Journal:

Article Title: Mycobacterium bovis BCG-Infected Mice Are More Susceptible to Staphylococcal Enterotoxin B-Mediated Toxic Shock than Uninfected Mice despite Reduced In Vitro Splenocyte Responses to Superantigens †

doi: 10.1128/IAI.70.8.4148-4157.2002

Figure Lengend Snippet: Infection with BCG results in accumulation and not deletion of superantigen-responsive CD4+ or CD8+ T cells. Unfractionated splenocytes (5 × 105 cells/tube) from SEB-exposed mice (7 days postinjection) (A) and PBS-injected controls or day-16-infected mice and PBS-injected controls (B) were stained with FITC-labeled anti-mouse Vβ8 TCR (A and B) and either PE-labeled anti-mouse CD4 or anti-mouse CD8. The percentages of Vβ8+ CD4+ T cells (white bars) or Vβ8+ CD8+ T cells (black bars) in the spleen were determined by flow cytometry. Means ± standard errors of the means of three experiments are shown. ∗, P < 0.05.

Article Snippet: In some experiments, splenocytes that were cultured for 24 or 72 h with SEB or BCG antigen were stained with FITC-conjugated rat anti-mouse CD4 (YTS 191.1) and rat anti-mouse CD8α (IM7.8.1; both from Cedarlane) and counterstained with propidium iodide (PI) (10 μg/ml; Sigma) to enumerate viable cells.

Techniques: Infection, Injection, Staining, Labeling, Flow Cytometry

Journal:

Article Title: Mycobacterium bovis BCG-Infected Mice Are More Susceptible to Staphylococcal Enterotoxin B-Mediated Toxic Shock than Uninfected Mice despite Reduced In Vitro Splenocyte Responses to Superantigens †

doi: 10.1128/IAI.70.8.4148-4157.2002

Figure Lengend Snippet: BCG infectious dose correlates with increased expression of the T-cell activation markers CD44, IL-2Rα, and IL-2Rβ. Unfractionated splenocytes (5 × 105 cells/tube) from each type of spleen were stained with FITC-labeled anti-mouse CD44, anti-mouse CD25/IL-2Rα, or anti-mouse CD122/IL-2Rβ and PE-labeled anti-mouse CD4 or anti-mouse CD8α. The percentages of CD4+ CD44hi, IL-2Rαhi, or IL-2Rβhi CD4+ (white bars) or CD8+ cells (black bars) in the spleen were determined by flow cytometry. Means ± standard errors of the means of three experiments are shown. ∗, P < 0.05; ∗∗, P < 0.01. Purified CD4+ T cells from the spleens of BCG HD-infected mice (day 16) were stained with FITC anti-mouse CD44 and were sorted in an EPICS Elite ESP (Beckman Coulter) into CD44low (naïve) and CD44hi (memory/effector) populations. Sorted cells were cocultured at 105 cells/well either with bone marrow-derived dendritic cells (5 × 104 cells/well) in the presence or absence of SEB (10 μg/ml) or with M12 B-lymphoma cells (5 × 104 cells/well) in the presence or absence of BCG-Ag (10 μg/ml). IFN-γ levels were determined as described in Materials and Methods. Mean ± standard deviation of triplicate wells is shown.

Article Snippet: In some experiments, splenocytes that were cultured for 24 or 72 h with SEB or BCG antigen were stained with FITC-conjugated rat anti-mouse CD4 (YTS 191.1) and rat anti-mouse CD8α (IM7.8.1; both from Cedarlane) and counterstained with propidium iodide (PI) (10 μg/ml; Sigma) to enumerate viable cells.

Techniques: Expressing, Activation Assay, Staining, Labeling, Flow Cytometry, Purification, Infection, Derivative Assay, Standard Deviation

Journal:

Article Title: Mycobacterium bovis BCG-Infected Mice Are More Susceptible to Staphylococcal Enterotoxin B-Mediated Toxic Shock than Uninfected Mice despite Reduced In Vitro Splenocyte Responses to Superantigens †

