th17 responses  (R&D Systems)

Journal: Frontiers in Immunology

Article Title: Bordetella pertussis infection activates the type I interferon signaling pathway to exacerbate respiratory tract inflammatory response

doi: 10.3389/fimmu.2025.1521970

Figure Lengend Snippet: Serological responses to PT, FHA, and PRN and cytokines in the supernatants of lung and spleen lymphocyte cultures from B. pertussis -infected WT and IFNAR1 -/- mice. Specific antibodies against B. pertussis and cytokines in the supernatants of lung and spleen lymphocyte cultures from B. pertussis -infected WT and IFNAR1 -/- mice. (A) Total serum IgG titers against pertussis toxoid (PT), filamentous hemagglutinin (FHA), and pertactin (PRN) at 28 dpi. (B, C) Splenic and pulmonary lymphocytes were isolated at 28 dpi. The cells were cultured at a concentration of 2 × 10 6 /ml at 37°C with 5% CO 2 and stimulated with the B. pertussis -specific antigens PT (2 μg/mL), FHA (2 μg/mL), and PRN (2 μg/mL). After incubation for 3 days, the culture supernatant was collected, and the levels of IFN-γ, IL-17A, and IL-4 were assayed via ELISA. Th1 response-associated cytokines (IFN-γ), Th17 response-associated cytokines (IL-17A), and (C) Th2 response-associated cytokines (IL-4) in the culture supernatants of splenic lymphocytes (B) and pulmonary lymphocytes (C) . The data are expressed as the means ± SEM. The P value is indicated as follows: * P < 0.05, ns, no significance (n=5).

Article Snippet: IFN-γ (Cat# VAL607, Novus Biologicals) levels were measured for Th1 responses; IL-17A (Cat# VAL610, Novus Biologicals) levels were tested for Th17 responses; and IL-4 (Cat# M4000B, R&D Systems) levels were detected for Th2 responses.

Techniques: Infection, Isolation, Cell Culture, Concentration Assay, Incubation, Enzyme-linked Immunosorbent Assay

Journal: Current Issues in Molecular Biology

Article Title: Inflammation, Autoinflammation and Autoimmunity in Inflammatory Bowel Diseases

doi: 10.3390/cimb45070350

Figure Lengend Snippet: Schematic illustration of the main differences between ulcerative colitis (UC, left ) and Crohn’s disease (CD, right ) in inflammation, autoinflammation and autoimmunity. The increased intestinal permeability of the epithelial cell layer allows in both diseases Danger (DAMPs) and Pathogen (PAMPs) associated molecular patterns (red dots) to easily reach the mucosa and stimulate innate immune cells. In UC neutrophil extracellular traps (NETs, green) are more abundant than in CD, activate M1 macrophages inducing TNF-α and IL-1β release. It is possible that NETs activate also the adaptive immune response causing ANCA autoantibodies production. The expression of inflammasomes, mainly NLRP1 and NLRP3, is induced in UC concurring in further enhancing IL-1β. CD4+ Th2 and CD8+ cells with IL-5, IL-13 and IFN-γ production prevails. In CD CD4+ Th1 cells prevails over Th2 with TNF-α and IFN-γ production. M1 macrophages produce mainly IL-12, while loss of function of the inflammasomes NLRP1 and NLRP3, due to NOD2 mutations and increased Immunity-related GTPase M (IRGM) that enhances NLRP3 degradation, occurs.

Article Snippet: Following exposure, the intestinal mucosal immune reaction (“immunome”) leads to inflammation and tissue damage accompanied by innate and adaptive immune response, with a predominant Th1 and Th17 response with IL-12, interferon (IFN)-γ and IL-17A production in CD [ ] and a Th2 response with IL-5 and IL-13 production in UC ( ) [ , ].

Techniques: Permeability, Expressing

Journal: Current Issues in Molecular Biology

Article Title: Inflammation, Autoinflammation and Autoimmunity in Inflammatory Bowel Diseases

doi: 10.3390/cimb45070350

Figure Lengend Snippet: Cytokines with a major role in IBD. Data reported in this table were obtained from [ 47 , 58 , 85 , 96 ].

