Project description:This SuperSeries is composed of the following subset Series: GSE18023: Mice with FIA develop anti-native and anti-in vitro citrullinated fibrinogen antibodies GSE18024: FIA plasma induces arthritis in naïve mice GSE18025: Fibrinogen-reactive T cells transfer disease to naïve mice Refer to individual Series
Project description:Splenocytes harvested from mice with FIA were cultured in vitro and stimulated with 0.01 mg/ml fibrinogen for 3 days. Enriched T cells were transferred into 6-week-old naïve SJL mice, which developed visible signs of arthritis within 2 weeks. Synovial array profiling of plasma from the arthritic recipient mice demonstrated autoreactive B cell responses against peptides representing native fibrinogen, and further spreading of the responses to target collagen type V, cartilage gp39, and citrullinated vimentin. Custom-spotted protein slides were with plasma samples from individual mice. Four slides were probed with plasma derived from naïve mice and five slides were probed with plasma derived from mice injected with FIA T cells.
Project description:Splenocytes harvested from mice with FIA were cultured in vitro and stimulated with 0.01 mg/ml fibrinogen for 3 days. Enriched T cells were transferred into 6-week-old naïve SJL mice, which developed visible signs of arthritis within 2 weeks. Synovial array profiling of plasma from the arthritic recipient mice demonstrated autoreactive B cell responses against peptides representing native fibrinogen, and further spreading of the responses to target collagen type V, cartilage gp39, and citrullinated vimentin.
Project description:Synovial antigen arrays were probed with 1:150 dilutions of plasma derived from SJL mice immunized with fibrinogen emulsified in CFA or with CFA alone. Autoantibody binding was detected with a Cy3-conjugated goat-anti-mouse IgG/M secondary antibody. SAM was applied to identify antigens with statistically significant differences in array reactivity between FIA and CFA control plasma (q < 0.01) obtained from mice before boosting. The SAM hits were subjected to hierarchical cluster analysis and are displayed as a heatmap. Synovial array profiling of FIA plasma demonstrated autoreactive B-cell responses against peptides representing native fibrinogen, and B-cell epitope spreading resulting in additional targeting of citrullinated fibrinogen in the samples obtained before boosting.
Project description:Synovial antigen arrays were probed with 1:150 dilutions of plasma derived from SJL mice immunized with fibrinogen emulsified in CFA or with CFA alone. Autoantibody binding was detected with a Cy3-conjugated goat-anti-mouse IgG/M secondary antibody. SAM was applied to identify antigens with statistically significant differences in array reactivity between FIA and CFA control plasma (q < 0.01) obtained from mice before boosting. The SAM hits were subjected to hierarchical cluster analysis and are displayed as a heatmap. Synovial array profiling of FIA plasma demonstrated autoreactive B-cell responses against peptides representing native fibrinogen, and B-cell epitope spreading resulting in additional targeting of citrullinated fibrinogen in the samples obtained before boosting. Custom-spotted protein slides were probed with plasma samples from individual mice. Four slides were probed with plasma derived from mice immunized with CFA and six slides were probed with plasma derived from mice immunized with fibrinogen emulsified in CFA.
Project description:Plasma collected from mice with FIA were pooled, and 0.3 ml was injected intravenously into 6-week-old naïve SJL mice on days 0 and 2. Synovial antigen array profiling of plasma from the arthritic recipient mice demonstrated autoreactive B-cell responses against peptides representing native fibrinogen and citrullinated fibrinogen, and further epitope spreading resulting in additional targeting of fibronectin, collagen type V, cartilage gp39, and clusterin.
Project description:Plasma collected from mice with FIA were pooled, and 0.3 ml was injected intravenously into 6-week-old naïve SJL mice on days 0 and 2. Synovial antigen array profiling of plasma from the arthritic recipient mice demonstrated autoreactive B-cell responses against peptides representing native fibrinogen and citrullinated fibrinogen, and further epitope spreading resulting in additional targeting of fibronectin, collagen type V, cartilage gp39, and clusterin. Custom-spotted protein slides were probed with plasma samples from individual mice. Four slides were probed with plasma derived from naïve mice and four slides were probed with plasma derived from mice injected with FIA plasma.
Project description:To examine the effects of GSK199 treatment on antibody responses, a synovial antigen array analysis was performed using day 35 sera derived from mice with CIA treated with 30 mg/kg qd, 30 mg/kg bid GSK199 or saline. The synovial antigen arrays contain nearly 200 peptides and proteins, both citrullinated and uncitrullinated, representing candidate antigens in RA. Significance Analysis of Microarrays identified 7 and 14 autoantibody reactivities that were statistically decreased in CIA mice treated with 30 mg/kg qd or 30 mg/kg bid GSK199 respectively, compared to PBS treatment. By comparison, over 150 native and citrullinated epitopes on the arrays did not exhibit significant differences in reactivity. Sera from mice treated with 30 mg/kg GSK199 qd exhibited decreased IgG reactivity to native epitopes such as peptides from human fibrinogen alpha and beta chains and keratin while sera from mice treated with 30 mg/kg bid GSK199 demonstrated decreased IgG reactivity to native epitopes including collagen IV, heat shock protein 65, peptides from human fibrinogen alpha and beta chains (two of which match those that were decreased in the 30 mg/kg qd group), and vimentin as compared to saline controls (q < 0.1%). Decreased autoantibody reactivity to citrullinated peptides from vimentin and fibromodulin were also observed in sera from mice treated with 30 mg/kg bid GSK199 compared to controls treated with saline (q < 0.1%), but not by GSK199 30 mg/kg qd. There were no differences between treatment with GSK199 or saline alone in the levels of autoantibodies to 38 other citrullinated proteins in the array. These data demonstrated that GSK199 treatment decreases the development of autoantibodies in CIA and impairs epitope spreading to a degree similar to that observed with pan-PAD inhibition.
Project description:To identify the targets recognized by anti-carbamylated protein (anti-CarP) antibodies in patients with early Rheumatoid Arthritis (RA), to study the cross-reactivity between anti-CarP and anti-citrullinated protein antibodies (ACPA) and to evaluate their prognostic value. 331 patients (184 RA and 147 other rheumatisms) from the Very Early Arthritis (VErA) French cohort were analyzed. We performed mass spectrometry analysis of RA sera displaying anti-CarP activity and epitope mapping of the carbamylated fibrinogen γ chain to identify immunodominant peptides. The specificity of these targets was studied using competition assays with the major antigens recognized by ACPA. The prognostic value of anti-carbamylated fibrinogen IgG antibodies (ACa-Fib IgG) was compared to anti-CCP and anti-CarP using an in-house ELISA. Besides the α chain, the γ chain of fibrinogen, particularly one immunodominant epitope that has a specific reactivity, was identified as a circulating carbamylated target in sera. Prevalence of ACa-Fib was 37% at baseline and 10.9% for anti-CCP-negative RA. In anti-CCP-negative patients, ACa-Fib positivity was associated with a more inflammatory and erosive disease at baseline but not with rapid radiological progression, which remains strongly related to anti-CCP antibodies. Fibrinogen seems to be one of the antigens recognized in vivo by the anti-CarP response, particularly 2 epitopes of the γ chain, one of which is not cross reactive with ACPA. This specificity might be associated with a distinct clinical phenotype since ACa-Fib IgG were shown to be linked to systemic inflammation in very early RA but not to rapid radiological progression.