Dysregulated endothelial cell markers in systemic lupus erythematosus: a systematic review and meta-analysis.

OBJECTIVES: To perform a systematic literature review and meta-analysis on endothelial cell (EC) markers that are involved and dysregulated in systemic lupus erythematosus (SLE) in relation to disease activity, as EC dysregulation plays a major role in the development of premature atherosclerosis in SLE. METHODS: Search terms were entered into Embase, MEDLINE, Web of Science, Google Scholar and Cochrane. Inclusion criteria were 1) studies published after 2000 reporting measurements of EC markers in serum and/or plasma of SLE patients (diagnosed according to ACR/SLICC criteria), 2) English language peer reviewed articles, and 3) disease activity measurement. For meta-analysis calculations, the Meta-Essentials tool by Erasmus Research Institute and of Management (ERIM) was used. Only those EC markers, which were 1) reported in at least two articles and 2) reported a correlation coefficient (i.e. Spearman's rank or Pearson's) between the measured levels of the EC marker and disease activity were included. For meta-analyses, a fixed effect model was used. RESULTS: From 2133 hits, 123 eligible articles were selected. The identified SLE-related endothelial markers were involved in EC activation, EC apoptosis, disturbed angiogenesis, defective vascular tone control, immune dysregulation and coagulopathy. Meta-analyses of primarily cross-sectional studies showed significant associations between marker levels and disease activity for the following endothelial markers: Pentraxin-3, Thrombomodulin, VEGF, VCAM-1, ICAM-1, IP-10 and MCP-1. Dysregulated EC markers without associations with disease activity were: Angiopoeitin-2, vWF, P-Selectin, TWEAK and E-Selectin. CONCLUSIONS: We provide a complete literature overview for dysregulated EC markers in SLE comprising a wide range of different EC functions. SLE-induced EC marker dysregulation was seen with, but also without, association with disease activity. This study provides some clarity in the eminent complex field of EC markers as biomarkers for SLE. Longitudinal data on EC markers in SLE are now needed to guide us more in unravelling the pathophysiology of premature atherosclerosis and cardiovascular events in SLE patients.

Longitudinal humoral response after SARS-CoV-2 vaccination in ocrelizumab treated MS patients: To wait and repopulate?

OBJECTIVE: The objective of this study was to measure humoral responses after SARS-CoV-2 vaccination in MS patients treated with ocrelizumab (OCR) compared to MS patients without disease modifying therapies (DMTs) in relation to timing of vaccination and B-cell count. METHODS: OCR treated patients were divided into an early and a late group (cut-off time 12 weeks between infusion and first vaccination). Patients were vaccinated with mRNA-1273 (Moderna). B-cells were measured at baseline (time of first vaccination) and SARS-CoV-2 antibodies were measured at baseline, day 28, 42, 52 and 70. RESULTS: 87 patients were included (62 OCR patients, 29 patients without DMTs). At day 70, seroconversion occurred in 39.3% of OCR patients compared to 100% of MS patients without DMTs. In OCR patients, seroconversion varied between 26% (early group) to 50% (late group) and between 27% (low B-cells) to 56% (at least 1 detectable B-cell/µL). CONCLUSIONS: Low B-cell counts prior to vaccination and shorter time between OCR infusion and vaccination may negatively influence humoral response but does not preclude seroconversion. We advise OCR treated patients to get their first vaccination as soon as possible. In case of an additional booster vaccination, timing of vaccination based on B-cell count and time after last infusion may be considered.

Spinning straw into gold: description of a disruptive rheumatology research platform inspired by the COVID-19 pandemic.

