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Objective: To assess the effects of tumour necrosis factor (TNF) antagonist therapy on B lymphocyte stimulator (BLyS) expression in patients with rheumatoid arthritis (RA)., Methods: Blood from 38 patients with RA from a single centre was collected prior to and following initiation of TNF antagonist therapy. Plasma BLyS protein levels, blood leukocyte BLyS mRNA levels and disease activity were longitudinally monitored. Twelve patients with RA who either refused or were felt not to be candidates for TNF antagonist therapy and five normal healthy volunteers served as TNF antagonist-naïve controls., Results: Baseline plasma BLyS protein levels, but not blood leukocyte BLyS mRNA levels, were elevated in patients with RA. Plasma BLyS protein levels declined following initiation of TNF antagonist therapy in good responders (GR) to TNF antagonist therapy but not in poor responders (PR). By contrast, the erythrocyte sedimentation rate (ESR) declined in response to TNF antagonist therapy in GR and PR. TNF antagonist therapy did not promote change in blood leukocyte BLyS mRNA levels in either GR or PR, suggesting that the TNF antagonist-associated changes in circulating BLyS protein levels reflected changes in local BLyS production in the affected joints rather than changes in systemic BLyS production. BLyS expression did not change over time in either the normal or RA control groups., Conclusions: A good clinical response to TNF antagonist therapy in patients with RA is associated with a decline in plasma BLyS protein levels. Increased BLyS expression in affected joints may contribute to ongoing disease activity, and reduction of such expression may help promote a favourable clinical response to TNF antagonist therapy.

Objective: To determine whether synovial fluid (SF) levels and cell-surface expression of B lymphocyte stimulator (BLyS) protein and SF levels of APRIL are elevated in patients with inflammatory arthritis (IA)., Methods: Same-day blood and SF samples from 89 patients with 103 knee effusions (81 knees with IA and 22 with noninflammatory arthritis [NIA]) were evaluated for BLyS protein and APRIL levels by enzyme-linked immunosorbent assay. Blood and SF mononuclear cells were double-stained for surface BLyS protein and surface CD14 (monocyte marker) and were analyzed by flow cytometry. Complete blood cell counts and SF nucleated cell counts were performed by the clinical hematology laboratory., Results: BLyS protein levels were higher in SF than in corresponding serum samples from both IA and NIA patients. SF BLyS protein levels, but not surface expression of BLyS protein, were disproportionately elevated in IA patients. APRIL levels were higher in SF than in corresponding serum samples from most IA patients but not NIA patients. SF BLyS protein and APRIL levels correlated with each other, and each correlated with SF monocyte, lymphocyte, neutrophil, and total nucleated cell counts. Although SF and serum BLyS protein levels correlated with each other, SF and serum APRIL levels did not, suggesting that SF BLyS protein levels are more dependent upon systemic factors than are SF APRIL levels. Moreover, in 8 patients who underwent sequential arthrocenteses, changes in SF BLyS protein levels did not immutably parallel changes in SF APRIL levels, indicating their differential regulation., Conclusion: BLyS protein and APRIL are locally produced in inflamed joints. Their respective SF levels are differentially regulated, suggesting that they serve different functions. Together, their local production may foster survival and/or expansion of B cells that produce pathogenic autoantibodies and/or promote local T cell activation and consequent joint destruction.

BLyS and APRIL are two members of the TNF superfamily that are secreted by activated myeloid cells and have costimulatory activity on B cells. BLyS and APRIL share two receptors, TACI and BCMA, whereas a third receptor, BAFF-R, specifically binds BLyS. Both BLyS and APRIL have been described as homotrimeric molecules, a feature common to members of the TNF superfamily. In this study, we show that APRIL and BLyS can form active heterotrimeric molecules when coexpressed and that circulating heterotrimers are present in serum samples from patients with systemic immune-based rheumatic diseases. These findings raise the possibility that active BLyS/APRIL heterotrimers may play a role in rheumatic and other autoimmune diseases and that other members of the TNF ligand superfamily may also form active soluble heterotrimers.

