Fig. 5

Systemic administration of PD-1-Fc to type II collagen (CII)-immunized mice accelerates arthritis onset, increases autoantibody production, and elevates Th1/Th17 responses. a Serum levels of anti-CII autoantibodies were determined by performing enzyme-linked immunosorbent assays in DBA/1 and collagen-induced arthritis (CIA) mice treated with phosphate-buffered saline or PD-1-Fc on day 50 after the second immunization. Values are the mean ± standard deviation (SD). *p < 0.05, **p < 0.01 versus controls. b Expression of T-bet, GATA3, RAR-related orphan receptor α (ROR-a), Bcl-6, interferon regulatory transcription factor (IRF)-6, interleukin (IL)-17 F, tumor necrosis factor (TNF)-α, interferon (IFN)-γ, and IL-4 was determined using real-time polymerase chain reactions in splenocytes obtained from each mouse and normalized to β-actin expression. Values are the mean ± SD (n = 3 samples per group) and are representative of at least three independent experiments with similar results. c and d Cells were isolated from the spleen on day 20 of the experiment shown in Fig. 4a and stimulated immediately with phorbol 12-myristate 13-acetate and ionomycin in the presence of GolgiStop before intracellular staining for IL-17A, IFN-γ, and IL-4. Cells were gated on the CD4⁺ T-cell population and analyzed by flow cytometry. Representative dot plots from four sets of similar results are shown. Bars show the mean and SD. *p < 0.05. PD-1 programmed cell death 1 protein, CXCR5 chemokine (C-X-C) motif receptor 5, ICOS inducible costimulatory molecule