Journal: Immunology

Article Title: Engagement of glycosylphosphatidylinositol-anchored proteins results in enhanced mouse and human invariant natural killer T cell responses

doi: 10.1111/j.1365-2567.2010.03369.x

Figure Lengend Snippet: Thy-1 cross-linking boosts α-galactosylceramide (αGC)-mediated cytokine secretion by primary mouse natural killer (NKT) cells. Hepatic lymphoid mononuclear cells were stained with a FITC-conjugated anti-T-cell receptor-β (TCR β) mAb, a phycoerythrin-conjugated anti-NK1.1 monoclonal antibody and either PBS-57-loaded or -unloaded CD1d tetramer (labelled with allophycocyanin). TCR-β + NK1.1 + cells were gated on (a; left panels) and assessed for their reactivity with CD1d tetramer reagents (a; right panels). Highly pure TCR β + NK1.1 + hepatic NKT cells were sorted and co-cultured with bone marrow-derived dendritic cells at a ratio of five i NKT cells to one bone marrow-derived dendritic cell. Cells were incubated with αGC alone (100 ng/ml), a low (5 μg/ml) or high (40 μg/ml) dose of G7, or a combination αGC and G7. After 48 hr, culture supernatants were harvested and interferon-γ (IFN-γ) levels (b) and interleukin-4 (IL-4) levels (c) were measured by ELISA. Statistical significance is denoted by asterisks, where ** and *** represent P

Article Snippet: The cytokine content of samples was quantified by ELISA using Ready-Set-Go kits for mouse IL-2, IL-4 and interferon-γ (IFN-γ) (eBioscience).

Techniques: Staining, Cell Culture, Derivative Assay, Incubation, Enzyme-linked Immunosorbent Assay

Journal: Immunology

Article Title: Engagement of glycosylphosphatidylinositol-anchored proteins results in enhanced mouse and human invariant natural killer T cell responses

doi: 10.1111/j.1365-2567.2010.03369.x

Figure Lengend Snippet: Thy-1 cross-linking augments α-galactosylceramide (αGC)-mediated cytokine secretion by mouse CD4 + invariant natural killer ( i NKT) cells. N38-2C12 cells were co-incubated with bone marrow-derived dendritic cells and stimulated with αGC alone (100 ng/ml), with a low (5 μg/ml) or high (40 μg/ml) dose of G7, or with a combination of αGC and G7. After 24 hr, culture supernatants were harvested and interleukin-2 (IL-2), interferon-γ (IFN-γ) and IL-4 levels were measured by ELISA. ** and *** represent P

Article Snippet: The cytokine content of samples was quantified by ELISA using Ready-Set-Go kits for mouse IL-2, IL-4 and interferon-γ (IFN-γ) (eBioscience).

Techniques: Incubation, Derivative Assay, Enzyme-linked Immunosorbent Assay

Journal: Immunology

Article Title: Engagement of glycosylphosphatidylinositol-anchored proteins results in enhanced mouse and human invariant natural killer T cell responses

doi: 10.1111/j.1365-2567.2010.03369.x

Figure Lengend Snippet: Thy-1 cross-linking enhances α-galactosylceramide (αGC)-mediated cytokine production by mouse CD4 − CD8 − invariant natural killer ( i NKT) cells. DN32.D3 cells were incubated with αGC alone (100 ng/ml), with a low (5 μg/ml) or high (40 μg/ml) dose of G7, or with a combination of αGC and G7. After 24 hr, culture supernatants were harvested and interleukin-2 (IL-2), interferon-γ (IFN-γ) and IL-4 levels were measured by ELISA (a). After 24 hr, cells were harvested and total RNA was extracted. RNA was reverse transcribed into cDNA, which was then amplified by quantitative PCR using primers specific for IL-2, IFN-γ and IL-4. Cytokine mRNA levels were normalized to glyceraldehyde 3-phosphate dehydrogenase and the normalized values were graphed as expression ratios. Expression ratios were relative to untreated cells, which were assigned a value of 1 (b). Statistical significance is denoted by asterisks, where ** and *** represent P

Article Snippet: The cytokine content of samples was quantified by ELISA using Ready-Set-Go kits for mouse IL-2, IL-4 and interferon-γ (IFN-γ) (eBioscience).

Techniques: Incubation, Enzyme-linked Immunosorbent Assay, Amplification, Real-time Polymerase Chain Reaction, Expressing

Journal: Frontiers in Immunology

Article Title: Endothelial Cell Amplification of Regulatory T Cells Is Differentially Modified by Immunosuppressors and Intravenous Immunoglobulin

doi: 10.3389/fimmu.2017.01761

Figure Lengend Snippet: The effect of mycophenolic acid (MPA), cyclosporine A (CsA), or intravenous immunoglobulin (IVIg) on endothelial cell (EC)-mediated allogeneic CD4 + -T cell polarization is maintained when PBMC have been exposed to immunosuppressors or IVIg. Figure shows the proportion of interleukin-17 producing cells [ (A) , n > 5 donors], interferon γ (IFN-γ) producing cells [ (B) , n > 5 donors], T memory cells [ (C) , n > 6 donors], and their proliferation [ (D) , n > 6 donors], Treg [ (E) , n > 6 donors], and Treg proliferation [ (F) , n > 6 donors]. Like ECs, PBMC were pretreated with MPA, CSA, or IVIg prior to coculture. In all conditions, PBMC and ECs were treated with the same immunomodulators. The concentrations used for stimulation of ECs and PBMC were 3 µg/ml for MPA, 5 µg/ml for CsA, and 1 mg/ml for IVIg. Results are expressed as the relative percentage of the control values corresponding to ECs treated with vehicle and PBMC treated by immunomodulators (represented by dotted lines). In all figures, horizontal lines show median values (* p

Article Snippet: ECs Pretreatment with Immunosuppressors Endothelial cells were cultured with interferon γ (IFN-γ) (200 IU/ml for 3 days; R & D Systems, Minneapolis, MN, USA) in tissue culture flasks and incubated, where indicated, with immunosuppressors: MPA (Sigma-Aldrich), CsA (Sandimmun® , Novartis), or with IVIg (Privigen® , CSL Behring) at the indicated concentrations.

Techniques:

Journal: Frontiers in Immunology

Article Title: Endothelial Cell Amplification of Regulatory T Cells Is Differentially Modified by Immunosuppressors and Intravenous Immunoglobulin

doi: 10.3389/fimmu.2017.01761

Figure Lengend Snippet: Effect of endothelial cell (EC) incubation with mycophenolic acid (MPA), cyclosporine A (CsA), and intravenous immunoglobulin (IVIg) on the proinflammatory activity of ECs. IL-6 secretion was quantified in the supernatants of cocultures where ECs was treated with MPA [ (A) , n > 19 donors], CsA [ (B) , n = 24 donors] or IVIg [ (C) , n > 6 donors]. IL-6 production by ECs incubated with vehicle alone is represented as 0. (D–F) show the expansion of the Th17 subset after a 7 day coculture of PBMC with EC pretreated with interferon γ (IFN-γ) and the indicated dose of MPA [ (D) , n = 7 donors], CsA [ (E) , n = 9 donors], or IVIg [ (F) , n > 18 donors]. Expansion of the Th1 subset under the same conditions is shown in [ (G) , n > 7 donors], [ (H) , n = 9 donors], and [ (I) , n > 18 donors]. Results are expressed as the relative percentage of the control values (ECs treated with vehicle alone, represented by dotted lines). Thick, horizontal lines represent median values in all the cocultures (* p

Article Snippet: ECs Pretreatment with Immunosuppressors Endothelial cells were cultured with interferon γ (IFN-γ) (200 IU/ml for 3 days; R & D Systems, Minneapolis, MN, USA) in tissue culture flasks and incubated, where indicated, with immunosuppressors: MPA (Sigma-Aldrich), CsA (Sandimmun® , Novartis), or with IVIg (Privigen® , CSL Behring) at the indicated concentrations.

