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  • 94
    TaKaRa primerarray cytokine cytokine receptor interaction
    Exercise with AICAR treatment resulted in a pro-inflammatory and pro-apoptotic FAP phenotype that promotes muscle regeneration. a – c Representative images of PDGFRα- and p16 INK4A -immunostained triceps surae (green and red, respectively) ( a ), and quantitative data of the number of FAPs ( b ) and the percentage of p16 INK4A + FAPs ( c ) in randomly chosen fields of view ( n = 3 for control, n = 4 for exercise and CIM). d Representative confocal images of Bcl-2-, p53-, IL-33-, and p16 INK4A -immunostained FAPs isolated from CIM with or without AICAR treatment in vitro. e Hierarchical clustering of differentially expressed <t>cytokine–cytokine</t> receptor gene expression was profiled by <t>PrimerArray</t> ® analysis in control, AMI, and CIM mice (the average value is that shown in Fig. 2d ; n = 3 per group), and control exercise, CIM exercise, CIM AICAR, and CIM exercise with AICAR ( n = 3 per group). Genes with higher expression are depicted in magenta, genes with lower expression are depicted in cyan, and genes with no difference are depicted in black. f Principal component analysis (PCA) of FAPs isolated from control, control exercise, CIM, CIM exercise, CIM AICAR, and CIM exercise with AICAR. g Proposed mechanism whereby exercise-induced FAP senescence promotes muscle regeneration. Data are shown as a dot plot for each specimen, and as means as well as medians with IQRs and 1.5 times the IQR by dot plots and box and whisker plots. P values were determined by the one-way ANOVA adjusted by the Holm method (* P
    Primerarray Cytokine Cytokine Receptor Interaction, supplied by TaKaRa, used in various techniques. Bioz Stars score: 94/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Becton Dickinson cytokine
    Sources of IL-2 (A) and IFN-γ (B) production in immunized mice. Spleens from immunized mice were harvested 3 weeks after the third immunization. Single-cell suspensions of CD4 + T cells, CD8 + T cells or both T-cell subpopulations were prepared and subjected to in vitro stimulation for 72 h with 10 PFU of UV-inactivated HSV-1 strain McKrae per cell. Cytokines in the supernatants were determined by ELISAs. Each value is the percentage of each <t>cytokine</t> that was produced by CD4 + T cells or CD8 + T cells in relation to the amount of each cytokine produced by both T cells. Values that were significantly different from each other as well as other vaccine groups ( P
    Cytokine, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 93/100, based on 2776 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Millipore cytokine
    Trif deficiency has no impact on peritoneal and serum <t>cytokine</t> levels during polymicrobial sepsis WT and Trif −/− mice underwent sham or CLP procedures. Twenty-four hours later, peritoneal lavage and blood were collected. Peritoneal lavage and serum IL-6, IL-10, TNFα, KC, MCP-1, and MIP-2 were measured with a multiplex fluorescent bead-based immunoassay. A . Peritoneal cytokines. B. Serum cytokines. Each error bar represents mean ± SE. The number of animals in each group: WT-Sham = 3, WT-CLP = 4, Trif −/− -Sham = 3, Trif −/− -CLP = 6. CLP = cecum ligation and puncture; IL = interleukin; KC = keratinocyte chemoattractant; MCP-1 = monocyte chemoattractant protein-1; MIP-2 = macrophage inflammatory protein-2; MyD88 = myeloid differentiation factor 88; TNF = tumor necrosis factor; Trif = TIR-domain-containing adaptor protein inducing interferon-β mediated transcription-factor; WT = wild type.
    Cytokine, supplied by Millipore, used in various techniques. Bioz Stars score: 94/100, based on 1067 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Pharmingen cytokine
    Trif deficiency has no impact on peritoneal and serum <t>cytokine</t> levels during polymicrobial sepsis WT and Trif −/− mice underwent sham or CLP procedures. Twenty-four hours later, peritoneal lavage and blood were collected. Peritoneal lavage and serum IL-6, IL-10, TNFα, KC, MCP-1, and MIP-2 were measured with a multiplex fluorescent bead-based immunoassay. A . Peritoneal cytokines. B. Serum cytokines. Each error bar represents mean ± SE. The number of animals in each group: WT-Sham = 3, WT-CLP = 4, Trif −/− -Sham = 3, Trif −/− -CLP = 6. CLP = cecum ligation and puncture; IL = interleukin; KC = keratinocyte chemoattractant; MCP-1 = monocyte chemoattractant protein-1; MIP-2 = macrophage inflammatory protein-2; MyD88 = myeloid differentiation factor 88; TNF = tumor necrosis factor; Trif = TIR-domain-containing adaptor protein inducing interferon-β mediated transcription-factor; WT = wild type.
    Cytokine, supplied by Pharmingen, used in various techniques. Bioz Stars score: 92/100, based on 271 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    95
    R&D Systems cytokine
    A-SAA production by NHEK stimulated with human IL-1α, IL-17A, IL-22, OSM, TNF-α, alone or in combination. (A) A-SAA mRNA expression and (B) protein secretion by NHEK stimulated with M5 were analyzed at different time-periods. (C) A-SAA mRNA expression and (D) protein secretion were determined 40 hours after <t>cytokine</t> activation. (E) A-SAA mRNA expression and (F) protein secretion by NHEK 40 hours after stimulation with four cytokines by sequentially subtracting either recombinant IL-1α, IL-17A, IL-22, OSM or TNF-α from M5. A-SAA mRNA and protein were quantified by RT-qPCR in NHEK and ELISA in supernatants, respectively. Data represent the mean ± SEM of three experiments with duplicates. Statistical comparisons were performed using 2way ANOVA test or t test (*p
    Cytokine, supplied by R&D Systems, used in various techniques. Bioz Stars score: 95/100, based on 1651 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    STEMCELL Technologies Inc cytokine
    Stat5 is not required for transformation mediated by KrasG12D. BM from pI:pC-treated control and MxCre; Stat5fl/fl mice were infected with retroviruses expressing Jak2V617F, BCR-ABL, or KrasG12D. Transduced BM cells (1 × 10 5 ) were then plated in methylcellulose medium without any <t>cytokine</t> (Methocult M3234). Cytokine-independent CFU-GM colonies (A) were counted after 7 days and CFU-E colonies (B) were counted after 2 days (n = 4). Asterisks show significant differences by t test (* P
    Cytokine, supplied by STEMCELL Technologies Inc, used in various techniques. Bioz Stars score: 92/100, based on 31 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    BioLegend cytokines
    Diminished neuroinflammation in Bhlhe40 −/− mice a , Flow cytometry of CNS myeloid cells from WT and Bhlhe40 −/− mice (n=3 per group) on day 16 after immunization. b, Percentage of MHC class II + microglia and infiltrating myeloid cells. c-e , Flow cytometry of CD4 + T cells in the CNS of WT and Bhlhe40 −/− mice (n=3 per group) on day 16 after immunization. ( c ) CD4 + T cell number. ( d ) Representative ICS for the indicated <t>cytokines.</t> ( e ) Frequency of CD4 + T cells secreting each of the indicated 8 possible combinations of GM-CSF, IL-17A, and IFN-γ. f, g , Flow cytometry of γδ T cells in the CNS of WT and Bhlhe40 −/− mice (n=3per group) on day 14 after immunization. ( f ) Representative ICS for IFN-γ, IL-17A, and GM-CSF. ( g ) Frequency of γδ T cells secreting IFN-γ, IL-17A, or GM-CSF.
    Cytokines, supplied by BioLegend, used in various techniques. Bioz Stars score: 99/100, based on 2026 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Lonza cytokines
    Diminished neuroinflammation in Bhlhe40 −/− mice a , Flow cytometry of CNS myeloid cells from WT and Bhlhe40 −/− mice (n=3 per group) on day 16 after immunization. b, Percentage of MHC class II + microglia and infiltrating myeloid cells. c-e , Flow cytometry of CD4 + T cells in the CNS of WT and Bhlhe40 −/− mice (n=3 per group) on day 16 after immunization. ( c ) CD4 + T cell number. ( d ) Representative ICS for the indicated <t>cytokines.</t> ( e ) Frequency of CD4 + T cells secreting each of the indicated 8 possible combinations of GM-CSF, IL-17A, and IFN-γ. f, g , Flow cytometry of γδ T cells in the CNS of WT and Bhlhe40 −/− mice (n=3per group) on day 14 after immunization. ( f ) Representative ICS for IFN-γ, IL-17A, and GM-CSF. ( g ) Frequency of γδ T cells secreting IFN-γ, IL-17A, or GM-CSF.
    Cytokines, supplied by Lonza, used in various techniques. Bioz Stars score: 94/100, based on 185 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Luminex cytokines
    <t>Cytokines</t> levels in pericardial fluid and plasma samples. Cytokines levels were determined before CDCs administration and 30 days post-administration using the Luminex xMAP technology. * Statistically significant differences ( p
    Cytokines, supplied by Luminex, used in various techniques. Bioz Stars score: 94/100, based on 6620 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Meso Scale Diagnostics LLC cytokines
    Inflammatory cytokine profile in response to respiratory introduction of predatory bacteria. qPCR analysis of IL-1β, IL-4, IL-6, IL-10, IL-12, IL-23, IFNγ, and TNF in response to intranasal inoculation of predatory bacteria relative to PBS control. Mice were inoculated intranasally with B. bacteriovorus 109J, HD100 or M. aeruginosavorus ARL-13. Expression of <t>cytokines</t> was assessed in the lung at ( A ) 1, ( B ) 24, and ( C ) 48 hours post-inoculation. Expression of cytokines was assessed in the spleen at ( D ) 24 and ( E ) 48 hours post-inoculation. Fold induction was calculated using the ΔΔC t by approximation method using an endogenous calibrator (β-actin). For the one hour experiment, 6 mice per predatory bacterial strain (and PBS) were used. Twelve mice per strain (and PBS) were used at each of the 24 and 48 hour time points, with the exception of the Lung/24 hour experiment, where 6 mice were used. Data for the one hour time point is from one experiment; data for the 24 and 48 hour time points are from two independent experiments. Data represent mean ± standard deviation. Student’s t -test was performed to compare each treated sample to their respective control sample, * P
    Cytokines, supplied by Meso Scale Diagnostics LLC, used in various techniques. Bioz Stars score: 92/100, based on 1157 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Miltenyi Biotec cytokines
    CNS Treg Cells are Stable and Express Blimp1 in Response to Proinflammatory <t>Cytokines</t> (A) Mononuclear cells were isolated from the CNS of EAE mice at the peak of disease and were stimulated with phorbol 12-myristate 13-acetate (PMA) and ionomycin to analyze the expression of IL-10, IL-17, and IFN-γ in Treg cells by flow cytometry. Mean of eight biological replicates ± SD, derived from three independent experiments. Symbols depict individual mice (bars, mean ± SD). (B) CD4 + Foxp3 + Treg cells were sorted from the CNS and spleen (SPL) of Foxp3 (GFP) reporter mice at the peak of EAE and subjected to RNA-seq. Principal-component analysis. (C) Foxp3 expression by intracellular staining of splenic Treg cells and CNS Treg cells at the peak of EAE. Mean of six biological replicates ± SD, derived from two independent experiments. Symbols depict individual mice (bars, mean ± SD), t test, p
    Cytokines, supplied by Miltenyi Biotec, used in various techniques. Bioz Stars score: 94/100, based on 467 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Prospec cytokines
    CNS Treg Cells are Stable and Express Blimp1 in Response to Proinflammatory <t>Cytokines</t> (A) Mononuclear cells were isolated from the CNS of EAE mice at the peak of disease and were stimulated with phorbol 12-myristate 13-acetate (PMA) and ionomycin to analyze the expression of IL-10, IL-17, and IFN-γ in Treg cells by flow cytometry. Mean of eight biological replicates ± SD, derived from three independent experiments. Symbols depict individual mice (bars, mean ± SD). (B) CD4 + Foxp3 + Treg cells were sorted from the CNS and spleen (SPL) of Foxp3 (GFP) reporter mice at the peak of EAE and subjected to RNA-seq. Principal-component analysis. (C) Foxp3 expression by intracellular staining of splenic Treg cells and CNS Treg cells at the peak of EAE. Mean of six biological replicates ± SD, derived from two independent experiments. Symbols depict individual mice (bars, mean ± SD), t test, p
    Cytokines, supplied by Prospec, used in various techniques. Bioz Stars score: 92/100, based on 108 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Roche cytokines
    The release of proinflammatory <t>cytokines</t> and prostaglandin E 2 are impaired in PKCε −/ − mice. (A–C) Thioglycollate-elicited peritoneal macrophages were activated for the indicated times with 200 ng/ml of LPS and 20 U/ml of IFNγ. The amount of TNF-α, IL-1β, and PGE 2 in the culture medium was then determined. (D–E) Bar graphs demonstrating the serum levels of TNF-α (1 h after injection) and IL-1β (4 h after injection) after the intraperitoneal injection of either 5 mg/kg or 40 mg/kg of LPS. (F) Apoptosis of thioglycollate-elicited peritoneal macrophages 24 h after activation by LPS (200 ng/ml), LPS plus IFNγ (200 ng/ml and 20 U/ml, respectively), or LPS, IFNγ, and 15dPGJ 2 (2 μM). Results show the mean ± SEM of three experiments. The * and ** denote P
    Cytokines, supplied by Roche, used in various techniques. Bioz Stars score: 92/100, based on 346 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    SABiosciences cytokines
    4% Lovaza ® decreases LPS induced <t>cytokines</t> in cultured splenocytes from (NZB×NZW)F1 female mice. Two-month-old mice were fed semi-purified diets containing 1% placebo, 4% placebo, 1% Lovaza ® , 4% Lovaza ® , 4% FO-18/12 and standard LC diet as a control. At eight months of age, splenocytes were isolated, cultured aseptically and stimulated with LPS for 24 h. 4% Lovaza ® fed mice showed significantly decreased (A)IL-1β, (B)IL-6, (C) TNF-α and (D) TGF-β in conditioned medium of splenocyte culture by ELISA (5 mice/group). Results are expressed as mean±SEM. * P
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    Shenandoah Biotechnology cytokines
    Th17 cells are resistant to apoptosis in the absence of IL2Rγ-chain <t>cytokines.</t> a Gating strategy for apoptosis assays b,c Apoptosis of Th17 cells (TRP-1), unpolarized CD4 + T cells (TRP-1), Tc17 cells (pmel-1), or unpolarized CD8 + T cells (pmel-1) was measured after culture with IL-2, IL-7, IL-15, or vehicle. Panel b shows representative samples from day 2. Panel c shows mean plus standard error of 3 independent experiments for the days indicated. d,e Antibody-mediated depletion of IL2Rγ-chain cytokines does not induce apoptosis of Th17 (TRP-1) cells over 2 day culture. Triangles represent independent experiments. p > .05 for all depletion conditions vs control by 2-tailed t-test. f Expression of Bcl-2 protein is greater in Th17 cells (dashed line) than in unpolarized CD8 + cells (solid line). Grey histogram is FMO control. Representative of two independent experiments
    Cytokines, supplied by Shenandoah Biotechnology, used in various techniques. Bioz Stars score: 90/100, based on 15 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    TaKaRa cytokines
    Expression of cytokine mRNA, and IL-10 protein levels in M-CSF- or GM-CSF-cultured BMCs. (A) The expression of proinflammatory <t>cytokines</t> (TNF-alpha, IL-6, IL-1beta, IL-12a, and IL12b) and anti-inflammatory cytokine (IL-10) mRNA in M-CSF- or GM-CSF-cultured BMCs. (B) IL-10 concentration in the supernatant of M-CSF- or GM-CSF-cultured BMCs. Data are expressed as mean ± SEM, *** P
    Cytokines, supplied by TaKaRa, used in various techniques. Bioz Stars score: 92/100, based on 296 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Thermo Fisher cytokines
    Serum cytokine induction. Levels of representative <t>cytokines</t> (CCL2, CSF3, CXCL1, TNFα, IL-5, IL-6, IFNγ, and CXCL10) were measured for each treatment group in the serum at 6–48 hours after adjuvant administration in 3–4 replicate experiments. Data is presented as the mean plus standard deviation of N = 8–12 for the 6 and 24 hour timepoints and N = 3 for the 48 hour timepoint.
    Cytokines, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 8988 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Bio-Rad cytokines
    Several <t>cytokines</t> are increased significantly in the sera of infected STAT1 KO mice. In WT mice, a multiplex assay showed small to moderate but significant changes for CCL4, CCL2, and IL-2 levels at day 3 postinfection (A) and for G-CSF, CCL2, IL-5, IL-12,
    Cytokines, supplied by Bio-Rad, used in various techniques. Bioz Stars score: 99/100, based on 4430 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cell Signaling Technology Inc cytokines
    Receptors for the pro-inflammatory <t>cytokines</t> TNFα and IL-1β are present on cells of the smooth muscle/myenteric plexus layer in vivo and in vitro . A , B , Images of fluorescence immunocytochemistry showing the presence of staining for TNF-R1 ( A ) and IL-1 R1 ( B ) on sections of control intestine. Staining was evident on smooth muscle cells of the CSM and longitudinal muscle (LSM) layers, as well as on neurons of the myenteric plexus (arrows). Scale bar, 100 μm. C , D , Depiction of the coculture model of myenteric neurons and intestinal smooth muscle cells. C , Nomarski image of culture appearance at 7 d in vitro , showing complex architecture. D , Fluorescence image of same field showing neurons and neurites among smooth muscle cells, as revealed by immunostaining for SNAP-25 (red) and the neuronal cell body marker HuD (green). Arrows indicate the location of neuronal cell bodies (yellow, merged color). Nuclei were stained with Hoechst (blue). Scale bar, 250 μm. E , F , Typical outcome of dual label immunocytochemistry showing expression of cytokine receptors on myenteric neurons in vitro . E , Images of positive staining for TNF-RI and the neuronal marker HuD (arrows) and their overlap (merged). Asterisks (*) indicate neurites. F , Similar outcome as E showing overlap of staining for IL1-RI with HuD (e.g., arrows). Scale bars, 100 μm.
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    Cellgenix cytokines
    Phenotype and migration of DC matured in IRX-2 or conventional <t>cytokines.</t> ( A ) DC obtained from HNSCC patients were matured for 48 h either with IRX-2 or the conventional maturation cocktail. While conventionally matured DC (white bars) expressed higher levels of CD80, CD83 and CD86 (*, p
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    Image Search Results


