Journal: European Journal of Immunology

Article Title: Termination of CD40L co‐stimulation promotes human B cell differentiation into antibody‐secreting cells

doi: 10.1002/eji.202249972

Figure Lengend Snippet: Termination of CD40L stimulation results in downregulation of NF‐kB and pSTAT3/STAT3 signaling Naïve B cells were sorted and 25.000 cells were stimulated with a human‐CD40L expressing 3T3 feeder layer and recombinant IL‐21 (50 ng/ml). After 48 hours CD40L blocking antibodies (13 ug/ml) were added or not. At indicated timepoints, cultures were harvested, stained for membrane markers, fixed, and stored until day 6 when all samples were stained intracellularly and analyzed by flow cytometry. ( A ) Schematic representation of CD40L and IL‐21 signaling pathways. ( B ) Representative histogram overlay (left) and quantification (right) of activated NF‐κB p65 GeoMFI per day under indicated conditions ( n = 4). ( C ) Representative histogram overlay (left) and quantification (right) of STAT3 GeoMFI per day under indicated conditions ( n = 4). ( D ) Representative histogram overlay (left) and quantification (right) of p‐STAT3 GeoMFI per day under indicated conditions ( n = 4). p ‐Values were calculated by comparing the red CD40L + IL‐21 condition and the blue CD40L stimulation terminated after 48 h condition, using two‐way ANOVA with Sidak's multiple comparison test Combined data from two independent experiments, each performed with two donors. Symbols represent the mean and error bars show ± SEM. * p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001, **** p ≤ 0.0001.

Article Snippet: Samples were then stained with anti‐CD27 BUV395 (BD), anti‐NF‐κB p65 (pS529) PE‐Cy7 (BD), anti‐STAT3 Percp‐Cy5.5 (BD), anti‐p‐STAT3 PE (BD), and anti‐C‐MYC AF647 (CST) diluted in PBS/0.1% BSA.

Techniques: Expressing, Recombinant, Blocking Assay, Staining, Membrane, Flow Cytometry, Comparison

Journal: European Journal of Immunology

Article Title: Termination of CD40L co‐stimulation promotes human B cell differentiation into antibody‐secreting cells

doi: 10.1002/eji.202249972

Figure Lengend Snippet: Schematic overview of CD40L stimulated and CD40L stimulation terminated B cells with phospho‐signaling and transcription factor profiles . Stimulation of CD40 by CD40L activates the NF‐κB pathway which in turn upregulates the expression of C‐MYC and IRF4. IL‐21 stimulation results in the phosphorylation of STAT3 which in turn can upregulate the expression of BLIMP1. Continuous CD40L stimulation results in an increased expression of NF‐κB, C‐MYC, IRF4, and PAX5 while preventing differentiation into ASCs. Upon termination of CD40L stimulation, NF‐κB, C‐MYC, and PAX5 expression is downregulated while expression of IRF4, BLIMP1, and XBP‐1s is increased, resulting in efficient ASC differentiation.

Article Snippet: Samples were then stained with anti‐CD27 BUV395 (BD), anti‐NF‐κB p65 (pS529) PE‐Cy7 (BD), anti‐STAT3 Percp‐Cy5.5 (BD), anti‐p‐STAT3 PE (BD), and anti‐C‐MYC AF647 (CST) diluted in PBS/0.1% BSA.

Techniques: Expressing

Journal: Frontiers in Immunology

Article Title: Expansion of effector memory Vδ2 neg γδ T cells associates with cytomegalovirus reactivation in allogeneic stem cell transplant recipients

doi: 10.3389/fimmu.2024.1397483

Figure Lengend Snippet: Differentiation of γδ T cells and subgroups in CMV reactivation and negative patients. (A) Representative flow cytometry plots illustrating the differentiation of γδ T cells and subgroups in CMV reactivation and negative patients. Cells were categorized into four subgroups based on the differential expression of CD27 and CD45RA: Naive T cells (T Naive , CD45RA + CD27 + ), Central Memory T cells (T CM , CD45RA - CD27 + ), Effector Memory T cells (T EM , CD45RA - CD27 - ), and Effector Memory CD45RA + T cells (T EMRA , CD45RA + CD27 - ). (B) Differentiation status of γδ T cells in CMV reactivation patients (CMV + ) and negative patients (CMV - ). (C) Differentiation status of Vδ2 + γδ T cells in CMV reactivation patients (CMV + ) and negative patients (CMV - ). (D) Differentiation status of Vδ2 neg γδ T cells in CMV reactivation patients (CMV + ) and negative patients (CMV - ). Paired t-tests were performed, with P < 0.05 considered statistically significant.

Article Snippet: Following PBS washing, cells were incubated at room temperature in the dark for 20 minutes with appropriate monoclonal antibodies, including APC-anti-CD3 (BD Bioscience), PE-anti-pan TCR γ/δ (BD Bioscience), FITC-anti-TCR Vδ2 (BioLegend), PE/Cy7-anti-CD45RA (BD Bioscience), and PerCP/Cy5.5-anti-CD27 (BD Bioscience).

Techniques: Flow Cytometry, Expressing

Journal: Allergy, Asthma, and Clinical Immunology : Official Journal of the Canadian Society of Allergy and Clinical Immunology

Article Title: The clinical, immunological and genetic features of 12 Chinese patients with STAT3 mutations

doi: 10.1186/s13223-020-00462-w

Figure Lengend Snippet: Lymphocyte subpopulation of three STAT3 deficiency patient

Article Snippet: The following B cell subsets were detected: anti-CD19 (APC), anti-human CD24 (PE), anti-CD27 (BV450), anti-CD38 (PerCP-Cy5.5) and anti-IgD (BV510) (BD Biosciences).

Techniques: