rmil 21  (R&D Systems)

 
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    Name:
    Recombinant Mouse IL 21 Protein CF
    Description:
    The Recombinant Mouse IL 21 Protein from R D Systems is derived from E coli The Recombinant Mouse IL 21 Protein has been validated for the following applications Bioactivity
    Catalog Number:
    594-ML-010/CF
    Price:
    329
    Category:
    Proteins and Enzymes
    Source:
    E. coli-derived Recombinant Mouse IL-21 Protein
    Applications:
    Bioactivity
    Purity:
    >97%, by SDS-PAGE under reducing conditions and visualized by silver stain
    Conjugate:
    Unconjugated
    Size:
    10 ug
    Buy from Supplier


    Structured Review

    R&D Systems rmil 21
    Recombinant Mouse IL 21 Protein CF
    The Recombinant Mouse IL 21 Protein from R D Systems is derived from E coli The Recombinant Mouse IL 21 Protein has been validated for the following applications Bioactivity
    https://www.bioz.com/result/rmil 21/product/R&D Systems
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rmil 21 - by Bioz Stars, 2021-06
    93/100 stars

    Images

    1) Product Images from "STAT3-Mediated Up-Regulation of BLIMP1 Is Coordinated with BCL6 Down-Regulation to Control Human Plasma Cell Differentiation 1"

    Article Title: STAT3-Mediated Up-Regulation of BLIMP1 Is Coordinated with BCL6 Down-Regulation to Control Human Plasma Cell Differentiation 1

    Journal: Journal of immunology (Baltimore, Md. : 1950)

    doi:

    Sustained STAT3 phosphorylation after IL-21 activation in primary human B cells. A , Peripheral blood (PB) CD19 + B cells were freshly isolated and rested at 37°C for 2 h. Cells were then stimulated for 15 min with either medium alone (−, thin histogram), or rmIL-21 (200 ng/ml, shaded histogram). Levels of phospho-STAT3 (Y705), phospho-STAT1 (Y701), or phospho-STAT5 (Y694) were determined by FACS. Isotype control staining is depicted by a dashed line. B , CD19 + B cells were activated for 36 h on CD40L-transfected mouse L cell fibroblasts (CD40L-L cells). Cells were then rested followed by stimulation with medium alone (thin line) or IL-21 (shaded) for 15 or 60 min and levels of phospho-STAT3, STAT1, and STAT5 were determined by flow cytometry. C , CD19 + B cells were cultured with CD40L stimulation with medium alone (thin line) or with IL-21 (shaded) and STAT activation was measured at the indicated times.
    Figure Legend Snippet: Sustained STAT3 phosphorylation after IL-21 activation in primary human B cells. A , Peripheral blood (PB) CD19 + B cells were freshly isolated and rested at 37°C for 2 h. Cells were then stimulated for 15 min with either medium alone (−, thin histogram), or rmIL-21 (200 ng/ml, shaded histogram). Levels of phospho-STAT3 (Y705), phospho-STAT1 (Y701), or phospho-STAT5 (Y694) were determined by FACS. Isotype control staining is depicted by a dashed line. B , CD19 + B cells were activated for 36 h on CD40L-transfected mouse L cell fibroblasts (CD40L-L cells). Cells were then rested followed by stimulation with medium alone (thin line) or IL-21 (shaded) for 15 or 60 min and levels of phospho-STAT3, STAT1, and STAT5 were determined by flow cytometry. C , CD19 + B cells were cultured with CD40L stimulation with medium alone (thin line) or with IL-21 (shaded) and STAT activation was measured at the indicated times.

    Techniques Used: Activation Assay, Isolation, FACS, Staining, Transfection, Flow Cytometry, Cytometry, Cell Culture

