Angiogenesis is a major contributor to the development of inflammation during Rheumatoid arthritis (RA), as the vascularization of the pannus provides nutrients and oxygen for the infiltrating immune cells and proliferating synoviocytes. Tocilizumab (TCZ) is an anti-IL-6 receptor antibody that is used in the treatment of RA patients, and has been shown to exert anti-inflammatory effects. However, its effects on angiogenesis are not fully elucidated, and the molecular mechanisms regulating this effect are unknown.We evaluated the concentrations of several pro- and anti-angiogenic factors and the expression levels of several microRNA molecules that are associated with RA and angiogenesis in serum samples obtained from 40 RA patients, before and 4 months after the initiation of TCZ treatment. Additionally, we used an in vitro co-culture system of fibroblasts (the HT1080 cell line) and monocytes (the U937 cell line) to explore the mechanisms of TCZ action.Serum samples from RA patients treated with TCZ exhibited reduced circulating levels of EMMPRIN/CD147, enhanced expression of circulating miR-146a-5p and miR-150-5p, and reduced the angiogenic potential as was manifested by the lower number of tube-like structures that were formed by EaHy926 endothelial cell line. In vitro, the accumulation in the supernatants of the pro-angiogenic factors EMMPRIN, VEGF and MMP-9 was increased by co-culturing the HT1080 fibroblasts and the U937 monocytes, while the accumulation of the anti-angiogenic factor thrombospondin-1 (Tsp-1) and the expression levels of miR-146a-5p were reduced. Transfection of HT1080 cells with the miR-146a-5p mimic, decreased the accumulation of EMMPRIN, VEGF and MMP-9. When we neutralized EMMPRIN with a blocking antibody, the supernatants derived from these co-cultures displayed reduced migration, proliferation and tube formation in the functional assays.Our findings implicate miR-146a-5p in the regulation of EMMPRIN and propose that TCZ affects angiogenesis through its effects on EMMPRIN and miR-146a-5p.
Objective Anti–citrullinated protein antibodies (ACPAs) are characteristic of rheumatoid arthritis (RA). However, their presence years before the onset of clinical RA is perplexing. Although multiple putative citrullinated antigens have been identified, no studies have demonstrated the specific capacity of these antigens to initiate inflammatory arthritis. This study was undertaken to recapitulate the transition from preclinical to clinical RA and to demonstrate the capacity of local citrullination to facilitate this transition. Methods We performed proteomic analysis of activated human neutrophils to identify citrullinated proteins, including those targeted as part of the RA immune response. Using enzyme‐linked immunosorbent assay, we compared RA and osteoarthritis synovial fluid for levels of citrullinated histone H2B and its immune complex. Using macrophage activation assays, we assessed the effect of histone citrullination on immunostimulatory capacity and evaluated the stimulatory capacity of native and citrullinated H2B immune complexes. Finally, we assessed the potential for anti–citrullinated H2B antibodies to mediate arthritis in vivo. Results We identified robust targeting of neutrophil‐derived citrullinated histones by the ACPA immune response. More than 90% of the RA patients had anti–citrullinated H2B antibodies. Histone citrullination increased innate immunostimulatory capacity, and immune complexes containing citrullinated histones activated macrophage cytokine production and propagated neutrophil activation. Finally, we demonstrated that immunization with H2B was arthritogenic, but only in the setting of underlying articular inflammation. Conclusion Our findings indicate that citrullinated histones, specifically citrullinated H2B, are an antigenic target of the ACPA immune response. Furthermore, local generation of citrullinated antigen during low‐grade articular inflammation provides a mechanistic model for the conversion from preclinical autoimmunity to inflammatory arthritis.
Abstract Objective To identify and characterize human T cells reactive with heterogeneous nuclear RNP A2 (hnRNP A2) antigen, and to determine the ability of hnRNP‐reactive T cells to assist in the production of human autoantibodies. Methods T cells from patients with high serum levels of anti‐hnRNP IgG autoantibody were stimulated with an hnRNP recombinant fusion protein, and the cells were cloned by limiting dilution. The surface phenotype and cytokine profiles of the T cells were examined by flow cytometry and enzyme‐linked immunosorbent assay (ELISA), respectively. T cell clones were cultured with highly purified autologous B cells, and the ability of T cells to enhance autoantibody production under a variety of conditions was measured by ELISA. Results Human T cells reactive with hnRNP antigen were cloned from 2 patients with systemic lupus erythematosus (SLE) and 1 patient with mixed connective tissue disease (MCTD). The T cells were CD4+ and had a Th1‐like functional phenotype. In coculture in vitro with autologous B cells, T cell clones augmented anti‐hnRNP autoantibody production and did so without the need for direct T cell–B cell contact. Conclusion This study provides direct evidence for a role of anti‐hnRNP–reactive T cells in autoantibody production in SLE and MCTD. These findings support the notion that hnRNP‐reactive T cells play a role in the pathogenesis of these diseases.
