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( A to C ) Human naïve B cells were isolated from PBMCs and cultured under indicated conditions for 7 days; pembrolizumab or isotype control IgG4 (IgG4 Iso) was added on day 2; n = 8. (A) Expression of CD38 and CD27 on B cells. Right: Percentages of CD27 + CD38 − and CD27 + CD38 + B cells. (B) Expression of CD138 on CD27 + CD38 + B cells. Right: Percentage of CD138 + CD27 + CD38 + B cells. (C) Different immunoglobulin isotypes were measured in the culture supernatant of (A) and (B) by multiplex assay. ( D to F ) B cells from Humanized PD-1 (HuPD-1) mice were isolated and cultured with LPS, <t>IL-4,</t> BAFF, or ODN 2006, anti-IgM, IL-21, IL-4 or R848, anti-IgM, anti-CD40, IL-21, and IFN-γ for 3 days; pembrolizumab or isotype control was added on day 1. (D) Expression of IgG2c on activated B cells. FSC-H, Forward scatter height. Right: Percentage of IgG2c + B cells; n = 5. (E) Expression of IgG1 on activated B cells. Right: Percentage of IgG1 + B cells; n = 5. (F) Different immunoglobulin isotypes in the supernatant were measured by multiplex assay; n = 5. Data in graphs represent mean ± SEM. Significance was tested by two-way ANOVA.
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( A to C ) Human naïve B cells were isolated from PBMCs and cultured under indicated conditions for 7 days; pembrolizumab or isotype control IgG4 (IgG4 Iso) was added on day 2; n = 8. (A) Expression of CD38 and CD27 on B cells. Right: Percentages of CD27 + CD38 − and CD27 + CD38 + B cells. (B) Expression of CD138 on CD27 + CD38 + B cells. Right: Percentage of CD138 + CD27 + CD38 + B cells. (C) Different immunoglobulin isotypes were measured in the culture supernatant of (A) and (B) by multiplex assay. ( D to F ) B cells from Humanized PD-1 (HuPD-1) mice were isolated and cultured with LPS, <t>IL-4,</t> BAFF, or ODN 2006, anti-IgM, IL-21, IL-4 or R848, anti-IgM, anti-CD40, IL-21, and IFN-γ for 3 days; pembrolizumab or isotype control was added on day 1. (D) Expression of IgG2c on activated B cells. FSC-H, Forward scatter height. Right: Percentage of IgG2c + B cells; n = 5. (E) Expression of IgG1 on activated B cells. Right: Percentage of IgG1 + B cells; n = 5. (F) Different immunoglobulin isotypes in the supernatant were measured by multiplex assay; n = 5. Data in graphs represent mean ± SEM. Significance was tested by two-way ANOVA.
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( A to C ) Human naïve B cells were isolated from PBMCs and cultured under indicated conditions for 7 days; pembrolizumab or isotype control IgG4 (IgG4 Iso) was added on day 2; n = 8. (A) Expression of CD38 and CD27 on B cells. Right: Percentages of CD27 + CD38 − and CD27 + CD38 + B cells. (B) Expression of CD138 on CD27 + CD38 + B cells. Right: Percentage of CD138 + CD27 + CD38 + B cells. (C) Different immunoglobulin isotypes were measured in the culture supernatant of (A) and (B) by multiplex assay. ( D to F ) B cells from Humanized PD-1 (HuPD-1) mice were isolated and cultured with LPS, IL-4, BAFF, or ODN 2006, anti-IgM, IL-21, IL-4 or R848, anti-IgM, anti-CD40, IL-21, and IFN-γ for 3 days; pembrolizumab or isotype control was added on day 1. (D) Expression of IgG2c on activated B cells. FSC-H, Forward scatter height. Right: Percentage of IgG2c + B cells; n = 5. (E) Expression of IgG1 on activated B cells. Right: Percentage of IgG1 + B cells; n = 5. (F) Different immunoglobulin isotypes in the supernatant were measured by multiplex assay; n = 5. Data in graphs represent mean ± SEM. Significance was tested by two-way ANOVA.

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 C ) Human naïve B cells were isolated from PBMCs and cultured under indicated conditions for 7 days; pembrolizumab or isotype control IgG4 (IgG4 Iso) was added on day 2; n = 8. (A) Expression of CD38 and CD27 on B cells. Right: Percentages of CD27 + CD38 − and CD27 + CD38 + B cells. (B) Expression of CD138 on CD27 + CD38 + B cells. Right: Percentage of CD138 + CD27 + CD38 + B cells. (C) Different immunoglobulin isotypes were measured in the culture supernatant of (A) and (B) by multiplex assay. ( D to F ) B cells from Humanized PD-1 (HuPD-1) mice were isolated and cultured with LPS, IL-4, BAFF, or ODN 2006, anti-IgM, IL-21, IL-4 or R848, anti-IgM, anti-CD40, IL-21, and IFN-γ for 3 days; pembrolizumab or isotype control was added on day 1. (D) Expression of IgG2c on activated B cells. FSC-H, Forward scatter height. Right: Percentage of IgG2c + B cells; n = 5. (E) Expression of IgG1 on activated B cells. Right: Percentage of IgG1 + B cells; n = 5. (F) Different immunoglobulin isotypes in the supernatant were measured by multiplex assay; n = 5. Data in graphs represent mean ± SEM. Significance was tested by two-way ANOVA.

Article Snippet: Condition 1: LPS (3 μg/ml; Sigma-Aldrich, catalog no. L2880-25MG) and recombinant mouse IL-4 (10 ng/ml; R&D Systems, catalog no. 404-ML-100/CF) plus recombinant human BAFF (20 ng/ml; BioLegend, catalog no. 7449534).

Techniques: Isolation, Cell Culture, Control, Expressing, Multiplex Assay