    STAT3 activation leads to BLIMP1 up-regulation and Ig secretion. A , PB CD19 + B cells were transduced with either LZRS-control-IRES-GFP (Control, □) or LZRS-STAT3ER-IRES-GFP (STAT3ER, ■). GFP + cells were sorted and cultured on CD40L-L cells in the presence of IL-2 (40U/ml) and in the presence (+) or absence (−) of 1 μ M 4-HT for 5 days. ivPC made from total PB CD19 + B cells (see Results ) and nontransduced CD19 + cells (None, ) cultured in the presence of IL-2 or rmIL-21 (50 ng/ml) for 5 days were included for comparison. mRNA expression of BLIMP1, IRF4 , XBP1, BCL6, and PAX5 were determined by quantitative (q)RT-PCR. Results represent means ± SD. *, p
    Figure Legend Snippet: STAT3 activation leads to BLIMP1 up-regulation and Ig secretion. A , PB CD19 + B cells were transduced with either LZRS-control-IRES-GFP (Control, □) or LZRS-STAT3ER-IRES-GFP (STAT3ER, ■). GFP + cells were sorted and cultured on CD40L-L cells in the presence of IL-2 (40U/ml) and in the presence (+) or absence (−) of 1 μ M 4-HT for 5 days. ivPC made from total PB CD19 + B cells (see Results ) and nontransduced CD19 + cells (None, ) cultured in the presence of IL-2 or rmIL-21 (50 ng/ml) for 5 days were included for comparison. mRNA expression of BLIMP1, IRF4 , XBP1, BCL6, and PAX5 were determined by quantitative (q)RT-PCR. Results represent means ± SD. *, p

    Techniques Used: Activation Assay, Transduction, Cell Culture, Expressing, Polymerase Chain Reaction

    IL-21 drives partial plasma cell differentiation in BCL6-transduced B cells. A , PB CD19 + B cells expressing BCL6-IRES-YFP were cultured in the presence of IL-2 and IL-4 or with rmIL-21 alone. At the indicated time points expression of BLIMP1 and ACTB were measured by qRT-PCR. BLIMP1 expression was normalized to ACTB expression within each sample and the means ± SD of triplicate measurements in two different donors is plotted. B , BCL6-YFP expressing B cells were cultured with CD40L in the presence of IL-2 and IL-4 or with IL-21 and BLIMP1, BCL6, and tubulin protein expression were determined by immunoblotting 3 or 7 days after IL-21 treatment. C , CD19 + B cells were transduced with LZRS-BCL6-IRES-ΔNGFR and cultured for 10 days on CD40L in the presence of IL-2 and IL-4 or with IL-21 alone. Cells were stained with anti-CD19, anti-NGFR, anti-CD38, and anti-CD20 and analyzed by flow cytometry. The rightmost four plots show CD38/CD20 expression on CD19 + BCL6–ΔNGFR– and CD19 + BCL6–ΔNGFR + cells. D , CD38/CD20 expression on BCL6-IRES-YFP-transduced B cells after 21 days of culture with IL-2 and IL-4 or with IL-21 alone. E , Surface Igλ and Igκ BCR expression on cells cultured as in C . F , Expression of CD138, HLA-DR, CD86, CD27, CD25, and CD3 on BCL6 + B cells cultured as in C . Dotted histogram is iso-type control. G , CD19 + BCL6-overexpressing B cells maintained on CD40L with either IL-2 and IL-4 (○)or with IL-21 (•). Cumulative expansion was calculated based on absolute cell numbers over a 17-day period. Data shown are means ± SD of three independent experiments involving different donors and constructs (BCL6-IRES-ΔNGFR or BCL6-IRES-GFP). H , BCL6-IRES-ΔNGFR + cells were cultured for 6 days on CD40L-L cells in the presence of IL-2 and IL-4 or with IL-21. Cell numbers were counted and IgM and IgG in culture supernatants were measured by ELISA to obtain per cell values of IgM and IgG. Data are representative of three independent experiments. I , Total PB CD19 + BCL6-GFP + cells were cultured with CD40L in the presence of IL-2 and IL-4 or with IL-21. After 7 days, equal numbers of cells were plated in serial dilutions and in triplicate and IgG and IgM production were determined by ELISPOT.
    Figure Legend Snippet: IL-21 drives partial plasma cell differentiation in BCL6-transduced B cells. A , PB CD19 + B cells expressing BCL6-IRES-YFP were cultured in the presence of IL-2 and IL-4 or with rmIL-21 alone. At the indicated time points expression of BLIMP1 and ACTB were measured by qRT-PCR. BLIMP1 expression was normalized to ACTB expression within each sample and the means ± SD of triplicate measurements in two different donors is plotted. B , BCL6-YFP expressing B cells were cultured with CD40L in the presence of IL-2 and IL-4 or with IL-21 and BLIMP1, BCL6, and tubulin protein expression were determined by immunoblotting 3 or 7 days after IL-21 treatment. C , CD19 + B cells were transduced with LZRS-BCL6-IRES-ΔNGFR and cultured for 10 days on CD40L in the presence of IL-2 and IL-4 or with IL-21 alone. Cells were stained with anti-CD19, anti-NGFR, anti-CD38, and anti-CD20 and analyzed by flow cytometry. The rightmost four plots show CD38/CD20 expression on CD19 + BCL6–ΔNGFR– and CD19 + BCL6–ΔNGFR + cells. D , CD38/CD20 expression on BCL6-IRES-YFP-transduced B cells after 21 days of culture with IL-2 and IL-4 or with IL-21 alone. E , Surface Igλ and Igκ BCR expression on cells cultured as in C . F , Expression of CD138, HLA-DR, CD86, CD27, CD25, and CD3 on BCL6 + B cells cultured as in C . Dotted histogram is iso-type control. G , CD19 + BCL6-overexpressing B cells maintained on CD40L with either IL-2 and IL-4 (○)or with IL-21 (•). Cumulative expansion was calculated based on absolute cell numbers over a 17-day period. Data shown are means ± SD of three independent experiments involving different donors and constructs (BCL6-IRES-ΔNGFR or BCL6-IRES-GFP). H , BCL6-IRES-ΔNGFR + cells were cultured for 6 days on CD40L-L cells in the presence of IL-2 and IL-4 or with IL-21. Cell numbers were counted and IgM and IgG in culture supernatants were measured by ELISA to obtain per cell values of IgM and IgG. Data are representative of three independent experiments. I , Total PB CD19 + BCL6-GFP + cells were cultured with CD40L in the presence of IL-2 and IL-4 or with IL-21. After 7 days, equal numbers of cells were plated in serial dilutions and in triplicate and IgG and IgM production were determined by ELISPOT.