Techniques: Incubation, Activity Assay

Journal: Frontiers in Immunology

Article Title: Endothelial Cell Amplification of Regulatory T Cells Is Differentially Modified by Immunosuppressors and Intravenous Immunoglobulin

doi: 10.3389/fimmu.2017.01761

Figure Lengend Snippet: Intravenous immunoglobulin (IVIg) pretreatment of endothelial cells (ECs) induces selective amplification of regulatory T cells in contrast with the commonly used immunosupressors mycophenolic acid (MPA) and cyclosporine A (CsA). (A,C,E) show the proportion of Treg expanded in cocultures of PBMC with EC pretreated with interferon γ (IFN-γ) and the indicated concentrations of MPA [ (A) , n = 8 donors], CsA [ (B) , n = 12 donors], or IVIg [ (C) , n > 13 donors]. [ (D) , n = 7 donors], [ (E) , n = 10 donors], and [ (F) , n > 12 donors] show the proliferation of Treg after MPA, CsA, or IVIg treatment of ECs, respectively. Results are expressed as the relative percentage of the control values (ECs treated with vehicle alone and represented by dotted lines). Thick, horizontal lines show the median values in each data set (* p

Article Snippet: ECs Pretreatment with Immunosuppressors Endothelial cells were cultured with interferon γ (IFN-γ) (200 IU/ml for 3 days; R & D Systems, Minneapolis, MN, USA) in tissue culture flasks and incubated, where indicated, with immunosuppressors: MPA (Sigma-Aldrich), CsA (Sandimmun® , Novartis), or with IVIg (Privigen® , CSL Behring) at the indicated concentrations.

Techniques: Amplification

Journal: Immunology

Article Title: Cooperative action of CD8 T lymphocytes and natural killer cells controls tumour growth under conditions of restricted T-cell receptor diversity

doi: 10.1111/j.1365-2567.2009.03150.x

Figure Lengend Snippet: P1A peptide dose-dependent proliferation, activation and interferon-γ (IFN-γ) secretion by TCRP1A DBA/2 and B10.D2 CD8 + T cells. (A) CFSE-labelled naïve lymph node (LN) CD8 T cells from TCRP1A DBA/2 (left) and TCRP1A B10.D2 (right)

Article Snippet: Interferon-γ (IFN-γ) in T-cell culture supernatants harvested following activation on day 3 was determined by enzyme-linked immunosorbent assay (ELISA) using mAb AN18 with biotin-conjugated R46A2 (PharMingen, San Diego, CA).

Techniques: Activation Assay

Journal: Immunology

Article Title: Early delayed-type hypersensitivity eosinophil infiltrates depend on T helper 2 cytokines and interferon-? via CXCR3 chemokines

doi: 10.1111/j.0019-2805.2004.01818.x

Figure Lengend Snippet: Keyhole limpet haemocyanin (KLH) delayed-type hypersensitivity (DTH) is dependent on interferon-γ (IFN-γ) on day 3 and day 5 after immunization. Left column: DTH was elicited on day 3 and 5 in IFN-γ +/+ mice (Group C and E), but

Article Snippet: Quantitative sandwich enzyme-linked immunosorbent assay (ELISA) was performed on ear extracts for IL-4, eotaxin, IL-5, interferon-γ (IFN-γ) and IP-10, using two different specific mAbs [all BD PharMingen, San Diego, CA, except for eotaxin (R & D, Minneapolis, MN) and IP-10 (Leinco Technologies, St Louis, MO)], in 0·1 m NaHCO3 (pH 8–9) at 4°.

Techniques: Mouse Assay

Journal: Immunology

Article Title: Early delayed-type hypersensitivity eosinophil infiltrates depend on T helper 2 cytokines and interferon-? via CXCR3 chemokines

doi: 10.1111/j.0019-2805.2004.01818.x

Figure Lengend Snippet: Production of interferon-γ (IFN-γ) and IP-10 [IFN-γ-inducible protein chemokine of 10 000 molecular weight (CXCL10)] in 24-hr keyhole limpet haemocyanin (KLH) delayed-type hypersensitivity (DTH) ear extracts, and dependency of

Article Snippet: Quantitative sandwich enzyme-linked immunosorbent assay (ELISA) was performed on ear extracts for IL-4, eotaxin, IL-5, interferon-γ (IFN-γ) and IP-10, using two different specific mAbs [all BD PharMingen, San Diego, CA, except for eotaxin (R & D, Minneapolis, MN) and IP-10 (Leinco Technologies, St Louis, MO)], in 0·1 m NaHCO3 (pH 8–9) at 4°.

Techniques: Molecular Weight

Journal: Journal of Veterinary Research

Article Title: Toxicological Evaluation of Flumequine in Pubertal Male Rats After Oral Administration for Six Weeks

doi: 10.1515/jvetres-2018-0012

Figure Lengend Snippet: Changes in plasma cytokines in Wistar male rats treated with corn oil (control) or flumequine (FLU) 53, 200, and 750 mg/kg for six weeks. IL-10 – interleukin-10; IL-6 – interleukin-6; TNF-α – tumour necrosis factor-α; IFN-γ – interferon-γ; GM-CSF – granulocyte-macrophage colony-stimulating factor; IL-1β – interleukin-1β; IL-2 – interleukin-2. Values are mean ±SD for seven rats in each group. ∗ P

Article Snippet: Serum concentrations of interleukin-10 (IL-10), interleukin-6 (IL-6), tumour necrosis factor-α (TNFα), interferon-γ (IFN-γ), granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-1β (IL-1β), and interleukin-2 (IL-2) were determined with a Bio-plex rat cytokine-9-plex A panel (Bio-Rad, USA).

Techniques:

Journal: Breast Cancer : Targets and Therapy

Article Title: A GM-CSF and CD40L bystander vaccine is effective in a murine breast cancer model

doi: 10.2147/BCTT.S89563

Figure Lengend Snippet: Immune response analysis. Notes: ( A ) Average ELISPOT counts for interferon-γ (IFN-γ) and interleukin-2 (IL-2) between the 4T1 alone (4T1) and 4T1 + BCG (BCG) harvested lymphocytes cocultured with 4T1 lysates (top). The ELISPOT images above each bar are representative of the spot counts for each of the four groups. ( B ) The IFN-γ average ELISPOT counts are shown in the LLC tumor antigen alone versus the BCG vaccinated LLC mice. All experiments were done in triplicate. Spot counts in the treatment groups were adjusted for background using the no-tumor mouse lymphocytes. Error bars are the SD’s for the groups. * P -value statistically significant. Abbreviations: ELISPOT, enzyme-linked immunospot; LLC, Lewis lung carcinoma; SD, standard deviation.