Clinical research projects often use traditional methods in which data collection and signing informed consent forms rely on patients' visits to the research institutes. However, during challenging times when the medical community is in dire need of information, such as the current COVID-19 pandemic, it becomes more urgent to use digital platforms that can rapidly collect data on large numbers of patients. In the current manuscript, we describe a novel digital rheumatology research platform, consisting of almost 5000 patients with autoimmune diseases and healthy controls, that was set up rapidly during the COVID-19 pandemic, but which is sustainable for the future. Using this platform, uniform patient data can be collected via questionnaires and stored in a single database readily available for analysis. In addition, the platform facilitates two-way communication between patients and researchers, so patients become true research partners. Furthermore, blood collection via a finger prick for routine and specific laboratory measurements has been implemented in this large cohort of patients, which may not only be applicable for research settings but also for clinical care. Finally, we discuss the challenges and potential future applications of our platform, including supplying tailored information to selected patient groups and facilitation of patient recruitment for clinical trials.

A Dutch consensus statement on the diagnosis and treatment of ANCA-associated vasculitis.

INTRODUCTION: Despite the availability of several guidelines on the diagnosis and treatment of antineutrophil cytoplasmic antibody-associated vasculitis (AAV), clinical routine practice will only improve when an implementation strategy is in place to support clinical decision making and adequate implementation of guidelines. We describe here an initiative to establish national and multidisciplinary consensus on broad aspects of the diagnosis and treatment of AAV relevant to daily clinical practice in the Netherlands. METHODS: A multidisciplinary working group of physicians in the Netherlands with expertise on AAV addressed the broad spectrum of diagnosis, terminology, and immunosuppressive and non-immunosuppressive treatment, including an algorithm for AAV patients. Based on recommendations from (inter)national guidelines, national consensus was established using a Delphi-based method during a conference in conjunction with a nationally distributed online consensus survey. Cut-off for consensus was 70% (dis)agreement. RESULTS: Ninety-eight professionals were involved in the Delphi procedure to assess consensus on 50 statements regarding diagnosis, treatment, and organisation of care for AAV patients. Consensus was achieved for 37/50 statements (74%) in different domains of diagnosis and treatment of AAV including consensus on the treatment algorithm for AAV. CONCLUSION: We present a national, multidisciplinary consensus on a diagnostic strategy and treatment algorithm for AAV patients as part of the implementation of (inter)national guideline-derived recommendations in the Netherlands. Future studies will focus on evaluating local implementation of treatment protocols for AAV, and assessments of current and future clinical practice variation in the care for AAV patients in the Netherlands.

Evidence for a pathogenic role of extrafollicular, IL-10-producing CCR6+B helper T cells in systemic lupus erythematosus.

Interleukin 10 (IL-10) is an antiinflammatory cytokine, but also promotes B cell responses and plays a pathogenic role in systemic lupus erythematosus (SLE). CD4+CCR6+IL-7R+T cells from human tonsils produced IL-10 following stimulation by naïve B cells, which promoted B cell immunoglobulin G (IgG) production. These tonsillar CCR6+B helper T cells were phenotypically distinct from follicular helper T (TFH) cells and lacked BCL6 expression. In peripheral blood, a CCR6+T cell population with similar characteristics was identified, which lacked Th17- and TFH-associated gene signatures and differentiation-associated surface markers. CD4+CCR6+T cells expressing IL-10, but not IL-17, were also detectable in the spleens of cytokine reporter mice. They provided help for IgG production in vivo, and expanded systemically in pristane-induced lupus-like disease. In SLE patients, CD4+CCR6+IL-7R+T cells were associated with the presence of pathogenic anti-dsDNA (double-stranded DNA) antibodies, and provided spontaneous help for autoantibody production ex vivo. Strikingly, IL-10-producing CCR6+T cells were highly abundant in lymph nodes of SLE patients, and colocalized with B cells at the margins of follicles. In conclusion, we identified a previously uncharacterized population of extrafollicular B helper T cells, which produced IL-10 and could play a prominent pathogenic role in SLE.

