone formation in the spine. In line with this, the disease has been characterized as both inflammatory and fibrotic. The current treatment dampens inflammation while new bone formation can progress. Therefore, there is an unmet therapeutic need for the treatment of new bone formation in SpA. Fibrosis is mediated by myofibroblasts and new bone formation is the result of increased osteoblast mineralization and decreased osteoclast resorption. Here, we evaluate the potential effect of the newly approved anti-fibrotic agent pirfenidone (PFD) on fibrosis and new bone formation in cell culture models of SpA. Methods Fibroblast-like synoviocytes (FLSs) were isolated from SpA patients (n = 6) while the osteoblast cell line Saos-2 was purchased. The cells were cultured with PFD at 0.25 0.5, or 1.0 mg/ml. The proliferation of FLSs was analyzed with light microscopy and flow cytometry. The differentiation and activation of FLSs was assessed with flow cytometry, a membrane-based antibody array and enzyme-linked immunosorbant assays. The mineralization capacity of osteoblasts was studied with an assay measuring deposition of hydroxyapatite. Results PFD reduced the Ki67 expression 7.1-fold in untreated FLSs (p = 0.001) and 11.0-fold in FLSs stimulated with transforming growth factor beta (TGFβ), tumor necrosis factor alpha (TNFα), and interferon gamma (IFNγ) (p = 0.022). There were no statistically significant changes in membrane expression of alpha smooth muscle actin (αSMA), intercellular adhesion molecule 1 (ICAM-1), or human leukocyte antigen DR (HLA-DR). In supernatants from FLSs stimulated with TGFβ, TNFα, and IFNγ, PFD decreased the secretion of 3 of 12 proteins more than 2-fold in the membrane-based antibody array. The changes in secretion of monocyte chemoattractant protein 1 (MCP-1) and chitinase-3-like protein 1 (CHI3L1, YKL-40) were validated with ELISA. PFD decreased the secretion of both Dickkopf-related protein 1 (DKK1) (p = 0.006) and osteoprotegerin (OPG) (p = 0.02) by SpA FLSs stimulated with TGFβ, TNFα, and IFNγ. Finally, PFD inhibited the deposition of hydroxyapatite by osteoblasts in a dose-dependent manner (p = 0.0001). Conclusions PFD inhibited SpA FLS proliferation and function and osteoblast mineralization in vitro. This encourages studies of the in vivo effect of PFD in SpA.

Background Resveratrol is a natural polyphenol found in berries, roots and wine that is well known to have anti-inflammatory and anti-oxidative properties. The anti-inflammatory effect has been reported for both immune cells and connective tissues, but only few studies have investigated effects on immune mediated inflammatory arthritis. None of which have studied this effect when combining resveratrol with methotrexate or adalimumab, two major drugs in the treatment of immune mediated inflammatory arthritis. We therefore aimed to investigate the anti-inflammatory effect of resveratrol alone and in combination with methotrexate or adalimumab in ex vivo models of immune mediated inflammatory arthritis. We furthermore aimed to describe any variations in this effect based on disease activity and cellular composition of the synovial fluid infiltrate. Methods Synovial fluid mononuclear cells from patients with rheumatoid arthritis (n = 7) and spondyloarthritis (n = 7) were cultured for either 48 h or 21 days. In both models, synovial fluid mononuclear cells were treated with resveratrol alone or in combination with methotrexate or adalimumab. Monocyte chemoattractant protein 1, matrix metalloproteinase 3 and tartrate resistant acidic phosphatase were measured to quantify inflammation, enzymatic degradation and osteoclast differentiation, respectively. Results Resveratrol reduced monocyte chemoattractant protein 1 production by synovial fluid mononuclear cells significantly (p = 0.005) compared to untreated controls. The effect of resveratrol was greatest in cultures from patients with low disease activity, i.e. DAS28CRP ≤ 3.2 (p = 0.022), and in cultures dominated by lymphocytes (p = 0.03). Further, the combination of methotrexate and resveratrol significantly reduced monocyte chemoattractant protein 1 levels compared with methotrexate alone in cultures from patients with low disease activity (p = 0.016), and in cultures with high lymphocyte count (p = 0.011). Resveratrol did not significantly affect matrix metalloproteinase 3 and tartrate resistant acidic phosphatase production. Conclusion Resveratrol has anti-inflammatory properties in our ex vivo model of immune mediated inflammatory arthritis. Results show an additive effect of resveratrol, when combined with methotrexate in samples dominated by lymphocytes and samples from patients with low disease activity. This suggests further investigations in vitro and whether this effect may also be present in a clinical setting. Electronic supplementary material The online version of this article (10.1186/s41927-018-0036-5) contains supplementary material, which is available to authorized users.