Article Snippet: IL-2 and IFN-γ ELISPOT Harvested lymph nodes from each mouse were subjected to ELISPOT analysis using cytokine ELISPOT kits for mouse interleukin-2 (IL-2) and interferon-γ (IFN-γ) from Mabtech (Cincinnati, OH, USA).

Techniques: Enzyme-linked Immunospot, Mouse Assay, Standard Deviation

Journal: Frontiers in Immunology

Article Title: Ultrasensitive Quantification of Cytokine Proteins in Single Lymphocytes From Human Blood Following ex-vivo Stimulation

doi: 10.3389/fimmu.2018.02462

Figure Lengend Snippet: Cytokine capture assay for IFN-γ and TNF-α in CD8 T cells. Examples of gating strategies for sorting cells that were either IFN-γ and TNF-α positive. Cells were labeled with CD8 and (A) IFN-γ using a bispecific CD45/IFN-γ capture reagent; (B) TNF-α using a bispecific CD45/TNF-α capture reagent; or (C) TNF-α using a TNF-α capture assay with TNF-α processing inhibitor (TAPI-0). Stained cells were acquired and sorted on BD-FACS-Aria sorter.

Article Snippet: Reagents The SiMoA HD-1 analyzer, SiMoA consumables, and IFN-γ (SiMoA™ IFN-γ,138 Kit) and TNF-α (SiMoA™ TNF-α 2.0, 208 Kit) were purchased from Quanterix, Lexington, MA.

Techniques: Labeling, Staining, FACS

Journal: Frontiers in Immunology

Article Title: Ultrasensitive Quantification of Cytokine Proteins in Single Lymphocytes From Human Blood Following ex-vivo Stimulation

doi: 10.3389/fimmu.2018.02462

Figure Lengend Snippet: IFN-γ protein quantification at different mean fluorescent intensities (MFI). (A) Flow cytometry dot plot showing gating strategy for sorting CD8+IFN-γ+ cells at different MFI's using bin gating. The gates were set as such that the MFI values fall in a range of 100,000 with 8 different intensities following a two-fold incremental order and covering from the brightest to the dimmest IFN-γ expression on CD8 cells (B) Measured amounts of IFN-γ from cells sorted from various levels of staining for cells surface IFN-γ expressed as mean ± SD fluorescence intensity channel numbers (MFI).

Article Snippet: Reagents The SiMoA HD-1 analyzer, SiMoA consumables, and IFN-γ (SiMoA™ IFN-γ,138 Kit) and TNF-α (SiMoA™ TNF-α 2.0, 208 Kit) were purchased from Quanterix, Lexington, MA.

Techniques: Flow Cytometry, Cytometry, Expressing, Staining, Fluorescence

Journal: Frontiers in Immunology

Article Title: Ultrasensitive Quantification of Cytokine Proteins in Single Lymphocytes From Human Blood Following ex-vivo Stimulation

doi: 10.3389/fimmu.2018.02462

Figure Lengend Snippet: Quantification of IFN-γ in single CD8 T cells. (A) Reproducibility of the SiMoA standard curves for IFN-γ. Standard concentrations of IFN-γ were used to generate curves to calculate the unknown concentrations in cells (B) Comparison of measured IFN-γ protein at 1, 2, 5, and10 cell levels in CD8 cells. The amount of IFN-γ measured in femtograms of lysates from cell surface IFN-γ-positive cells or cells that were cell surface IFN-γ-negative. Left are data from IFN-γ-positive cells, right are data from IFN-γ-negative cells. Horizontal axis is the number of cells sorted into each lysate. (C) Competitive inhibition assay for IFN-γ assay. Five and ten cells were sorted in 25 μl of lysis buffer and the lysates were incubated with or without anti-IFN-γ inhibition antibody for 45 mins at 4°C. Lysates were transferred to V bottom SiMoA plates and IFN-γ assay was performed. Data are represented as mean and mean ± SD and differences among means were calculated using Mann–Whitney test. P

Article Snippet: Reagents The SiMoA HD-1 analyzer, SiMoA consumables, and IFN-γ (SiMoA™ IFN-γ,138 Kit) and TNF-α (SiMoA™ TNF-α 2.0, 208 Kit) were purchased from Quanterix, Lexington, MA.

Techniques: Inhibition, Lysis, Incubation, MANN-WHITNEY

Journal: BMC Immunology

Article Title: Lupus-like oral mucosal lesions in mercury-induced autoimmune response in Brown Norway rats

doi: 10.1186/1471-2172-14-47

Figure Lengend Snippet: Tissue expression of interleukin-4 (IL-4) and interferon-γ (IFN-γ) mRNA in the tongue of the mercury-treated rats by real-time reverse transcription-polymerase chain reaction (qRT-PCR). A and B : qRT-PCR analyses examine expression levels of mRNA encoding IL-4 (A) and IFN-γ (B) normalized by those of glyceraldehyde-3-phosphate dehydrogenase (G3PDH). Data represents mean ± standard error (SE) of pooled data derived from three to five independent experiments. *, significantly different at p

Article Snippet: Predesigned primers and probe reagents for rat interleukin-4 (IL-4), interferon-γ (IFN-γ), and glyceraldehyde-3-phosphate dehydrogenase (G3PDH) were commercially obtained from Roche Diagnostics.

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Quantitative RT-PCR, Derivative Assay

Journal: Stem Cells and Development

Article Title: Nuclear Receptors Nur77 and Nurr1 Modulate Mesenchymal Stromal Cell Migration

doi: 10.1089/scd.2011.0076

Figure Lengend Snippet: Increased cytokine production in Nur77- and Nurr1-transduced FBMSC does not influence T-cell proliferation. (A) Four days after transduction, IL-6 and IL-8 protein secreted in supernatant of unstimulated FBMSC cultures were measured by ELISA after 24 h incubation and mRNA levels of HGF , TGF-β1 , and IDO were analyzed by RQ-PCR. Basal protein levels of IL-6 and IL-8 and HGF mRNA expression were significantly increased in Nur77- or Nurr1-transduced FBMSC compared with mock-transduced FBMSC. Data (mean±SD) normalized to unstimulated mock-transduced MSC. (B) Transduced FBMSC were stimulated with TNF-α and IFN-γ for 24 h or left untreated. Again the highest levels of IL-6 and IL-8 protein and HGF mRNA were observed in Nur77- and Nurr1-tranduced FBMSC. Data (mean±SD) normalized to unstimulated mock-transduced MSC. (C, D) Transduced FBMSC were cocultured with CD3/CD28 bead-activated PBMC. FBMSC with enhanced expression of Nur77 (C) or Nurr1 (D) have at least a similar capacity to inhibit T-cell proliferation compared with mock-transduced FBMSC. Beads were added to PBMC as negative control (CCPM count 49.157). Data represent pooled data (mean±SD) of 3 individual FBMSC donors cocultured with the same allogeneic PBMC. IL-6, interleukin-6; IL-8, interleukin-8; HGF, hepatocyte growth factor; TGF-β1, transforming growth factor-β1; IDO, indoleamine-2,3-dioxygenase; TNF-α, tumor necrosis factor-α; IFN-γ, interferon-γ; PBMC, peripheral blood mononuclear cells; CCPM, radioactivity counts per minute. * P ≤0.05.