Article Snippet: Following exposure, the intestinal mucosal immune reaction (“immunome”) leads to inflammation and tissue damage accompanied by innate and adaptive immune response, with a predominant Th1 and Th17 response with IL-12, interferon (IFN)-γ and IL-17A production in CD [ ] and a Th2 response with IL-5 and IL-13 production in UC ( ) [ , ].

Techniques: Activation Assay, Immunopeptidomics, Cell Differentiation, Blocking Assay

Journal: Scandinavian Journal of Immunology

Article Title: Pathogenic NKT cells attenuate urogenital chlamydial clearance and enhance infertility

doi: 10.1111/sji.13263

Figure Lengend Snippet: Pro‐inflammatory signalling is dampened in CD1d‐deficent mice. Female BALB/c mice (n = 5/group, in 1 experiment) were vaginally infected with Chlamydia muridarum for 6 d and oviducts were collected and pooled. RNA was extracted and qRT‐PCR was performed in duplicate using a Mouse Th17 response RT2‐PCR array. Results were normalized to uninfected oviducts. Fold change expression of inflammatory cytokines (A, B), chemokines (C), surface markers (D) and signalling pathways (E) was determined and represented as a heatmap.

Article Snippet: Mouse Th17 Response RT 2 Profiler PCR Array (PAMM‐073Z) (Qiagen, Clayton, VIC, AUS) was performed as per the manufacturer's instructions on an ABI 7900 HT plate PCR (Applied Biosystems).

Techniques: Infection, Quantitative RT-PCR, Expressing

Journal: The Journal of Experimental Medicine

Article Title: STING agonist promotes CAR T cell trafficking and persistence in breast cancer

doi: 10.1084/jem.20200844

Figure Lengend Snippet: Th/Tc17 CAR T cells exhibit enhanced early control of tumor growth over 7/15 CAR T cells owing to enhanced persistence in the tumor. (A) Schematic of the LH28z CAR cassette encoding the scFv (7.16.4), hinge, and transmembrane domain from CD8, and intracellular domains from CD28 and CD3ζ. (B) Expression of CAR T receptor on transduced murine 7/15 CD4 + (left) and CD8 + (right) T cells. (C) Representative flow cytometry histograms depicting viability of Neu + NT2 tumor cells in vitro after 3 d of coculture with 7/15 CAR T cells. Tumor cells were prelabeled with CFSE and plated before the addition of CAR T cells at a 1:1 ratio with tumor cells. (D) Intracellular staining illustrating IFN-γ and TNF production by 7/15 CAR T cells after coculture with Neu + cells or Neu − 3T3 cells at a 2:1 ratio. (E) Tumor area change (tumor area before therapy subtracted from area after therapy) was determined and compared in FVB-neu mice that received 7/15 CAR T cells or mock-transduced T cells (Mock Ts). (F) Expression of CAR T receptor on transduced Th17 (left) or Tc17 (right) cells. (G) In vitro killing of NT2 tumor cells by 7/15 CAR T and Th/Tc17 CAR T cells after overnight culture. (H) Histogram flow plots of IL-17A, TNF, and IFN-γ secretion by Th/Tc17 CAR T and 7/15 CAR T compared with mock T cells after coculture with Neu + cells at 3:1 ratio for 6 h. (I) Tumor area change was calculated and compared in FVB-neu mice that received Th/Tc17 CAR T cells or 7/15 CAR T cells. (J) Detection of CD4 + and CD8 + CAR T cells in spleen or tumor by flow cytometry 5 d after injection of 7/15 CAR T cells or Th/Tc17 CAR T cells. Data are shown as mean ± SD; *, P < 0.05; **, P < 0.01; significance was determined by Student’s t test or two-way ANOVA. n ≥ 5 mice per group with data from at least two independent experiments with the presented data pooled.