IL-10 producing regulatory and helper T-cells in systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is a highly heterogeneous autoimmune disease characterised by the production of pathogenic autoantibodies against nuclear self-antigens. The anti-inflammatory and tolerogenic cytokine Interleukin-10 appears to play a paradoxical pathogenic role in SLE and is therefore currently therapeutically targeted in clinical trials. It is generally assumed that the pathogenic effect of IL-10 in SLE is due to its growth and differentiation factor activity on autoreactive B-cells, but effects on other cells might also play a role. To date, a unique cellular source of pathogenic IL-10 in SLE has not been identified. In this review, we focus on the contribution of different CD4+T-cell subsets to IL-10 and autoantibody production in SLE. In particular, we discuss that IL-10 produced by different subsets of adaptive regulatory T-cells, follicular helper T-cells and extra-follicular B-helper T-cells is likely to have different effects on autoreactive B-cell responses. A better understanding of the role of IL-10 in B-cell responses and lupus would allow to identify the most promising therapies for individual SLE patients in the future.

IgG4:IgG RNA ratio differentiates active disease from remission in granulomatosis with polyangiitis: a new disease activity marker? A cross-sectional and longitudinal study.

OBJECTIVES: An important limitation in granulomatosis with polyangiitis (GPA) is the lack of disease activity markers. Immunoglobulin G4-positive (IgG4+) B cells and plasma cells are implicated in the pathogenesis of GPA. We hypothesized that the presence of these cells in peripheral blood could serve as disease activity parameter in GPA. METHODS: We included 35 proteinase 3-antineutrophil cytoplasmic antibodies-positive patients with GPA in a cross-sectional study. Active disease was defined as Birmingham Vasculitis Activity Score (BVAS) ≥ 3 (n = 15), remission as BVAS of 0 (n = 17), and low disease activity (LDA) as BVAS of 1-2 and clinical remission (n = 3). Healthy subjects (n = 10), patients with systemic lupus erythematosus (n = 24), and patients with rheumatoid arthritis (n = 19) functioned as control subjects. An additional longitudinal study was performed in ten patients with GPA. Using a validated qPCR test, we measured the IgG4:IgG RNA ratio in all groups and compared the results with known biomarkers. RESULTS: The median qPCR score was higher in active GPA (21.4; IQR 12.1-29.6) than in remission/LDA (3.3; IQR 1.6-5.6) (Mann-Whitney U test, p < 0.0001) and outperformed other known disease activity parameters in detecting activity. A cutoff qPCR score of 11.2% differentiated active disease from remission/LDA accurately (AUC 0.993). The qPCR test correlated well with the BVAS (Spearman r = 0.77, p < 0.0001). In the longitudinal study, a decrease in BVAS correlated with qPCR score reduction (paired t test, p < 0.05). CONCLUSIONS: The IgG4:IgG RNA ratio in GPA accurately distinguishes active disease from remission and correlates well with disease activity in these single-center studies. If these results are confirmed in larger longitudinal studies, this test might help to steer treatment decisions in patients with GPA.

Endothelial cells: From innocent bystanders to active participants in immune responses.

The endothelium is crucially important for the delivery of oxygen and nutrients throughout the body under homeostatic conditions. However, it also contributes to pathology, including the initiation and perpetuation of inflammation. Understanding the function of endothelial cells (ECs) in inflammatory diseases and molecular mechanisms involved may lead to novel approaches to dampen inflammation and restore homeostasis. In this article, we discuss the various functions of ECs in inflammation with a focus on pathological angiogenesis, attraction of immune cells, antigen presentation, immunoregulatory properties and endothelial-to-mesenchymal transition (EndMT). We also review the current literature on approaches to target these processes in ECs to modulate immune responses and advance anti-inflammatory therapies.

Ticking the right boxes: classification of patients suspected of Lyme borreliosis at an academic referral center in the Netherlands.

