Journal: medRxiv

Article Title: Inflammatory arthritis immune related adverse events represent a unique autoimmune disease entity primarily driven by T cells, but likely not autoantibodies

doi: 10.1101/2025.06.06.25328991

Figure Lengend Snippet: Pembrolizumab does not substantially affect B cell activation and antibody production ex vivo (A-C) Human naïve B cells were isolated from PBMCs and cultured under indicated conditions for 7 days; Pembrolizumab or isotype control IgG4 was added on Day 2, n = 8. (A) Expression of CD38 and CD27 on B cells. Right, percentages of CD27 + CD38 - , 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-B) by multiplex assay. (D-F) B cells from HuPD-1 mice were isolated and cultured with LPS, IL4, BAFF, or ODN2006, anti-IgM, IL-21, IL-4 or R848, anti-IgM, anti-CD40, IL-21, IFN-γ for 3 days, pembrolizumab or isotype control was added on Day 1. (D) Expression of IgG2c on activated B cells. 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 of were measured by multiplex assay, n = 5. Data in graphs represent mean ± SEM, Significance was tested by Two-way ANOVA.

Article Snippet: Condition 1: 2.5 μg/mL anti-human Ig (M + G + A) (Jackson Immunoresearch, Cat# 109-006-064), 2.5 μg/mL CpG ODN (Invivogen, Cat# tlrl-2006-1), 10 μg/mL anti-human CD40 (BioXcell, Cat# BE0189), 20 ng/mL rhIL-21 (Peprotech, Cat# 200-21-50UG), 10 ng/mL rhIL-4 (Biolegend, Cat# 574004), 10 ng/mL rhIL-2 (Peprotech, Cat#200-02-250UG).

Techniques: Activation Assay, Ex Vivo, Isolation, Cell Culture, Control, Expressing, Multiplex Assay

Journal: medRxiv

Article Title: Inflammatory arthritis immune related adverse events represent a unique autoimmune disease entity primarily driven by T cells, but likely not autoantibodies

doi: 10.1101/2025.06.06.25328991

Figure Lengend Snippet: PD-1 inhibition does not have substantial impact on B cells ex vivo (A) Human naïve B cells were isolated from healthy donor PBMCs and stimulated with different conditions; Pembrolizumab (Keytruda) or isotype control IgG4 was added into the culture media on day 2, mean fluorescence intensity (MFI) of CD86 on B cells was measured, n = 8. (B-F) B cells were isolated from HuPD-1 mice, labeled with CTV, and cultured under 3 different conditions: LPS, rmIL-4, BAFF or Anti-IgM, CpG ODN, rmIL-21, rmIL-4, 100 rhIL-2 or R848, anti-CD40, anti-IgM, rmIL-21, rmIFN-γ for 3 days. Pembrolizumab or isotype control IgG4 was added on day 1, n = 5. (B) Summary of human PD-1 MFI on B cells from different groups. (C) Expression of CD138 on activated B cells. Right, percentage of CD138 + B cells. (D) Summaries of activation marker CD69 and CD86 MFIs on B cells from different groups. (E) Summaries of CD71 and CD98 MFIs on B cells from different groups. (F) Representative flow cytometry plot of CTV dilution on B cells. Right, a summary of the proliferation index of B cells from different groups. Data in graphs represent mean ± SEM, Significance was tested by Two-way ANOVA.

Article Snippet: Condition 1: 2.5 μg/mL anti-human Ig (M + G + A) (Jackson Immunoresearch, Cat# 109-006-064), 2.5 μg/mL CpG ODN (Invivogen, Cat# tlrl-2006-1), 10 μg/mL anti-human CD40 (BioXcell, Cat# BE0189), 20 ng/mL rhIL-21 (Peprotech, Cat# 200-21-50UG), 10 ng/mL rhIL-4 (Biolegend, Cat# 574004), 10 ng/mL rhIL-2 (Peprotech, Cat#200-02-250UG).

