Journal: Science Advances

Article Title: Inflammatory arthritis irAE may represent a unique autoimmune disease primarily driven by T cells but likely not autoantibodies

doi: 10.1126/sciadv.aea4262

Figure Lengend Snippet: ( A to G ) PBMCs from the patients with irAE were cultured in the plate coated with anti-CD3 and anti-CD28 (10 μg/ml) in the presence of IgG1 isotype control or anti–human IL-6R (50 μg/ml), anti–human IL-12p40 (50 μg/ml), and anti–human IFNAR1 (50 μg/ml) for 3 days; n = 9. (A) Expression of CD38 and CD127 on CD8 + T cells. Right: Percentage of CD38 + CD127 − CD8 + T cells. (B and C) CD8 + T cells activated for 5 days were restimulated with PMA, ionomycin, and monensin for 5 hours. (B) Expression of granzyme B and IFN-γ. Right: Percentage of granzyme B + IFN-γ + CD8 + T cells. (C) Expression of perforin and IFN-γ. Right: Percentage of perforin + IFN-γ + CD8 + T cells. [(D) and (E)] MFIs of GluCy5 (D), TMRM, and MTDR (E) in CD8 + T cells were presented. [(F) and (G)] CD4 + T cells activated for 5 days were restimulated with PMA, ionomycin, and monensin for 5 hours. (F) Expression of IL-21 and IL-2. Right: Percentage of IL-21 + IL-2 + CD4 + T cells. (G) Expression of CD38 and CXCR5 on CD4 + T cells. Right: Percentage of CD38 + CXCR5 − CD4 + T cells. Data in graphs represent mean ± SEM. Significance was tested paired Student’s t test [(A) to (G)].

Article Snippet: PBMCs from patients with irAE were thawed and rested in a complete medium for 1 hour and then activated with plate-coated anti–human CD3 and anti–human CD28 (10 μg/ml) with IgG1 isotype control (Bio X Cell, catalog no. CP174) or a combination of anti–human IL-6R 50 μg/ml; Bio X Cell, catalog no. SIM0014), anti–human IL-12p40 (50 μg/ml; Bio X Cell, catalog no. SIM0020), and anti–human IFNAR1 (50 μg/ml; Bio X Cell, catalog no. SIM0022) for 3 days.

Techniques: Cell Culture, Control, Expressing

Journal: European Journal of Immunology

Article Title: CD4 + T‐Cell‐Intrinsic IL‐6 Is Critical for Th17 Differentiation and Dampened Responsiveness to CD8 + T Cell‐Mediated Suppression

doi: 10.1002/eji.70150

Figure Lengend Snippet: Th17 cultures containing IL‐21+ TGF‐β1 (Th17.d) develop independently of T cell–intrinsic IL‐6 . WT or Il6 KO CD4 + T cells were differentiated under Th17.d conditions for 7 days, and Il17A expression was assessed by ELISA (A) and qPCR (B). Responsiveness to CD8 + T‐cell‐mediated suppression was evaluated in coculture assays (C). In parallel, WT cells were cultured with or without anti–IL6R antibody and subjected to the same functional readouts (D–F). Data are from three independent experiments ( n = 6 per group); each point represents an individual donor. Error bars indicate SEM. Statistical analysis was performed using paired or unpaired Student's t ‐test (* p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001; ** p ≤ 0.0001).

Article Snippet: IL6/STAT3 Pathway Blockade: CD4+ naive T‐cells were isolated, treated with human anti‐IL6R alpha antibody (R&D Systems, MAB227‐100) to block the IL‐6 receptor, and differentiated into Th17 subtypes in the presence of human recombinant cytokines as stated above.

Techniques: Expressing, Enzyme-linked Immunosorbent Assay, Cell Culture, Functional Assay