    Techniques Used: Cell Differentiation, Expressing, Cell Culture, Quantitative RT-PCR, Transduction, Staining, Flow Cytometry, Cytometry, Construct, Enzyme-linked Immunosorbent Assay, Enzyme-linked Immunospot

    Related Articles

    Incubation:

    Article Title: Dual regulation of IRF4 function in T and B cells is required for the coordination of T-B cell interactions and the prevention of autoimmunity
    Article Snippet: .. 4 h later, 5 µg/ml monoclonal anti-CD40 (BD) in the presence or absence of 50 ng/ml mouse IL-21 (R & D Systems) was added, and the cells were further incubated for various time periods depending on the assay. .. For plasma cell studies, B cells (106 /well) were stimulated with αIgM (5 µg/ml; Jackson ImmunoResearch Laboratories) and αCD40 (2 µg/ml) for 96 h in the presence or absence of recombinant mouse IL-21 (50 ng/ml).

    Recombinant:

    Article Title: IL-21-stimulated human plasmacytoid dendritic cells secrete granzyme B, which impairs their capacity to induce T-cell proliferation
    Article Snippet: Oligodeoxynucleotides CpG-A (ODN2216) and CpG-B (ODN2006) and R848 (Invivogen) were used at 10 μg/mL. .. For IL-21 stimulation, we used mouse recombinant IL-21 (25 ng/mL, R & D Systems), which is cross-reactive between mouse and human. .. The Z-AAD-CMK–specific inhibitor of GrB (Enzo Life Sciences, New York) was used at 5 μg/mL.

    Article Title: Upregulated IL-21 and IL-21 receptor expression is involved in experimental autoimmune uveitis (EAU)
    Article Snippet: Cells were resuspended at a density of 2×106 cells/ml in RPMI 1640 medium (Gibco, Grand Island, NY) containing L-glutamine (2 mM), penicillin/streptomycin (100 U/ml), and 10% fetal calf serum. .. For the cytokine assay cells were cultured with or without recombinant mouse interleukin-21 (rmIL-21; 100 ng/ml) and recombinant transforming growth factor-β (rTGF-β; 5 ng/ml; both from R & D Systems, Minneapolis, MN) for 72 h at 37 °C, 100% humidity, and 5% CO2. .. Reverse transcription polymerase chain reaction Total RNA was extracted from the splenocytes and DLN cells, using the RNAprep Cell kit (Tiangen Biotech, Beijing, China), according to the manufacturer’s instructions.

    other:

    Article Title: Interleukin-21 differentially affects human natural killer cell subsets
    Article Snippet: This appears to be even more attractive as a mouse model with combination therapy with IL-21 and low-dose IL-2 resulted in long-term tumour-free survival by improving the function of tumour-antigen-specific CD8+ T cells.

