Journal: The Turkish Journal of Gastroenterology

Article Title: Butyrate Ameliorates Intestinal Epithelial Barrier Injury Via Enhancing Foxp3+ Regulatory T-Cell Function in Severe Acute Pancreatitis Model

doi: 10.5152/tjg.2022.21307

Figure Lengend Snippet: The effect of butyrate on the percentage of CD4 + CD25 + Foxp3 + Treg cells in the lamina propria. Control: Control group, SAP: SAP group, SAP + butyrate: SAP + butyrate group. (A) The percentages of CD4 + CD25 + Foxp3 + Treg cells detected by flow cytometry analysis. (B) Quantification of the percentages of CD4 + CD25 + Foxp3 + Treg cells in each group. Control group versus SAP group: P < .001, Control group versus SAP + butyrate group: P < .01, SAP group versus SAP + butyrate group: P < .01. n = 8 in each group. SAP, severe acute pancreatitis.

Article Snippet: Subsequently, the cells were stained with monoclonal antibodies, that is, anti-rat CD4 (mouse IgG, Catlog: 11-0010-82, eBioscience), anti-rat CD25 (mouse IgG, Catlog: 12-0390-82, eBioscience), and anti-rat Foxp3 (rat IgG, Catlog: 56-5773-82, eBioscience).

Techniques: Control, Flow Cytometry

Journal: Immunopharmacology and immunotoxicology

Article Title: DAB 389 IL-2 recombinant fusion toxin effect on lymphocyte- and macrophage- producing cytokine subpopulation cells in experimentally induced demyelinating disease in mice

doi: 10.1080/08923973.2017.1369099

Figure Lengend Snippet: FACS analysis * of T cell subpopulations from different organs in acute-EAE mice treated with PBS or DAB 389 IL-2 at day 21.

Article Snippet: Immunostaining and FACS analysis Fluorescein isothiocyanate (FITC)-labeled anti-rat CD3 (mouse IgG 3 , G4.18 clone), CD4 (mouse IgG 2a , OX-35), CD8 (mouse IgG 1 , OX-8) mAbs, phycoerythrin (PE)-labeled anti-rat αβ TCR (mouse IgG 1 , R73), FITC-labeled anti-rat IL-2, IFNγ, TNF-α, IL-4 and IL-10 from PharMingen (San Diego, CA), and biotinylated anti-rat CD3, CD4, CD8, CD25 (mouse IgG 1 , OX39) mAbs were purchased from PharMingen (San Diego, CA).

Techniques:

Journal: Immunopharmacology and immunotoxicology

Article Title: DAB 389 IL-2 recombinant fusion toxin effect on lymphocyte- and macrophage- producing cytokine subpopulation cells in experimentally induced demyelinating disease in mice

doi: 10.1080/08923973.2017.1369099

Figure Lengend Snippet: FACS analysis * of T cell subpopulations from different organs in chronic-EAE mice treated with PBS or DAB 389 IL-2 at day 24.

Article Snippet: Immunostaining and FACS analysis Fluorescein isothiocyanate (FITC)-labeled anti-rat CD3 (mouse IgG 3 , G4.18 clone), CD4 (mouse IgG 2a , OX-35), CD8 (mouse IgG 1 , OX-8) mAbs, phycoerythrin (PE)-labeled anti-rat αβ TCR (mouse IgG 1 , R73), FITC-labeled anti-rat IL-2, IFNγ, TNF-α, IL-4 and IL-10 from PharMingen (San Diego, CA), and biotinylated anti-rat CD3, CD4, CD8, CD25 (mouse IgG 1 , OX39) mAbs were purchased from PharMingen (San Diego, CA).

Techniques:

Journal: Immunopharmacology and immunotoxicology

Article Title: DAB 389 IL-2 recombinant fusion toxin effect on lymphocyte- and macrophage- producing cytokine subpopulation cells in experimentally induced demyelinating disease in mice

doi: 10.1080/08923973.2017.1369099

Figure Lengend Snippet: FACS analysis * of T cell subpopulations from different organs in chronic-EAE mice treated with PBS or DAB 389 IL-2 at day 16.

