Journal: Frontiers in Immunology

Article Title: Age-related increase of mitochondrial content in human memory CD4+ T cells contributes to ROS-mediated increased expression of proinflammatory cytokines

doi: 10.3389/fimmu.2022.911050

Figure Lengend Snippet: Capacity of cytokine production in memory CD4 + T cell. Cells were activated with PMA/ionomycin for 5 h, adding BFA for the last 3 hours for intracellular cytokine expression. (A) Representative example of cytokine expression in memory CD4 + T cells from young and aged donors after stimulation with PMA/ionomycin. (B) Frequencies in % and geometric mean fluorescence intensity (gMFI) of IL-2, IL-4, IL-10, IL-17, IFN-γ, and TNF-α producing memory CD4 + T cells (mean ± SEM; n = 7).

Article Snippet: Incubation was conducted by dissolving 1x 10 6 cells/ml in RPMI 1640 medium (Thermo Fisher Scientific) with 5% human AB serum (Sigma Aldrich) and 1% penicillin/streptomycin (Thermo Fisher Scientific) at 37˚C in a 5% CO 2 atmosphere Cells were collected, washed, fixed, permeabilized (Inside staining kit, Miltenyi Biotec) and stained with a combination of monoclonal antibodies including anti-human-IFNγ (PERCP-Cy5.5, Biolegend, clone 4SB3, Cat.# 502526), anti-human-IL-2 (APC-vio770, Miltenyi Biotec, clone N7.48 A, Cat.# 130-097-011), anti-human-IL-4 (PE, Miltenyi Biotec, clone 7A3-3, Cat.# 130-091-647), anti-human-IL-17A (FITC, Miltenyi Biotec, clone CZ8-23G1, Cat.# 130-094-520), and anti-human-TNF-α (PE-Vio770 (Miltenyi Biotec, clone cA2, Cat.# 130-096-755).

Techniques: Expressing, Fluorescence

Journal: Mediators of Inflammation

Article Title: Analysis of Th17 and Tc17 Frequencies and Antiviral Defenses in Gut-Associated Lymphoid Tissue of Chronic HIV-1 Positive Patients

doi: 10.1155/2015/395484

Figure Lengend Snippet: Activation markers, IFN- γ and IL-17A expression in CD4 + and CD8 + T cells subsets in blood and in gut. Panel (a): representative flow cytometry plots outlining CD38 and HLA-DR expression on naïve, effector, and central memory CD4 + and CD8 + lymphocytes derived from peripheral blood mononuclear cells (PBMCs) and gut-associated lymphoid tissue (GALT). Panel (b): representative flow cytometry plots outlining IFN- γ and IL-17A intracitoplasmatic expression on naïve, effector, and central memory CD4 + (Th1 and Th17) and CD8 + (Tc1 and Tc17) lymphocytes derived from PBMCs and GALT.

Article Snippet: Cultured cells were fixed, permeabilized (BD Cytofix/Cytoperm, Becton Dickinson, San Jose, CA, USA), and stained with combinations of fluorochrome-labeled monoclonal antibodies: CD3-PerCP, CD4-APC-Vio770, CD8-FITC, CD45RO-PE-Vio770, CD27-VioBlue, IFN- γ -APC, and IL-17A-PE (Miltenyi Biotec, Bergisch Gladbach, Germany).

Techniques: Activation Assay, Expressing, Flow Cytometry, Derivative Assay

Journal: Mediators of Inflammation

Article Title: Disruption of Tumor Necrosis Factor Receptor-Associated Factor 5 Exacerbates Murine Experimental Colitis via Regulating T Helper Cell-Mediated Inflammation

doi: 10.1155/2016/9453745

Figure Lengend Snippet: The gene-specific primer pairs list in the study.

Article Snippet: The following primary antibodies were used: anti-IFN- γ (1 : 200 dilution; Proteintech, USA), anti-IL-4 (1 : 100 dilution; BioLegend, USA), anti-IL-17a (1 : 200 dilution; Proteintech, USA), anti-p65 (1 : 100 dilution; Cell Signaling Technology, USA), anti-CD4 (1 : 100 dilution; BioLegend, USA), and anti-CK18 (1 : 100 dilution; Goodbio Technology Co., China).

