Short- and long-term effects of anti-CD20 treatment on B cell ontogeny in bone marrow of patients with rheumatoid arthritis

It has been known for a long time that B cells play a role in rheumatoid arthritis (RA). By production of autoantibodies, presentation of auto-antigens and by producing cytokines B cells may contribute to the pathogenesis of RA. In recent years it has been shown that anti-B cell therapy is a powerful tool in the treatment of RA. The aim of this thesis was to a) investigate the effect on B cell ontogeny following B cell depletion therapy, b) during B cell depletion therapy evaluate serological and humoral immune responses and finally, c) try to establish a connection between Epstein-Barr virus (EBV) infection, CD25+ B cells and outcome of B cell deletion therapy. In paper I we could show that in bone marrow of RA patients following anti-CD20 treatment with rituximab (RTX) IgD expressing naïve cells are depleted whereas immature and memory B cells where still detectable. However, the long-term effects clearly showed a reduction of memory B cells in bone marrow. The examination of rheumatoid factor (RF) production revealed that RFs decline short after treatment but returned to baseline levels concurrently with the IgD expressing B cells when patients where subjected to an additional course. In paper II the cellular and humoral immune responses were evaluated by immunisation of RA patients before or during RTX treatment with a protein vaccine against influenza and a pneumococcal polysaccharide vaccine. The results suggest that both cellular and humoral immune responses are affected in patients receiving RTX treatment and we therefore suggest that immunisation should be performed before RTX treatment. In paper III we investigate the effects of EBV on selected B cell subsets and how infection may affect the clinical response to RTX treatment. The phenotypical study showed that B cells are more mature in EBV infected patients and the CD25+ B cell subset was more mature as compared to the CD25- B cell population. The evaluation of clinical response to RTX treatment with regard to B cell subsets showed that non-responding EBV+ patients had a significantly larger CD25+ plasma cell population. When investigating the effects of EBV stimulation in vitro we found that the CD25+ B cell population developed into antibody-producing cells to a higher extent than did the corresponding CD25- B cell population. The results of our studies indicate that that B cells play an essential role in the pathogenesis of RA. During RTX treatment we suggest that the IgD expressing population may harbour the autoantibody producing B cells. We also claim that that there are subsets of B cells (i.e. CD25+ B cells) that may have significant impact on the pathogenesis of RA, and the clinical outcome following RTX treatment

Survivin Measurement improves Clinical Prediction of Transition From Arthralgia to RA—Biomarkers to Improve Clinical Sensitivity of Transition From Arthralgia to RA

Background: Arthralgia often predates development of rheumatoid arthritis (RA). A set of joint symptoms commonly found in patients during their transition from arthralgia to RA, has been recently proposed.Aim: To combine clinical and serological markers and to improve recognition of imminent rheumatoid arthritis (RA) among patients with arthralgia.Methods: The total of 1,743 first-visit patients attending the rheumatology ward in Gothenburg for joint symptoms were identified during 12 consecutive months. Among those, 63 patients were classified as RA, 73 had undifferentiated arthritis and 180 had unexplained arthralgia. New RA cases, which prospectively developed during 48 months, comprised the preclinical (pre) RA group. The joint symptoms of the first-visit were analyzed aiming to distinguish patients with arthralgia and arthritis, and patients with pre-RA, who later developed the disease. The receiver operating characteristics curves were constructed. In the model, symptoms with the odds ratio >2.0 between the arthralgia and pre-RA were combined with information about RA-specific antibodies, C-reactive protein (CRP), and survivin in serum.Results: The proposed set of clinical symptoms distinguished the arthralgia patients from RA and pre-RA. Presence of survivin in serum showed strong association with clinical joint symptoms in arthralgia. A combination of symptoms in several small joint areas, increasing number of joints with symptoms, and patient's experience of swelling in small hand joints at the first visit identified pre-RA cases with 93% specificity. Grouping those symptoms with information about survivin, RA-specific antibodies, and CRP (or gender) in the final algorithm achieved 91% specificity and 55.2% of positive prediction for transition from arthralgia to RA.Conclusion: Clinical and serological parameters in combination aid recognition of imminent RA among arthralgia patients with appropriate sensitivity