doi: 10.1128/IAI.70.8.4148-4157.2002

Figure Lengend Snippet: BCG infection induces potent APCs that stimulate rather than inhibit IFN-γ production to SEB and BCG-Ag by purified CD4+ T cells. Sixteen days after injection of mice with PBS or BCG LD or BCG HD, adherent cells were obtained from bulk splenocyte suspensions by panning as described in Materials and Methods. Adherent cell populations (5 × 105 cells/well) were cocultured with purified CD4+ T cells (105 cells/well) from the same mice in the presence of either SEB (10 μg/ml) (A) or BCG-Ag (10 μg/ml) (B). (C) KLH-specific HDK1 type 1 clone was cocultured at 105 cells/well with different numbers of unfractionated splenocytes from PBS control mice or BCG LD- or HD-infected mice in the presence of KLH (10 μg/ml). IFN-γ levels were determined as described in the Fig. ​Fig.11 legend. Mean ± standard deviation of triplicate wells is shown. (D) BCG-infected spleens have more MHC class II+ and Mac1+ cells. Unfractionated splenocytes (5 × 105 cells/tube) from mice that were infected for 2 weeks with BCG LD- or BCG HD-infected mice or PBS-injected controls were stained with FITC-labeled anti-mouse I-Ad/I-Ed/MHC class II and PE-labeled anti-mouse Mac1/CD11b. The percentages of MHC class II+ (white bars) and Mac1+ (black bars) APCs in the spleen were determined by flow cytometry. Means ± standard errors of the means of three experiments are shown.

Article Snippet: In some experiments, splenocytes that were cultured for 24 or 72 h with SEB or BCG antigen were stained with FITC-conjugated rat anti-mouse CD4 (YTS 191.1) and rat anti-mouse CD8α (IM7.8.1; both from Cedarlane) and counterstained with propidium iodide (PI) (10 μg/ml; Sigma) to enumerate viable cells.

Techniques: Infection, Purification, Injection, Standard Deviation, Staining, Labeling, Flow Cytometry

Journal:

Article Title: Mycobacterium bovis BCG-Infected Mice Are More Susceptible to Staphylococcal Enterotoxin B-Mediated Toxic Shock than Uninfected Mice despite Reduced In Vitro Splenocyte Responses to Superantigens †

doi: 10.1128/IAI.70.8.4148-4157.2002

Figure Lengend Snippet: HD splenocytes have a decreased percentage of T cells and show reduced viability early after culture with BCG-Ag and SEB. (A) Unfractionated splenocytes (5 × 105 cells/tube) from mice that were infected for 2 or 3 weeks with BCG LD or BCG HD or from PBS-injected controls were stained with FITC-labeled anti-mouse CD4 and PE-labeled anti-mouse CD8α. The percentages of CD4+ (white bars) or CD8+ cells (black bars) in the spleen were determined by flow cytometry. Means ± standard errors of the means of three experiments are shown. ∗, P < 0.05; ∗∗, P < 0.01. (B) Unfractionated splenocytes were subsequently stimulated with BCG-Ag (10 μg/ml) or SEB (10 μg/ml) at 5 × 105 cells/well in parallel cultures. At 24 h, cells from triplicate wells were pooled and stained with both FITC-labeled anti-mouse CD4 and anti-mouse CD8α and were counterstained with PI (10 μg/ml). The total percentage of nonviable (PI+) T cells was determined by flow cytometry.

Article Snippet: In some experiments, splenocytes that were cultured for 24 or 72 h with SEB or BCG antigen were stained with FITC-conjugated rat anti-mouse CD4 (YTS 191.1) and rat anti-mouse CD8α (IM7.8.1; both from Cedarlane) and counterstained with propidium iodide (PI) (10 μg/ml; Sigma) to enumerate viable cells.

Techniques: Infection, Injection, Staining, Labeling, Flow Cytometry

Journal: PLoS ONE

Article Title: Focal Radiation Therapy Combined with 4-1BB Activation and CTLA-4 Blockade Yields Long-Term Survival and a Protective Antigen-Specific Memory Response in a Murine Glioma Model

doi: 10.1371/journal.pone.0101764

Figure Lengend Snippet: A, B) Influx of CD4 + and CD8 + tumor infiltrating lymphocytes respectively was higher in brains of animals treated with immunotherapy with 4-1BB activation and CTLA-4 blockade and triple therapy compared to brains from non-tumor bearing mice ( p <0.05). C, D) Cervical lymph nodes from naïve mice showed a higher baseline of CD4 + and CD8 + T cells which resulted in comparable densities of T cells between all groups. Error bars represent standard error of the mean (SEM) and p values were calculated with the student t-test.

Article Snippet: Hamster and rat IgG to serve as control antibodies were purchased from Rockland Immunochemicals Inc. CD4-FITC and CD8-PE antibodies were used for FACS analysis (BD-Pharmingen).