Techniques:

Journal: Mediators of Inflammation

Article Title: Disruption of Tumor Necrosis Factor Receptor-Associated Factor 5 Exacerbates Murine Experimental Colitis via Regulating T Helper Cell-Mediated Inflammation

doi: 10.1155/2016/9453745

Figure Lengend Snippet: Analysis of the mRNA expression of cytokines and transcription factors for Th cells in the colons of TRAF5 KO and WT mice. (a) TNF- α , (b) IFN- γ , (c) IL-4, (d) IL-17a, (e) IL-22, (f) T-bet, (g) GATA-3, (h) ROR- α , and (i) ROR- γ t mRNA expression levels were determined by real-time PCR. The data are representative of 3 independent experiments (mean ± SD). N = 8 per group. ∗ p < 0.05 versus the DSS-treated WT group; ∗∗ p < 0.01 versus the DSS-treated WT group. NS: not significant versus the DSS-treated WT group.

Article Snippet: The following primary antibodies were used: anti-IFN- γ (1 : 200 dilution; Proteintech, USA), anti-IL-4 (1 : 100 dilution; BioLegend, USA), anti-IL-17a (1 : 200 dilution; Proteintech, USA), anti-p65 (1 : 100 dilution; Cell Signaling Technology, USA), anti-CD4 (1 : 100 dilution; BioLegend, USA), and anti-CK18 (1 : 100 dilution; Goodbio Technology Co., China).

Techniques: Expressing, Real-time Polymerase Chain Reaction

Journal: Mediators of Inflammation

Article Title: Disruption of Tumor Necrosis Factor Receptor-Associated Factor 5 Exacerbates Murine Experimental Colitis via Regulating T Helper Cell-Mediated Inflammation

doi: 10.1155/2016/9453745

Figure Lengend Snippet: Deletion of TRAF5 increases the protein expressions of the cytokines IFN- γ , IL-4, and IL-17a in the colons of mice induced by DSS. (a) The production of IFN- γ , IL-4, and IL-17a in the colons of DSS-induced mice or control mice was examined by ELISA ( N = 6 per group). (b) Representative immunofluorescence staining for IFN- γ , IL-4, and IL-17a in the colons of TRAF5 KO and WT mice (magnification 400x). (c) Quantification of positive cells ( N = 4 per group). The data are representative of 3 independent experiments (mean ± SD). ∗ p < 0.05 versus the DSS-treated WT group; ∗∗ p < 0.01 versus the DSS-treated WT group.

Article Snippet: The following primary antibodies were used: anti-IFN- γ (1 : 200 dilution; Proteintech, USA), anti-IL-4 (1 : 100 dilution; BioLegend, USA), anti-IL-17a (1 : 200 dilution; Proteintech, USA), anti-p65 (1 : 100 dilution; Cell Signaling Technology, USA), anti-CD4 (1 : 100 dilution; BioLegend, USA), and anti-CK18 (1 : 100 dilution; Goodbio Technology Co., China).

Techniques: Control, Enzyme-linked Immunosorbent Assay, Immunofluorescence, Staining

Journal: Mediators of Inflammation

Article Title: Disruption of Tumor Necrosis Factor Receptor-Associated Factor 5 Exacerbates Murine Experimental Colitis via Regulating T Helper Cell-Mediated Inflammation

doi: 10.1155/2016/9453745

Figure Lengend Snippet: Increased proportions of Th2 and IFN- γ /IL-17a-coproducing CD4 + T cells in the colons of TRAF5-deficient animals induced by DSS. LPMCs obtained from the colons of DSS-fed KO and WT mice were stimulated with PMA/ionomycin and subjected to surface staining for CD4. After fixation and permeabilization, intracellular staining for IL-4, IFN- γ , and IL-17a was performed as described in . CD4 + T cells were then gated and analyzed. (a) Representative flow plots. The percentages of (b) Th2 cells (CD4 + IL-4 + ), (c) IFN- γ /IL-17a-coproducing CD4 + T cells (CD4 + IFN- γ + IL-17a + ), (d) classical Th1 cells (CD4 + IFN- γ + IL-17a − ), and (e) classical Th17 cells (CD4 + IL-17a + IFN- γ − ) are presented as the mean ± SD. The data are representative of 3 independent experiments. N = 6 per group. ∗∗ p < 0.01 versus the DSS-treated WT group. NS: not significant versus the DSS-treated WT group.

Article Snippet: The following primary antibodies were used: anti-IFN- γ (1 : 200 dilution; Proteintech, USA), anti-IL-4 (1 : 100 dilution; BioLegend, USA), anti-IL-17a (1 : 200 dilution; Proteintech, USA), anti-p65 (1 : 100 dilution; Cell Signaling Technology, USA), anti-CD4 (1 : 100 dilution; BioLegend, USA), and anti-CK18 (1 : 100 dilution; Goodbio Technology Co., China).

Techniques: Staining