Article Snippet: After 24 h, the medium was refreshed and cells were stimulated with tumor necrosis factor-α (TNF-α) (1,000 U/mL; Peprotech) or interferon-γ (IFN-γ) (1,000 U/mL; Peprotech) or left untreated.

Techniques: Transduction, Enzyme-linked Immunosorbent Assay, Incubation, Polymerase Chain Reaction, Expressing, Negative Control, Radioactivity

Journal: Life Science Alliance

Article Title: Absence of MHC-II expression by lymph node stromal cells results in autoimmunity

doi: 10.26508/lsa.201800164

Figure Lengend Snippet: Selective abrogation of endogenous MHCII expression in LNSCs. (A) Schematic representation of K14tgpIV KO mice. Briefly, pIV KO were crossed with transgenic mice expressing the full CIITA cDNA under the control of the keratin 14 (K14) promoter ( K14tg ). (B) Relative mRNA expression of K14 by LECs, BECs, FRCs, B cells and cTEC from K14tgpIV KO mice, measured by qPCR and normalized to GAPDH. Data are representative of two experiments with 10 mice pooled/group (C) Flow cytometry histograms showing the expression of MHCII molecules by cTECs (gated on CD45 neg EpCAM + Ly5.1 hi cells) and mTECs (gated on CD45 neg EpCAM + Ly5.1 int cells) from indicated mice. (D) Flow cytometry dot plots showing CD8 + and CD4 + T-cell frequencies in LN of indicated mice (gated on CD3 + cells). (C, D) Data are representative of two experiments with three mice/group. (E) Histograms showing MHCII expression (MFI) by LECs, BECs, and FRCs from LN of indicated mice, either naive (filled) or injected s.c. with IFN-γ 24 h before (hatched). (F) Graphs showing MHCII expression (MFI) by CD11b + cDCs (CD11c hi CD11b + ), CD8α + DCs (CD11c hi CD8α + ), pDCs (CD11c int PDCA-1 + ), B cells (CD19 + ), and monocytes/macrophages (CD11c neg CD11b + ) isolated from LNs of indicated mice. (E, F) Data are representative of two experiments with 3-4 mice/group.

Article Snippet: When indicated, IFN-γ (Peprotech) was injected subcutaneously (both flanks and neck, 1 μg/50 μl), and FTY720 (Sigma-Aldrich) (20 μg) was injected every day for 6 d before the experiment.

Techniques: Expressing, Mouse Assay, Transgenic Assay, Real-time Polymerase Chain Reaction, Flow Cytometry, Cytometry, Injection, Isolation

Journal: Life Science Alliance

Article Title: Absence of MHC-II expression by lymph node stromal cells results in autoimmunity

doi: 10.26508/lsa.201800164

Figure Lengend Snippet: T cells isolated from mice lacking MHCII expression in LNSCs induce autoimmunity upon transfer into immunodeficient mice. (A–E) T cells isolated from LNs and spleen of 18-mo-old K14tgpIV KO (white) and control mice (black) were adoptively transferred into Rag2 −/− recipient mice that were analysed 4 mo later. (A) Mouse weight. Data are pooled from four mice and representative of two experiments. (B) Sections of small intestines from recipient Rag2 −/− mice were stained with antibodies against CD45 (green), CD3 (red), and with DAPI (blue). Pictures show representative tissue sections. Graphs represent the quantification of CD3 + T cells/area and are pooled from 12 tissue sections from three individual mice/group. (C–E) Flow cytometry dot plots showing the frequency of CD62L hi CD44 lo CD4 + and CD8 + T cells (C), the frequency of CD4 + T cell–producing IFN-γ or IL-17 and the frequency of CD8 + T cell–producing IFN-γ in LNs (D) and Treg frequencies (E) in LNs of transferred Rag2 −/− recipient mice. Data are representative of two experiments with 2–4 mice/group each. (C–E) * P

Article Snippet: When indicated, IFN-γ (Peprotech) was injected subcutaneously (both flanks and neck, 1 μg/50 μl), and FTY720 (Sigma-Aldrich) (20 μg) was injected every day for 6 d before the experiment.

Techniques: Isolation, Mouse Assay, Expressing, Staining, Flow Cytometry, Cytometry

Journal: Life Science Alliance

Article Title: Absence of MHC-II expression by lymph node stromal cells results in autoimmunity

doi: 10.26508/lsa.201800164

Figure Lengend Snippet: K14tgpIV KO mice exhibit enhanced T-cell activation and impaired Treg frequencies in LNs upon aging. K14tgpIV KO (white) and K14tg control mice (black) were analysed at 4 and 18 mo. (A) Frequencies of naive (CD62L hi CD44 lo ) and activated/memory (CD62L lo CD44 hi ) CD4 + and CD8 + T cells. (B) Frequencies of PD-1 + , IFN-γ, and IL-17 producing cells among CD4 + and/or CD8 + T cells. (C, D) Foxp3 + Treg identification by flow cytometry (CD4 + CD25 + Foxp3 + ) (C) and IHC staining (Foxp3 + ) (D) in lymph nodes of indicated mice. (A–D) Data are representative of three experiments with 3–7 mice/group. * P

Article Snippet: When indicated, IFN-γ (Peprotech) was injected subcutaneously (both flanks and neck, 1 μg/50 μl), and FTY720 (Sigma-Aldrich) (20 μg) was injected every day for 6 d before the experiment.

Techniques: Mouse Assay, Activation Assay, Flow Cytometry, Cytometry, Immunohistochemistry, Staining

Journal: Life Science Alliance

Article Title: Absence of MHC-II expression by lymph node stromal cells results in autoimmunity

doi: 10.26508/lsa.201800164

Figure Lengend Snippet: LNSCs upte MHCII and MHCII-associated molecules in response to IFN-γ. (A) Flow cytometry and plotted histograms showing MHCII expression (MFI) by LECs, BECs, and FRCs from LN of indicated naive mice. MHCII coming from DC (blue arrow)- or endogenous (red arrow)-origin, is indicated. (B) WT or pIV CIITA–deficient mice (pIV KO ) mice were injected subcutaneously with PBS or 1 μg of IFN-γ; LNSCs were sorted by flow cytometry 24 h later. Histograms represent I-A β (upper panel) and H-2Mα (lower panel) relative mRNA expression by LECs, BECs, and FRCs from indicated mice, measured by qPCR and normalized to GAPDH. Data are representative of two experiments with 10 mice pooled/group. (C) LNSCs and DCs from draining LNs were analysed 24 h after IFN-γ injection (as previously) by flow cytometry. Dot plots represent the expression of MHCII and H-2M by LECs, BECs, FRCs, and DCs from indicated mice. Frequency of double positive fractions are highlighted in red. Data are representative of two experiments with three mice/group.

Article Snippet: When indicated, IFN-γ (Peprotech) was injected subcutaneously (both flanks and neck, 1 μg/50 μl), and FTY720 (Sigma-Aldrich) (20 μg) was injected every day for 6 d before the experiment.

