Children with extended oligoarticular and polyarticular juvenile idiopathic arthritis have alterations in B and T follicular cell subsets in peripheral blood and a cytokine profile sustaining B cell activation.

OBJECTIVES: The main goal of this study was to characterise the frequency and phenotype of B, T follicular helper (Tfh) and T follicular regulatory (Tfr) cells in peripheral blood and the cytokine environment present in circulation in children with extended oligoarticular juvenile idiopathic arthritis (extended oligo JIA) and polyarticular JIA (poly JIA) when compared with healthy controls, children with persistent oligoarticular JIA (persistent oligo JIA) and adult JIA patients. METHODS: Blood samples were collected from 105 JIA patients (children and adults) and 50 age-matched healthy individuals. The frequency and phenotype of B, Tfh and Tfr cells were evaluated by flow cytometry. Serum levels of APRIL, BAFF, IL-1ß, IL-2, IL-4, IL-6, IL-10, IL-17A, IL-21, IL-22, IFN-γ, PD-1, PD-L1, sCD40L, CXCL13 and TNF were measured by multiplex bead-based immunoassay and/or ELISA in all groups included. RESULTS: The frequency of B, Tfh and Tfr cells was similar between JIA patients and controls. Children with extended oligo JIA and poly JIA, but not persistent oligo JIA, had significantly lower frequencies of plasmablasts, regulatory T cells and higher levels of Th17-like Tfh cells in circulation when compared with controls. Furthermore, APRIL, BAFF, IL-6 and IL-17A serum levels were significantly higher in paediatric extended oligo JIA and poly JIA patients when compared with controls. These immunological alterations were not found in adult JIA patients in comparison to controls. CONCLUSIONS: Our results suggest a potential role and/or activation profile of B and Th17-like Tfh cells in the pathogenesis of extended oligo JIA and poly JIA, but not persistent oligo JIA.

B-cell phenotype and IgD-CD27- memory B cells are affected by TNF-inhibitors and tocilizumab treatment in rheumatoid arthritis.

BACKGROUND: The use of TNF-inhibitors and/or the IL-6 receptor antagonist, tocilizumab, in rheumatoid arthritis (RA) have pleiotropic effects that also involve circulating B-cells. The main goal of this study was to assess the effect of TNF-inhibitors and tocilizumab on B-cell phenotype and gene expression in RA. METHODS: Blood samples were collected from untreated early RA (ERA) patients, established RA patients under methotrexate treatment, established RA patients before and after treatment with TNF-inhibitors and tocilizumab, and healthy donors. B-cell subpopulations were characterized by flow cytometry and B-cell gene expression was analyzed by real-time PCR on isolated B-cells. Serum levels of BAFF, CXCL13 and sCD23 were determined by ELISA. RESULTS: The frequency of total CD19+ B cells in circulation was similar between controls and all RA groups, irrespective of treatment, but double negative (DN) IgD-CD27- memory B cells were significantly increased in ERA and established RA when compared to controls. Treatment with TNF-inhibitors and tocilizumab restored the frequency of IgD-CD27- B-cells to normal levels, but did not affect other B cell subpopulations. TACI, CD95, CD5, HLA-DR and TLR9 expression on B-cells significantly increased after treatment with either TNF-inhibitors and/ or tocilizumab, but no significant changes were observed in BAFF-R, BCMA, CD69, CD86, CXCR5, CD23, CD38 and IgM expression on B-cells when comparing baseline with post-treatment follow-ups. Alterations in B-cell gene expression of BAFF-R, TACI, TLR9, FcγRIIB, BCL-2, BLIMP-1 and ß2M were found in ERA and established RA patients, but no significant differences were observed after TNF-inhibitors and tocilizumab treatment when comparing baseline and follow-ups. Serum levels of CXCL13, sCD23 and BAFF were not significantly affected by treatment with TNF-inhibitors and tocilizumab. CONCLUSIONS: In RA patients, the use of TNF-inhibitors and/ or tocilizumab treatment affects B-cell phenotype and IgD-CD27- memory B cells in circulation, but not B-cell gene expression levels.

BAFF and TACI gene expression are increased in patients with untreated very early rheumatoid arthritis.

OBJECTIVE: B cells play important roles in rheumatoid arthritis (RA). Given the beneficial effect of B cell depletion therapy in RA as well as the observed alterations in B cell subpopulations in this disease, we evaluated whether changes in the expression of genes related to B cell survival and activation were already present in patients with untreated very early RA (VERA; < 6 weeks of disease duration). METHODS: The expression of a group of B cell-related activation and survival genes was quantified in peripheral blood mononuclear cells from patients with VERA by real-time PCR and compared with untreated early RA (< 1 year), established treated RA, and other untreated early arthritis conditions. Serum B cell-activating factor belonging to the tumor necrosis factor family (BAFF) was quantified by ELISA. RESULTS: BAFF gene expression and serum levels were highest in patients with VERA. The expression of BAFF receptor (BAFF-R) increased with disease progression, while transmembrane activator and calcium modulator and cyclophilin ligand interactor (TACI) was elevated since the first weeks of RA onset. Paired box 5 gene expression was also increased at all RA stages. Chemokine (C-X-C motif) receptor 5 was elevated only in established RA. No differences were observed in B cell maturation antigen, activation-induced cytidine deaminase, B lymphocyte-induced maturation protein, and B cell lymphoma 2 expression. CONCLUSION: Disturbances in the expression of B cell-related activation and survival genes, particularly BAFF and TACI, occur from the onset of RA and precede changes in BAFF-R. These alterations can lead to the development of autoreactive B cells from the first weeks of RA onset.