    Exercise with AICAR treatment resulted in a pro-inflammatory and pro-apoptotic FAP phenotype that promotes muscle regeneration. a – c Representative images of PDGFRα- and p16 INK4A -immunostained triceps surae (green and red, respectively) ( a ), and quantitative data of the number of FAPs ( b ) and the percentage of p16 INK4A + FAPs ( c ) in randomly chosen fields of view ( n = 3 for control, n = 4 for exercise and CIM). d Representative confocal images of Bcl-2-, p53-, IL-33-, and p16 INK4A -immunostained FAPs isolated from CIM with or without AICAR treatment in vitro. e Hierarchical clustering of differentially expressed cytokine–cytokine receptor gene expression was profiled by PrimerArray ® analysis in control, AMI, and CIM mice (the average value is that shown in Fig. 2d ; n = 3 per group), and control exercise, CIM exercise, CIM AICAR, and CIM exercise with AICAR ( n = 3 per group). Genes with higher expression are depicted in magenta, genes with lower expression are depicted in cyan, and genes with no difference are depicted in black. f Principal component analysis (PCA) of FAPs isolated from control, control exercise, CIM, CIM exercise, CIM AICAR, and CIM exercise with AICAR. g Proposed mechanism whereby exercise-induced FAP senescence promotes muscle regeneration. Data are shown as a dot plot for each specimen, and as means as well as medians with IQRs and 1.5 times the IQR by dot plots and box and whisker plots. P values were determined by the one-way ANOVA adjusted by the Holm method (* P

    Journal: Nature Communications

    Article Title: Exercise enhances skeletal muscle regeneration by promoting senescence in fibro-adipogenic progenitors

    doi: 10.1038/s41467-020-14734-x

    Figure Lengend Snippet: Exercise with AICAR treatment resulted in a pro-inflammatory and pro-apoptotic FAP phenotype that promotes muscle regeneration. a – c Representative images of PDGFRα- and p16 INK4A -immunostained triceps surae (green and red, respectively) ( a ), and quantitative data of the number of FAPs ( b ) and the percentage of p16 INK4A + FAPs ( c ) in randomly chosen fields of view ( n = 3 for control, n = 4 for exercise and CIM). d Representative confocal images of Bcl-2-, p53-, IL-33-, and p16 INK4A -immunostained FAPs isolated from CIM with or without AICAR treatment in vitro. e Hierarchical clustering of differentially expressed cytokine–cytokine receptor gene expression was profiled by PrimerArray ® analysis in control, AMI, and CIM mice (the average value is that shown in Fig. 2d ; n = 3 per group), and control exercise, CIM exercise, CIM AICAR, and CIM exercise with AICAR ( n = 3 per group). Genes with higher expression are depicted in magenta, genes with lower expression are depicted in cyan, and genes with no difference are depicted in black. f Principal component analysis (PCA) of FAPs isolated from control, control exercise, CIM, CIM exercise, CIM AICAR, and CIM exercise with AICAR. g Proposed mechanism whereby exercise-induced FAP senescence promotes muscle regeneration. Data are shown as a dot plot for each specimen, and as means as well as medians with IQRs and 1.5 times the IQR by dot plots and box and whisker plots. P values were determined by the one-way ANOVA adjusted by the Holm method (* P

    Article Snippet: The PrimerArray® Cytokine–Cytokine Receptor Interaction Kit (Mouse) (PN001; Takara Bio Inc.) was used with TB Green Premix Ex Taq II (Tli RNaseH Plus; Takara Bio Inc.) with the Applied Biosystems 7500 Real-Time PCR System (Applied Biosystems) under the following cycling conditions: 95 °C for 30 s, followed by 40 cycles of amplification (95 °C for 5 s and 64 °C for 34 s).

