Objectives The exact underlying mechanism of rituximab treatment in patients with RA is poorly defined and knowledge about the effect of B cell depletion on immune cells in secondary lymphoid organs is lacking. We analysed lymphoid tissue responses to rituximab in RA patients. Methods Fourteen RA patients received 2 × 1000 mg rituximab intravenously, and lymph node (LN) biopsies were obtained before and 4 weeks after the first infusion. Tissues were examined by flow cytometry, immunohistochemistry and quantitative PCR. LN biopsies from five healthy individuals (HC) served as controls. Results LN biopsies of RA patients showed increased frequencies of CD21 + CD23 + IgD high IgM variable follicular B cells and CD3 + CD25 + CD69 + early activated, tissue resident T cells when compared with HCs. After treatment, there was incomplete depletion of LN B cells. There was a significant decrease in CD27 − IgD + naïve B cells, and CD27 + IgD + unswitched memory B cells including the CD27 + IgD + IgM + subset and follicular B cells. Strikingly, CD27 + IgD − switched memory B cells persisted in LN biopsies after rituximab treatment. In the T cell compartment, a significant decrease was observed in the frequency of early activated, tissue resident T cells after rituximab treatment, but late activated T cells persisted. B cell proliferation inducing cytokine IL-21 was higher expressed in LN biopsies of RA patients compared with HC and expression was not affected by rituximab treatment. Conclusion Rituximab does not cure RA, possibly due to persistence of switched memory B cells in lymphoid tissues suggesting that factors promoting B cell survival and differentiation need to be additionally targeted.

Objective Innate lymphoid cells (ILCs) are emerging mediators of immunity, and accumulation of inflammatory ILC populations can occur in inflammatory‐mediated conditions. Since early lymph node (LN) activation has been shown in rheumatoid arthritis (RA), we aimed to investigate the frequency and distribution of ILCs in LN biopsy specimens obtained during the earliest phases of RA. Methods Twelve patients with early RA, 12 individuals with IgM rheumatoid factor and/or anti–citrullinated protein antibodies without arthritis (RA risk group), and 7 healthy controls underwent ultrasound‐guided inguinal LN biopsy. ILC subsets and the expression of vascular cell adhesion molecule (VCAM) and intercellular adhesion molecule (ICAM) by LN endothelial cells and fibroblasts were analyzed by flow cytometry. Results Although no differences in the frequencies of total ILCs (Lin−CD45 +/low CD127+) were found, the distribution of the ILC subpopulations differed among groups. RA patients showed lower numbers of lymphoid tissue–inducer (LTi) cells (c‐Kit+NKp44− ILCs) and increased ILC1 (c‐Kit−NKp44− ILCs) and ILC3 (c‐Kit+NKp44+ ILCs) numbers compared with controls ( P < 0.001, P < 0.050, and P < 0.050, respectively). Individuals at risk of RA exhibited an increased frequency of ILC1 compared with controls ( P < 0.01). LTi cells paralleled the expression of adhesion molecules on endothelial cells and fibroblasts. Conclusion Our findings indicate that during the at‐risk and earliest phases of RA, the ILC distribution in LN changes from a homeostatic profile toward a more inflammatory profile, thereby providing evidence of a role for ILCs in RA pathogenesis.

BackgroundSystemic autoimmunity can be present years before clinical onset of rheumatoid arthritis (RA). Adaptive immunity is initiated in lymphoid tissue where lymph node stromal cells (LNSCs) regulate immune responses through their intimate connection with leucocytes. We postulate that malfunctioning of LNSCs creates a microenvironment in which normal immune responses are not properly controlled, possibly leading to autoimmune disease. In this study we established an experimental model for studying the functional capacities of human LNSCs during RA development.MethodsTwenty-four patients with RA, 23 individuals positive for autoantibodies but without clinical disease (RA risk group) and 14 seronegative healthy control subjects underwent ultrasound-guided inguinal lymph node (LN) biopsy. Human LNSCs were isolated and expanded in vitro for functional analyses. In analogous co-cultures consisting of LNSCs and peripheral blood mononuclear cells, αCD3/αCD28-induced T-cell proliferation was measured using carboxyfluorescein diacetate succinimidyl ester dilution.ResultsFibroblast-like cells expanded from the LN biopsy comprised of fibroblastic reticular cells (gp38+CD31−) and double-negative (gp38−CD31−) cells. Cultured LNSCs stably expressed characteristic adhesion molecules and cytokines. Basal expression of C-X-C motif chemokine ligand 12 (CXCL12) was lower in LNSCs from RA risk individuals than in those from healthy control subjects. Key LN chemokines C-C motif chemokine ligand (CCL19), CCL21 and CXCL13 were induced in LNSCs upon stimulation with tumour necrosis factor-α and lymphotoxin α1β2, but to a lesser extent in LNSCs from patients with RA. The effect of human LNSCs on T-cell proliferation was ratio-dependent and altered in RA LNSCs.ConclusionsOverall, we developed an experimental model to facilitate research on the role of LNSCs during the earliest phases of RA. Using this innovative model, we show, for the first time to our knowledge, that the LN stromal environment is changed during the earliest phases of RA, probably contributing to deregulated immune responses early in disease pathogenesis.Electronic supplementary materialThe online version of this article (10.1186/s13075-018-1529-8) contains supplementary material, which is available to authorized users.