Techniques: Activation Assay

Journal: PLoS ONE

Article Title: Focal Radiation Therapy Combined with 4-1BB Activation and CTLA-4 Blockade Yields Long-Term Survival and a Protective Antigen-Specific Memory Response in a Murine Glioma Model

doi: 10.1371/journal.pone.0101764

Figure Lengend Snippet: Kaplan Meier survival curves for untreated animals, triple therapy (no depletion), triple therapy with depleted CD4 + T cells and triple therapy with depleted CD8 + T cells ( n = 5–7 mice/group). CD4 + T cell depletion abolishes the survival benefit of triple therapy ( p <0.001). Depletion of CD8 + T cells did not interfere with the efficacy of triple therapy and resulted in long-term survival.

Article Snippet: Hamster and rat IgG to serve as control antibodies were purchased from Rockland Immunochemicals Inc. CD4-FITC and CD8-PE antibodies were used for FACS analysis (BD-Pharmingen).

Techniques:

Journal: Frontiers in Immunology

Article Title: Characterization of atypical T cells generated during ex vivo expansion process for T cell-based adoptive immunotherapy

doi: 10.3389/fimmu.2024.1202017

Figure Lengend Snippet: Antibody panels used for cytotoxic-associated CD4+ T cell phenotypic evaluation.

Article Snippet: CD4 , BV510 , BD Biosciences , 562970 , FITC , Diaclone , 954.031.010.

Techniques:

Journal: Frontiers in Pharmacology

Article Title: A Retinoid X Receptor Agonist Directed to the Large Intestine Ameliorates T-Cell-Mediated Colitis in Mice

doi: 10.3389/fphar.2021.715752

Figure Lengend Snippet: NEt-3IB inhibits differentiation of inflammatory T cells and induces Foxp3 + cells in Th1 and Th17 conditions. (A, B) Naïve T cells were cultured in Th1 conditions, with or without NEt-3IB treatment for 5 days and analyzed for T-bet and IFN-γ expression (A) , and for Foxp3 expression in the CD4 + T cell population (B) . (C) Naïve T cells were labeled with CellTrace Violet and cultured in Th1 conditions with or without NEt-3IB treatment for 5 days. Cultured cells were analyzed for their degree of proliferation in the CD4 + T-bet + IFN-γ + population. * p < 0.05, ** p < 0.01, *** p < 0.005.

Article Snippet: For Treg cell analysis of the colitis model, the following monoclonal antibodies were used: BV510-conjugated anti-mouse CD45 (30-F11; BioLegend), redFluor-710-conjugated anti-mouse CD4 (RM4-5; Tonbo Biosciences, San Diego, CA, United States), FITC-conjugated anti-mouse TCRβ (H57-597; BD Biosciences), and eFluor 660-conjugated anti-mouse Foxp3 (FJK-16s; ThermoFisher Scientific).

Techniques: Cell Culture, Expressing, Labeling

Journal: Frontiers in Pharmacology

Article Title: A Retinoid X Receptor Agonist Directed to the Large Intestine Ameliorates T-Cell-Mediated Colitis in Mice

doi: 10.3389/fphar.2021.715752

Figure Lengend Snippet: NEt-3IB ameliorates T-cell-dependent experimental colitis. (A–H) Experimental colitis was induced by adoptive transfer of CD4 + CD45RB high T cells into C.B-17 scid/scid mice fed with control or NEt-3IB-containing diet for 7 weeks, and bodyweight loss (A) and diarrhea score (B) were observed. Data shown are the average. Area under the curve of bodyweight change and fecal score. (C–F) Colon length, colon weight, and colon thickening were observed and measured in week 7. Scale bar: 1 cm. (G) Colonic specimens were stained with hematoxylin and eosin (HE). (H) Histological scores of colonic specimens. (I) Total CD45 + cells in the colon lamina propria of the CD4 + CD45RB high T cell transfer model. * p < 0.05, ** p < 0.01.

Article Snippet: For Treg cell analysis of the colitis model, the following monoclonal antibodies were used: BV510-conjugated anti-mouse CD45 (30-F11; BioLegend), redFluor-710-conjugated anti-mouse CD4 (RM4-5; Tonbo Biosciences, San Diego, CA, United States), FITC-conjugated anti-mouse TCRβ (H57-597; BD Biosciences), and eFluor 660-conjugated anti-mouse Foxp3 (FJK-16s; ThermoFisher Scientific).

Techniques: Adoptive Transfer Assay, Control, Staining