    Techniques: Isolation, In Vitro, Expressing, Mouse Assay, Whisker Assay

    FAPs derived from chronic inflammatory myopathy model mice acquired features of apoptosis resistance. a Cytotoxicity assay shows the LDH release level after stimulation with different concentrations of TNF-α in FAPs isolated from AMI and CIM ( n = 3 per group). b , c Representative image of FITC-Annexin V (green) and ethidium homodimer III (red) after stimulation with 1, 10, and 100 ng/mL of TNF-α, and the quantified data. The number of Annexin V+, apoptotic FAPs increased in a dose-dependent fashion with the addition of TNF-α in AMI, but not in CIM mice ( n = 3 per group). d , e Hierarchical clustering of differentially expressed cytokine–cytokine receptor gene expression was profiled by PrimerArray ® analysis in control, AMI, and CIM mice ( n = 3 per group) ( d ). Genes with higher expression are depicted in magenta, genes with lower expression are depicted in cyan, and genes with no difference are depicted in black ( d ). Scatterplots of gene expression changes in AMI-FAPs ( n = 3) compared with CIM-FAPs ( n = 3) ( e ). f , g Representative images of PDGFRα- and active caspase-3-immunostained triceps surae in control, AMI, and CIM mice ( f ), and quantification of the percentage of active caspase-3-positive FAPs in randomly chosen fields of view ( g ). h Representative confocal images of Bcl-2- and p53-immunostained FAPs isolated from AMI and CIM. i mRNA expression of Cd274 , Pdcd1lg2 , and Cd47 in FAPs isolated from AMI and CIM mice ( n = 3 per group). j Correlation of Cd274 , Pdcd1lg2 , and Cd47 mRNA expression with Cdkn2a mRNA expression in FAPs isolated from AMI and CIM mice. Quantitative data for each specimen are shown in a dot plot. P values were determined by one-way ANOVA adjusted by the Holm method (* P

    Journal: Nature Communications

    Article Title: Exercise enhances skeletal muscle regeneration by promoting senescence in fibro-adipogenic progenitors

    doi: 10.1038/s41467-020-14734-x

    Figure Lengend Snippet: FAPs derived from chronic inflammatory myopathy model mice acquired features of apoptosis resistance. a Cytotoxicity assay shows the LDH release level after stimulation with different concentrations of TNF-α in FAPs isolated from AMI and CIM ( n = 3 per group). b , c Representative image of FITC-Annexin V (green) and ethidium homodimer III (red) after stimulation with 1, 10, and 100 ng/mL of TNF-α, and the quantified data. The number of Annexin V+, apoptotic FAPs increased in a dose-dependent fashion with the addition of TNF-α in AMI, but not in CIM mice ( n = 3 per group). d , e Hierarchical clustering of differentially expressed cytokine–cytokine receptor gene expression was profiled by PrimerArray ® analysis in control, AMI, and CIM mice ( n = 3 per group) ( d ). Genes with higher expression are depicted in magenta, genes with lower expression are depicted in cyan, and genes with no difference are depicted in black ( d ). Scatterplots of gene expression changes in AMI-FAPs ( n = 3) compared with CIM-FAPs ( n = 3) ( e ). f , g Representative images of PDGFRα- and active caspase-3-immunostained triceps surae in control, AMI, and CIM mice ( f ), and quantification of the percentage of active caspase-3-positive FAPs in randomly chosen fields of view ( g ). h Representative confocal images of Bcl-2- and p53-immunostained FAPs isolated from AMI and CIM. i mRNA expression of Cd274 , Pdcd1lg2 , and Cd47 in FAPs isolated from AMI and CIM mice ( n = 3 per group). j Correlation of Cd274 , Pdcd1lg2 , and Cd47 mRNA expression with Cdkn2a mRNA expression in FAPs isolated from AMI and CIM mice. Quantitative data for each specimen are shown in a dot plot. P values were determined by one-way ANOVA adjusted by the Holm method (* P

    Article Snippet: The PrimerArray® Cytokine–Cytokine Receptor Interaction Kit (Mouse) (PN001; Takara Bio Inc.) was used with TB Green Premix Ex Taq II (Tli RNaseH Plus; Takara Bio Inc.) with the Applied Biosystems 7500 Real-Time PCR System (Applied Biosystems) under the following cycling conditions: 95 °C for 30 s, followed by 40 cycles of amplification (95 °C for 5 s and 64 °C for 34 s).

    Techniques: Derivative Assay, Mouse Assay, Cytotoxicity Assay, Isolation, Expressing

    Sources of IL-2 (A) and IFN-γ (B) production in immunized mice. Spleens from immunized mice were harvested 3 weeks after the third immunization. Single-cell suspensions of CD4 + T cells, CD8 + T cells or both T-cell subpopulations were prepared and subjected to in vitro stimulation for 72 h with 10 PFU of UV-inactivated HSV-1 strain McKrae per cell. Cytokines in the supernatants were determined by ELISAs. Each value is the percentage of each cytokine that was produced by CD4 + T cells or CD8 + T cells in relation to the amount of each cytokine produced by both T cells. Values that were significantly different from each other as well as other vaccine groups ( P

    Journal: Journal of Virology

    Article Title: Comparison of Adjuvant Efficacy of Herpes Simplex Virus Type 1 Recombinant Viruses Expressing TH1 and TH2 Cytokine Genes

    doi: 10.1128/JVI.77.10.5774-5783.2003

    Figure Lengend Snippet: Sources of IL-2 (A) and IFN-γ (B) production in immunized mice. Spleens from immunized mice were harvested 3 weeks after the third immunization. Single-cell suspensions of CD4 + T cells, CD8 + T cells or both T-cell subpopulations were prepared and subjected to in vitro stimulation for 72 h with 10 PFU of UV-inactivated HSV-1 strain McKrae per cell. Cytokines in the supernatants were determined by ELISAs. Each value is the percentage of each cytokine that was produced by CD4 + T cells or CD8 + T cells in relation to the amount of each cytokine produced by both T cells. Values that were significantly different from each other as well as other vaccine groups ( P

    Article Snippet: Cell-free culture supernatants were assayed for IL-2, IL-4, and IFN-γ using an ELISA specific for each cytokine (BD PharMingen, San Diego, Calif.).

    Techniques: Mouse Assay, In Vitro, Produced

    Cytokine production by splenocytes of immunized mice. BALB/c mice were immunized once or three times as described in Materials and Methods. Three weeks after the final immunization, mice were euthanized, spleens from three mice per group were harvested, and single-cell suspensions of splenocytes were prepared and stimulated in vitro for 72 h with UV-inactivated HSV-1 strain McKrae, as described in Materials and Methods. The concentrations of IL-2 (A), IL-4 (B), and IFN-γ (C) in the supernatants were measured by ELISA. Each point represents the mean of the titers from three experiments ± standard error (error bar).

    Journal: Journal of Virology

    Article Title: Comparison of Adjuvant Efficacy of Herpes Simplex Virus Type 1 Recombinant Viruses Expressing TH1 and TH2 Cytokine Genes

    doi: 10.1128/JVI.77.10.5774-5783.2003

    Figure Lengend Snippet: Cytokine production by splenocytes of immunized mice. BALB/c mice were immunized once or three times as described in Materials and Methods. Three weeks after the final immunization, mice were euthanized, spleens from three mice per group were harvested, and single-cell suspensions of splenocytes were prepared and stimulated in vitro for 72 h with UV-inactivated HSV-1 strain McKrae, as described in Materials and Methods. The concentrations of IL-2 (A), IL-4 (B), and IFN-γ (C) in the supernatants were measured by ELISA. Each point represents the mean of the titers from three experiments ± standard error (error bar).

    Article Snippet: Cell-free culture supernatants were assayed for IL-2, IL-4, and IFN-γ using an ELISA specific for each cytokine (BD PharMingen, San Diego, Calif.).

    Techniques: Mouse Assay, In Vitro, Enzyme-linked Immunosorbent Assay

    The effect of P2X 7 receptor inhibition in E. coli -induced sepsis . ARD6 was injected iv into anesthetized balb/cj mice (low dose—41 million). Two hours prior to ARD6 injection the animals were administered the P2X 7 antagonist BBG or vehicle (50 mg kg −1 , subcutaneously). (A) Kaplan-Meier plot shows survival, n = 6 for controls and 10–12 for ARD6 with or without BBG. (B) Absorbance of plasma as an indication of plasma haemolysis 2.5 h after ARD6 injection, n = 13 for all four groups. (C) Colony forming units from blood samples 2.5 h after a low number of bacteria (D) Levels of the cytokines TNFα, KC, IL-1β, and IL-6 2.5 h after ARD6 injection, n = 14 for ARD6 and 13 for ARD6 + BBG. * p

    Journal: Frontiers in Cellular and Infection Microbiology

    Article Title: P2X1, P2X4, and P2X7 Receptor Knock Out Mice Expose Differential Outcome of Sepsis Induced by α-Haemolysin Producing Escherichia coli

    doi: 10.3389/fcimb.2017.00113

    Figure Lengend Snippet: The effect of P2X 7 receptor inhibition in E. coli -induced sepsis . ARD6 was injected iv into anesthetized balb/cj mice (low dose—41 million). Two hours prior to ARD6 injection the animals were administered the P2X 7 antagonist BBG or vehicle (50 mg kg −1 , subcutaneously). (A) Kaplan-Meier plot shows survival, n = 6 for controls and 10–12 for ARD6 with or without BBG. (B) Absorbance of plasma as an indication of plasma haemolysis 2.5 h after ARD6 injection, n = 13 for all four groups. (C) Colony forming units from blood samples 2.5 h after a low number of bacteria (D) Levels of the cytokines TNFα, KC, IL-1β, and IL-6 2.5 h after ARD6 injection, n = 14 for ARD6 and 13 for ARD6 + BBG. * p

    Article Snippet: CBA flex sets for measuring cytokines were from BD Biosciences.

    Techniques: Inhibition, Injection, Mouse Assay

    Trif deficiency has no impact on peritoneal and serum cytokine levels during polymicrobial sepsis WT and Trif −/− mice underwent sham or CLP procedures. Twenty-four hours later, peritoneal lavage and blood were collected. Peritoneal lavage and serum IL-6, IL-10, TNFα, KC, MCP-1, and MIP-2 were measured with a multiplex fluorescent bead-based immunoassay. A . Peritoneal cytokines. B. Serum cytokines. Each error bar represents mean ± SE. The number of animals in each group: WT-Sham = 3, WT-CLP = 4, Trif −/− -Sham = 3, Trif −/− -CLP = 6. CLP = cecum ligation and puncture; IL = interleukin; KC = keratinocyte chemoattractant; MCP-1 = monocyte chemoattractant protein-1; MIP-2 = macrophage inflammatory protein-2; MyD88 = myeloid differentiation factor 88; TNF = tumor necrosis factor; Trif = TIR-domain-containing adaptor protein inducing interferon-β mediated transcription-factor; WT = wild type.

    Journal: Anesthesiology

    Article Title: MyD88 and Trif signaling play distinct roles in cardiac dysfunction and mortality during endotoxin shock and polymicrobial sepsis

    doi: 10.1097/ALN.0b013e31822a22f7

    Figure Lengend Snippet: Trif deficiency has no impact on peritoneal and serum cytokine levels during polymicrobial sepsis WT and Trif −/− mice underwent sham or CLP procedures. Twenty-four hours later, peritoneal lavage and blood were collected. Peritoneal lavage and serum IL-6, IL-10, TNFα, KC, MCP-1, and MIP-2 were measured with a multiplex fluorescent bead-based immunoassay. A . Peritoneal cytokines. B. Serum cytokines. Each error bar represents mean ± SE. The number of animals in each group: WT-Sham = 3, WT-CLP = 4, Trif −/− -Sham = 3, Trif −/− -CLP = 6. CLP = cecum ligation and puncture; IL = interleukin; KC = keratinocyte chemoattractant; MCP-1 = monocyte chemoattractant protein-1; MIP-2 = macrophage inflammatory protein-2; MyD88 = myeloid differentiation factor 88; TNF = tumor necrosis factor; Trif = TIR-domain-containing adaptor protein inducing interferon-β mediated transcription-factor; WT = wild type.