Article Snippet: Quantitative RT-PCR was performed according to the manufacturer’s instructions with the RT 2 Profiler PCR Array Mouse Th17 Response (PAMM-073ZE-4; Qiagen) and performed on the QuantStudio 6K (Applied Biosystems).

Techniques: Control, Expressing, Flow Cytometry, In Vitro, Staining, Injection

Journal: The Journal of Experimental Medicine

Article Title: STING agonist promotes CAR T cell trafficking and persistence in breast cancer

doi: 10.1084/jem.20200844

Figure Lengend Snippet: Th17 CAR T cells are indispensable for DMXAA therapeutic efficacy, which enhances the number and cytotoxicity of Th17 CAR T cells in the TME. (A and B) Tables for the percentage of T cell subsets (A) and macrophage and myeloid cell subsets (B) calculated from scRNA-seq data. (C) Table for the percentage of CD8 + T cells expressing indicated genes calculated from scRNA-seq data. (D–F) Assessment of CAR expression on tumor-infiltrating T cells isolated 7 d after therapy by flow cytometry. (D) Determination of change in tumor area 7 d after injection of Tc17 or Th/Tc17 CAR T cells. (E) Assessment of intratumoral CD8 + CAR T cells was performed 7 d after therapy. (F) Absolute number of CD8 + CAR T cells/mm 2 tumor 7 d after therapy. Data in G–M data were derived from the characterization of T cells expressing CD4 from scRNA-seq result. (G) CD4 + T cells were subdivided based on absence of Cd44 (naive T cells), expression of Cd4 and Cd44 (activated CD4 T cells), Cd4 and Foxp3 (T reg cells). CD4 + T cells were compared between the +DMXAA and −DMXAA treatment groups, and each subpopulation was examined further as described below. (H) Table for the percentage of CD4 + T cells expressing the indicated genes from the scRNA-seq data. (I) t-SNE plot of CD4 + T cells for Th17-related genes. (J) t-SNE plot of CD4 + T cells for Th17/Th1-related genes. (K) Violin plot shows the distribution and change of indicated gene expression in B and C. (L) Validation of single-cell data using flow cytometry to detect CD4 + CAR T cells. (M) Analysis of the correlation between tumor growth change and absolute cell number of CAR-expressing CD4 T cells/cm 2 tumor. Data (excluding scRNA-seq) represent one of two independent experiments ( n ≥ 5 mice per group). Single-cell sequencing data represent three mice/treatment group, with statistical significance determined by differential GSA with the Partek flow workstation. *, q < 0.05; **, q < 0.01, in the change of gene expression level. Fold change is +DMXAA versus −DMXAA. For percentage/number change, please refer to . Flow cytometry data were pooled from two independent experiments and shown as mean ± SD. Statistics analysis for cytometric analysis is determined by Student’s t test; *, P < 0.05; **, P < 0.01; ****, P < 0.0001.

Article Snippet: Quantitative RT-PCR was performed according to the manufacturer’s instructions with the RT 2 Profiler PCR Array Mouse Th17 Response (PAMM-073ZE-4; Qiagen) and performed on the QuantStudio 6K (Applied Biosystems).

Techniques: Drug discovery, Expressing, Isolation, Flow Cytometry, Injection, Derivative Assay, Gene Expression, Biomarker Discovery, Sequencing

Journal: The Journal of Experimental Medicine

Article Title: STING agonist promotes CAR T cell trafficking and persistence in breast cancer