Techniques: Flow Cytometry, Cytometry, Expressing, Mouse Assay, Injection, Real-time Polymerase Chain Reaction

Journal: Life Science Alliance

Article Title: Absence of MHC-II expression by lymph node stromal cells results in autoimmunity

doi: 10.26508/lsa.201800164

Figure Lengend Snippet: Deletion of MHCII on LECs decreases Treg proliferation. (A) MHCII and IFN-γ receptor (IFNγR) expression (MFI) by LECs, BECs, and FRCs from LN of WT mice of the indicated age. Data are representative of two experiments with three mice/group each. (B–D) LN sections depicting LECs (Lyve-1, white) and proliferating Tregs (red arrows) (Ki67, red; and Foxp3, green) from 18-mo-old K14 tg and K14 tg pIV KO mice (B), 3-mo-old WT and K14 tg and K14 tg pIV KO (C), and 3-mo-old Tamoxifen-treated Prox-1-Cre ERT2 MHCII fl and MHCII fl control mice (D). In (C) and (D), animals were treated with FTY720 every day for the last 6 d, and with IFN-γ 6 d before harvesting the LNs. Graphs represent the percentages of Ki67 + among Foxp3 + cells in contact with LECs from LN of the indicated mice. (B–D) Data are pooled from at least nine tissue sections from three individual mice/group. * P

Article Snippet: When indicated, IFN-γ (Peprotech) was injected subcutaneously (both flanks and neck, 1 μg/50 μl), and FTY720 (Sigma-Aldrich) (20 μg) was injected every day for 6 d before the experiment.

Techniques: Expressing, Mouse Assay

Journal: The Journal of investigative dermatology

Article Title: Human Natural Killer T Cells Infiltrate into the Skin at Elicitation Sites of Allergic Contact Dermatitis

doi: 10.1038/sj.jid.5701199

Figure Lengend Snippet: Infiltrating NKT cells in ACD express both transcripts for the cytokines IFN-γ and IL-4 ACD skin-biopsy specimens studied by ISH: double-labeled with antisense oligoprobes for Vα24 (left panel, green; middle panel, cytokine ( a ) (IFN-γ) or ( b ) IL-4 (red); right panel, overlays) (bar=10 μm).

Article Snippet: Our ISH studies ( ) indicate that NKT cells contribute to this cytokine profile, in that we detected both IFN-γ and IL-4 gene expression in the infiltrating NKT cells in the elicitation phase of ACD.

Techniques: In Situ Hybridization, Labeling

Journal: The Journal of investigative dermatology

Article Title: Human Natural Killer T Cells Infiltrate into the Skin at Elicitation Sites of Allergic Contact Dermatitis

doi: 10.1038/sj.jid.5701199

Figure Lengend Snippet: CD1d+ Monocytic APCs, but not CD1d+ KC, induce cytokine gene expression and secretion by NKT-cells in vitro A polyclonal NKT cell line was cultured with the CD1d+ cell line, THP-1 or CD1d+ KC (differentiated or undifferentiated, and treated with IFN-γ, see Methods) in the presence ( a and b ) or absence (data not shown) of α-galactosylceramide for 24 hours, and supernatants were harvested for specific luminex assay. ( a ) IFN-γ; ( b ) IL-4. In another experiment, the NKT cells were isolated after 6 hours of incubation with the above cell lines, and RNA was extracted, cDNA was synthesized, and gene expression was studied by ( c ) real-time PCR IFN-γ; ( d ) IL-4. (* P

Article Snippet: Our ISH studies ( ) indicate that NKT cells contribute to this cytokine profile, in that we detected both IFN-γ and IL-4 gene expression in the infiltrating NKT cells in the elicitation phase of ACD.

Techniques: Expressing, In Vitro, Cell Culture, Luminex, Isolation, Incubation, Synthesized, Real-time Polymerase Chain Reaction

Journal: Vaccines

Article Title: Protective Cancer Vaccine Using Genetically Modified Hematopoietic Stem Cells

doi: 10.3390/vaccines6030040

Figure Lengend Snippet: Development of Ag-specific CD8 + T cells by genetic modification of HSCs following in vivo priming. TCR gene-transduced HSCs (GFP + ) in PBS were sorted, before being adoptively transferred into Thy1.1 congenic mice. On the following week, mice were i.p. injected with agonistic α-Notch2 Ab, rIL-7 and rFlt3L or a mouse IgG/PBS control. ( A ) After another two weeks, Thy1.2 + TCRVβ5 + cells from the pooled LNs and spleen were analyzed by flow cytometry, after gating on CD8 + T cell population (upper panels). A representative image from mice receiving the HSCs transduced with the Mig-TCR is shown, which was obtained after IgG control or agonistic α-Notch2 Ab, rIL-7 and rFlt3L protein injections. ( B ) Expression of CD25, CD69 and CD62L was analyzed by flow cytometry, after gating on CD8 + Thy1.2 + TCRVβ5 + T cells from the pooled LNs and spleen (dark lines; shaded areas indicate isotype controls). Data are representative of three independent experiments. ( C ) IL-2 and IFN-γ production (dark lines; shaded areas indicate isotype controls). The pooled LNs and spleen were stimulated with OVA 257–264 peptide and analyzed by intracellular cytokine staining, after gating on Thy1.2 + TCRVβ5 + cells. Data are representative of three independent experiments.

Article Snippet: PE, PE/Cy7, PerCP, PerCP/Cy5.5, or APC conjugated Thy1.2 (Clone 53-2.1), TCRVβ5 (MR9-4), TCRVα2 TCR (B20.1), CD44 (Clone IM7), IFN-γ (Clone XMG1.2) and IL-2 (JES6-5H4) were purchased from Biolegend (San Diego, CA, USA).

Techniques: Modification, In Vivo, Mouse Assay, Injection, Flow Cytometry, Cytometry, Transduction, Expressing, Staining

Journal: Vaccines

Article Title: Protective Cancer Vaccine Using Genetically Modified Hematopoietic Stem Cells

doi: 10.3390/vaccines6030040

Figure Lengend Snippet: Immunization using genetic modification of HSCs following in vivo priming induces strong Ag-specific T cell memory. T cell development and tumor injection were performed as described in Figure 4 . After various days, memory progenitors or memory T cells from the spleen and LNs were analyzed. Five mice were used for each time point. ( A ) The frequencies of Thy1.2 + at day 35, gating on CD8 + cells (left panels). Data are representative of three independent experiments ( n = 5). ( B ) Functional analysis using IFN-γ at day 35. Splenocytes were stimulated with OVA peptide for intracellular IFN-γ staining, gating on Thy1.2 + cells. Data are representative of three independent experiments ( n = 5). ( C ) Numbers of CD8 + Thy1.2 + T cells on indicated day. Data are represented as the mean ± SEM. ( D ) The absolute numbers of CD8 + Thy1.2 + T cells at day 35. Data are represented as the mean ± SEM from three independent experiments ( n = 5) ( p > 0.05, Student’s unpaired t -test).

Article Snippet: PE, PE/Cy7, PerCP, PerCP/Cy5.5, or APC conjugated Thy1.2 (Clone 53-2.1), TCRVβ5 (MR9-4), TCRVα2 TCR (B20.1), CD44 (Clone IM7), IFN-γ (Clone XMG1.2) and IL-2 (JES6-5H4) were purchased from Biolegend (San Diego, CA, USA).