During alcoholic hepatitis (AH) monocytes traverse the vascular boundaries and massively invade the liver. In principle, tissue extravasation can be limited through shedding of CD18 integrins from leukocytes, including monocytes. The soluble (s) product sCD18 conceals adhesion receptors on the endothelium, which reduces monocyte extravasation. In AH, monocytes are dysfunctional, but whether this involves their self-generated anti-migration is unknown. Our aim was, therefore, to investigate monocyte CD18 dynamics in AH.We studied 50 AH patients and 20 healthy controls. We measured monocyte expression and conformational activation of CD18, plasma (P)-sCD18, stimulated in vitro CD18 shedding and P-sCD18 in a short-term chronic-binge mouse model.AH-derived monocytes had a 30-60% higher expression of active CD18 receptors (p 0.01), but the sCD18 concentration per monocyte was reduced in vivo by 30% and in vitro by 120% (p 0.01). Ethanol reduced the in vitro shedding of CD18 in the patients only. TNFα increased sCD18 concentration per monocyte, but less so in the patients (p 0.04). P-sCD18 per monocyte was inversely related to disease severity. In early alcoholic liver disease, P-sCD18 was decreased in the mouse model.The monocyte CD18 integrins are highly activated in AH and the single monocyte shedding of CD18 was decreased favoring tissue extravasation. Alcohol in itself and altered monocyte responsiveness to TNFα may explain this lowered shedding.The contribution of this mechanism to the excessive monocyte liver infiltration in AH should be further explored as it may serve as a potential therapeutic target to limit liver inflammation.

Background Targeting intracellular pathways with oral small molecules is an attractive therapeutic approach for treating immune mediated inflammatory diseases. The mitogen-activated protein kinase (MAPK) pathway is activated by environmental stressors, growth factors and inflammatory cytokines. However, the inhibition of central MAPK proteins has so far had undesirable side effects. The MAPK-activated protein kinase 2 (MK2) is a downstream mediator in the MAPK signalling pathway and could therefore be inhibited without the same side effects (see figure 1). Objectives The objective of this study was to study the effects of a small molecule inhibiting MK2 on inflammation and structural changes in ex vivo models of immune mediated inflammatory arthritis. Methods Synovial fluid mononuclear cells (SFMCs), fibroblast like synovial cells (FLSs) and peripheral blood mononuclear cells (PBMCs) were obtained from a study population consisting of patients with active RA or peripheral SpA with at least one swollen joint (for obtaining synovial fluid) (n=14). SFMCs were cultured for 48 hours with and without addition of a MK2 inhibitor (Celgene) at 1000 nM, 333 nM and 111 nM and supernatants were analysed by the Olink proseek multiplex interferon panel and commercially available ELISA assays. Because FLSs are only found in small amounts among SFMCs, autologous co-cultures of FLS and PBMCs and SFMCs were also used. SFMCs cultured for 21 days were used to study inflammatory macrophage differentiation and osteoclastogenesis. Results In SFMCs cultured for 48 hours, the MK2 inhibitor decreased the production of CXCL9 (p Conclusions This study reveals the effects of a MK2 inhibitor in ex vivo models of immune mediated inflammatory arthritis. The MK2 inhibitor changed the secretory profile of SFMCs and decreased inflammatory osteoclastogenesis. Taken together, this points to a role of this MK2 inhibitor in attenuating inflammatory and destructive arthritis. Disclosure of Interest T. Kragstrup: None declared, A. Mellemkjaer: None declared, M. Nielsen: None declared, L. Heftdal: None declared, P. Schafer Shareholder of: Celgene, Employee of: Celgene, B. Deleuran: None declared