Article Snippet: Immunostaining and FACS analysis Fluorescein isothiocyanate (FITC)-labeled anti-rat CD3 (mouse IgG 3 , G4.18 clone), CD4 (mouse IgG 2a , OX-35), CD8 (mouse IgG 1 , OX-8) mAbs, phycoerythrin (PE)-labeled anti-rat αβ TCR (mouse IgG 1 , R73), FITC-labeled anti-rat IL-2, IFNγ, TNF-α, IL-4 and IL-10 from PharMingen (San Diego, CA), and biotinylated anti-rat CD3, CD4, CD8, CD25 (mouse IgG 1 , OX39) mAbs were purchased from PharMingen (San Diego, CA).

Techniques:

Journal: International journal of molecular medicine

Article Title: Streptococcus pneumoniae fructose-1,6-bisphosphate aldolase, a protein vaccine candidate, elicits Th1/Th2/Th17-type cytokine responses in mice.

doi: 10.3892/ijmm.2016.2512

Figure Lengend Snippet: Figure 2. Memory CD4+ T-cell proliferation in response to recombinant fructose-1,6-bisphosphate aldolase (rFBA) re-stimulation. The extent of pro liferation, determined with a BrdU ELISA kit, was examined in the following groups of cells: i) CD4+ T-cells from mice immunized with rFBA + com plete Freund's adjuvant (CFA)/incomplete Freund's adjuvant (IFA)/IFA (FBA CD4+ T-cells). All the following groups served as control groups: ii) CD4+ T-cells derived from mice immunized with an adjuvant only (con trol CD4+ T-cells). In addition, mononuclear cells depleted of CD4+ T-cells, derived from iii) mice immunized with rFBA (control FBA CD4- T-cells) or from iv) mice immunized with an adjuvant only (control CD4- T-cells). All cells were co-cultured with mitomycin-treated spleen-derived naïve antigen- presenting cells (APCs) and incubated for 72 h with increasing concentrations of rFBA (0-2 µg/ml). The values represent the means of quadruplicate wells ± standard deviation, and are expressed as the stimulation index, namely, the ratio of the extent of proliferation in the presence of rFBA to that in the absence of rFBA. Significant differences were found with the Student's t-test between the FBA CD4+ T-cells group and the control groups (*p≤0.05 and ***p<0.001). Results are the summary of two independent experiments.

Article Snippet: The depletion of CD4+ and of CD8+ T-cells was verified with a FITC-conjugated rat anti-mouse CD4 and rat anti-mouse CD8 (YTS169) respectively, detected using a FITC-conjugated AffiniPure F(ab')2 fragment of mouse antirat IgG (H+L) (Jackson ImmunoResearch Laboratories, Inc., West Grove, PA, USA).

Techniques: Recombinant, Enzyme-linked Immunosorbent Assay, Adjuvant, Control, Derivative Assay, Cell Culture, Incubation, Standard Deviation

Journal: International journal of molecular medicine

Article Title: Streptococcus pneumoniae fructose-1,6-bisphosphate aldolase, a protein vaccine candidate, elicits Th1/Th2/Th17-type cytokine responses in mice.

doi: 10.3892/ijmm.2016.2512

Figure Lengend Snippet: Figure 3. T helper (Th)1-type cytokine secretion from memory CD4+ T-cells in response to recombinant fructose-1,6-bisphosphate aldolase (rFBA) re-stimulation. CD4+ T-cells were obtained from BALB/c mice immunized with rFBA + complete Freund's adjuvant (CFA)/incomplete Freund's adjuvant (IFA)/IFA [‘FBA’ cultures; the legend for the bar fill format that appears in (A) also applies to (B and C)], or from mice immunized with adjuvant alone (‘CFA’ cultures), followed by negative selection. CD4+ T-cells that were obtained from BALB/c mice immunized with rFBA and CFA/IFA were used for the ‘CD4+ only’ control culture; BMDC cultures were used for the ‘DC only’ control cultures; and CD4+ T-cells that were obtained from mice immunized with the adjuvant alone and stimulated with concanavalin A (ConA) were used for the ‘CD4+ConA’ control culture. CD4+ T-cells in the control groups were stimulated with ConA. All CD4+ T-cells were co- incubated with rFBA-pulsed BMDCs for 72 h. The levels of cytokine secretion [(A) IFN-γ; (B) interleukin (IL)-2; (C) tumor necrosis factor (TNF)-α] was measured by multiplex ELISA. The results represent the means ± standard deviations of triplicate wells. A two-tailed Student's t-test found significant differences (p<0.05) between the ‘FBA’ cultures and ‘CFA’: cultures (*p<0.05, **p<0.01 and ***p<0.001). Results show one representative experiment out of two independent experiments.

