B lymphocytes from patients with systemic lupus erythematosus (SLE) exhibit defective intracellular signaling, hyperactivity, and autoantibody production. Recent evidence indicates a reduced expression of Lyn kinase, a negative regulator of B cell signaling, and reduced translocation of Lyn into membrane signaling domains in SLE. The present study was undertaken to investigate the causes of this altered regulation of Lyn by assessing the expression levels of regulatory molecules and their translocation into the signaling domains of SLE B lymphocytes.

Splice variants of CD74 differentially modulate the activity of cathepsin L (CTSL). As CD74 and CTSL participate in the pathogenesis of inflammatory diseases such as rheumatoid arthritis (RA), we determined whether splice variants of CD74 could be biomarkers of disease activity. Gene expression was measured in mice with collagen-induced arthritis using quantitative PCR (qPCR). In vitro studies using murine macrophage/DC-lineage cells determined the relative influence of macrophage phenotype on isoform expression and the potential to produce CTSL in response to TNF. CD74 splice variants were measured in human RA synovium and RA patients' monocytes. In arthritic mice, the expression of the p41 CD74 isoform was significantly higher in severely affected paws compared with unaffected paws or the paws of naïve mice; the p41 isoform significantly correlated with the expression of TNF in arthritic paws. Compared with M2-like macrophages, M1-like macrophages expressed increased levels of CD74 and had higher expression, secretion and activity of CTSL. RA patients that responded to TNF blockade had significantly higher expression levels of CD74 in circulating monocytes after treatment, compared with non-responders. The expression of the human CD74 isoform a was significantly higher in RA synovia, compared with osteoarthritis synovia, and was associated with CSTL enzymatic activity. This study is the first to demonstrate differential expression of the CD74 p41 isoform in an auto-immune disorder and in response to therapy. The differential expression of CD74 splice variants indicates an association, and potentially a mechanistic role, in the pathogenesis of RA.

The role of the innate immune system has been established in the initiation and perpetuation of inflammatory disease, but less attention has been paid to its role in the resolution of inflammation and return to homeostasis. Toll-like receptor (TLR) expression profiles were analysed in tissues with differing disease status in rheumatoid arthritis (RA), ankylosing spondylitis (AS), and in experimental arthritis. TLR gene expression was measured in whole blood and monocytes, before and after TNF blockade. In RA and osteoarthritis synovia, the expression of TLRs was quantified by standard curve qPCR. In addition, four distinct stages of disease were defined and validated in collagen-induced arthritis (CIA), the gold standard animal model for RA - pre-onset, early disease, late disease and immunised mice that were resistant to the development of disease. TLR expression was measured in spleens, lymph nodes, blood cells, liver and the paws (inflamed and unaffected). In RA whole blood, the expression of TLR1, 4 and 6 was significantly reduced by TNF blockade but the differences in TLR expression profiles between responders and non-responders were less pronounced than the differences between RA and AS patients. In RA non-responders, monocytes had greater TLR2 expression prior to therapy compared to responders. The expression of TLR1, 2, 4 and 8 was higher in RA synovium compared to control OA synovium. Circulating cytokine levels in CIA resistant mice were similar to naïve mice, but anti-collagen antibodies were similar to arthritic mice. Distinct profiles of inflammatory gene expression were mapped in paws and organs with differing disease status. TLR expression in arthritic paws tended to be similar in early and late disease, with TLR1 and 2 moderately higher in late disease. TLR expression in unaffected paws varied according to gene and disease status but was generally lower in resistant paws. Disease status-specific profiles of TLR expression were observed in spleens, lymph nodes, blood cells and the liver. Notably, TLR2 expression rose then fell in the transition from naïve to pre-onset to early arthritis. TLR gene expression profiles are strongly associated with disease status. In particular, increased expression in the blood precedes clinical manifestation.

CD24(high)CD38(high) transitional B cells represent cells at a key stage in their developmental pathway. In addition, these B cells have been widely ascribed regulatory functions and involvement in the control of chronic inflammatory diseases. However, the phenotypic and functional overlap between these cells and regulatory B cells remains controversial.

Genome-wide association studies have identified various susceptibility variants and loci associated with incidence of rheumatoid arthritis (RA) in different populations. One of these is T cell activation Rho GTPase activating protein (TAGAP). The present study sought to measure the expression of TAGAP in RA patients, CD4+ T cells subsets from healthy humans and in mice with collagen-induced arthritis. Peripheral blood mononuclear cells (PBMC) from RA patients and tissues of arthritic mice at different stages of the disease were used for the evaluation of TAGAP mRNA expression. Increased TAGAP expression was observed in RA patients compared to healthy controls, and there were differences in the expression level of TAGAP in the tissues of mice with experimental arthritis. Gene expression in CD4+ T cells from healthy humans was greatest 4 h after activation and protein expression was greatest after 24 h. The expression of TAGAP was not correlated with CD4+ lymphocyte subsets which were enriched for functionally defined subsets (Th17, Treg, Th1), further indicating its utility as an indicator of lymphocyte activation. These findings indicate that increased TAGAP expression is a distinguishing feature of inflammatory disease and further highlight the role of TAGAP in RA susceptibility.

To evaluate the efficacy of orally-administered alendronate compared with intravenously-administered zoledronate.

Biologic TNFα inhibitors are a mainstay treatment option for patients with rheumatoid arthritis (RA) refractory to other treatment options. However, many patients either do not respond or relapse after initially responding to these agents. This study was carried out to identify biomarkers that can distinguish responder from non-responder patients before the initiation of treatment. The level of cytokines in plasma and those produced by ex vivo T cells, B cells and monocytes in 97 RA patients treated with biologic TNFα inhibitors was measured before treatment and after 1 and 3 months of treatment by multiplex analyses. The frequency of T cell subsets and intracellular cytokines were determined by flow cytometry. The results reveal that pre-treatment, T cells from patients who went on to respond to treatment with biologic anti-TNFα agents produced significantly more GM-CSF than non-responder patients. Furthermore, immune cells from responder patients produced higher levels of IL-1β, TNFα and IL-6. Cytokine profiling in the blood of patients confirmed the association between high levels of GM-CSF and responsiveness to biologic anti-TNFα agents. Thus, high blood levels of GM-CSF pre-treatment had a positive predictive value of 87.5% (61.6 to 98.5% at 95% CI) in treated RA patients. The study also shows that cells from most anti-TNFα responder patients in the current cohort produced higher levels of GM-CSF and TNFα pre-treatment than non-responder patients. Findings from the current study and our previous observations that non-responsiveness to anti-TNFα is associated with high IL-17 levels suggest that the disease in responder and non-responder RA patients is likely to be driven/sustained by different inflammatory pathways. The use of biomarker signatures of distinct pro-inflammatory pathways could lead to evidence-based prescription of the most appropriate biological therapies for different RA patients.