    Article Title: Interleukin-21 differentially affects human natural killer cell subsets
    Article Snippet: The additive effect mediated by IL-21 and low-dose IL-2 may be of great interest for clinical applications.

    Cytokine Assay:

    Article Title: Upregulated IL-21 and IL-21 receptor expression is involved in experimental autoimmune uveitis (EAU)
    Article Snippet: Cells were resuspended at a density of 2×106 cells/ml in RPMI 1640 medium (Gibco, Grand Island, NY) containing L-glutamine (2 mM), penicillin/streptomycin (100 U/ml), and 10% fetal calf serum. .. For the cytokine assay cells were cultured with or without recombinant mouse interleukin-21 (rmIL-21; 100 ng/ml) and recombinant transforming growth factor-β (rTGF-β; 5 ng/ml; both from R & D Systems, Minneapolis, MN) for 72 h at 37 °C, 100% humidity, and 5% CO2. .. Reverse transcription polymerase chain reaction Total RNA was extracted from the splenocytes and DLN cells, using the RNAprep Cell kit (Tiangen Biotech, Beijing, China), according to the manufacturer’s instructions.

    Cell Culture:

    Article Title: Upregulated IL-21 and IL-21 receptor expression is involved in experimental autoimmune uveitis (EAU)
    Article Snippet: Cells were resuspended at a density of 2×106 cells/ml in RPMI 1640 medium (Gibco, Grand Island, NY) containing L-glutamine (2 mM), penicillin/streptomycin (100 U/ml), and 10% fetal calf serum. .. For the cytokine assay cells were cultured with or without recombinant mouse interleukin-21 (rmIL-21; 100 ng/ml) and recombinant transforming growth factor-β (rTGF-β; 5 ng/ml; both from R & D Systems, Minneapolis, MN) for 72 h at 37 °C, 100% humidity, and 5% CO2. .. Reverse transcription polymerase chain reaction Total RNA was extracted from the splenocytes and DLN cells, using the RNAprep Cell kit (Tiangen Biotech, Beijing, China), according to the manufacturer’s instructions.

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  • 93
    R&D Systems recombinant murine il 21
    IRF4 is important for <t>IL-21-mediated</t> Foxp3 suppression. Flow cytometry of naïve Irf4 −/− or WT CD4 + T cells differentiated as described in A and stained for intracellular Foxp3 ( A ) and of CD4 + WT cells nucleofected, cultured
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    KW3110 or <t>IL-10</t> treatment suppresses caspase-1 activation in LPS/ATP-stimulated J774A.1 cells. (A) J774A.1 cells were treated with KW3110 (5 μg/mL) for 24 h, and treated with LPS (10 μg/mL) for 4 h and ATP (2 mM) for 1 h. (B) Cells were treated with IL-10 (1 μg/mL) and LPS (10 μg/mL) for 4 h and ATP (2 mM) for 1 h. Caspase-Glo 1 inflammasome assays measured caspase-1 activity in the supernatants from treated cells. Values indicate means ± SEM (triplicate data). Statistical differences were analyzed by ANOVA followed by Tukey’s test (**, P
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    R&D Systems recombinant il 12
    pmel-1 undergo limited homeostatic proliferation in irradiated wild-type hosts (A) Pmel-1.Rag T cells were isolated from lymph nodes and spleens and further purified by MACS negative selection for CD44 low CD8+. After CFSE labelling, pmel-1.Rag were injected into irradiated B6 hosts. For indicated animals, recombinant <t>IL-12</t> was administered on days 1, 2 and 3 after adoptive transfer. CFSE dilution of donor T cells (upper panels) and CD44 expression levels on donor and host CD8 T cells (lower panels) was assessed in the spleen at 10 days and 6 weeks after adoptive transfer. (B) Naïve pmel-1 were adoptively transferred into irradiated B6 mice. One day after transfer, mice were given i.v injections of either PBS, IL-15 or, IL-15/Rα. Mice were harvested 3 weeks later and CFSE dilution of donor T cells (upper panels) and CD44 expression levels on donor and host CD8 T cells (lower panels) was assessed. Results are representative of 3 independent experiments. In all cases, donor cells were identified by Thy-1 congenic markers.
    Recombinant Il 12, supplied by R&D Systems, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    R&D Systems anti mouse il 12 antibody
    Co-treatment with neutralizing <t>anti-IL-12</t> antibody reverses both rIL-12 and silibinin-mediated protection against UVB-induced apoptosis in JB6 cells. (A–D) JB6 cells were irradiated with 50 mJ/cm 2 UVB and treated with rIL-12 in presence or absence
    Anti Mouse Il 12 Antibody, supplied by R&D Systems, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    IRF4 is important for IL-21-mediated Foxp3 suppression. Flow cytometry of naïve Irf4 −/− or WT CD4 + T cells differentiated as described in A and stained for intracellular Foxp3 ( A ) and of CD4 + WT cells nucleofected, cultured