Techniques: Modification, In Vivo, Injection, Mouse Assay, Functional Assay, Staining

Journal: The Journal of Experimental Medicine

Article Title: Chronic Inflammation and Susceptibility to Bacterial Infections in Mice Lacking the Polypeptide (p)105 Precursor (NF-?B1) but Expressing p50

doi:

Figure Lengend Snippet: Cytokine production in T cells or macrophages. ( A ) Secretion of IL-2, IL-4, and TNF-α is decreased in p105 −/− T cells. Splenic T cells purified from 3-wk-old control ( closed bars ) or p105 −/− ( open bars ) untreated ( 1 ) or treated with anti-CD3 ( 2 ) or anti-CD3 plus anti-CD28 ( 3 ) for 24 h. The cytokine levels in the supernatants were determined by ELISA. ( B ) Cytokines released from macrophages. Peritoneal macrophages purified from 3-wk-old mice untreated (−) or treated with (+) LPS and IFN-γ for 72 h. The cytokine levels are indicated by mean values ± SD.

Article Snippet: Purified T cells (5 × 105 /ml) were incubated with or without coated anti-CD3 antibody or coated anti-CD3 plus anti-CD28 antibodies for 24 h. Purified peritoneal macrophages (5 × 105 /ml) were incubated in the presence or absence of 1 μg/ml of LPS and 100 U/ml of IFN-γ (Genzyme Corp., Cambridge, MA) for 72 h. Cytokine levels for IL-1β, IL-4, IL-6, IL-10, GM-CSF, and TNF-α in supernatants were measured by ELISA (R & D Systems, Inc.); levels of IFN-γ and IL-2 were also determined by ELISA (BIOSOURCE, Camarillo, CA).

Techniques: Purification, Enzyme-linked Immunosorbent Assay, Mouse Assay

Journal: International journal of cancer. Journal international du cancer

Article Title: Lack of HLA class II antigen expression in microsatellite unstable colorectal carcinomas is caused by mutations in HLA class II regulatory genes

doi: 10.1002/ijc.25106

Figure Lengend Snippet: Restoration of HLA class II antigen expression in HDC108 cells by stable transfection with wild type RFX5 (left) and incubation with IFN-γ (500 IU/ml) for 48 h at 37 °C. Cells transfected with an empty vector (right) were used as a control.

Article Snippet: IFN-γ was obtained from PromoCell (Heidelberg, Germany).

Techniques: Expressing, Stable Transfection, Incubation, Transfection, Plasmid Preparation

Journal: Science signaling

Article Title: Simultaneous Profiling of 194 Distinct Receptor Transcripts in Human Cells

doi: 10.1126/scisignal.2003624

Figure Lengend Snippet: Stimulus-dependent changes in receptome profiles are dependent on cell type. ( A )Receptor abundances in 293T embryonic kidney cells and MCF7 breast carcinoma cells after stimulation with 100 ng ml −1 EGF for 4 hr, 200 U ml −1 IFN-γ for 4 hr, 5 Gy IR for 2 hr, or 20 ng ml −1 TNF for 4 hr. One-way hierarchical clustering was done using a Euclidean distance metric with Ward’s linkage after normalization to GAPDH as a loading control. Data were centered on the cell type-matched untreated (No tx) condition or the median observed abundance across both cell types if the receptor was absent for one of the untreated conditions. ( B )Plate-matched qRT-PCR quantification of the indicated receptor transcripts in 293T or MCF7 cells treated with TNF or IFN-γ, normalized so that the geometric-mean relative abundance of the indicated transcript in unstimulated cells equals one. For (A), data are shown as the median cycle threshold (approximate log 2 relative abundance) of three independent biological samples. For (B), data are shown as the geometric mean ± log-transformed s.e.m. of three independent biological samples. Asterisk indicates statistical significance ( P

Article Snippet: 293T cells were plated at 50,000 cells cm −2 and MCF7 cells at 25,000 cells cm−2 for 24 hr before stimulation with 100 ng ml−1 EGF (Peprotech) for 4 hr, 200 U ml−1 IFN- γ (Roche) for 4 hr, 5 Gy IR (60 Co) for 2 hr, or 20 ng ml−1 INF (Peprotech) for 4 hr.

Techniques: Quantitative RT-PCR, Transformation Assay

Journal: Science signaling

Article Title: Simultaneous Profiling of 194 Distinct Receptor Transcripts in Human Cells

doi: 10.1126/scisignal.2003624

Figure Lengend Snippet: qRT-PCR receptome profiling is significantly more sensitive for detecting receptor transcripts than conventional oligonucleotide microarrays. ( A )Present-absent calls for 177 receptor transcripts monitored on Affymetrix U133A microarrays were compared to receptome-profiling results for HT-29 cells treated with 200 U ml −1 IFN-γ for 24 hr. Statistical significance was assessed by Fisher’s exact test. ( B )Detection of FAS in HT-29 cells with or without IFN-γ sensitization. ( C )Caspase-3 cleavage in IFN-γ-treated HT-29 cells after FAS crosslinking with 1 µg ml −1 anti-APO for 24 hr. ( D )Replicated densitometry of caspase-3 cleavage in HT-29 cells. Data are shown as the mean relative abundance of cleaved caspase-3 (normalized so that the mean vehicle control equals one) ± s.e.m. of three independent samples, and asterisk indicates statistical significance ( P

Article Snippet: 293T cells were plated at 50,000 cells cm −2 and MCF7 cells at 25,000 cells cm−2 for 24 hr before stimulation with 100 ng ml−1 EGF (Peprotech) for 4 hr, 200 U ml−1 IFN- γ (Roche) for 4 hr, 5 Gy IR (60 Co) for 2 hr, or 20 ng ml−1 INF (Peprotech) for 4 hr.

Techniques: Quantitative RT-PCR

Journal: Science signaling

Article Title: Simultaneous Profiling of 194 Distinct Receptor Transcripts in Human Cells

doi: 10.1126/scisignal.2003624

Figure Lengend Snippet: qRT-PCR receptome profiling is more sensitive for detecting receptor transcripts than exon arrays. ( A )Receiver operating characteristic (ROC) curves relating exon array expression index to qRT-PCR present-absent calls for the indicated cells. HT-29 cells were treated with 200 U ml −1 IFN-γ for 24 hr. In all three ROC curves, the dashed line indicates expression index = 20, a representative threshold that properly distinguishes CSF1R expression in MCF10A and MDA-MB-436 cells. The area under the ROC curve (AUC, integrated from zero to one false-positive rate) indicates the overall quality of the present-absent classification based on exon array data, with AUC = 1 indicating perfect classification and AUC = 0.5 indicating random guessing. ( B )Detection of IL2RG in HT-29, MCF10A, and MDA-MB-436 cells but not in A375 cells. Cells were immunoblotted for IL2RG with tubulin used as a loading control. Immunoblots are representative of three independent experiments.

Article Snippet: 293T cells were plated at 50,000 cells cm −2 and MCF7 cells at 25,000 cells cm−2 for 24 hr before stimulation with 100 ng ml−1 EGF (Peprotech) for 4 hr, 200 U ml−1 IFN- γ (Roche) for 4 hr, 5 Gy IR (60 Co) for 2 hr, or 20 ng ml−1 INF (Peprotech) for 4 hr.