Techniques: Inhibition, Ex Vivo, Isolation, Control, Fluorescence, Labeling, Cell Culture, Expressing, Activation Assay, Marker, Flow Cytometry

Journal: medRxiv

Article Title: Inflammatory arthritis immune related adverse events represent a unique autoimmune disease entity primarily driven by T cells, but likely not autoantibodies

doi: 10.1101/2025.06.06.25328991

Figure Lengend Snippet: Inflammatory signatures enriched in irAE patients reduce antibody production (A-F) Beads based multiplex assays were used to measure plasma concentration of IL-6, and IL-12p70 (A), TNF-α, IFN-γ and IL-1β (B), HC (n = 19), irAE (n = 34), RAC (n = 45), ICI (n = 9). IP-10 (CXCL10), CXCL11, and CXCL9 (C, HC, n = 17; irAE, n = 33; RAC, n = 46; ICI, n = 17), CCL20 (D), CX3CL1 (E), and CCL2 (F). (G-J) Human naïve B cells were isolated and cultured with 0.5 μg/mL anti-human CD40, 2.5 μg/mL anti-human Ig (M+G+A), and 20 ng/mL rhIL-21 with 100 ng/mL IFN-α, 100 ng/mL IL-6, 100 ng/mL IL-12, control, or the combination of IFN-α, IL-6, and IL-12 for 7 days. Cells and culture supernatants were analyzed. (G) Representative flow plot of CD38 and CD138 expression on CD27 hi CD38 hi ASCs. Right, a summary of the percentage of CD138 + ASCs, n = 6. (H) Expression of CD11c and CD27 on CD27 - IgD - ASCs. Right, percentage of CD11c + IgD - CD27 - B cells, n = 6. (I) Expression of active-caspase-3 in B cells. Right, percentage of active-caspase-3 + B cells from different groups, n = 3. (J) Different immunoglobulin isotype levels in the culture supernatants from G-H were measured by the multiplex assay, n = 6. Data in graphs represent mean ± SEM, Significance was tested by One-way ANOVA (A-I), and paired Student’s t-test (J).

Article Snippet: Condition 1: 2.5 μg/mL anti-human Ig (M + G + A) (Jackson Immunoresearch, Cat# 109-006-064), 2.5 μg/mL CpG ODN (Invivogen, Cat# tlrl-2006-1), 10 μg/mL anti-human CD40 (BioXcell, Cat# BE0189), 20 ng/mL rhIL-21 (Peprotech, Cat# 200-21-50UG), 10 ng/mL rhIL-4 (Biolegend, Cat# 574004), 10 ng/mL rhIL-2 (Peprotech, Cat#200-02-250UG).

Techniques: Multiplex Assay, Clinical Proteomics, Concentration Assay, Isolation, Cell Culture, Control, Expressing

Journal: eLife

Article Title: Molecular tracking devices quantify antigen distribution and archiving in the murine lymph node

doi: 10.7554/eLife.62781

Figure Lengend Snippet: ( a ) Bone marrow- derived dendritic cells (BMDCs) were grown for 7 days and cultured in GM-CSF. After 7 days, 5 mg of either pDNA, psDNA, ova-pDNA, or ova-psDNA were added to the culture media, and 1, 3, or 7 days after addition, media was removed and qPCR was performed on using primers against the DNA added. 3–5 wells were evaluated per group on 2–3 independent occasions. ( b ) As in ( a ) except with murine lymph node lymphatic endothelial cells. ( c ) As in Figure 1h, I , BMDCs were treated with ovalbumin (ova) (5 μg) ± polyI:C and anti-CD40 (20 μg each) or ova-psDNA ± polyI:C and anti-CD40 overnight and then co-cultured with Carboxyfluorescein Succinimidyl Ester (CFSE)-labeled OT1 T cells for 3 days before evaluating CFSE dilution by flow cytometry. Experiments were performed three times with 3–5 wells per sample with similar results.

Article Snippet: Antibody , Anti-mouse CD40 (Rat monoclonal) , BioXcell , Cat#BE0016-2 , .

Techniques: Derivative Assay, Cell Culture, Labeling, Flow Cytometry

Journal: eLife

Article Title: Molecular tracking devices quantify antigen distribution and archiving in the murine lymph node

doi: 10.7554/eLife.62781

Figure Lengend Snippet:

Article Snippet: Antibody , Anti-mouse CD40 (Rat monoclonal) , BioXcell , Cat#BE0016-2 , .

Techniques: Recombinant