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: IRF4 is essential for IL-21-mediated induction, amplification, and stabilization of the Th17 phenotype

    doi: 10.1073/pnas.0809077106

    Figure Lengend Snippet: IRF4 is important for IL-21-mediated Foxp3 suppression. Flow cytometry of naïve Irf4 −/− or WT CD4 + T cells differentiated as described in A and stained for intracellular Foxp3 ( A ) and of CD4 + WT cells nucleofected, cultured

    Article Snippet: Some cultures also received 2 ng/ml recombinant human TGFβ1 (R & D Systems), 50 ng/ml recombinant murine IL-6 (Peprotech), 50 ng/ml recombinant murine IL-21 (R & D Systems), or combinations of these stimuli.

    Techniques: Flow Cytometry, Cytometry, Staining, Cell Culture

    IRF4 regulates IL-21-mediated Th17 development in WT cells. CD4 + WT cells were nucleofected without any siRNA (NF), with scrambled (Scr) or one (si1) or three different (siMix) siRNA molecules targeting IRF4 and further cultured. ( A ) Immunoblot of IRF4

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: IRF4 is essential for IL-21-mediated induction, amplification, and stabilization of the Th17 phenotype

    doi: 10.1073/pnas.0809077106

    Figure Lengend Snippet: IRF4 regulates IL-21-mediated Th17 development in WT cells. CD4 + WT cells were nucleofected without any siRNA (NF), with scrambled (Scr) or one (si1) or three different (siMix) siRNA molecules targeting IRF4 and further cultured. ( A ) Immunoblot of IRF4

    Article Snippet: Some cultures also received 2 ng/ml recombinant human TGFβ1 (R & D Systems), 50 ng/ml recombinant murine IL-6 (Peprotech), 50 ng/ml recombinant murine IL-21 (R & D Systems), or combinations of these stimuli.

    Techniques: Cell Culture

    IRF4 deficiency abrogates IL-21-mediated Th17 differentiation. ( A ) Flow cytometry of naïve Irf4 −/− or WT CD4 + cells differentiated in the presence of indicated cytokines, re-stimulated, and stained intracellularly by anti-IL-17PE

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: IRF4 is essential for IL-21-mediated induction, amplification, and stabilization of the Th17 phenotype

    doi: 10.1073/pnas.0809077106

    Figure Lengend Snippet: IRF4 deficiency abrogates IL-21-mediated Th17 differentiation. ( A ) Flow cytometry of naïve Irf4 −/− or WT CD4 + cells differentiated in the presence of indicated cytokines, re-stimulated, and stained intracellularly by anti-IL-17PE

    Article Snippet: Some cultures also received 2 ng/ml recombinant human TGFβ1 (R & D Systems), 50 ng/ml recombinant murine IL-6 (Peprotech), 50 ng/ml recombinant murine IL-21 (R & D Systems), or combinations of these stimuli.

    Techniques: Flow Cytometry, Cytometry, Staining

    IRF4 is essential for IL-21 production. ( a and b ) Real-time RT-PCR analysis for Il21 mRNA: ( A ) in naïve Irf4 −/− (filled columns) or WT (open columns) CD4 + cells differentiated for 48 h under the indicated conditions. The expression

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: IRF4 is essential for IL-21-mediated induction, amplification, and stabilization of the Th17 phenotype

    doi: 10.1073/pnas.0809077106

    Figure Lengend Snippet: IRF4 is essential for IL-21 production. ( a and b ) Real-time RT-PCR analysis for Il21 mRNA: ( A ) in naïve Irf4 −/− (filled columns) or WT (open columns) CD4 + cells differentiated for 48 h under the indicated conditions. The expression

    Article Snippet: Some cultures also received 2 ng/ml recombinant human TGFβ1 (R & D Systems), 50 ng/ml recombinant murine IL-6 (Peprotech), 50 ng/ml recombinant murine IL-21 (R & D Systems), or combinations of these stimuli.