Cytokine pattern in very early rheumatoid arthritis favours B-cell activation and survival.

OBJECTIVES: B cells play an important role in the perpetuation of RA, particularly as autoantibody-producing cells. The ICs that further develop deposit in the joints and aggravate the inflammatory process. However, B-cell contribution in the very early stage of the disease remains unknown. The main goal of this work was to determine the concentration of cytokines potentially relevant for B-cell activation in serum from very early polyarthritis patients, with <6 weeks of disease duration, who latter on evolved into very early RA (VERA). METHODS: A proliferation-inducing ligand (APRIL), B-cell activating factor (BAFF) and IL-21 levels were measured by ELISA in the serum of VERA, other very early arthritis (VEA), established RA patients and controls. SF samples of established RA were also analysed. RESULTS: VERA patients have higher levels of APRIL and BAFF as compared with VEA, established RA and controls. Furthermore, APRIL and BAFF levels are also significantly elevated in RA-SF when compared with serum. CONCLUSIONS: The increased levels of APRIL and BAFF in VERA patients suggests that B-cell activation and the development of autoreactive B-cell responses might be crucial in early phases of RA. Therefore, APRIL and BAFF could be promising targets for therapy in the early phase of RA.

B-cell-activating factor receptor expression on naive and memory B cells: relationship with relapse in patients with rheumatoid arthritis following B-cell depletion therapy.

OBJECTIVES: To examine the expression of B-cell-activating factor receptor (BAFF-R) on naive CD27- and memory CD27+ B cells in normal individuals and patients with rheumatoid arthritis (RA) undergoing B-cell depletion therapy with rituximab. PATIENTS AND METHODS: BAFF-R expression on B-cell subsets was determined in normal controls (NC; n = 11), active patients with RA pre-rituximab (pre-RX; n = 15), relapsing patients either concordant for B-cell repopulation (C-R, n = 13) or discordant, with relapse more than 3 months after repopulation (D-R, n = 11) and patients in remission over 3 months postrepopulation (discordant non-relapsing (D-NR), n = 5). Serum BAFF was measured by ELISA and analysed using Mann-Whitney. RESULTS: There was no significant difference between NC, pre-RX and D-NR patients in %BAFF-R-positive B cells or mean fluorescence intensity (MFI) in naive and memory B cells. Relapsing patients had significantly lower MFI and %BAFF-R-positive cells in both naive and memory compartments from NC and pre-RX (C-R and D-R; p < 0.01). BAFF levels in pre-RX patients were within the normal range and did not correlate with BAFF-R expression in any patient group. D-NR patients had relatively lower proportions of pre and postswitch CD27+ B cells than pre-RX patients (D-NR vs pre-RX; p < 0.05 for both) and also lower numbers of postswitch B cells than D-R patients (D-NR vs D-R, p < 0.05). CONCLUSION: BAFF-R expression was significantly reduced on both naive and memory B cells in patients at relapse, regardless of the relationship with B-cell repopulation or serum BAFF levels. Re-establishment of active disease was also associated with an increase in class-switch recombination. Factors responsible for lower levels of BAFF-R may relate to altered thresholds for autoreactive B-cell generation at relapse in patients with RA.

Alterations on peripheral blood B-cell subpopulations in very early arthritis patients.

OBJECTIVE: To characterize circulating B-cell subpopulations of arthritis patients with <6 weeks of disease duration. METHODS: Peripheral blood samples were collected from very early untreated polyarthritis patients, with <6 weeks of disease duration, for flow cytometric evaluation of B-cell subpopulations. Samples from patients who were later diagnosed as RA [very early RA (VERA)] were also collected 4-6 weeks after starting a low dose of prednisone (5-10 mg) and 4 months after reaching the minimum effective dose of MTX. A matched healthy group was used as a control. RESULTS: VERA patients have a lower percentage of total peripheral blood memory B cells (CD19(+)CD27(+)) and a significant decrease in the frequency of circulating pre-switch memory B cells (CD19(+)IgD(+)CD27(+)) as compared with controls. Therapy with corticosteroids or MTX was unable to restore the normal frequencies of these B-cell subpopulations. A significant decrease in peripheral pre-switch memory B cells is equally observed in other early arthritis patients. Furthermore, no significant differences are found in the frequencies of CD4(+) and CD8(+) T cells in all patient groups. CONCLUSIONS: In very early polyarthritis patients, there is a reduction in circulating pre-switch memory B cells. The reasons that may account for this effect are still unknown. Short-term corticosteroids and MTX do not seem to have a direct effect on circulating B-cell subpopulations in VERA patients.