doi: 10.1084/jem.20200844

Figure Lengend Snippet: DMXAA treatment enhances IFN-γ–dependent antitumor function of Th/Tc17 CAR T cells, but T cell exhaustion limits the cumulative beneficial effect of DMXAA on Th/Tc17 CAR T therapy. Mice received Th/Tc17 CAR T cells in the presence or absence of DMXAA therapy. 7 d later, RNA from tumors was isolated and analyzed by microarray for Th17/Th1 response. (A) Heatmap depicts genes where fold-change was significant, with a threefold increase. Each column represents an individual mouse. (B) Quantification of significant changes (P < 0.05) in TME for Th17 or Th1 response following DMXAA treatment. (C) Mice were treated i.p. with anti–IFN-γ (250 µg/mouse) twice a week after Th/Tc17 CAR T therapy. Significance was determined by Student’s t test or two-way ANOVA. Studies involved at least four mice per independent experiment. Data from C represent two independent experiments with four mice per group. (D–G) T cell cluster selected based on expression of Cd3e by unsupervised clustering of tumor-infiltrating immune cells from 7 and 10 d after Th/Tc17 CAR + D therapy. (D) Table shows the percentage of T cells expressing the indicated genes calculated from scRNA-seq data. (E) t-SNE plot (left) and violin plot (right) of T cells for genes associated with T cell exhaustion. (F) Validation of PD-1 expression using flow-based method to detect high levels of PD-1 expression on T cells from mice receiving Th/Tc17 CAR T or mock T cell therapy. Representative flow cytometry histograms (left); percentage of CAR T (right) within the CD4 or CD8 T cell group ( n = 4/group). (G) t-SNE plot (left) and violin plot (right) of T cells for genes associated with T cell apoptosis. Data (excluding scRNA-seq) represent one of two independent experiments ( n ≥ 5 mice per group). Single-cell sequencing data represent two to three mice/treatment group. Statistical significance was determined by differential GSA with the Partek flow workstation. *, q < 0.05; **, q < 0.01; ***, q < 0.001; ****, q < 0.0001 in the change of gene expression level. Fold change is 7 versus 10 d. For percentage/number change, please refer to . Flow cytometric analysis is shown as mean ± SD. Statistics analysis for cytometric analysis was determined by Student’s t test; *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.

Article Snippet: Quantitative RT-PCR was performed according to the manufacturer’s instructions with the RT 2 Profiler PCR Array Mouse Th17 Response (PAMM-073ZE-4; Qiagen) and performed on the QuantStudio 6K (Applied Biosystems).

Techniques: Isolation, Microarray, Expressing, Biomarker Discovery, Flow Cytometry, Sequencing, Gene Expression

Journal: Immunology

Article Title: Glycogen synthase kinase‐3 promotes T helper type 17 differentiation by promoting interleukin‐9 production

doi: 10.1111/imm.13199

Figure Lengend Snippet: IL‐17A+ CD4+ (T helper type 17; Th17) cells and the expression of several Th17 differentiation‐related genes are lower in GSK3β −/− (knockout; KO) CD4 cells compared with wild‐type (WT) CD4 cells. WT (GSK3β Flox/Flox) and glycogen synthase kinase‐3β (GSK3β) KO (GSK3β Flox/Flox CD4‐CRE) CD4 cells were differentiated in vitro towards Th17 cells for 2–4 days. (a) IL‐17A+ CD4+ and IRF4+ CD4+ cells after 4 days of differentiation, analyzed by flow cytometry. (b) Gene expression after 2 days of differentiation towards Th17 cells, identified using Th17 RT2 profiler PCR array. (c) Fold changes of gene expression of GSK3β −/− CD4 cells after 2 days of differentiation towards Th17 cells, analyzed by RT‐PCR. Fold change was calculated as (2−Δ Ct of a sample)/(average 2−Δ Ct of WT CD4 cells). Gapdh was used as the endogenous control gene to normalize individual gene expression. Each symbol in (c) represents an individual GSK3β −/− mouse (n = 3). *P < 0·05, **P < 0·01, unpaired t‐test compared with respective gene level in WT CD4 cells.

Article Snippet: Real‐time PCR was performed on ‘mouse Th17 response’ RT 2 profiler PCR arrays (Qiagen) according to the manufacturer’s instructions.