Background Immune mediated inflammatory arthritis including rheumatoid arthritis (RA), psoriatic arthritis (PsA) and spondyloarthritis (SpA) all characterised by joint synovitis. Disease-modifying antirheumatic drugs (bDMARDs) targeting specific components of the pathogenesis have radically improved the treatment of the diseases. However, a fair proportion of patients are non-responders. Today, the first choice of DMARD is dependent on market pricing, regardless of the immunological target. This is due to the rather similar efficacy profile of the different DMARDs. Therefore, there is a need for stratification of patients suffering from Immune mediated inflammatory arthritis in order to reduce the fraction of DMARD non-responders. Objectives The objective of this study was to study the effects of various DMARDs on different synovial cell subsets using several human ex vivo models of immune mediated inflammatory arthritis. This could potentially guide future studies of personalising DMARDs in these diseases. Methods Synovial fluid was obtained from a study population of patients with active rheumatoid arthritis (RA) or peripheral spondyloarthritis (SpA). Synovial fluid mononuclear cells (SFMCs) containing primarily synovial monocytes and lymphocytes cultured for 48 hours (“Macrophage and Lymphocyte model”) were used to study the effect of different biological agents on secretion of monocyte chemoattractant protein-1 (MCP-1) (n=14). Further, fibroblast-like synovial cells (FLSs) were co-cultured with autologous PBMCs (“FLS model”) to study the effects of the same biological agents (n=6) in cultures dominated by synovial FLSs. Finally, SFMCs cultured for 21 days (“Osteoclast model”) were studied to assess the effects on inflammatory osteoclastogenesis (n=10) measured by tartrate-resistant acid phosphatase (TRAP). The DMARDs investigated are shown in table 1. Results “Macrophage and Lymphocyte model”; In SFMCs cultured for 48 hours, all DMARDs included, except anakinra, had the ability to decrease the production of MCP-1. The two TNF inhibitors (adalimumab and etanercept) (p “FLS model”; In the FLS+PBMCs cultured for 48 hours, tocilizumab (p “Osteoclast model”; In SFMCs cultured for 21 days, only the two TNF inhibitors, adalimumab and etanercept were able to significantly reduce the secretion of TRAP from adherent macrophage like synovial cells by roughly 25% (p Conclusions This study reveals that most DMARDs have effects in the “Macrophage and Lymphocyte model” whereas tocilizumab, tofacitinib and baricitinib were superior in the “FLS model” and only the two TNF inhibitors were effective in the “Osteoclast model”. The findings in the “FLS model” reveals a possible beneficial effect of tocilizumab and JAK inhibitors to patients with fibroblast dominated arthritis. This study could potentially guide future studies of personalising DMARDs to treat immune mediated inflammatory arthritis. Disclosure of Interest None declared

Background Apremilast (Otezla) is a phosphodiesterase 4 (PDE4) inhibitor approved for the treatment of psoriasis and psoriatic arthritis (PsA), but the reason why apremilast shows clinical effect in PsA is not fully understood. Objectives The objective of this study was to study the downstream effects of apremilast on cells of the inflamed joint in ex vivo models of immune mediated inflammatory arthritis. First, we tested the effect of apremilast on the secretion of several cytokines, chemokines and growth factors by synovial fluid mononuclear cells (SFMCs). Then, we tested whether apremilast affect factors involved in structural changes by studying fibroblast-like synovial cells (FLSs), osteoclasts, synovial macrophages, and osteoblasts. Methods Synovial fluid was obtained from a study population consisting of patients with active rheumatoid arthritis (RA), psoriatic arthritis (PsA) or peripheral spondyloarthritis (SpA) with at least one swollen joint (n=18). Synovial fluid mononuclear cells (SFMCs) cultured for 48 hours were used to study the effect of apremilast on secretion of a large panel of cytokines, chemokines and growth factors using the Olink Proseek Multiplex interferon I panel. These effects were compared with the effects of the tumour necrosis factor alpha (TNFα) inhibitor adalimumab. Further, fibroblast-like synovial cells (FLSs) were used to study metalloproteinase secretion, SFMCs cultured for 21 days were used to study inflammatory osteoclastogenesis and macrophage differentiation, an osteoclast pit formation assay was used to study osteoclast activity, and a mineralization assay was used to study new bone formation. Results In SFMCs cultured for 48 hours apremilast decreased the production of IL-12/IL-23p40 (the shared subunit of IL-12 and IL-23) (p Conclusions This study reveals the downstream effects of apremilast in ex vivo models of arthritis with a strong inhibition of IL-12/IL-23p40. Our findings could explain some of the efficacy of apremilast seen in IL-12/IL-23 driven immune mediated inflammatory diseases such as psoriasis and psoriatic arthritis. Disclosure of Interest T. Kragstrup: None declared, M. Adams: None declared, S. Lomholt: None declared, M. Nielsen: None declared, L. Heftdal: None declared, P. Schafer Shareholder of: Celgene, Employee of: Celgene, B. Deleuran: None declared