To provide better care for patients suspected of having Lyme borreliosis (LB) we founded the Amsterdam Multidisciplinary Lyme borreliosis Center (AMLC). The AMLC reflects a collaborative effort of the departments of internal medicine/infectious diseases, rheumatology, neurology, dermatology, medical microbiology and psychiatry. In a retrospective case series, characteristics of 200 adult patients referred to the AMLC were recorded, and patients were classified as having LB, post-treatment LB syndrome (PTLBS), persistent Borrelia burgdorferi sensu lato (s.l.) infection despite antibiotic treatment or no LB. In addition, LB, PTLBS and persistent B. burgdorferi s.l. infection cases were classified as 'definite,' 'probable' or 'questionable.' Of the 200 patients, 120 (60%) did not have LB and 31 (16%) had a form of localized or disseminated LB, of which 12 were classified as definite, six as probable and 13 as questionable. In addition, 34 patients (17%) were diagnosed with PTLBS, of which 22 (11%) were probable and 12 (6%) questionable. A total of 15 patients (8%) were diagnosed with persistent B. burgdorferi s.l. infection, of which none was classified as definite, three as probable and 12 as questionable. In conclusion, in line with previous studies, the number of definite and probable (persisting) LB cases was low. The overall high number of questionable cases illustrates the fact that it can sometimes be challenging to either rule out or demonstrate an association with a B. burgdorferi s.l. infection, even in an academic setting. Finally, we were able to establish alternative diagnoses in a large proportion of patients.

Biologics for rare inflammatory diseases: TNF blockade in the SA PHO syndrome.

INTRODUCTION: SAPHO is an invalidating syndrome characterised by Synovitis, Acne, Pustulosis, Hyperostosis and Osteitis. The low prevalence and heterogeneous presentation often leads to a significant diagnostic delay. Here, we provide an up-to-date overview of current insights into the pathogenesis and different treatment options. In addition, we describe the effects of anti-TNF treatment in three refractory cases. CASE REPORTS: Patient A is a 25-year-old female with hidradenitis suppurativa, inflammatory back pain and painful joints. After diagnosis, anti-TNF treatment was started resulting in clinical improvement. Patient B is a 44-year-old woman who presented with acne, palmoplantar pustulosis and anterior chest wall pain. Bone scintigraphy showed increased uptake at the anterior chest wall. Treatment with bisphosphonates resulted in temporary improvement and subsequent treatment with anti-TNF induced long-term clinical improvement. Patient C is a 37-year-old woman with palmoplantar psoriasis, relapsing hidradenitis and inflammatory back pain. MRI revealed osteitis of the pubic bone. Anti-TNF was started for SAPHO syndrome. However, despite a clinical response, our patient discontinued treatment, resulting in rapid deterioration. Anti-TNF treatment was re-introduced followed by clinical improvement. CONCLUSION: These case reports illustrate, consistent with the current literature, that TNF blockers can be considered for treatment of refractory SAPHO syndrome.

Inhibition of forkhead box class O family member transcription factors in rheumatoid synovial tissue.

OBJECTIVE: Phosphatidylinositol 3-kinase-dependent activation of protein kinase B (PKB) has been observed in rheumatoid arthritis (RA) synovial tissue, and mechanisms that interfere with this process are protective in animal models of arthritis. PKB can regulate cell survival and proliferation via phosphorylation-dependent inactivation of forkhead box class O (FoxO) transcription factors. The present study was undertaken to examine whether FoxO transcription factors are differentially inactivated in RA synovial tissue, and whether this inactivation correlates with laboratory and clinical parameters of disease activity. METHODS: The expression and phosphorylation of FoxO family members were assessed in synovial biopsy tissue from 12 patients with RA and 9 patients with inflammatory osteoarthritis (OA), by immunohistochemistry and quantitative computer-assisted image analysis. Immunoblotting was used to assess the interleukin-1beta (IL-1beta)- and tumor necrosis factor alpha (TNFalpha)-induced phosphorylation of FoxO1 and FoxO4 in cultured fibroblast-like synoviocytes (FLS) and macrophages. RESULTS: FoxO1, FoxO3a, and FoxO4 were expressed and phosphorylated in synovial tissue from both RA patients and OA patients. In RA synovial tissue, phosphorylation of FoxO1 was observed in both FLS and macrophages, FoxO3a in T lymphocytes, and FoxO4 in macrophages alone. Following stimulation with IL-1beta and TNFalpha, FoxO1 and FoxO4 were phosphorylated in both RA and OA FLS and synovial macrophages, respectively. Inactivation of FoxO4 was significantly enhanced in the RA as compared with the OA synovial sublining. There was a strong negative correlation between inactivation of FoxO4 in RA synovial tissue and increased serum C-reactive protein levels and a raised erythrocyte sedimentation rate in RA patients. CONCLUSION: All 3 FoxO family members examined were phosphorylated in both RA and OA synovial tissue; in particular, inactivation of FoxO4 was significantly enhanced in macrophages from RA synovial tissue. Thus, cell-specific inactivation of FoxO family members appears to differentially regulate cell survival and proliferation in the RA synovium.