Techniques: Expressing, Knock-Out, In Vitro, Flow Cytometry, Reverse Transcription Polymerase Chain Reaction

Journal: Immunology

Article Title: Glycogen synthase kinase‐3 promotes T helper type 17 differentiation by promoting interleukin‐9 production

doi: 10.1111/imm.13199

Figure Lengend Snippet: Effects of glycogen synthase kinase‐3 (GSK3) inhibitors on gene expression during T helper type 17 (Th17) cell differentiation. Wild‐type (WT) CD4 cells were differentiated toward Th17 cells in the presence of (a) LiCl (10 mm); (b) CHIR99021 (1 μm); or (c) TDZD8 (1 μm) for 2 days. Fold change was calculated as (2−Δ Ct of a sample)/(average 2−Δ Ct of untreated CD4 cells). Gapdh was used as the endogenous control gene to normalize individual gene expression. Each symbol represents an independent cell differentiation from the pool of CD4+ cells from two mice (n = 4). *P < 0·05, **P < 0·01, unpaired t‐test as compared with respective gene levels in untreated CD4 controls.

Article Snippet: Real‐time PCR was performed on ‘mouse Th17 response’ RT 2 profiler PCR arrays (Qiagen) according to the manufacturer’s instructions.

Techniques: Expressing, Cell Differentiation

Journal: Immunology

Article Title: Glycogen synthase kinase‐3 promotes T helper type 17 differentiation by promoting interleukin‐9 production

doi: 10.1111/imm.13199

Figure Lengend Snippet: Glycogen synthase kinase‐3 (GSK3) inhibitors still inhibit T helper type 17 (Th17) ‐dependent genes in RORγT−/− CD4 cells. RORγT−/− CD4 cells were differentiated toward Th17 cells for 2 days with or without treatment with LiCl (10 mm) or CHIR99021 (1 μm). Gene expression of (a) Ccl20, (b) Ccr6, (c) IL‐9, (d) Irf4, or (e) Runx1. Fold change was calculated as (2−Δ Ct of a sample)/(average 2−Δ Ct of wild‐type CD4 cells). Gapdh was used as the endogenous control gene to normalize individual gene expressions. Each symbol represents an independent cell differentiation from the pool of CD4+ cells from two RORγT−/− mice (n = 3 or n = 4). One‐way analysis of variance F(2, 9) = 154·2 (a), F(2, 9) = 62·31 (b), F(2, 8) = 36·75 (c), F(2, 8) = 14·76 (d), F(2, 9) = 1·940 (e). *P < 0·05, **P < 0·01, Tukey’s post‐hoc test, compared with the respective gene levels in wild‐type CD4 cells, # P < 0·05, ## P < 0·05 compared with the gene expression levels in untreated RORγT−/− CD4 cells.

Article Snippet: Real‐time PCR was performed on ‘mouse Th17 response’ RT 2 profiler PCR arrays (Qiagen) according to the manufacturer’s instructions.

Techniques: Expressing, Cell Differentiation

Journal: Immunology

Article Title: Glycogen synthase kinase‐3 promotes T helper type 17 differentiation by promoting interleukin‐9 production

doi: 10.1111/imm.13199

Figure Lengend Snippet: Effects of interleukin‐9 (IL‐9) and/or glycogen synthase kinase‐3 (GSK3) inhibitors on T helper type 17 (Th17) cell differentiation. (a) IL‐17A+ CD4+ cells at day 4 of culture of cells from wild‐type (WT) mice. WT CD4 cells were differentiated towards Th17 cells and treated or not with IL‐9 (40 ng/ml) and/or GSK3 inhibitors [LiCl (10 mm), CHIR99021 (1 μm), and TDZD8 (1 μm)] for 4 days. (b) WT (GSK3β Flox/Flox) and GSK3β −/− CD4 cells were differentiated toward Th17 cells with or without IL‐9 (40 ng/ml) for 4 days. IL‐17A+CD4+ cells were analyzed by flow cytometry and were gated on CD4+ cells. Fold change was calculated as percentage of cells of a sample/average percentage of cells of untreated WT (GSK3β Flox/Flox) CD4 cells. WT CD4 cells cultured without GSK3 inhibitors and IL‐9 were used as controls. Each symbol represents an independent cell differentiation (n = 7 to n = 9 for a, n = 3 to n = 7 for b). Two‐way analysis of variance F(1, 28) = 22·68 (a – GSK3 inhibitors), F(1, 28) = 23·74 (a – IL‐9); F(1, 18) = 63·93 (b – mice type), F(1, 18) = 159·6 (b – IL9), F(1, 18) = 17·53 (b – interaction), *P < 0·05, **P < 0·01, Tukey’s post‐hoc test, compared with controls.