Metabolic signature and proteasome activity controls synovial migration of CDC42hiCD14+ cells in rheumatoid arthritis

ObjectiveActivation of Rho-GTPases in macrophages causes inflammation and severe arthritis in mice. In this study, we explore if Rho-GTPases define the joint destination of pathogenic leukocytes, the mechanism by which they perpetuate rheumatoid arthritis (RA), and how JAK inhibition mitigates these effects.MethodsCD14+ cells of 136 RA patients were characterized by RNA sequencing and cytokine measurement to identify biological processes and transcriptional regulators specific for CDC42hiCD14+ cells, which were summarized in a metabolic signature (MetSig). The effect of hypoxia and IFN-γ signaling on the metabolic signature of CD14+ cells was assessed experimentally. To investigate its connection with joint inflammation, the signature was translated into the single-cell characteristics of CDC42hi synovial tissue macrophages. The sensitivity of MetSig to the RA disease activity and the treatment effect were assessed experimentally and clinically.ResultsCDC42hiCD14+ cells carried MetSig of genes functional in the oxidative phosphorylation and proteasome-dependent cell remodeling, which correlated with the cytokine-rich migratory phenotype and antigen-presenting capacity of these cells. Integration of CDC42hiCD14+ and synovial macrophages marked with MetSig revealed the important role of the interferon-rich environment and immunoproteasome expression in the homeostasis of these pathogenic macrophages. The CDC42hiCD14+ cells were targeted by JAK inhibitors and responded with the downregulation of immunoproteasome and MHC-II molecules, which disintegrated the immunological synapse, reduced cytokine production, and alleviated arthritis.ConclusionThis study shows that the CDC42-related MetSig identifies the antigen-presenting CD14+ cells that migrate to joints to coordinate autoimmunity. The accumulation of CDC42hiCD14+ cells discloses patients perceptive to the JAKi treatment

Inhibition of CCL3 abrogated precursor cell fusion and bone erosions in human osteoclast cultures and murine collagen-induced arthritis

Objective Macrophage inflammatory protein 1-alpha (CCL3) is a chemokine that regulates macrophage trafficking to the inflamed joint. The agonistic effect of CCL3 on osteolytic lesions in patients with multiple myeloma is recognized; however, its role in skeletal damage during inflammatory arthritis has not been established. The aim of the study was to explore the role of osteoclast-associated CCL3 upon bone resorption, and to test its pharmacological blockade for protecting against bone pathology during inflammatory arthritis. Methods CCL3 production was studied during osteoclast differentiation from osteoclast precursor cells: human CD14-positive mononuclear cells. Mice with CIA were treated with an anti-CCL3 antibody. The effect of CCL3 blockade through mAb was studied through osteoclast number, cytokine production and bone resorption on ivory disks, and in vivo through CIA progression (clinical score, paw diameter, synovial inflammation and bone damage). Results Over time, CCL3 increased in parallel with the number of osteoclasts in culture. Anti-CCL3 treatment achieved a concentration-dependent inhibition of osteoclast fusion and reduced pit formation on ivory disks (P ⩽ 0.05). In CIA, anti-CCL3 treatment reduced joint damage and significantly decreased multinucleated tartrate-resistant acid phosphatase-positive osteoclasts and erosions in the wrists (P < 0.05) and elbows (P < 0.05), while also reducing joint erosions in the hind (P < 0.01) and fore paws (P < 0.01) as confirmed by X-ray. Conclusion Inhibition of osteoclast-associated CCL3 reduced osteoclast formation and function whilst attenuating arthritis-associated bone loss and controlling development of erosion in murine joints, thus uncoupling bone damage from inflammation. Our findings may help future innovations for the diagnosis and treatment of inflammatory arthritis

Epstein–Barr virus in bone marrow of rheumatoid arthritis patients predicts response to rituximab treatment

Objectives. Viruses may contribute to RA. This prompted us to monitor viral load and response to anti-CD20 therapy in RA patients