    Techniques: Quantitative RT-PCR, Expressing

    IL-21 induces normal STAT3 activation but increased Ifng expression in Irf4 −/− Th cells. ( A ) Immunoblot of pSTAT3, total STAT3, and β-actin in whole-cell lysates prepared from naïve Irf4 −/− or WT CD4 + cells

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: IRF4 is essential for IL-21-mediated induction, amplification, and stabilization of the Th17 phenotype

    doi: 10.1073/pnas.0809077106

    Figure Lengend Snippet: IL-21 induces normal STAT3 activation but increased Ifng expression in Irf4 −/− Th cells. ( A ) Immunoblot of pSTAT3, total STAT3, and β-actin in whole-cell lysates prepared from naïve Irf4 −/− or WT CD4 + cells

    Article Snippet: Some cultures also received 2 ng/ml recombinant human TGFβ1 (R & D Systems), 50 ng/ml recombinant murine IL-6 (Peprotech), 50 ng/ml recombinant murine IL-21 (R & D Systems), or combinations of these stimuli.

    Techniques: Activation Assay, Expressing

    KW3110 or IL-10 treatment suppresses caspase-1 activation in LPS/ATP-stimulated J774A.1 cells. (A) J774A.1 cells were treated with KW3110 (5 μg/mL) for 24 h, and treated with LPS (10 μg/mL) for 4 h and ATP (2 mM) for 1 h. (B) Cells were treated with IL-10 (1 μg/mL) and LPS (10 μg/mL) for 4 h and ATP (2 mM) for 1 h. Caspase-Glo 1 inflammasome assays measured caspase-1 activity in the supernatants from treated cells. Values indicate means ± SEM (triplicate data). Statistical differences were analyzed by ANOVA followed by Tukey’s test (**, P

    Journal: PLoS ONE

    Article Title: Lactic acid bacterium, Lactobacillus paracasei KW3110, suppresses inflammatory stress-induced caspase-1 activation by promoting interleukin-10 production in mouse and human immune cells

    doi: 10.1371/journal.pone.0237754

    Figure Lengend Snippet: KW3110 or IL-10 treatment suppresses caspase-1 activation in LPS/ATP-stimulated J774A.1 cells. (A) J774A.1 cells were treated with KW3110 (5 μg/mL) for 24 h, and treated with LPS (10 μg/mL) for 4 h and ATP (2 mM) for 1 h. (B) Cells were treated with IL-10 (1 μg/mL) and LPS (10 μg/mL) for 4 h and ATP (2 mM) for 1 h. Caspase-Glo 1 inflammasome assays measured caspase-1 activity in the supernatants from treated cells. Values indicate means ± SEM (triplicate data). Statistical differences were analyzed by ANOVA followed by Tukey’s test (**, P

    Article Snippet: The list of purchased items includes: ultrapure lipopolysaccharide from E . coli 0111:B4 strain (LPS) (Invivogen, San Diego, CA, USA), adenosine 5′-triphosphate (ATP) and cytochalasin D (Sigma, St. Louis, MO, USA), recombinant mouse and human IL-10 proteins (R & D Systems, MN, USA), anti-mouse caspase-1 (p20) (Adipogen Life Sciences (San Diego, CA, USA); catalog number: AG-20B-0042-C100; mouse-monoclonal antibody), anti-mouse IL-1β (catalog number: 12507S; rabbit-monoclonal antibody) and anti-mouse β-actin (catalog number: 3700; mouse-monoclonal antibody) (Cell Signaling Technology, Danvers, MA, USA) and secondary antibodies, anti-mouse IgG horseradish peroxidase-linked whole antibody from sheep (catalog number: NA931) and anti-rabbit IgG horseradish peroxidase-linked whole antibody from donkey (catalog number: NA934) (GE Healthcare, Chicago, IL, USA).