    Article Snippet: Briefly, antibody for each cytokine was covalently immobilized to a set of fluorescent microspheres by manufacturer (Millipore, Billerica, MA).

    Techniques: Mouse Assay, Multiplex Assay, Bead-based Assay, Ligation

    Effect of MyD88- and Trif-deficiency on endotoxin-induced cytokine production in serum WT, MyD88 −/− , and Trif −/− mice were administered with LPS (15 mg/kg, intra-peritoneal injection) or saline. Six hours later, blood was collected. Serum IL-6, TNFα, and IL-1β were measured with a multiplex fluorescent bead-based immunoassay. Each error bar represents mean ± SE. The number of animals in each group: WT-Saline = 5, WT-LPS = 7, MyD88 −/− -Saline = 5, MyD88 −/− -LPS = 5, Trif −/− -Saline = 3, Trif −/− -LPS = 6. * P

    Journal: Anesthesiology

    Article Title: MyD88 and Trif signaling play distinct roles in cardiac dysfunction and mortality during endotoxin shock and polymicrobial sepsis

    doi: 10.1097/ALN.0b013e31822a22f7

    Figure Lengend Snippet: Effect of MyD88- and Trif-deficiency on endotoxin-induced cytokine production in serum WT, MyD88 −/− , and Trif −/− mice were administered with LPS (15 mg/kg, intra-peritoneal injection) or saline. Six hours later, blood was collected. Serum IL-6, TNFα, and IL-1β were measured with a multiplex fluorescent bead-based immunoassay. Each error bar represents mean ± SE. The number of animals in each group: WT-Saline = 5, WT-LPS = 7, MyD88 −/− -Saline = 5, MyD88 −/− -LPS = 5, Trif −/− -Saline = 3, Trif −/− -LPS = 6. * P

    Article Snippet: Briefly, antibody for each cytokine was covalently immobilized to a set of fluorescent microspheres by manufacturer (Millipore, Billerica, MA).

    Techniques: Mouse Assay, Injection, Multiplex Assay, Bead-based Assay

    MyD88 −/− mice had attenuated peritoneal lavage and serum cytokine levels during polymicrobial sepsis WT and MyD88 −/− mice underwent sham or CLP procedures. Twenty-four hours later, peritoneal lavage and blood were collected. Six cytokines, namely IL-6, IL-10, TNFα, KC, MCP-1, and MIP-2, in the peritoneal lavage and sera were measured using a multiplex fluorescent bead-based immunoassay. A . Peritoneal lavage cytokines. B. Serum cytokines. Each error bar represents mean ± SE. The number of animals in each group: WT-Sham = 4, WT-CLP = 4, MyD88 −/− -Sham = 3, MyD88 −/− -CLP = 3. CLP = cecum ligation and puncture; IL = interleukin; TNF = tumor necrosis factor; KC = keratinocyte chemoattractant; MCP-1 = monocyte chemoattractant protein-1; MIP-2 = macrophage inflammatory protein-2; MyD88 = myeloid differentiation factor 88; Trif = TIR-domain-containing adaptor protein inducing interferon-β mediated transcription-factor; WT = wild type. * P

    Journal: Anesthesiology

    Article Title: MyD88 and Trif signaling play distinct roles in cardiac dysfunction and mortality during endotoxin shock and polymicrobial sepsis

    doi: 10.1097/ALN.0b013e31822a22f7

    Figure Lengend Snippet: MyD88 −/− mice had attenuated peritoneal lavage and serum cytokine levels during polymicrobial sepsis WT and MyD88 −/− mice underwent sham or CLP procedures. Twenty-four hours later, peritoneal lavage and blood were collected. Six cytokines, namely IL-6, IL-10, TNFα, KC, MCP-1, and MIP-2, in the peritoneal lavage and sera were measured using a multiplex fluorescent bead-based immunoassay. A . Peritoneal lavage cytokines. B. Serum cytokines. Each error bar represents mean ± SE. The number of animals in each group: WT-Sham = 4, WT-CLP = 4, MyD88 −/− -Sham = 3, MyD88 −/− -CLP = 3. CLP = cecum ligation and puncture; IL = interleukin; TNF = tumor necrosis factor; KC = keratinocyte chemoattractant; MCP-1 = monocyte chemoattractant protein-1; MIP-2 = macrophage inflammatory protein-2; MyD88 = myeloid differentiation factor 88; Trif = TIR-domain-containing adaptor protein inducing interferon-β mediated transcription-factor; WT = wild type. * P

    Article Snippet: Briefly, antibody for each cytokine was covalently immobilized to a set of fluorescent microspheres by manufacturer (Millipore, Billerica, MA).

    Techniques: Mouse Assay, Multiplex Assay, Bead-based Assay, Ligation

    A-SAA production by NHEK stimulated with human IL-1α, IL-17A, IL-22, OSM, TNF-α, alone or in combination. (A) A-SAA mRNA expression and (B) protein secretion by NHEK stimulated with M5 were analyzed at different time-periods. (C) A-SAA mRNA expression and (D) protein secretion were determined 40 hours after cytokine activation. (E) A-SAA mRNA expression and (F) protein secretion by NHEK 40 hours after stimulation with four cytokines by sequentially subtracting either recombinant IL-1α, IL-17A, IL-22, OSM or TNF-α from M5. A-SAA mRNA and protein were quantified by RT-qPCR in NHEK and ELISA in supernatants, respectively. Data represent the mean ± SEM of three experiments with duplicates. Statistical comparisons were performed using 2way ANOVA test or t test (*p

    Journal: PLoS ONE

    Article Title: Interleukin-17A-induced production of acute serum amyloid A by keratinocytes contributes to psoriasis pathogenesis

    doi: 10.1371/journal.pone.0181486

    Figure Lengend Snippet: A-SAA production by NHEK stimulated with human IL-1α, IL-17A, IL-22, OSM, TNF-α, alone or in combination. (A) A-SAA mRNA expression and (B) protein secretion by NHEK stimulated with M5 were analyzed at different time-periods. (C) A-SAA mRNA expression and (D) protein secretion were determined 40 hours after cytokine activation. (E) A-SAA mRNA expression and (F) protein secretion by NHEK 40 hours after stimulation with four cytokines by sequentially subtracting either recombinant IL-1α, IL-17A, IL-22, OSM or TNF-α from M5. A-SAA mRNA and protein were quantified by RT-qPCR in NHEK and ELISA in supernatants, respectively. Data represent the mean ± SEM of three experiments with duplicates. Statistical comparisons were performed using 2way ANOVA test or t test (*p

    Article Snippet: NHEK were stimulated for different time-periods with human recombinant cytokines alone or in combination (IL-1α, IL-17A, IL-22, OSM, TNF-α: M5), as described previously (final concentration 10 ng/ml of each cytokine; R & D Systems) [ ] or with human rA-SAA (10 μg/ml, purity > 98% and endotoxin level is < 0.1 ng/μg of protein, Peprotech,) associated or not with rIL-17A (10 ng/ml, R & D Systems).

    Techniques: Expressing, Activation Assay, Recombinant, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay

    Modulation of the A549 phenotype by IFN-γ and/or TNF-α (in percentage) . The A549 cells were incubated during 24 hrs with human recombinant IFN-γ (100 ng/ml, I100), TNF-α (50 ng/ml, T50) or both cytokines (50 ng/ml IFN-γ and 25 ng/ml TNF-α, I50/T25). The phenotype was analyzed by flow cytometry. The horizontal bars represent the medians of the results from seven independent experiments (each performed in triplicate). Mean values and medians are represented as dots and bars respectively. A value of P

    Journal: Respiratory Research

    Article Title: Phenotypic characteristics of human type II alveolar epithelial cells suitable for antigen presentation to T lymphocytes

    doi: 10.1186/1465-9921-12-15

    Figure Lengend Snippet: Modulation of the A549 phenotype by IFN-γ and/or TNF-α (in percentage) . The A549 cells were incubated during 24 hrs with human recombinant IFN-γ (100 ng/ml, I100), TNF-α (50 ng/ml, T50) or both cytokines (50 ng/ml IFN-γ and 25 ng/ml TNF-α, I50/T25). The phenotype was analyzed by flow cytometry. The horizontal bars represent the medians of the results from seven independent experiments (each performed in triplicate). Mean values and medians are represented as dots and bars respectively. A value of P

    Article Snippet: In parallel to resting conditions, the cells were in vitro stimulated with human recombinant IFN-γ (100 ng/ml), human recombinant TNF-α (50 ng/ml) or a combination of the two cytokines (IFN-γ 50 ng/ml and TNF-α 25 ng/ml) (R & D Systems Europe) during 24 hours before analyzing their phenotype by flow cytometry.

    Techniques: Incubation, Recombinant, Flow Cytometry, Cytometry

    Modulation of the phenotypic markers density of fresh AECII by inflammatory cytokines . These results were obtained from the same samples as those presented in Figure 4 but relative medians of fluorescence intensity (MFI) were analyzed. Results are represented by the medians and the inter-quartile ranges obtained with four independent experiments. A value of P

    Journal: Respiratory Research

    Article Title: Phenotypic characteristics of human type II alveolar epithelial cells suitable for antigen presentation to T lymphocytes

    doi: 10.1186/1465-9921-12-15

    Figure Lengend Snippet: Modulation of the phenotypic markers density of fresh AECII by inflammatory cytokines . These results were obtained from the same samples as those presented in Figure 4 but relative medians of fluorescence intensity (MFI) were analyzed. Results are represented by the medians and the inter-quartile ranges obtained with four independent experiments. A value of P

    Article Snippet: In parallel to resting conditions, the cells were in vitro stimulated with human recombinant IFN-γ (100 ng/ml), human recombinant TNF-α (50 ng/ml) or a combination of the two cytokines (IFN-γ 50 ng/ml and TNF-α 25 ng/ml) (R & D Systems Europe) during 24 hours before analyzing their phenotype by flow cytometry.

    Techniques: Fluorescence

    Modulation of the phenotypic markers density of A549 by inflammatory cytokines . These results were obtained from the same samples as those presented in Figure 2 but relative medians of fluorescence intensity (MFI) were analyzed. Results are represented by the medians and the inter-quartile ranges obtained for seven independent experiments. A value of P

    Journal: Respiratory Research

    Article Title: Phenotypic characteristics of human type II alveolar epithelial cells suitable for antigen presentation to T lymphocytes

    doi: 10.1186/1465-9921-12-15

    Figure Lengend Snippet: Modulation of the phenotypic markers density of A549 by inflammatory cytokines . These results were obtained from the same samples as those presented in Figure 2 but relative medians of fluorescence intensity (MFI) were analyzed. Results are represented by the medians and the inter-quartile ranges obtained for seven independent experiments. A value of P

    Article Snippet: In parallel to resting conditions, the cells were in vitro stimulated with human recombinant IFN-γ (100 ng/ml), human recombinant TNF-α (50 ng/ml) or a combination of the two cytokines (IFN-γ 50 ng/ml and TNF-α 25 ng/ml) (R & D Systems Europe) during 24 hours before analyzing their phenotype by flow cytometry.