OBJECTIVES: During alcoholic hepatitis (AH) monocytes traverse the vascular boundaries and massively invade the liver. In principle, tissue extravasation can be limited through shedding of CD18 integrins from leukocytes, including monocytes. The soluble (s) product sCD18 conceals adhesion receptors on the endothelium, which reduces monocyte extravasation. In AH, monocytes are dysfunctional, but whether this involves their self-generated anti-migration is unknown. Our aim was, therefore, to investigate monocyte CD18 dynamics in AH.METHODS: We studied 50 AH patients and 20 healthy controls. We measured monocyte expression and conformational activation of CD18, plasma (P)-sCD18, stimulated in vitro CD18 shedding and P-sCD18 in a short-term chronic-binge mouse model.RESULTS: AH-derived monocytes had a 30-60% higher expression of active CD18 receptors (p CONCLUSIONS: The monocyte CD18 integrins are highly activated in AH and the single monocyte shedding of CD18 was decreased favoring tissue extravasation. Alcohol in itself and altered monocyte responsiveness to TNFα may explain this lowered shedding.TRANSLATIONAL IMPACT: The contribution of this mechanism to the excessive monocyte liver infiltration in AH should be further explored as it may serve as a potential therapeutic target to limit liver inflammation.

IntroductionThe pathophysiology of osteoarthritis (OA) involves wear and tear, and a state of low-grade inflammation. Wear and tear leads to tissue degradation and tissue repair responses, including tissue growth factor beta (TGFβ)-induced myofibroblast production of extracellular matrix (ECM). Fibronectins are an essential part of the ECM, and injection of fibronectin fragments into rabbit joints is a previously established animal model of OA. Alternatively-spliced fibronectin contains the ED-A domain (ED-A FN) and has been shown to activate Toll-like receptor 4. In this study, we tested the hypothesis that FN fragments containing the ED-A domain could be one mechanism transducing mechanical events into inflammatory signals in OA.MethodsSamples of synovial membrane and cartilage were obtained from patients with knee OA undergoing joint replacement surgery. Immunostaining for ED-A FN and the myofibroblast marker alpha smooth muscle actin (αSMA) was performed on synovial membranes and fibroblast-like synovial cells (FLS). FLS were stimulated with TGFβ, TNFα, lipopolysaccharide, IL-6, OA synovial fluid, or chondrocyte lysate, and analyzed for ED-A FN. Synovial cells isolated by enzymatic digestion and human monocyte-derived macrophages (MDM) were incubated with recombinant ED-A FN, plasmin, cellular FN, or cellular FN digested with plasmin; and culture supernatants were analyzed for MCP-1 and TNFα.ResultsWe hypothesized that ED-A FN is produced by OA FLS in response to factors found in the OA synovial joint. Indeed, the production of ED-A FN by OA FLS was increased by TGFβ, OA synovial fluid, and lysed chondrocytes in all experiments (n=3). ED-A FN co- localized with the myofibroblast marker αSMA in both the OA FLS (n=3) and in the OA synovial membranes (n=8). We further hypothesized that ED-A FN expression is associated with cellular density and expression of inflammatory molecules in OA. ED-A FN staining was associated with both number of lining layer cells (rho=0.85 and p=0.011) and sublining cells (rho=0.88 and p=0.007) in the OA synovium (n=8), and co-localized with both MCP-1 and TNFα (n=5). Recombinant ED-A FN increased the production of both MCP-1 and TNFα by MDM (n=3) and OA FLS (n=3). Finally, we demonstrated that the FN fragments containing the ED-A domain generated the same production of both MCP-1 and TNFα as recombinant ED-A FN.ConclusionThe disease process in OA shares features with the chronic wound healing response including myofibroblast differentiation and production of mediators that promote myofibroblast production of ED-A FN. We show that recombinant and plasmin-derived ED-A fragments stimulate FLS and MDM to produce pro-inflammatory mediators. Our findings support utilizing ED-A FN for drug delivery or therapeutic targeting of the formation of ED- A FN or the enzymatic fragmentation of FN to reduce pro-inflammatory responses in OA.