Macrophages from patients with SLE and rheumatoid arthritis have defective adhesion in vitro, while only SLE macrophages have impaired uptake of apoptotic cells.

BACKGROUND: It has been suggested that defective handling of apoptotic cells by macrophages plays a key role in the development of systemic lupus erythematosus (SLE). The relative contribution of intrinsic defects and serum factors remains controversial. OBJECTIVE: To compare monocytes from SLE patients, patients with rheumatoid arthritis, and healthy controls for their ability to differentiate in vitro into macrophages and to bind/engulf apoptotic cells. METHODS: Peripheral blood derived monocytes from healthy donors or from patients with SLE or rheumatoid arthritis were allowed to differentiate into macrophages. The in vitro uptake of apoptotic cells by macrophages was evaluated by a flow cytometry assay that allowed discrimination between binding and internalisation. RESULTS: Monocytes from SLE and rheumatoid patients showed a striking defect in adherence to plastic compared with healthy donors. Absence or heat inactivation of serum resulted in a reduction in the binding and engulfment of apoptotic cells by macrophages. Macrophages from rheumatoid and SLE patients had similar percentages of apoptotic cells bound to their surface compared with normal controls. However, macrophages from SLE patients showed a significant defect in the internalisation of apoptotic cells compared with those from healthy controls, even in the presence of normal human serum. CONCLUSIONS: Monocytes from patients with SLE and rheumatoid arthritis have a similar defect in their capacity to adhere to plastic. However, only macrophages from SLE patients showed an impaired ability to engulf apoptotic cells, which indicates that an intrinsic cellular defect may be responsible for this phenomenon.

Enhanced gene transfer to arthritic joints using adeno-associated virus type 5: implications for intra-articular gene therapy.

BACKGROUND: Gene therapy of the joint has great potential as a new therapeutic approach for the treatment of rheumatoid arthritis (RA). The vector chosen is of crucial importance for clinical success. OBJECTIVE: To investigate the tropism and transduction efficiency in arthritic joints in vivo, and in synovial cells in vitro, using five different serotypes of recombinant adeno-associated virus (rAAV) encoding beta-galactosidase or green fluorescent protein genes. METHODS: rAAV was injected into the ankle joints of rats with adjuvant arthritis after the onset of disease. Synovial tissue was examined at different time points for beta-galactosidase protein and gene expression by in situ staining and polymerase chain reaction (PCR) analysis, respectively. In addition, the ability of rAAV to transduce primary human fibroblast-like synoviocytes from patients with RA was investigated in vitro. RESULTS: Intra-articular injection of the rAAV5 serotype resulted in the highest synovial transduction, followed by much lower expression using rAAV2. Expression of the transgene was already detectable 7 days after injection and lasted for at least 4 weeks. Only background staining was seen for serotypes 1, 3, and 4. Importantly, there was a minimal humoral immune response to rAAV5 compared with rAAV2. Additionally, it was found that both rAAV2 and rAAV5 can efficiently transduce human fibroblast-like synoviocytes obtained from patients with RA. CONCLUSION: Intra-articular rAAV mediated gene therapy in RA might be improved by using rAAV5 rather than other serotypes.

Complement receptor 1/CD35 is a receptor for mannan-binding lectin.