Techniques: Quantitative RT-PCR, Expressing, Multiple Displacement Amplification, Western Blot

Journal: PLoS ONE

Article Title: Bifidobacterium longum Alleviates Dextran Sulfate Sodium-Induced Colitis by Suppressing IL-17A Response: Involvement of Intestinal Epithelial Costimulatory Molecules

doi: 10.1371/journal.pone.0079735

Figure Lengend Snippet: B. longum JCM 1222 T (B.l) suppresses the expression of costimulatory molecules in Colon-26 cells. (a) mRNA expression of CD80 and CD40 was analyzed by quantitative real-time PCR in Colon-26 cells after stimulation with IFN-γ. Levels of mRNA were normalized to β-actin mRNA, and expressed relative to before stimulation (0 h). Results are expressed as means ± standard error (n = 4). * p

Article Snippet: The cells (5 × 104 cells/well) were cultured in 24-well plates containing DMEM medium without antibiotics overnight and pretreated with JCM 1222T (1 × 105 cfu/well) for 2 h. Then, the bacterial growth was stopped by adding penicillin and streptomycin, and the cells were stimulated with 20 ng/ml IFN-γ (Miltenyi Biotec).

Techniques: Expressing, Real-time Polymerase Chain Reaction

Journal: Nucleic Acids Research

Article Title: Expression of RAB4B, a protein governing endocytic recycling, is co-regulated with MHC class II genes

doi: 10.1093/nar/gkl980

Figure Lengend Snippet: Expression of RAB4B is controlled by CIITA and is induced by IFN-γ. mRNA levels for the RAB4B , HLA-DRA and RAB4A genes were determined for ( A ) wt B cells (Raji), CIITA-/- B cells (RJ2.2.5) and CIITA-/- cells complemented with expression vectors encoding the three isoforms of CIITA (CIITA I, III and IV), ( B ) Me67.8 melanoma cells (Me) induced with IFN-γ for 0, 12, 24 and 48 h and ( C ) HUVEC cells induced with IFN-γ for 0, 12, 24 and 48 h. Results were normalized using TBP mRNA. Values for RAB4B and RAB4A are expressed as % of the levels found in wt B cells ( A ) or relative to uninduced cells ( B and C ). Values for HLA–DRA mRNA are expressed as % of the levels found in wt B cells ( A ) or cells induced with IFN-γ for 48 h ( B and C ).

Article Snippet: HUVEC were prepared as described ( ) and induced with 1000 U/ml of IFN-γ (Invitrogen).

Techniques: Expressing

Journal: Nucleic Acids Research

Article Title: Expression of RAB4B, a protein governing endocytic recycling, is co-regulated with MHC class II genes

doi: 10.1093/nar/gkl980

Figure Lengend Snippet: Occupation of the RAB4B promoter by the MHC-II regulatory machinery in DC and IFN-γ induced cells. ( A ) Binding of CIITA to the RAB4B , HLA-DRA and TBP promoters was analyzed by ChIP in DC before (iDC) and after (mDC) maturation with LPS. Results are represented as % of the occupation observed at the HLA-DRA promoter in iDC. The TBP promoter was used as a negative control. ( B and C ) ChIP was used to measure occupancy of the RAB4B promoter by CIITA ( B ) and RFX5 ( C ) in Me67.8 melanoma cells induced with IFN-γ for 0, 2, 4, 6 and 12 h. Results are represented as % of the occupation observed at the HLA–DRA promoter after 12 h of induction.

Article Snippet: HUVEC were prepared as described ( ) and induced with 1000 U/ml of IFN-γ (Invitrogen).

Techniques: Binding Assay, Chromatin Immunoprecipitation, Negative Control

Journal: Experimental and Therapeutic Medicine

Article Title: CD80 and CD86 knockdown in dendritic cells regulates Th1/Th2 cytokine production in asthmatic mice

doi: 10.3892/etm.2016.2989

Figure Lengend Snippet: IFN-γ and IL-4 levels in the supernatant of an mDC and T cell co-culture system as detected by enzyme-linked immunosorbent assay. The IFN-γ level in the siRNA group (transfected with CD80 and CD80 siRNA) was significantly increased compared

Article Snippet: IFN-γ and IL-4 levels were analyzed using ELISA kits specific for IFN-γ and IL-4 (Dakewe Biotech Co., Ltd.) according to the manufacturer's instructions.

Techniques: Co-Culture Assay, Enzyme-linked Immunosorbent Assay, Transfection

Journal: Oncotarget

Article Title: Tumor-derived granzyme B-expressing neutrophils acquire antitumor potential after lipid A treatment

doi: 10.18632/oncotarget.25342

Figure Lengend Snippet: Tumor cytokines and LipA together induced strong expression of granzyme B in spleen and tumor neutrophils ( A ) Spleen neutrophils from rats were purified and incubated or not with LipA (24 h) before staining for granzyme B. The GZMB expression was determined by microscopy. ( B ) Spleen neutrophils from untreated or LipA-treated mice (24 h) were purified before analysis of GZMB by flow cytometry analysis. (A, B) 3 independent experiments. ( C , D ) Spleens and tumors from control rats were removed at day 17. (C) Expression levels of il-2 , il-12 , il-21 and ifn- γ mRNA were evaluated by RT-PCR and expressed as the mean of percentage of change compared to the level of gapdh expression. (D) Neutrophils from spleen were purified as in (A) and incubated for 24 h in vitro without or with a cytokine mix (ILs, IL-2 + IL-12 + IL-21) alone, IFN-γ alone or a combination of both (ILs + IFN-γ), with or without of 10 µg/ml LipA for 6 h. Concentration of granzyme B (GZMB) in conditioned media was measured by ELISA. Significant difference in Mann–Whitney U test, * p

Article Snippet: The following cytokines and inhibitors were added to neutrophil cultures: recombinant rat IL-2 (Biolegend), recombinant mouse IL-21 (eBioscience), IL-12 and IFN-γ (R & D Systems) and p38 MAP Kinase Inhibitor (SB203580, Invivogen).

Techniques: Expressing, Purification, Incubation, Staining, Microscopy, Mouse Assay, Flow Cytometry, Cytometry, Reverse Transcription Polymerase Chain Reaction, In Vitro, Concentration Assay, Enzyme-linked Immunosorbent Assay, MANN-WHITNEY

Journal: Journal of Virology

Article Title: Transient Mobilization of Human Immunodeficiency Virus (HIV)-Specific CD4 T-Helper Cells Fails To Control Virus Rebounds during Intermittent Antiretroviral Therapy in Chronic HIV Type 1 Infection

doi: 10.1128/JVI.75.1.234-241.2001

Figure Lengend Snippet: Patient 1. (A) Effect of intermittent interruptions of antiretroviral therapy on viral load and CD4 or CD8 counts. The HIV-1 RNA level was measured by RT-PCR with a limit of sensitivity of 20 copies per ml. Viral loads are expressed as numbers of HIV-1 RNA copies per milliliter of plasma; CD4 and CD8 counts were analyzed by flow cytometry with fluorescent beads as an internal standard. The values (mean ± standard deviation) in healthy people are as follows: CD4 + cells, 858 ± 260 cells/μl of plasma; CD8 + cells, 482 ± 164/μl plasma. White areas at top of the figure indicate periods of treatment (nevirapine, stavudine, and lamivudine); interruptions are in grey and are also shown in panels A to D. (B) Effect of intermittent interruptions of antiretroviral therapy on Ki67 antigen expression. Ki67 antigen expression was analyzed in CD4 + and CD8 + T cells by flow cytometry after intracellular staining. The values in healthy people are 2.5% ± 0.6% for CD4 + Ki67 + cells and 2% ± 0.6% for CD8 + Ki67 + cells. (C) Effect of intermittent interruptions of antiretroviral therapy on T-helper cell responses to HIV-1 p24 protein. HIV-1 p24-stimulated IFN-γ production by CD8-depleted PBMC was measured by enzyme-linked immunosorbent assay in the 2-day culture supernatants and is expressed as 10 −1 picograms per milliliter; T cell proliferative responses against HIV-1 (p24) were measured on CD8-depleted PBMC, and the results are expressed as a stimulation index. (D) Effect of intermittent interruptions of antiretroviral therapy on CD8 + cell responses to HIV-1 protein. The frequency of HIV-specific CD8 + T cells was tested by a recombinant vaccinia virus ELISPOT assay. PBMC were infected with wild-type vaccinia virus or recombinant vaccinia virus Gag, Pol, Env, or Nef, and IFN-γ-producing SFC were enumerated in an 18-h ELISPOT assay. Results are expressed as the number of IFN-γ SFC per 10 6 PBMC.