Article Snippet: Real‐time PCR was performed on ‘mouse Th17 response’ RT 2 profiler PCR arrays (Qiagen) according to the manufacturer’s instructions.

Techniques: Cell Differentiation, Flow Cytometry, Cell Culture

Journal: Immunology

Article Title: Glycogen synthase kinase‐3 promotes T helper type 17 differentiation by promoting interleukin‐9 production

doi: 10.1111/imm.13199

Figure Lengend Snippet: Proteins binding to the IL‐17a promoter determined by chromatin immunoprecipitation (ChIP). Wild‐type (WT) CD4 cells were differentiated towards Th17 cells and treated or not with interleukin‐9 (IL‐9; 40 ng/ml) and/or glycogen synthase kinase‐3 (GSK3) inhibitors [LiCl (10 mm), CHIR99021 (1 μm), and TDZD8 (1 μm)] for 4 days. ChIP was performed by immunoprecipitating with either signal transducer and activator of transcription 3 (STAT3) (a), STAT5 (b), or GSK3β (c), and DNA from the IL‐17a promoter region was amplified. All results were normalized by respective input values. Each symbol represents an independent cell differentiation (n = 3 to n = 6). Two‐way analysis of variance for a–c, (a) F(1, 13) = 7·561 (GSK3 inhibitors), F(1, 13) = 7·236 (IL‐9), F(1, 13) = 5·910 (interaction); (b) F(1, 13) = 38·16 (GSK3 inhibitors), F(1, 13) = 4·487 (IL9); (c) F(1, 21) = 13·44 (GSK3 inhibitors), F(1, 21) = 32·09 (IL‐9), F(1, 21) = 9·510 (interaction). *P < 0·05, **P < 0·01, Tukey’s post‐hoc test for (a)–(c).

Article Snippet: Real‐time PCR was performed on ‘mouse Th17 response’ RT 2 profiler PCR arrays (Qiagen) according to the manufacturer’s instructions.

Techniques: Binding Assay, Chromatin Immunoprecipitation, Amplification, Cell Differentiation

Journal: Immunology

Article Title: Glycogen synthase kinase‐3 promotes T helper type 17 differentiation by promoting interleukin‐9 production

doi: 10.1111/imm.13199

Figure Lengend Snippet: Signal transducer and activator of transcription 3 (STAT3) is required for the interleukin‐9 (IL‐9) effect on T helper type 17 (Th17) cell differentiation. Wild‐type (WT) CD4 cells were transfected with 50 moi of Ad‐GFP (GFP) or 50 moi of Ad‐STAT3C (STAT3C) and differentiated towards Th17 cells with or without IL‐9 (40 ng/ml) and/or glycogen synthase kinase‐3 (GSK3) inhibitors [LiCl (10 mm), CHIR99021 (1 μm), and TDZD8 (1 μm)] for 4 days. IL‐17A+ CD4+ cells were analyzed by flow cytometry and gated on CD4+ cells. Each symbol represents an independent cell differentiation from the pool of CD4+ cells from three mice (n = 3–4). One‐way analysis of variance F(7, 18) = 34·80,**P < 0·01, Tukey’s post‐hoc test, compared with GFP CD4 cells.

Article Snippet: Real‐time PCR was performed on ‘mouse Th17 response’ RT 2 profiler PCR arrays (Qiagen) according to the manufacturer’s instructions.

Techniques: Cell Differentiation, Transfection, Flow Cytometry