Mannan-binding lectin (MBL), a member of the collectin family, is known to have opsonic function, although identification of its cellular receptor has been elusive. Complement C1q, which is homologous to MBL, binds to complement receptor 1 (CR1/CD35), and thus we investigated whether CR1 also functions as the MBL receptor. Radioiodinated MBL bound to recombinant soluble CR1 (sCR1) that had been immobilized on plastic with an apparent equilibrium dissociation constant of 5 nM. N-acetyl-d-glucosamine did not inhibit sCR1-MBL binding, indicating that the carbohydrate binding site of MBL is not involved in binding CR1. C1q inhibited MBL binding to immobilized sCR1, suggesting that MBL and C1q might bind to the same or adjacent sites on CR1. MBL binding to polymorphonuclear leukocytes (PMNs) was associated positively with changes in CR1 expression induced by phorbol myristate acetate. Finally, CR1 mediated the adhesion of human erythrocytes to immobilized MBL and functioned as a phagocytic receptor on PMNs for MBL-immunoglobulin G opsonized bacteria. Thus, MBL binds to both recombinant sCR1 and cellular CR1, which supports the role of CR1 as a cellular receptor for the collectin MBL.

Intercellular adhesion molecule 1 and beta2 integrins in C1q-stimulated superoxide production by human neutrophils: an example of a general regulatory mechanism governing acute inflammation.

OBJECTIVE: To investigate the role of intercellular adhesion molecule 1 (ICAM-1) and beta2 integrins in the production of superoxide (O2-) by C1q-stimulated human polymorphonuclear leukocytes (PMN). METHODS: PMN were pretreated with F(ab')2 fragments of monoclonal antibodies (mAb) that blocked or did not block beta2 integrin-mediated adhesion. The cells were added to wells coated with C1q, and the production of O2- was monitored kinetically as a color change due to reduction of cytochrome c. In some experiments, C1q was co-immobilized with purified ICAM-1. RESULTS: Blocking mAb to the shared beta2 integrin subunit, CD18, completely inhibited the O2- response triggered by immobilized C1q, while blocking mAb to the alpha subunits of the beta2 integrins each partially blocked the O2- response. PMN treated with C1q were found to activate the beta2 integrins lymphocyte function-associated antigen 1 and CR3 for binding to ICAM-1. Co-immobilization of ICAM-1 with C1q cooperatively triggered O2- production by PMN. CONCLUSION: beta2 integrin binding to an ICAM provided an essential costimulatory signal for O2-production triggered by C1q in PMN. Our findings suggest a model for PMN activation in which 2 stimuli are required for O2- production: a first signal that also activates PMN beta2 integrins, followed by a second, beta2 integrin-mediated signal, which occurs physiologically upon PMN binding to ICAM-1. The requirement for this dual signal for PMN generation of O2- would serve as a regulatory mechanism to limit the production of O2- to a tissue environment where C1q, or some other stimulus, is colocalized with stromal cells bearing up-regulated ICAM-1. This mechanism may explain why all tissues can express ICAM-1 and may explain in part why inhibitors of tumor necrosis factor alpha, a major physiologic stimulus of ICAM-1 up-regulation, are potent antiinflammatory agents.

C1q and C4b bind simultaneously to CR1 and additively support erythrocyte adhesion.

Previously, we showed that soluble C1q bound specifically to CR1 on transfected cells. If the CR1-C1q interaction were to participate in immune complex clearance, then this interaction should support E adhesion. Using a tip plate adhesion assay, we found that immobilized C1q mediated adhesion of human E. E binding to C1q was specifically inhibited by polyclonal anti-CR1 Fab fragments. Intact C1 was not efficient as an adherence ligand until it was treated with EDTA or the C1 inhibitor to remove the C1r2C1s2 complex from C1, leaving C1q. Titration of C1q alone, C4b alone, and C1q + C4b indicated that the two complement ligands were additive in their ability to support CR1-mediated adhesion of E. Analysis of binding to immobilized CR1 using a BIAcore instrument documented that C1q, C4b, and C3b binding were independent events. Additionally, C1q-dependent binding of immune complexes and heat-aggregated IgG to E was documented. These experiments confirm that the immune adherence receptor in humans, CR1, is the single receptor for all of the opsonic ligands of complement, provide evidence for a single C1q binding site on LHR-D of CR1, and suggest that C1q may participate in immune clearance.