DR3 is a death domain-containing receptor that is upregulated during T cell activation and whose overexpression induces apoptosis and NF-kappaB activation in cell lines. Here we show that an endothelial cell-derived TNF-like factor, TL1A, is a ligand for DR3 and decoy receptor TR6/DcR3 and that its expression is inducible by TNF and IL-1alpha. TL1A induces NF-kappaB activation and apoptosis in DR3-expressing cell lines, while TR6-Fc protein antagonizes these signaling events. Interestingly, in T cells, TL1A acts as a costimulator that increases IL-2 responsiveness and secretion of proinflammatory cytokines both in vitro and in vivo. Our data suggest that interaction of TL1A with DR3 promotes T cell expansion during an immune response, whereas TR6 has an opposing effect.

B lymphocyte stimulator (BLyS) is a novel member of the TNF family of proteins expressed by myeloid cells as membrane-bound and soluble forms. BLyS was shown to act specifically on B cells, inducing proliferation and immunoglobulin production both in vitro and in vivo. The present study was undertaken to characterize binding of radiolabeled BLyS to its cognate receptor on human B lymphocytes and examine intracellular events initiated by BLyS binding. Similar to other TNF family members, BLyS is present in solution as a homotrimer as determined by gel filtration chromatography and light scattering analysis. BLyS binding to B cells is specific as other TNF family members tested did not compete for(125)I-BLyS binding. Analysis of equilibrium binding of(125)I-labeled BLyS to purified human tonsillar B cells demonstrated saturable binding. Scatchard analysis of the binding data revealed a single class of high-affinity binding on human B cells with approximately 2600 binding sites per cell and an apparent dissociation constant (K(D)) of about 0.1 nM. In addition we report that BLyS binding to B cells results in the activation of NF-kappaB and the Ets family transcription factor, ELF-1, and in the induction of mRNA for Polo-like kinase (PLK)., (Copyright 2001 Academic Press.)

B-lymphocyte stimulator (BLyS) is a recently identified novel member of the tumor necrosis factor ligand superfamily shown to exist in a membrane-bound and soluble form. BLyS was found to be specifically expressed on cells of myeloid lineage and to selectively stimulate B-lymphocyte proliferation and immunoglobulin production. The expression of a cytokine involved in potentiation of humoral immune responses, such as BLyS, is expected to be strictly controlled. The goal of the present study was to examine regulation of BLyS levels in monocytic cells in response to cytokines and during their differentiation to macrophages and dendritic cells. The presence of BLyS on the cell surface and in the culture medium of both normal blood monocytes and on tumor cells of myelomonocytic origin was demonstrated. BLyS gene expression and levels of membrane-associated and soluble BLyS were found to be regulated by cytokines, in particular interferon (IFN)-gamma and to a lesser extent interleukin-10 (IL-10). The expression of BLyS on monocyte membranes was retained following differentiation into macrophages, but detection on the surface of monocyte-derived dendritic cells required stimulation with IFN-gamma. Both IFN-gamma and IL-10 enhanced the release of soluble BLyS that was active in B-cell proliferation assays. Cells transfected with BLyS complementary DNA mutated in a predicted cleavage site failed to release BLyS into the culture medium, thereby suggesting that soluble BLyS was derived from the membrane form. These results provide further support for an important role for BLyS expressed in myeloid cells in B-cell expansion and antibody responses.

An expression cloning approach was employed to identify the receptor for B-lymphocyte stimulator (BLyS) and identified the tumor necrosis factor receptor superfamily member TACI as a BLyS-binding protein. Expression of TACI in HEK293T cells confers on the cells the ability to bind BLyS with subnanomolar affinity. Furthermore, a TACI-Fc fusion protein recognizes both the cleaved, soluble form of BLyS as well as the membrane BLyS present on the cell surface of a recombinant cell line. TACI mRNA is found predominantly in B-cells and correlates with BLyS binding in a panel of B-cell lines. We also demonstrate that TACI interacts with nanomolar affinity with the BLyS-related tumor necrosis factor homologue APRIL for which no clear in vivo role has been described. BLyS and APRIL are capable of signaling through TACI to mediate NF-kappaB responses in HEK293 cells. We conclude that TACI is a receptor for BLyS and APRIL and discuss the implications for B-cell biology.