Article Snippet: Cell culture supernatants were harvested on day 2 from CD8-depleted PBMC cultured at 106 /ml with p24 (0.25 μg/ml) (Intracell) or medium alone and measured by an enzyme immunoassay for IFN-γ (Diaclone, Besançon, France).

Techniques: Reverse Transcription Polymerase Chain Reaction, Flow Cytometry, Cytometry, Standard Deviation, Expressing, Staining, Enzyme-linked Immunosorbent Assay, Recombinant, Enzyme-linked Immunospot, Infection

Journal: PLoS ONE

Article Title: Excretory/Secretory Products from Trichinella spiralis Adult Worms Ameliorate DSS-Induced Colitis in Mice

doi: 10.1371/journal.pone.0096454

Figure Lengend Snippet: Treatment with the T. spiralis AES reduced the levels of the DSS-induced pro-inflammatory cytokines IFN-γ, IL-6 and IL-17 in the spleens, MLN and colon lymphocytes. The data are presented as the mean ± SE. The asterisks* indicate statistical significance at P

Article Snippet: The cells were subjected to a cytokine assay using IFN-γ, IL-4, IL-6, IL-10 (BD Biosciences, USA) and IL-17 (R & D Systems, USA) ELISPOT sets according to the manufacturer's instructions.

Techniques:

Journal: Frontiers in Immunology

Article Title: Hantavirus-Driven PD-L1/PD-L2 Upregulation: An Imperfect Viral Immune Evasion Mechanism

doi: 10.3389/fimmu.2018.02560

Figure Lengend Snippet: Control of PD-L1 expression by inflammatory stimuli. (A) Immature DCs were exposed to the following inflammatory stimuli before staining for PD-L1: type I IFN (IFN-α at 1,000 U/ml), type II IFN (IFN-γ at 1,000 U/ml), poly(dA:dT), UV-inactivated VSV or poly(I:C) for 24 h. The results shown are representative of three independent experiments using three different donors. (B) Huh7.5 cells (control), Huh7.5 cells permanently expressing a constitutively active form of RIG-I (RIG-CA) or Huh7.5 cells stimulated with IFN-γ at 1,000 U/ml for 24 h were stained for PD-L1 and analyzed by flow cytometry. Results are derived from three independent experiments, error bars represent the mean ± SEM ( * p

Article Snippet: Cytokines And Pathogen-Associated Molecular Patterns (PAMPs) IFN-α, IFN-β, and IFN-γ were provided by ImmunoTools.

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

Journal: Cancer immunology, immunotherapy : CII

Article Title: Deficiency of activated STAT1 in head and neck cancer cells mediates TAP1-dependent escape from cytotoxic T lymphocytes

doi: 10.1007/s00262-010-0961-7

Figure Lengend Snippet: IFN-γ-mediated-STAT1 binding to the TAP1 promoter and APM protein expression is independent of STAT1:STAT3 heterodimerization. a PCI-13 and b SCC90 cells were untreated, treated with IL-6 (50 ng/ml, 60 min), IFN-γ (1,000 U/ml, 30 min)

Article Snippet: Recombinant human IL-6 was purchased from R & D systems (Minneapolis, MN), and IFN-γ was purchased from InterMune (Brisbane, CA).

Techniques: Binding Assay, Expressing

Journal: Cancer immunology, immunotherapy : CII

Article Title: Deficiency of activated STAT1 in head and neck cancer cells mediates TAP1-dependent escape from cytotoxic T lymphocytes

doi: 10.1007/s00262-010-0961-7

Figure Lengend Snippet: SCCHN cells express low basal pSTAT1 and high basal pSTAT3 that are inducible by treatment with IFN-γ and IL-6. Cells were treated with IFN-γ (100 U/ml, 15 min) or IL-6 (50 ng/ml, 15 min) and assayed for pSTAT1 (Tyrosine 701) or pSTAT3

Article Snippet: Recombinant human IL-6 was purchased from R & D systems (Minneapolis, MN), and IFN-γ was purchased from InterMune (Brisbane, CA).

Techniques:

Journal: Cancer immunology, immunotherapy : CII

Article Title: Deficiency of activated STAT1 in head and neck cancer cells mediates TAP1-dependent escape from cytotoxic T lymphocytes

doi: 10.1007/s00262-010-0961-7

Figure Lengend Snippet: IFN-γ-mediated pSTAT1, APM expression and CTL recognition in STAT3-depleted SCCHN cells. a PCI-13 and b SCC90 cells were transfected with the indicated doses of siRNA, and knockdown of STAT3 protein was assessed by immunoblot and densitometry

Article Snippet: Recombinant human IL-6 was purchased from R & D systems (Minneapolis, MN), and IFN-γ was purchased from InterMune (Brisbane, CA).

Techniques: Expressing, CTL Assay, Transfection

Journal: Cancer immunology, immunotherapy : CII

Article Title: Deficiency of activated STAT1 in head and neck cancer cells mediates TAP1-dependent escape from cytotoxic T lymphocytes

doi: 10.1007/s00262-010-0961-7

Figure Lengend Snippet: STAT1 signaling is intact in SCCHN cells. a SCCHN cells isolated from SCCHN tumors were tested for basal IFN-γR expression by flow cytometry. PCI-13 cells were treated with IFN-γ (100 U/ml) for either 15 min or 48 h and then analyzed by

Article Snippet: Recombinant human IL-6 was purchased from R & D systems (Minneapolis, MN), and IFN-γ was purchased from InterMune (Brisbane, CA).

Techniques: Isolation, Expressing, Flow Cytometry, Cytometry

Journal: Cancer immunology, immunotherapy : CII

Article Title: Deficiency of activated STAT1 in head and neck cancer cells mediates TAP1-dependent escape from cytotoxic T lymphocytes

doi: 10.1007/s00262-010-0961-7

Figure Lengend Snippet: IL-6 and IFN-γ increase pSTAT1:pSTAT3 heterodimerization. Cells were treated with either IL-6 (50 ng/ml, 15 min) or IFN-γ (100 U/ml, 15 min). Whole cell lysates were prepared and immunoprecipitated with anti-STAT1 or anti-STAT3 or an irrelevant

Article Snippet: Recombinant human IL-6 was purchased from R & D systems (Minneapolis, MN), and IFN-γ was purchased from InterMune (Brisbane, CA).

Techniques: Immunoprecipitation