    Techniques: Activation Assay, Activity Assay

    KW3110 treatment suppresses active form of caspase-1 expression in LPS/ATP-stimulated J774A.1 cells through IL-10 signaling. (A) J774A.1 cells were treated with KW3110 (5 μg/mL) for 24 h, and treated with LPS (10 μg/mL) for 4 h and ATP (2 mM) for 1 h. The supernatant and cell lysate were analyzed by western blotting to detect matured IL-1β, pro-IL-1β, and β-actin. (B) The cells were transfected with IL-10 or IL-10 receptor α siRNAs and treated with KW3110 (10 μg/mL) for 24 h, and then treated with LPS (10 μg/mL) for 4 h and ATP (2 mM) for 1 h. The supernatant and cell lysate were analyzed by western blotting to detect caspase-1 (p20), pro-caspase-1, and β-actin. The black arrows are pointing to bands representing matured IL-1β or caspase-1 (p20). All blot images are spliced along with black lines. Sup; supernatant, Cell; cell lysate, siNT ; siRNA for non-targeting, siIL-10 ; siRNA for IL-10 , siIL-10R ; siRNA for IL-10 receptor α , mIL-1β; mature IL-1β.

    Journal: PLoS ONE

    Article Title: Lactic acid bacterium, Lactobacillus paracasei KW3110, suppresses inflammatory stress-induced caspase-1 activation by promoting interleukin-10 production in mouse and human immune cells

    doi: 10.1371/journal.pone.0237754

    Figure Lengend Snippet: KW3110 treatment suppresses active form of caspase-1 expression in LPS/ATP-stimulated J774A.1 cells through IL-10 signaling. (A) J774A.1 cells were treated with KW3110 (5 μg/mL) for 24 h, and treated with LPS (10 μg/mL) for 4 h and ATP (2 mM) for 1 h. The supernatant and cell lysate were analyzed by western blotting to detect matured IL-1β, pro-IL-1β, and β-actin. (B) The cells were transfected with IL-10 or IL-10 receptor α siRNAs and treated with KW3110 (10 μg/mL) for 24 h, and then treated with LPS (10 μg/mL) for 4 h and ATP (2 mM) for 1 h. The supernatant and cell lysate were analyzed by western blotting to detect caspase-1 (p20), pro-caspase-1, and β-actin. The black arrows are pointing to bands representing matured IL-1β or caspase-1 (p20). All blot images are spliced along with black lines. Sup; supernatant, Cell; cell lysate, siNT ; siRNA for non-targeting, siIL-10 ; siRNA for IL-10 , siIL-10R ; siRNA for IL-10 receptor α , mIL-1β; mature IL-1β.

    Article Snippet: The list of purchased items includes: ultrapure lipopolysaccharide from E . coli 0111:B4 strain (LPS) (Invivogen, San Diego, CA, USA), adenosine 5′-triphosphate (ATP) and cytochalasin D (Sigma, St. Louis, MO, USA), recombinant mouse and human IL-10 proteins (R & D Systems, MN, USA), anti-mouse caspase-1 (p20) (Adipogen Life Sciences (San Diego, CA, USA); catalog number: AG-20B-0042-C100; mouse-monoclonal antibody), anti-mouse IL-1β (catalog number: 12507S; rabbit-monoclonal antibody) and anti-mouse β-actin (catalog number: 3700; mouse-monoclonal antibody) (Cell Signaling Technology, Danvers, MA, USA) and secondary antibodies, anti-mouse IgG horseradish peroxidase-linked whole antibody from sheep (catalog number: NA931) and anti-rabbit IgG horseradish peroxidase-linked whole antibody from donkey (catalog number: NA934) (GE Healthcare, Chicago, IL, USA).

    Techniques: Expressing, Western Blot, Transfection

    KW3110 treatment induces IL-10 production and suppresses IL-1β production and caspase-1 activity in human monocytes. (A-C) Human monocytes were treated with KW3110 (100 μg/mL) for 24 h. IL-10 levels in the supernatant were measured by ELISA (A). The cells were then treated with IL-10 (1 μg/mL) and LPS (10 μg/mL) for 4 h and ATP (2 mM) for 1 h. IL-1β levels in the supernatant were measured by ELISA (B) and the supernatant was analyzed by Caspase-Glo 1 inflammasome assay (C). Values indicate means ± SEM (triplicate data). Statistical differences were analyzed by unpaired t -tests for (A) and ANOVA followed by Tukey’s test for (B, C) (**, P