    Techniques: Fluorescence

    Stat5 is not required for transformation mediated by KrasG12D. BM from pI:pC-treated control and MxCre; Stat5fl/fl mice were infected with retroviruses expressing Jak2V617F, BCR-ABL, or KrasG12D. Transduced BM cells (1 × 10 5 ) were then plated in methylcellulose medium without any cytokine (Methocult M3234). Cytokine-independent CFU-GM colonies (A) were counted after 7 days and CFU-E colonies (B) were counted after 2 days (n = 4). Asterisks show significant differences by t test (* P

    Journal: Blood

    Article Title: Critical requirement for Stat5 in a mouse model of polycythemia vera

    doi: 10.1182/blood-2011-03-345215

    Figure Lengend Snippet: Stat5 is not required for transformation mediated by KrasG12D. BM from pI:pC-treated control and MxCre; Stat5fl/fl mice were infected with retroviruses expressing Jak2V617F, BCR-ABL, or KrasG12D. Transduced BM cells (1 × 10 5 ) were then plated in methylcellulose medium without any cytokine (Methocult M3234). Cytokine-independent CFU-GM colonies (A) were counted after 7 days and CFU-E colonies (B) were counted after 2 days (n = 4). Asterisks show significant differences by t test (* P

    Article Snippet: To detect cytokine-independent CFU-E and CFU-GM colonies, BM (1 × 105 ) or spleen (1 × 105 ) cells were plated in duplicate in methylcellulose medium without any cytokine (Methocult M3234; StemCell Technologies).

    Techniques: Transformation Assay, Mouse Assay, Infection, Expressing

    Erk, Akt and Stat3 are dispensable for hematopoietic transformation induced by Jak2V617F. (A) Left panel: inhibition of Erk1/2 phosphorylation in the BM of Jak2V617F knockin mice by MEK inhibitor U0126. Right panel: BM cells from Jak2V617F knockin mice were plated in the methylcellulose medium in the absence of cytokine with indicated concentrations of U0126. CFU-E colonies were counted after 2 days. (B) Left panel: inhibition of Akt phosphorylation in the BM of Jak2V617F knockin mice by Akt inhibitor wortmannin. Right panel: BM cells from Jak2V617F knockin mice were plated in the methylcellulose medium in the absence of cytokine with indicated concentrations of wortmannin. CFU-E colonies were counted after 2 days. (C) Effect of dominant negative Stat3 (DN-Stat3) on phosphorylation/activation of Stat3 in the BM of Jak2V617F knockin mice and erythroid transformation mediated by Jak2V617F. BM from induced MxCre;V617F/+ mice were transduced with retrovirus expressing either DN-Stat3 or vector. After selection with puromycin (2 μg/mL) for 2 days, BM cells (1 × 10 5 ) were plated in methylcellulose medium without any cytokine (Methocult M3234). CFU-E colonies were counted after 2 days. Notably, expression of DN-Stat3 in the BM of Jak2V617F knockin mice almost completely inhibited phosphorylation of Stat3 (left panel); however, erythroid transformation induced by Jak2V617F was only partially inhibited (right panel). Asterisks show significant differences (* P

    Journal: Blood

    Article Title: Critical requirement for Stat5 in a mouse model of polycythemia vera

    doi: 10.1182/blood-2011-03-345215

    Figure Lengend Snippet: Erk, Akt and Stat3 are dispensable for hematopoietic transformation induced by Jak2V617F. (A) Left panel: inhibition of Erk1/2 phosphorylation in the BM of Jak2V617F knockin mice by MEK inhibitor U0126. Right panel: BM cells from Jak2V617F knockin mice were plated in the methylcellulose medium in the absence of cytokine with indicated concentrations of U0126. CFU-E colonies were counted after 2 days. (B) Left panel: inhibition of Akt phosphorylation in the BM of Jak2V617F knockin mice by Akt inhibitor wortmannin. Right panel: BM cells from Jak2V617F knockin mice were plated in the methylcellulose medium in the absence of cytokine with indicated concentrations of wortmannin. CFU-E colonies were counted after 2 days. (C) Effect of dominant negative Stat3 (DN-Stat3) on phosphorylation/activation of Stat3 in the BM of Jak2V617F knockin mice and erythroid transformation mediated by Jak2V617F. BM from induced MxCre;V617F/+ mice were transduced with retrovirus expressing either DN-Stat3 or vector. After selection with puromycin (2 μg/mL) for 2 days, BM cells (1 × 10 5 ) were plated in methylcellulose medium without any cytokine (Methocult M3234). CFU-E colonies were counted after 2 days. Notably, expression of DN-Stat3 in the BM of Jak2V617F knockin mice almost completely inhibited phosphorylation of Stat3 (left panel); however, erythroid transformation induced by Jak2V617F was only partially inhibited (right panel). Asterisks show significant differences (* P

    Article Snippet: To detect cytokine-independent CFU-E and CFU-GM colonies, BM (1 × 105 ) or spleen (1 × 105 ) cells were plated in duplicate in methylcellulose medium without any cytokine (Methocult M3234; StemCell Technologies).

    Techniques: Transformation Assay, Inhibition, Knock-In, Mouse Assay, Dominant Negative Mutation, Activation Assay, Transduction, Expressing, Plasmid Preparation, Selection

    Molecular and functional subdivision of the HSC compartment using the Ikaros-GFP reporter. ( a ) Analysis of BM HSCs for Ikaros-GFP expression. Lineage-negative BM cells were examined for expression of Sca-1, c-Kit (LSK-left plot) and Ikaros-GFP (right histogram). The percentage of the LSKs is indicated (left) as well as the percentage of LSK progenitors falling within the two -GFP neg—lo and -GFP + gates (right). Sorted LSK -GFP neg—lo and -GFP + subsets were used for subsequent studies. ( b ) Semi-quantitative RT-PCR analysis of endogenous Zfpn1a1 ( Ikaros ) expression with primers spanning exons 3–8. Ikaros isoforms are indicated on the right side of the panel. Analysis is representative of two independent studies. ( c ) Semi-quantitative RT-PCR analysis for lineage-specific genes in LSK -GFP subsets. C, normalization and identity controls; GM, granulocyte-macrophage; EMk, erythro-megakaryocyte; Ly, lymphoid. Data is representative of at least four independent studies. ( d ) Myelo-erythroid differentiation potential of 100 sorted cells from LSK -GFP subpopulations revealed by colony assays performed in the presence of Epo or mixed cytokines (Mix). The percentage of colonies obtained from each subset is shown. Colonies were scored from day 2–4 for the Epo cultures, and from day 5–17 for the Mix cultures. CFU-E, colony-forming unit erythroid; BFU-E, burst forming unit erythroid; EMk, erythoid-megakaryocyte; Mk, megakaryocyte; G, granulocyte; GM, granulocyte-macrophage; M, macrophage; Mc, mast cells; Mix, mixed colonies contain more than three lineages usually G, M, E and Mk. The colony assays shown are from one representative out of more than five independent experiments with similar results. ( e ) Microscopic examination of LSK -GFP colonies. A 50x magnification of a representative mix-lineage colony from the LSK -GFP neg—lo cultures (upper left) and a GM colony from the LSK -GFP + cultures (upper right) visualized by fluorescence microscopy. A 600x magnified image of a mix-colony from the LSK -GFP neg—lo culture (lower left) and a GM colony from the LSK -GFP + culture (lower right) stained by May-Giemsa and visualized by bright field microscopy. ( f ) Lymphoid activities of LSK -GFP neg—lo and -GFP + subsets. 200–500 sorted cells were co-cultured on OP9-GFP (top) and OP9-DL1 (middle and bottom) stromal cells. Differentiating cells were analyzed by expression of lymphoid markers and their percentages are shown in each quadrant. One representative out of three independent studies is shown. ( g online).

    Journal: Nature immunology

    Article Title: Early hemopoietic lineage restrictions directed by Ikaros

    doi: 10.1038/ni1314

    Figure Lengend Snippet: Molecular and functional subdivision of the HSC compartment using the Ikaros-GFP reporter. ( a ) Analysis of BM HSCs for Ikaros-GFP expression. Lineage-negative BM cells were examined for expression of Sca-1, c-Kit (LSK-left plot) and Ikaros-GFP (right histogram). The percentage of the LSKs is indicated (left) as well as the percentage of LSK progenitors falling within the two -GFP neg—lo and -GFP + gates (right). Sorted LSK -GFP neg—lo and -GFP + subsets were used for subsequent studies. ( b ) Semi-quantitative RT-PCR analysis of endogenous Zfpn1a1 ( Ikaros ) expression with primers spanning exons 3–8. Ikaros isoforms are indicated on the right side of the panel. Analysis is representative of two independent studies. ( c ) Semi-quantitative RT-PCR analysis for lineage-specific genes in LSK -GFP subsets. C, normalization and identity controls; GM, granulocyte-macrophage; EMk, erythro-megakaryocyte; Ly, lymphoid. Data is representative of at least four independent studies. ( d ) Myelo-erythroid differentiation potential of 100 sorted cells from LSK -GFP subpopulations revealed by colony assays performed in the presence of Epo or mixed cytokines (Mix). The percentage of colonies obtained from each subset is shown. Colonies were scored from day 2–4 for the Epo cultures, and from day 5–17 for the Mix cultures. CFU-E, colony-forming unit erythroid; BFU-E, burst forming unit erythroid; EMk, erythoid-megakaryocyte; Mk, megakaryocyte; G, granulocyte; GM, granulocyte-macrophage; M, macrophage; Mc, mast cells; Mix, mixed colonies contain more than three lineages usually G, M, E and Mk. The colony assays shown are from one representative out of more than five independent experiments with similar results. ( e ) Microscopic examination of LSK -GFP colonies. A 50x magnification of a representative mix-lineage colony from the LSK -GFP neg—lo cultures (upper left) and a GM colony from the LSK -GFP + cultures (upper right) visualized by fluorescence microscopy. A 600x magnified image of a mix-colony from the LSK -GFP neg—lo culture (lower left) and a GM colony from the LSK -GFP + culture (lower right) stained by May-Giemsa and visualized by bright field microscopy. ( f ) Lymphoid activities of LSK -GFP neg—lo and -GFP + subsets. 200–500 sorted cells were co-cultured on OP9-GFP (top) and OP9-DL1 (middle and bottom) stromal cells. Differentiating cells were analyzed by expression of lymphoid markers and their percentages are shown in each quadrant. One representative out of three independent studies is shown. ( g online).

    Article Snippet: Cytokines were purchased from StemCell Technologies and R & D systems.

    Techniques: Functional Assay, Expressing, Quantitative RT-PCR, Fluorescence, Microscopy, Staining, Cell Culture

    Diminished neuroinflammation in Bhlhe40 −/− mice a , Flow cytometry of CNS myeloid cells from WT and Bhlhe40 −/− mice (n=3 per group) on day 16 after immunization. b, Percentage of MHC class II + microglia and infiltrating myeloid cells. c-e , Flow cytometry of CD4 + T cells in the CNS of WT and Bhlhe40 −/− mice (n=3 per group) on day 16 after immunization. ( c ) CD4 + T cell number. ( d ) Representative ICS for the indicated cytokines. ( e ) Frequency of CD4 + T cells secreting each of the indicated 8 possible combinations of GM-CSF, IL-17A, and IFN-γ. f, g , Flow cytometry of γδ T cells in the CNS of WT and Bhlhe40 −/− mice (n=3per group) on day 14 after immunization. ( f ) Representative ICS for IFN-γ, IL-17A, and GM-CSF. ( g ) Frequency of γδ T cells secreting IFN-γ, IL-17A, or GM-CSF.