T helper 17 (Th17) cells produce interleukin (IL) 17-A. In addition, Th17 cells produce IL-21 and IL-22. Th17 cells have a disease-promoting role in Crohn’s disease (CD). We investigated the effects of anti-TNFαtreatment on mucosal gene expression (qPCR) of IL-17A, IL-21, and IL-22 as well as on the frequency of lamina propria (LP) T cell subsets producing these cytokines (flow cytometry) in 12 active CD patients before and after 4 weeks of anti-TNFαtreatment with adalimumab. At baseline, in inflamed mucosa we found increased gene expression of IL-17A and IL-22 but not IL-21 when compared to noninflamed mucosa. There were increased frequencies of IL-21-producing LP T cells but no differences in the frequencies of IL-17A- or IL-22-producing LP T cells when comparing inflamed versus noninflamed mucosa at baseline. There were no changes in the mucosal gene expression of IL-17A, IL-21, and IL-22 or the frequencies of IL-17A-, IL-21- and IL-22-producing LP T cell subsets between baseline and following 4 weeks of adalimumab initiation. Our results do not support the hypothesis that anti-TNFαtreatment has an early effect on the mucosal levels of IL-17A, IL-21, and IL-22 or LP T cell production of these cytokines in CD.

Summary The pathogenesis of sepsis involves a dual inflammatory response, with a hyperinflammatory phase followed by, or in combination with, a hypoinflammatory phase. The adhesion molecules lymphocyte function-associated antigen (LFA-1) (CD11a/CD18) and macrophage-1 (Mac-1) (CD11b/CD18) support leucocyte adhesion to intercellular adhesion molecules and phagocytosis through complement opsonization, both processes relevant to the immune response during sepsis. Here, we investigate the role of soluble (s)CD18 in sepsis with emphasis on sCD18 as a mechanistic biomarker of immune reactions and outcome of sepsis. sCD18 levels were measured in 15 septic and 15 critically ill non-septic patients. Fifteen healthy volunteers served as controls. CD18 shedding from human mononuclear cells was increased in vitro by several proinflammatory mediators relevant in sepsis. sCD18 inhibited cell adhesion to the complement fragment iC3b, which is a ligand for CD11b/CD18, also known as Mac-1 or complement receptor 3. Serum sCD18 levels in sepsis non-survivors displayed two distinct peaks permitting a partitioning into two groups, namely sCD18 ‘high’ and sCD18 ‘low’, with median levels of sCD18 at 2158 mU/ml [interquartile range (IQR) 2093–2811 mU/ml] and 488 mU/ml (IQR 360–617 mU/ml), respectively, at the day of intensive care unit admission. Serum sCD18 levels partitioned sepsis non-survivors into one group of ‘high’ sCD18 and low CRP and another group with ‘low’ sCD18 and high C-reactive protein. Together with the mechanistic data generated in vitro, we suggest the partitioning in sCD18 to reflect a compensatory anti-inflammatory response syndrome and hyperinflammation, respectively, manifested as part of sepsis.

Spondyloarthritis (SpA) is characterized by inflammation and new bone formation and can be treated by inhibition of the proinflammatory cytokines TNF-α and IL-17A. IL-26 is considered a proinflammatory cytokine, predominantly related to Th17 cells. In the present study, we investigate IL-26 expression in SpA patients, and examine the in vitro production of IL-26 by synovial cells and the effects of IL-26 on human osteoblasts. IL-26 was measured by ELISA in plasma and synovial fluid (SF) of 15 SpA patients and in plasma samples from 12 healthy controls. Facet joints from axial SpA patients were stained for IL-26 and analyzed by fluorescence microscopy. Synovial fluid mononuclear cells, C-C motif chemokine receptor 6 memory Th17 cells, and fibroblast-like synoviocytes (FLSs) were isolated, and supernatants were analyzed for IL-26 content by ELISA. FLSs were further stained for IL-26 production and the myofibroblast marker α-smooth-muscle-actin (αSMA) and analyzed by flow cytometry. Human osteoblasts were cultured in the presence of IL-26, and the degree of mineralization was quantified. We found that IL-26 levels in SF were increased compared with plasma (P < 0.0001). Moreover, IL-26 expression was found in facet joints of axial SpA patients within the bone marrow. IL-26 secretion was primarily found in αSMA(+) myofibroblasts. In contrast, Th17 cells did not produce detectable amounts of IL-26. Human osteoblasts treated with IL-26 showed increased mineralization compared with untreated osteoblasts (P = 0.02). In conclusion, IL-26 seems to be produced by myofibroblasts in the inflamed synovium and could be a possible facilitator of bone mineralization in SpA.KEY MESSAGES: IL-26 levels are higher in synovial fluid compared to plasma in spondyloarthritis. IL-26 was identified in axial facet joints of spondyloarthritis patients. Myofibroblasts from the spondyloarthritis synovium produce large amounts of IL-26. IL-26 induces bone mineralization in human osteoblasts.