Article Snippet: The depletion of CD4+ and of CD8+ T-cells was verified with a FITC-conjugated rat anti-mouse CD4 and rat anti-mouse CD8 (YTS169) respectively, detected using a FITC-conjugated AffiniPure F(ab')2 fragment of mouse antirat IgG (H+L) (Jackson ImmunoResearch Laboratories, Inc., West Grove, PA, USA).

Techniques: Recombinant, Adjuvant, Selection, Control, Incubation, Multiplex Assay, Enzyme-linked Immunosorbent Assay, Two Tailed Test

Journal: International journal of molecular medicine

Article Title: Streptococcus pneumoniae fructose-1,6-bisphosphate aldolase, a protein vaccine candidate, elicits Th1/Th2/Th17-type cytokine responses in mice.

doi: 10.3892/ijmm.2016.2512

Figure Lengend Snippet: Figure 4. Pro-inflammatory-type cytokine secretion from memory CD4+ T-cells in response to recombinant fructose-1,6-bisphosphate aldolase (rFBA) re- stimulation. CD4+ T-cells were obtained from BALB/c mice immunized with rFBA + complete Freund's adjuvant (CFA)/incomplete Freund's adjuvant (IFA)/IFA [‘FBA’ cultures; the legend for the bar fill format that appears in (A) applies also to (C-E)] or from mice immunized with the adjuvant alone (‘CFA’ cultures), followed by negative selection. CD4+ T-cells that were obtained from BALB/c mice immunized with rFBA and CFA/IFA were used for the ‘CD4+ only’ control cultures, and CD4+ T-cells that were obtained from mice immunized with the adjuvant alone were stimulated with 5 µg/ml of concanavalin A (ConA) and used for the ‘CD4+ConA’ control cultures. CD4+ T-cells were co-incubated with rFBA-pulsed BMDCs for 72 h for determination of the level of the cytokine secretion measured by multiplex ELISA. The results represent the means ± standard deviation of triplicate wells. A two-tailed Student's t-test revealed no significant differ ences between the FBA and CFA cultures for (A) granulocyte-macrophage colony-stimulating factor (GM-CSF) (C) IL-1β, (D) IL-12p40. (E) By contrast IL-12p70 demonstrated high levels at time 0 h that decreased at 12 h and significantly increased at 24, 48 and 78 h (p<0.05). Results are from one representative experiment out of two independent experiments. (B) GM-CSF/APC. CD4+ T-cells were obtained from BALB/c mice immunized with rFBA + CFA/IFA/IFA (‘FBA’ cultures), or from mice immunized with the adjuvant alone (‘CFA’ cultures), followed by negative selection. CD4+ T-cells were co-incubated with rFBA-pulsed spleen- derived naïve antigen-presenting cells (APCs) for 72 h. The results represent the mean ± standard deviation of triplicate wells. Significant differences (p<0.05) were determined by a two-tailed Student's t-test between the ‘FBA’ cultures and the ‘CFA’ cultures. Results are combined from two independent experiments.

Article Snippet: The depletion of CD4+ and of CD8+ T-cells was verified with a FITC-conjugated rat anti-mouse CD4 and rat anti-mouse CD8 (YTS169) respectively, detected using a FITC-conjugated AffiniPure F(ab')2 fragment of mouse antirat IgG (H+L) (Jackson ImmunoResearch Laboratories, Inc., West Grove, PA, USA).