    Journal: Nature communications

    Article Title: Bhlhe40 controls cytokine production by T cells and is essential for pathogenicity in autoimmune neuroinflammation

    doi: 10.1038/ncomms4551

    Figure Lengend Snippet: Diminished neuroinflammation in Bhlhe40 −/− mice a , Flow cytometry of CNS myeloid cells from WT and Bhlhe40 −/− mice (n=3 per group) on day 16 after immunization. b, Percentage of MHC class II + microglia and infiltrating myeloid cells. c-e , Flow cytometry of CD4 + T cells in the CNS of WT and Bhlhe40 −/− mice (n=3 per group) on day 16 after immunization. ( c ) CD4 + T cell number. ( d ) Representative ICS for the indicated cytokines. ( e ) Frequency of CD4 + T cells secreting each of the indicated 8 possible combinations of GM-CSF, IL-17A, and IFN-γ. f, g , Flow cytometry of γδ T cells in the CNS of WT and Bhlhe40 −/− mice (n=3per group) on day 14 after immunization. ( f ) Representative ICS for IFN-γ, IL-17A, and GM-CSF. ( g ) Frequency of γδ T cells secreting IFN-γ, IL-17A, or GM-CSF.

    Article Snippet: For TH cell polarization, magnetically purified total CD4+ T cells (Invitrogen Dynabeads FlowComp Mouse CD4 kit) or naïve CD4+ T cells (Stemcell Technologies Easysep Mouse Naïve CD4+ T Cell Isolation kit, typical purity ~90-96%) were cultured in cIMDM with plate bound anti-CD3 and anti-CD28 antibodies in the presence of cytokines (BioLegend except TGF-β3) and neutralizing antibodies (BioLegend except anti-IL-12) as follows.

    Techniques: Mouse Assay, Flow Cytometry, Cytometry

    T H cells require Bhlhe40 for normal cytokine production in vitro a-d, CD4 + T cells from WT and Bhlhe40 −/− mice (n=3 per group) were polarized in T H 1, T H 2, or T H 17 culture conditions for 4 days. ( a ) Representative ICS for the indicated cytokines. ( b ) Frequencies of IFN-γ + , IL-4 + , IL-17A + , GM-CSF + , and IL-10 + cells in T H 1, T H 2, and T H 17 cultures. ( c ) T H 1, T H 2, and T H 17 cells were stimulated for 24 hours with anti-CD3 and anti-CD28. GM-CSF was measured in the supernatant by ELISA. ( d ) Representative ICS for the indicated cytokines. e , CD4 + T cells from the indicated congenic mice were mixed prior to culturing in non-polarizing conditions for 4 days, followed by ICS. Cells were gated based on their expression of CD45.1 or CD45.2. Representative ICS for IL-10 and GM-CSF is shown.

    Journal: Nature communications

    Article Title: Bhlhe40 controls cytokine production by T cells and is essential for pathogenicity in autoimmune neuroinflammation

    doi: 10.1038/ncomms4551

    Figure Lengend Snippet: T H cells require Bhlhe40 for normal cytokine production in vitro a-d, CD4 + T cells from WT and Bhlhe40 −/− mice (n=3 per group) were polarized in T H 1, T H 2, or T H 17 culture conditions for 4 days. ( a ) Representative ICS for the indicated cytokines. ( b ) Frequencies of IFN-γ + , IL-4 + , IL-17A + , GM-CSF + , and IL-10 + cells in T H 1, T H 2, and T H 17 cultures. ( c ) T H 1, T H 2, and T H 17 cells were stimulated for 24 hours with anti-CD3 and anti-CD28. GM-CSF was measured in the supernatant by ELISA. ( d ) Representative ICS for the indicated cytokines. e , CD4 + T cells from the indicated congenic mice were mixed prior to culturing in non-polarizing conditions for 4 days, followed by ICS. Cells were gated based on their expression of CD45.1 or CD45.2. Representative ICS for IL-10 and GM-CSF is shown.

    Article Snippet: For TH cell polarization, magnetically purified total CD4+ T cells (Invitrogen Dynabeads FlowComp Mouse CD4 kit) or naïve CD4+ T cells (Stemcell Technologies Easysep Mouse Naïve CD4+ T Cell Isolation kit, typical purity ~90-96%) were cultured in cIMDM with plate bound anti-CD3 and anti-CD28 antibodies in the presence of cytokines (BioLegend except TGF-β3) and neutralizing antibodies (BioLegend except anti-IL-12) as follows.

    Techniques: In Vitro, Mouse Assay, Enzyme-linked Immunosorbent Assay, Expressing

    Cytokines levels in pericardial fluid and plasma samples. Cytokines levels were determined before CDCs administration and 30 days post-administration using the Luminex xMAP technology. * Statistically significant differences ( p

    Journal: PLoS ONE

    Article Title: Intrapericardial Delivery of Cardiosphere-Derived Cells: An Immunological Study in a Clinically Relevant Large Animal Model

    doi: 10.1371/journal.pone.0149001

    Figure Lengend Snippet: Cytokines levels in pericardial fluid and plasma samples. Cytokines levels were determined before CDCs administration and 30 days post-administration using the Luminex xMAP technology. * Statistically significant differences ( p

    Article Snippet: Again, the limited availability of porcine-specific reagents for cytokines was an important limitation and Luminex technology allowed us the quantification of only nine cytokines.

    Techniques: Luminex

    Inflammatory cytokine profile in response to respiratory introduction of predatory bacteria. qPCR analysis of IL-1β, IL-4, IL-6, IL-10, IL-12, IL-23, IFNγ, and TNF in response to intranasal inoculation of predatory bacteria relative to PBS control. Mice were inoculated intranasally with B. bacteriovorus 109J, HD100 or M. aeruginosavorus ARL-13. Expression of cytokines was assessed in the lung at ( A ) 1, ( B ) 24, and ( C ) 48 hours post-inoculation. Expression of cytokines was assessed in the spleen at ( D ) 24 and ( E ) 48 hours post-inoculation. Fold induction was calculated using the ΔΔC t by approximation method using an endogenous calibrator (β-actin). For the one hour experiment, 6 mice per predatory bacterial strain (and PBS) were used. Twelve mice per strain (and PBS) were used at each of the 24 and 48 hour time points, with the exception of the Lung/24 hour experiment, where 6 mice were used. Data for the one hour time point is from one experiment; data for the 24 and 48 hour time points are from two independent experiments. Data represent mean ± standard deviation. Student’s t -test was performed to compare each treated sample to their respective control sample, * P

    Journal: Scientific Reports

    Article Title: Examining the safety of respiratory and intravenous inoculation of Bdellovibrio bacteriovorus and Micavibrio aeruginosavorus in a mouse model

    doi: 10.1038/srep12899

    Figure Lengend Snippet: Inflammatory cytokine profile in response to respiratory introduction of predatory bacteria. qPCR analysis of IL-1β, IL-4, IL-6, IL-10, IL-12, IL-23, IFNγ, and TNF in response to intranasal inoculation of predatory bacteria relative to PBS control. Mice were inoculated intranasally with B. bacteriovorus 109J, HD100 or M. aeruginosavorus ARL-13. Expression of cytokines was assessed in the lung at ( A ) 1, ( B ) 24, and ( C ) 48 hours post-inoculation. Expression of cytokines was assessed in the spleen at ( D ) 24 and ( E ) 48 hours post-inoculation. Fold induction was calculated using the ΔΔC t by approximation method using an endogenous calibrator (β-actin). For the one hour experiment, 6 mice per predatory bacterial strain (and PBS) were used. Twelve mice per strain (and PBS) were used at each of the 24 and 48 hour time points, with the exception of the Lung/24 hour experiment, where 6 mice were used. Data for the one hour time point is from one experiment; data for the 24 and 48 hour time points are from two independent experiments. Data represent mean ± standard deviation. Student’s t -test was performed to compare each treated sample to their respective control sample, * P

    Article Snippet: Cytokines were measured using a V-Plex Proinflammatory Panel1 (mouse) Kit (K15048D-1; Meso Scale Discovery) according to manufacturer’s instructions, and read on a SECTOR Imager 2400 (Meso Scale Discovery).

    Techniques: Real-time Polymerase Chain Reaction, Mouse Assay, Expressing, Standard Deviation

    Inflammatory cytokine profile in response to intravenous injection of B. bacteriovorus 109J. ( A – C ) qPCR analysis of IL-6, IL-12, IFNγ, and TNF in response to tail vein injection of B. bacteriovorus 109J relative to PBS control. Expression of cytokines was assessed in the ( A ) kidney, ( B ) liver and ( C ) spleen at 3 and 18 hours post-injection. Fold induction was calculated using the ΔΔC t by approximation method using an endogenous calibrator (β-actin). Five mice per treatment group were used at each time point. Data represent mean ± standard deviation. ( D ) ELISA analysis of IL-1β, IL-6, IL-10, IL-12, CXCL-1/KC, IFNγ, and TNF in response to tail vein injection of B. bacteriovorus 109J relative to PBS control. Inflammatory proteins were assessed in the blood at 3 and 18 hours post-injection. Five mice per treatment group were used at each time point. Data represent mean ± standard error of the mean. Student’s t -test was performed to compare each treated sample to their respective control sample, * P

    Journal: Scientific Reports

    Article Title: Examining the safety of respiratory and intravenous inoculation of Bdellovibrio bacteriovorus and Micavibrio aeruginosavorus in a mouse model

    doi: 10.1038/srep12899

    Figure Lengend Snippet: Inflammatory cytokine profile in response to intravenous injection of B. bacteriovorus 109J. ( A – C ) qPCR analysis of IL-6, IL-12, IFNγ, and TNF in response to tail vein injection of B. bacteriovorus 109J relative to PBS control. Expression of cytokines was assessed in the ( A ) kidney, ( B ) liver and ( C ) spleen at 3 and 18 hours post-injection. Fold induction was calculated using the ΔΔC t by approximation method using an endogenous calibrator (β-actin). Five mice per treatment group were used at each time point. Data represent mean ± standard deviation. ( D ) ELISA analysis of IL-1β, IL-6, IL-10, IL-12, CXCL-1/KC, IFNγ, and TNF in response to tail vein injection of B. bacteriovorus 109J relative to PBS control. Inflammatory proteins were assessed in the blood at 3 and 18 hours post-injection. Five mice per treatment group were used at each time point. Data represent mean ± standard error of the mean. Student’s t -test was performed to compare each treated sample to their respective control sample, * P

    Article Snippet: Cytokines were measured using a V-Plex Proinflammatory Panel1 (mouse) Kit (K15048D-1; Meso Scale Discovery) according to manufacturer’s instructions, and read on a SECTOR Imager 2400 (Meso Scale Discovery).