Patients with rheumatoid arthritis (RA) are at increased risk of cardiovascular disease. Dyslipidemia is a known adverse effect of tocilizumab (TCZ), an anti-interleukin-6 receptor antibody used in RA treatment. We aimed to assess the effect of TCZ on lipid profile and adipokine levels in RA patients. Height, weight, disease activity scores, lipid profile and atherogenic indices (AI), leptin, adiponectin, resistin, interleukin-6, and high-sensitivity C-reactive protein (CRP) were measured before and four months after initiation of TCZ in 40 RA patients and 40 healthy controls. Following TCZ treatment, total cholesterol, high density lipoprotein (HDL), and triglycerides were significantly elevated, but no significant changes in weight, body mass index (BMI), low density lipoprotein (LDL), and AI were observed. Compared with controls, significantly higher adiponectin levels were measured in the RA group at baseline. Following TCZ treatment, resistin levels and the leptin-to-adiponectin ratio increased, adiponectin levels decreased, and leptin levels remained unchanged. No correlation was found between the change in adipokine serum levels and changes in the disease activity indices, nor the lipid profile. In conclusion, the changes observed suggest a protective role for TCZ on the metabolic and cardiovascular burden associated with RA, but does not provide a mechanistic explanation for this phenomenon.
Abstract BackgroundAngiogenesis is an important contributor to the development of Rheumatoid arthritis (RA). Tocilizumab (TCZ), an anti-IL-6 receptor antibody, is used in the treatment of RA patients, and has been shown to exert anti-inflammatory effects. However, its effects on angiogenesis are not fully elucidated, and the molecular mechanisms regulating this effect are unknown. MethodsWe evaluated the concentrations of several pro- and anti-angiogenic factors and the expression levels of several microRNA molecules that are associated with RA and angiogenesis in serum samples obtained from 40 RA patients, before and 4 months after the initiation of TCZ treatment. Additionally, we used an in vitro co-culture system of fibroblasts (the HT1080 cell line) and monocytes (the U937 cell line) to explore the mechanisms of TCZ action. ResultsSerum samples from RA patients treated with TCZ exhibited reduced levels of EMMPRIN/CD147, enhanced expression of miR-146a-5p and miR-150-5p, and reduced angiogenesis as was manifested by the reduced number of tube-like structures formed by EaHy926 endothelial cell line. In vitro , the accumulation of the pro-angiogenic factors EMMRPIN, VEGF and MMP-9 in the supernatants was increased by co-culturing the HT1080 fibroblasts and the U937 monocytes, while the accumulation of the anti-angiogenic factor thrombospondin-1 (Tsp-1) and the expression levels of miR-146a-5p were reduced. Transfection of HT1080 cells with the miR-146a-5p mimic, decreased the accumulation of EMMPRIN, VEGF and MMP-9. When EMMPRIN was neutralized with a blocking antibody, supernatants derived from these co-cultures exhibited reduced migration, proliferation and tube formation in functional assays. ConclusionsOur findings implicate miR-146a-5p in the regulation of EMMPRIN and propose that TCZ affects angiogenesis through its effects on EMMRPIN and miR-146a-5p.
The data on the risk of herpes zoster (HZ) in spondyloarthropathy (SpA) patients are sparse, especially regarding its association with the novel mRNA COVID-19 vaccines and immunosuppressants. We aimed to evaluate whether SpA diagnosis and/or immunosuppressant use affect HZ risk and the influence of mRNA COVID-19 vaccination. We assessed the association between SpA (psoriatic arthritis (PsA) and ankylosing spondylitis (AS)) diagnoses and HZ in a large population database with patients matched by age and sex to controls. We also assessed the association between the COVID-19 vaccine and new-onset HZ using two nested case–control studies, identifying all new HZ cases diagnosed from 1 January–31 December 2021 within the SpA and general population cohorts, matched randomly by sex, age and HZ index date to controls without HZ. Exposure to mRNA COVID-19 vaccination was ascertained in the 6 weeks prior to the index date both in cases and controls. In our results, the incidence rate of HZ was higher in PsA patients vs. the general population, at 1.03 vs. 0.64 per 100 person-years, respectively (adjusted HR = 1.55; 95%CI, 1.19–2.02). Within the SpA group, Jak-I treatment was associated with a higher risk of developing new-onset HZ (adjusted OR = 3.79; 1.15–12.5). Multivariable conditional logistic regression models we used showed no association between COVID-19 vaccination and new-onset HZ among the SpA patients (OR = 1.46; 0.68–3.14).