Techniques: Recombinant, Adjuvant, Selection, Control, Incubation, Multiplex Assay, Enzyme-linked Immunosorbent Assay, Standard Deviation, Two Tailed Test, Derivative Assay

Journal: International journal of molecular medicine

Article Title: Streptococcus pneumoniae fructose-1,6-bisphosphate aldolase, a protein vaccine candidate, elicits Th1/Th2/Th17-type cytokine responses in mice.

doi: 10.3892/ijmm.2016.2512

Figure Lengend Snippet: Figure 5. T helper (Th)2- and Th17-type cytokine secretion from memory CD4+ T-cells in response to recombinant fructose-1,6-bisphosphate aldolase (rFBA) re-stimulation. CD4+ T-cells were obtained from BALB/c mice immunized with rFBA + complete Freund's adjuvant (CFA)/incomplete Freund's adjuvant (IFA) booster/IFA booster [‘FBA’ cultures; the legend for the bar fill format in (A) applies also to (B-D)], or from mice immunized with adjuvant alone (‘CFA’ cultures), followed by negative selection. CD4+ T-cells that were obtained from BALB/c mice immunized with rFBA and CFA/IFA were used for the ‘CD4+ only’ control cultures, and CD4+ T-cells that were obtained from mice immunized with the adjuvant alone were used for the ‘CD4+ConA’ control cultures. CD4+ T-cells stimulated with 5 µg/ml of concanavalin A (ConA) served as CD4+ ConA group. CD4+ T-cells were co-incubated with rFBA-pulsed BMDCs for 72 h. The levels of cytokine [(A) IL-4; (B) IL-5; (C) IL-10; and (D) IL-17] secretion were measured by multiplex ELISA. The results represent the means ± standard deviation of triplicate wells. Significant differences between groups were calculated with a two-tailed Student's t-test (*p<0.05, **p<0.01 and ***p<0.001). Results are from one representative experiment out of two independent experiments.

Article Snippet: The depletion of CD4+ and of CD8+ T-cells was verified with a FITC-conjugated rat anti-mouse CD4 and rat anti-mouse CD8 (YTS169) respectively, detected using a FITC-conjugated AffiniPure F(ab')2 fragment of mouse antirat IgG (H+L) (Jackson ImmunoResearch Laboratories, Inc., West Grove, PA, USA).

Techniques: Recombinant, Adjuvant, Selection, Control, Incubation, Multiplex Assay, Enzyme-linked Immunosorbent Assay, Standard Deviation, Two Tailed Test

Journal: Cardiovascular research

Article Title: Regression of atherosclerosis with anti-CD3 antibody via augmenting a regulatory T-cell response in mice.

doi: 10.1093/cvr/cvu002

Figure Lengend Snippet: Figure 5 DepletionofCD4+CD25+Foxp3+ Tregs by ip injectionofCD25-Ababolished thebeneficial effectsofCD3-Abontheregressionofatheroscler- osis. (A) Tregdepletionstudy design. (B) Representative photomicrographs of oil red O staining. The black bar represents 200 mm. (C) Quantitative analysis of atherosclerotic lesion size in the aortic sinus of LDLR2/2 mice at 14 and 18 weeks of age. (D) Graphs represent the number of CD4+ T cells in the spleen and LNs. (E) Quantitative analysis of immunohistochemical CD4 staining in atherosclerotic lesions. (F) Graphs represent the percentage of CD25+Foxp3+ Tregs withintheCD4+ T-cellpopulationinthespleenandLNs.(G)QuantitativeanalysisofimmunohistochemicalFoxp3staininginatheroscleroticlesions.Alldatain Figure5arepresentedasmeans+SEM;n ¼ 6in(C),(E),and(G),andn ¼ 5in(D)and(F)foreachgroup. #P , 0.05vs.baseline.*P , 0.05vs.HamsterIgGplus rat IgG. }P , 0.05 vs. Hamster IgG plus CD25-Ab. †P , 0.05 vs. CD3-Ab plus rat IgG. §P , 0.05 vs. CD3-Ab plus CD25-Ab.

Article Snippet: In several experiments, mice received ip injection of 100 mg of anti-CD25 antibody (CD25-Ab) (clone PC61; Bio X cell) to deplete CD4+CD25+Foxp3+ Tregs or 100 mg of isotype-matched rat IgG (Bio X cell) a week after the first injection of CD3-Ab or control IgG.

Techniques: Staining, Immunohistochemical staining