    Techniques: Injection, Real-time Polymerase Chain Reaction, Expressing, Mouse Assay, Standard Deviation, Enzyme-linked Immunosorbent Assay

    CNS Treg Cells are Stable and Express Blimp1 in Response to Proinflammatory Cytokines (A) Mononuclear cells were isolated from the CNS of EAE mice at the peak of disease and were stimulated with phorbol 12-myristate 13-acetate (PMA) and ionomycin to analyze the expression of IL-10, IL-17, and IFN-γ in Treg cells by flow cytometry. Mean of eight biological replicates ± SD, derived from three independent experiments. Symbols depict individual mice (bars, mean ± SD). (B) CD4 + Foxp3 + Treg cells were sorted from the CNS and spleen (SPL) of Foxp3 (GFP) reporter mice at the peak of EAE and subjected to RNA-seq. Principal-component analysis. (C) Foxp3 expression by intracellular staining of splenic Treg cells and CNS Treg cells at the peak of EAE. Mean of six biological replicates ± SD, derived from two independent experiments. Symbols depict individual mice (bars, mean ± SD), t test, p

    Journal: Cell Reports

    Article Title: Blimp1 Prevents Methylation of Foxp3 and Loss of Regulatory T Cell Identity at Sites of Inflammation

    doi: 10.1016/j.celrep.2019.01.070

    Figure Lengend Snippet: CNS Treg Cells are Stable and Express Blimp1 in Response to Proinflammatory Cytokines (A) Mononuclear cells were isolated from the CNS of EAE mice at the peak of disease and were stimulated with phorbol 12-myristate 13-acetate (PMA) and ionomycin to analyze the expression of IL-10, IL-17, and IFN-γ in Treg cells by flow cytometry. Mean of eight biological replicates ± SD, derived from three independent experiments. Symbols depict individual mice (bars, mean ± SD). (B) CD4 + Foxp3 + Treg cells were sorted from the CNS and spleen (SPL) of Foxp3 (GFP) reporter mice at the peak of EAE and subjected to RNA-seq. Principal-component analysis. (C) Foxp3 expression by intracellular staining of splenic Treg cells and CNS Treg cells at the peak of EAE. Mean of six biological replicates ± SD, derived from two independent experiments. Symbols depict individual mice (bars, mean ± SD), t test, p

    Article Snippet: All cytokines were purchased from Miltenyi Biotec and R & D Systems.

    Techniques: Isolation, Mouse Assay, Expressing, Flow Cytometry, Cytometry, Derivative Assay, RNA Sequencing Assay, Staining

    The release of proinflammatory cytokines and prostaglandin E 2 are impaired in PKCε −/ − mice. (A–C) Thioglycollate-elicited peritoneal macrophages were activated for the indicated times with 200 ng/ml of LPS and 20 U/ml of IFNγ. The amount of TNF-α, IL-1β, and PGE 2 in the culture medium was then determined. (D–E) Bar graphs demonstrating the serum levels of TNF-α (1 h after injection) and IL-1β (4 h after injection) after the intraperitoneal injection of either 5 mg/kg or 40 mg/kg of LPS. (F) Apoptosis of thioglycollate-elicited peritoneal macrophages 24 h after activation by LPS (200 ng/ml), LPS plus IFNγ (200 ng/ml and 20 U/ml, respectively), or LPS, IFNγ, and 15dPGJ 2 (2 μM). Results show the mean ± SEM of three experiments. The * and ** denote P

    Journal: The Journal of Experimental Medicine

    Article Title: Protein Kinase C? Is Required for Macrophage Activation and Defense Against Bacterial Infection

    doi:

    Figure Lengend Snippet: The release of proinflammatory cytokines and prostaglandin E 2 are impaired in PKCε −/ − mice. (A–C) Thioglycollate-elicited peritoneal macrophages were activated for the indicated times with 200 ng/ml of LPS and 20 U/ml of IFNγ. The amount of TNF-α, IL-1β, and PGE 2 in the culture medium was then determined. (D–E) Bar graphs demonstrating the serum levels of TNF-α (1 h after injection) and IL-1β (4 h after injection) after the intraperitoneal injection of either 5 mg/kg or 40 mg/kg of LPS. (F) Apoptosis of thioglycollate-elicited peritoneal macrophages 24 h after activation by LPS (200 ng/ml), LPS plus IFNγ (200 ng/ml and 20 U/ml, respectively), or LPS, IFNγ, and 15dPGJ 2 (2 μM). Results show the mean ± SEM of three experiments. The * and ** denote P

    Article Snippet: Cytokines were from Roche.

    Techniques: Mouse Assay, Injection, Activation Assay

    4% Lovaza ® decreases LPS induced cytokines in cultured splenocytes from (NZB×NZW)F1 female mice. Two-month-old mice were fed semi-purified diets containing 1% placebo, 4% placebo, 1% Lovaza ® , 4% Lovaza ® , 4% FO-18/12 and standard LC diet as a control. At eight months of age, splenocytes were isolated, cultured aseptically and stimulated with LPS for 24 h. 4% Lovaza ® fed mice showed significantly decreased (A)IL-1β, (B)IL-6, (C) TNF-α and (D) TGF-β in conditioned medium of splenocyte culture by ELISA (5 mice/group). Results are expressed as mean±SEM. * P

    Journal: Experimental biology and medicine (Maywood, N.J.)

    Article Title: Concentrated fish oil (Lovaza®) extends lifespan and attenuates kidney disease in lupus-prone short-lived (NZBxNZW)F1 mice

    doi: 10.1177/1535370213489485

    Figure Lengend Snippet: 4% Lovaza ® decreases LPS induced cytokines in cultured splenocytes from (NZB×NZW)F1 female mice. Two-month-old mice were fed semi-purified diets containing 1% placebo, 4% placebo, 1% Lovaza ® , 4% Lovaza ® , 4% FO-18/12 and standard LC diet as a control. At eight months of age, splenocytes were isolated, cultured aseptically and stimulated with LPS for 24 h. 4% Lovaza ® fed mice showed significantly decreased (A)IL-1β, (B)IL-6, (C) TNF-α and (D) TGF-β in conditioned medium of splenocyte culture by ELISA (5 mice/group). Results are expressed as mean±SEM. * P

    Article Snippet: Multi-Analyte ELISArray™ was used analysed for cytokines (SA Biosciences Inc. San Diego, CA) in supernatant.

    Techniques: Cell Culture, Mouse Assay, Purification, Isolation, Enzyme-linked Immunosorbent Assay

    Th17 cells are resistant to apoptosis in the absence of IL2Rγ-chain cytokines. a Gating strategy for apoptosis assays b,c Apoptosis of Th17 cells (TRP-1), unpolarized CD4 + T cells (TRP-1), Tc17 cells (pmel-1), or unpolarized CD8 + T cells (pmel-1) was measured after culture with IL-2, IL-7, IL-15, or vehicle. Panel b shows representative samples from day 2. Panel c shows mean plus standard error of 3 independent experiments for the days indicated. d,e Antibody-mediated depletion of IL2Rγ-chain cytokines does not induce apoptosis of Th17 (TRP-1) cells over 2 day culture. Triangles represent independent experiments. p > .05 for all depletion conditions vs control by 2-tailed t-test. f Expression of Bcl-2 protein is greater in Th17 cells (dashed line) than in unpolarized CD8 + cells (solid line). Grey histogram is FMO control. Representative of two independent experiments

    Journal: Cancer immunology, immunotherapy : CII

    Article Title: Murine Th17 cells utilize IL-2 receptor gamma chain cytokines but are resistant to cytokine withdrawal-induced apoptosis

    doi: 10.1007/s00262-017-1965-3

    Figure Lengend Snippet: Th17 cells are resistant to apoptosis in the absence of IL2Rγ-chain cytokines. a Gating strategy for apoptosis assays b,c Apoptosis of Th17 cells (TRP-1), unpolarized CD4 + T cells (TRP-1), Tc17 cells (pmel-1), or unpolarized CD8 + T cells (pmel-1) was measured after culture with IL-2, IL-7, IL-15, or vehicle. Panel b shows representative samples from day 2. Panel c shows mean plus standard error of 3 independent experiments for the days indicated. d,e Antibody-mediated depletion of IL2Rγ-chain cytokines does not induce apoptosis of Th17 (TRP-1) cells over 2 day culture. Triangles represent independent experiments. p > .05 for all depletion conditions vs control by 2-tailed t-test. f Expression of Bcl-2 protein is greater in Th17 cells (dashed line) than in unpolarized CD8 + cells (solid line). Grey histogram is FMO control. Representative of two independent experiments

    Article Snippet: Cytokines were obtained from Shenandoah Biotechnology unless otherwise noted.

    Techniques: Expressing

    Th17 cells respond to IL2Rγ-chain cytokines in vitro . a,b TRP-1 Th17 cells phosphorylate STAT5, MEK, Akt, and S6 following IL-2 (100ng/ml, solid lines) or IL-15 (100ng/ml, dotted lines) stimulation (gray histograms = no cytokine). Graphs show mean plus standard error of 8 independent experiments, *p

    Journal: Cancer immunology, immunotherapy : CII

    Article Title: Murine Th17 cells utilize IL-2 receptor gamma chain cytokines but are resistant to cytokine withdrawal-induced apoptosis

    doi: 10.1007/s00262-017-1965-3

    Figure Lengend Snippet: Th17 cells respond to IL2Rγ-chain cytokines in vitro . a,b TRP-1 Th17 cells phosphorylate STAT5, MEK, Akt, and S6 following IL-2 (100ng/ml, solid lines) or IL-15 (100ng/ml, dotted lines) stimulation (gray histograms = no cytokine). Graphs show mean plus standard error of 8 independent experiments, *p

    Article Snippet: Cytokines were obtained from Shenandoah Biotechnology unless otherwise noted.

    Techniques: In Vitro

    IL2Rγ-chain cytokines affect the ability of adoptively transferred Th17 cells to control tumor growth. a Depletion of IL-2 and IL-7 decreases efficacy of Th17 adoptive transfer. B6 mice bearing established B16 melanomas received adoptive transfer of 5×10 5 TRP-1 Th17 cells. The indicated IL2Rγ-chain cytokines were then depleted from the mice and tumor area was measured 3 times per week. Each line represents one mouse. b Survival curves for deaths due to any cause (top) or only deaths attributable to tumor burden (tumor area > 400mm 2 ) (bottom), survival data was pooled from 2 independent experiments with a total of 14–16 mice or 8–12 mice per group for all cause and tumor specific mortality respectively, *p

    Journal: Cancer immunology, immunotherapy : CII

    Article Title: Murine Th17 cells utilize IL-2 receptor gamma chain cytokines but are resistant to cytokine withdrawal-induced apoptosis

    doi: 10.1007/s00262-017-1965-3

    Figure Lengend Snippet: IL2Rγ-chain cytokines affect the ability of adoptively transferred Th17 cells to control tumor growth. a Depletion of IL-2 and IL-7 decreases efficacy of Th17 adoptive transfer. B6 mice bearing established B16 melanomas received adoptive transfer of 5×10 5 TRP-1 Th17 cells. The indicated IL2Rγ-chain cytokines were then depleted from the mice and tumor area was measured 3 times per week. Each line represents one mouse. b Survival curves for deaths due to any cause (top) or only deaths attributable to tumor burden (tumor area > 400mm 2 ) (bottom), survival data was pooled from 2 independent experiments with a total of 14–16 mice or 8–12 mice per group for all cause and tumor specific mortality respectively, *p

    Article Snippet: Cytokines were obtained from Shenandoah Biotechnology unless otherwise noted.

    Techniques: Adoptive Transfer Assay, Mouse Assay

    Expression of cytokine mRNA, and IL-10 protein levels in M-CSF- or GM-CSF-cultured BMCs. (A) The expression of proinflammatory cytokines (TNF-alpha, IL-6, IL-1beta, IL-12a, and IL12b) and anti-inflammatory cytokine (IL-10) mRNA in M-CSF- or GM-CSF-cultured BMCs. (B) IL-10 concentration in the supernatant of M-CSF- or GM-CSF-cultured BMCs. Data are expressed as mean ± SEM, *** P

    Journal: PLoS ONE

    Article Title: GM-CSF Treated F4/80+ BMCs Improve Murine Hind Limb Ischemia Similar to M-CSF Differentiated Macrophages

    doi: 10.1371/journal.pone.0106987

    Figure Lengend Snippet: Expression of cytokine mRNA, and IL-10 protein levels in M-CSF- or GM-CSF-cultured BMCs. (A) The expression of proinflammatory cytokines (TNF-alpha, IL-6, IL-1beta, IL-12a, and IL12b) and anti-inflammatory cytokine (IL-10) mRNA in M-CSF- or GM-CSF-cultured BMCs. (B) IL-10 concentration in the supernatant of M-CSF- or GM-CSF-cultured BMCs. Data are expressed as mean ± SEM, *** P

    Article Snippet: Real-time PCR expression profile of cytokines was performed in M-CSF- or GM-CSF-cultured BMCs at day 7 using SYBR Green (TaKaRa, Japan) and LightCycler 2.0 systems (Roche, Basel, Schweiz), and relative quantification analysis was performed with LightCycler Software Version 4.1.

    Techniques: Expressing, Cell Culture, Concentration Assay

    Serum cytokine induction. Levels of representative cytokines (CCL2, CSF3, CXCL1, TNFα, IL-5, IL-6, IFNγ, and CXCL10) were measured for each treatment group in the serum at 6–48 hours after adjuvant administration in 3–4 replicate experiments. Data is presented as the mean plus standard deviation of N = 8–12 for the 6 and 24 hour timepoints and N = 3 for the 48 hour timepoint.

    Journal: PLoS ONE

    Article Title: Molecular and Cellular Response Profiles Induced by the TLR4 Agonist-Based Adjuvant Glucopyranosyl Lipid A

    doi: 10.1371/journal.pone.0051618

    Figure Lengend Snippet: Serum cytokine induction. Levels of representative cytokines (CCL2, CSF3, CXCL1, TNFα, IL-5, IL-6, IFNγ, and CXCL10) were measured for each treatment group in the serum at 6–48 hours after adjuvant administration in 3–4 replicate experiments. Data is presented as the mean plus standard deviation of N = 8–12 for the 6 and 24 hour timepoints and N = 3 for the 48 hour timepoint.

    Article Snippet: However, we did not see significant transcriptional activation of these cytokines at the injection site although alum obtained from Pierce was reported to do so .

    Techniques: Standard Deviation

    Several cytokines are increased significantly in the sera of infected STAT1 KO mice. In WT mice, a multiplex assay showed small to moderate but significant changes for CCL4, CCL2, and IL-2 levels at day 3 postinfection (A) and for G-CSF, CCL2, IL-5, IL-12,

    Journal: Journal of Virology

    Article Title: Mice Deficient in STAT1 but Not STAT2 or IRF9 Develop a Lethal CD4+ T-Cell-Mediated Disease following Infection with Lymphocytic Choriomeningitis Virus

    doi: 10.1128/JVI.07147-11

    Figure Lengend Snippet: Several cytokines are increased significantly in the sera of infected STAT1 KO mice. In WT mice, a multiplex assay showed small to moderate but significant changes for CCL4, CCL2, and IL-2 levels at day 3 postinfection (A) and for G-CSF, CCL2, IL-5, IL-12,

    Article Snippet: To determine cytokine levels in serum, a Luminex assay for 23 cytokines (Bioplex Mouse Cytokine 23-Plex Panel; Bio-Rad, Gladesville, Australia) was performed according to the manufacturer's instructions.

    Techniques: Infection, Mouse Assay, Multiplex Assay

    Receptors for the pro-inflammatory cytokines TNFα and IL-1β are present on cells of the smooth muscle/myenteric plexus layer in vivo and in vitro . A , B , Images of fluorescence immunocytochemistry showing the presence of staining for TNF-R1 ( A ) and IL-1 R1 ( B ) on sections of control intestine. Staining was evident on smooth muscle cells of the CSM and longitudinal muscle (LSM) layers, as well as on neurons of the myenteric plexus (arrows). Scale bar, 100 μm. C , D , Depiction of the coculture model of myenteric neurons and intestinal smooth muscle cells. C , Nomarski image of culture appearance at 7 d in vitro , showing complex architecture. D , Fluorescence image of same field showing neurons and neurites among smooth muscle cells, as revealed by immunostaining for SNAP-25 (red) and the neuronal cell body marker HuD (green). Arrows indicate the location of neuronal cell bodies (yellow, merged color). Nuclei were stained with Hoechst (blue). Scale bar, 250 μm. E , F , Typical outcome of dual label immunocytochemistry showing expression of cytokine receptors on myenteric neurons in vitro . E , Images of positive staining for TNF-RI and the neuronal marker HuD (arrows) and their overlap (merged). Asterisks (*) indicate neurites. F , Similar outcome as E showing overlap of staining for IL1-RI with HuD (e.g., arrows). Scale bars, 100 μm.

    Journal: The Journal of Neuroscience

    Article Title: The Pro-Inflammatory Cytokines IL-1β and TNFα Are Neurotrophic for Enteric Neurons

    doi: 10.1523/JNEUROSCI.3564-12.2013

    Figure Lengend Snippet: Receptors for the pro-inflammatory cytokines TNFα and IL-1β are present on cells of the smooth muscle/myenteric plexus layer in vivo and in vitro . A , B , Images of fluorescence immunocytochemistry showing the presence of staining for TNF-R1 ( A ) and IL-1 R1 ( B ) on sections of control intestine. Staining was evident on smooth muscle cells of the CSM and longitudinal muscle (LSM) layers, as well as on neurons of the myenteric plexus (arrows). Scale bar, 100 μm. C , D , Depiction of the coculture model of myenteric neurons and intestinal smooth muscle cells. C , Nomarski image of culture appearance at 7 d in vitro , showing complex architecture. D , Fluorescence image of same field showing neurons and neurites among smooth muscle cells, as revealed by immunostaining for SNAP-25 (red) and the neuronal cell body marker HuD (green). Arrows indicate the location of neuronal cell bodies (yellow, merged color). Nuclei were stained with Hoechst (blue). Scale bar, 250 μm. E , F , Typical outcome of dual label immunocytochemistry showing expression of cytokine receptors on myenteric neurons in vitro . E , Images of positive staining for TNF-RI and the neuronal marker HuD (arrows) and their overlap (merged). Asterisks (*) indicate neurites. F , Similar outcome as E showing overlap of staining for IL1-RI with HuD (e.g., arrows). Scale bars, 100 μm.

    Article Snippet: Alternatively, cultures were treated with cytokines (50 ng/ml) for 15–120 min, and then processed for Western blot analysis to detect phosphorylated p65, ERK, and AKT (1:1000; Cell Signaling Technology).

    Techniques: In Vivo, In Vitro, Fluorescence, Immunocytochemistry, Staining, Immunostaining, Marker, Expressing

    Pro-inflammatory cytokines are neurotrophic for myenteric neurons in vitro . A , Neuron number was unchanged in cocultures treated with the pro-inflammatory cytokines TNFα, IL-1β, or a combination of both cytokines for 48 h ( p > 0.05; n = 9, 8, and 4). B , No effect on neuronal survival with 48 h exposure to TH1 cytokines (IFNγ, IL-6), TH2 cytokines (IL-4, IL-10) at 100 ng/ml, or TGFβ (50 ng/ml) ( p > 0.05, n = 4). C , D , Images showing increased neurite outgrowth caused by TNFα in vitro . Representative images of immunofluorescence outcomes of control coculture ( C ) and coculture treated with 50 ng/ml TNFα for 48 h ( D ) labeled with anti-SNAP-25 antibodies, showing large increase in neurite number with cytokine treatment. Inset, view of neuronal cell bodies in main panels from dual label immunocytochemistry for the neuronal cell body marker HuD. Arrows indicate representative neurons in each pair of images. Scale bar, 100 μm. E , Quantification of neurite number in cocultures treated with TNFα or IL-1β for 48 h compared with untreated cohort controls, showing concentration-dependent increases in neurite number in each case ( n = 3–9; * p

    Journal: The Journal of Neuroscience

    Article Title: The Pro-Inflammatory Cytokines IL-1β and TNFα Are Neurotrophic for Enteric Neurons

    doi: 10.1523/JNEUROSCI.3564-12.2013

    Figure Lengend Snippet: Pro-inflammatory cytokines are neurotrophic for myenteric neurons in vitro . A , Neuron number was unchanged in cocultures treated with the pro-inflammatory cytokines TNFα, IL-1β, or a combination of both cytokines for 48 h ( p > 0.05; n = 9, 8, and 4). B , No effect on neuronal survival with 48 h exposure to TH1 cytokines (IFNγ, IL-6), TH2 cytokines (IL-4, IL-10) at 100 ng/ml, or TGFβ (50 ng/ml) ( p > 0.05, n = 4). C , D , Images showing increased neurite outgrowth caused by TNFα in vitro . Representative images of immunofluorescence outcomes of control coculture ( C ) and coculture treated with 50 ng/ml TNFα for 48 h ( D ) labeled with anti-SNAP-25 antibodies, showing large increase in neurite number with cytokine treatment. Inset, view of neuronal cell bodies in main panels from dual label immunocytochemistry for the neuronal cell body marker HuD. Arrows indicate representative neurons in each pair of images. Scale bar, 100 μm. E , Quantification of neurite number in cocultures treated with TNFα or IL-1β for 48 h compared with untreated cohort controls, showing concentration-dependent increases in neurite number in each case ( n = 3–9; * p

    Article Snippet: Alternatively, cultures were treated with cytokines (50 ng/ml) for 15–120 min, and then processed for Western blot analysis to detect phosphorylated p65, ERK, and AKT (1:1000; Cell Signaling Technology).

    Techniques: In Vitro, Immunofluorescence, Labeling, Immunocytochemistry, Marker, Concentration Assay

    The cytokines TNFα and IL-1β induce neurite outgrowth via GDNF expression in cocultures and in CSM cultures. A , qPCR data showing increased mRNA for GDNF in cocultures exposed to cytokines 17 h previously ( p

    Journal: The Journal of Neuroscience

    Article Title: The Pro-Inflammatory Cytokines IL-1β and TNFα Are Neurotrophic for Enteric Neurons

    doi: 10.1523/JNEUROSCI.3564-12.2013

    Figure Lengend Snippet: The cytokines TNFα and IL-1β induce neurite outgrowth via GDNF expression in cocultures and in CSM cultures. A , qPCR data showing increased mRNA for GDNF in cocultures exposed to cytokines 17 h previously ( p

    Article Snippet: Alternatively, cultures were treated with cytokines (50 ng/ml) for 15–120 min, and then processed for Western blot analysis to detect phosphorylated p65, ERK, and AKT (1:1000; Cell Signaling Technology).

    Techniques: Expressing, Real-time Polymerase Chain Reaction

    Phenotype and migration of DC matured in IRX-2 or conventional cytokines. ( A ) DC obtained from HNSCC patients were matured for 48 h either with IRX-2 or the conventional maturation cocktail. While conventionally matured DC (white bars) expressed higher levels of CD80, CD83 and CD86 (*, p

    Journal: PLoS ONE

    Article Title: IRX-2, a Novel Immunotherapeutic, Enhances Functions of Human Dendritic Cells

    doi: 10.1371/journal.pone.0047234

    Figure Lengend Snippet: Phenotype and migration of DC matured in IRX-2 or conventional cytokines. ( A ) DC obtained from HNSCC patients were matured for 48 h either with IRX-2 or the conventional maturation cocktail. While conventionally matured DC (white bars) expressed higher levels of CD80, CD83 and CD86 (*, p

    Article Snippet: To mature iDC, IRX-2 or a conventional maturation cocktail containing TNF-α (50 ng/ml), IL-1β (25 ng/ml) and IL-6 (10 ng/ml) (all cytokines from Cellgenix) diluted in ex vivo 10 medium (Lonza) were added.

    Techniques: Migration