    Journal: PLoS ONE

    Article Title: Lactic acid bacterium, Lactobacillus paracasei KW3110, suppresses inflammatory stress-induced caspase-1 activation by promoting interleukin-10 production in mouse and human immune cells

    doi: 10.1371/journal.pone.0237754

    Figure Lengend Snippet: KW3110 treatment induces IL-10 production and suppresses IL-1β production and caspase-1 activity in human monocytes. (A-C) Human monocytes were treated with KW3110 (100 μg/mL) for 24 h. IL-10 levels in the supernatant were measured by ELISA (A). The cells were then treated with IL-10 (1 μg/mL) and LPS (10 μg/mL) for 4 h and ATP (2 mM) for 1 h. IL-1β levels in the supernatant were measured by ELISA (B) and the supernatant was analyzed by Caspase-Glo 1 inflammasome assay (C). Values indicate means ± SEM (triplicate data). Statistical differences were analyzed by unpaired t -tests for (A) and ANOVA followed by Tukey’s test for (B, C) (**, P

    Article Snippet: The list of purchased items includes: ultrapure lipopolysaccharide from E . coli 0111:B4 strain (LPS) (Invivogen, San Diego, CA, USA), adenosine 5′-triphosphate (ATP) and cytochalasin D (Sigma, St. Louis, MO, USA), recombinant mouse and human IL-10 proteins (R & D Systems, MN, USA), anti-mouse caspase-1 (p20) (Adipogen Life Sciences (San Diego, CA, USA); catalog number: AG-20B-0042-C100; mouse-monoclonal antibody), anti-mouse IL-1β (catalog number: 12507S; rabbit-monoclonal antibody) and anti-mouse β-actin (catalog number: 3700; mouse-monoclonal antibody) (Cell Signaling Technology, Danvers, MA, USA) and secondary antibodies, anti-mouse IgG horseradish peroxidase-linked whole antibody from sheep (catalog number: NA931) and anti-rabbit IgG horseradish peroxidase-linked whole antibody from donkey (catalog number: NA934) (GE Healthcare, Chicago, IL, USA).

    Techniques: Activity Assay, Enzyme-linked Immunosorbent Assay

    Knockdown of IL-10 signaling components reduces KW3110-induced IL-1β suppression in LPS/ATP-stimulated J774A.1 cells. (A) J774A.1 cells were treated with KW3110 (1.25, 2.5, or 5 μg/mL) for 24 h. IL-10 levels in the supernatant were measured by ELISA. (B) The cells were treated with IL-10 (1 μg/mL) and LPS (10 μg/mL) for 4 h and ATP (2 mM) for 1 h. IL-1β levels in the supernatant were measured by ELISA. (C) Total RNA from cells transfected with IL-10 siRNA and treated with KW3110 (10 μg/mL) for 6 h was extracted and reverse-transcribed. Quantitative RT-PCR was performed to amplify mouse GAPDH and IL-10 . The IL-10 values were normalized to the expression of GAPDH . (D, E) Cells were transfected with IL-10 or IL-10 receptor α siRNAs and treated with KW3110 (10 μg/mL) for 24 h. IL-10 levels in the supernatant were measured by ELISA for (D). The cells were then treated with LPS (10 μg/mL) for 4 h and ATP (2 mM) for 1 h. IL-1β levels in the supernatant were measured by ELISA for (E). siNT ; siRNA for non-targeting, siIL-10 ; siRNA for IL-10 , siIL-10R ; siRNA for IL-10 receptor α. Values indicate means ± SEM (triplicate data). Statistical differences were analyzed by ANOVA followed by Dunnett’s test for (A) and Tukey’s test for (B-E) (**, P

    Journal: PLoS ONE

    Article Title: Lactic acid bacterium, Lactobacillus paracasei KW3110, suppresses inflammatory stress-induced caspase-1 activation by promoting interleukin-10 production in mouse and human immune cells

    doi: 10.1371/journal.pone.0237754

    Figure Lengend Snippet: Knockdown of IL-10 signaling components reduces KW3110-induced IL-1β suppression in LPS/ATP-stimulated J774A.1 cells. (A) J774A.1 cells were treated with KW3110 (1.25, 2.5, or 5 μg/mL) for 24 h. IL-10 levels in the supernatant were measured by ELISA. (B) The cells were treated with IL-10 (1 μg/mL) and LPS (10 μg/mL) for 4 h and ATP (2 mM) for 1 h. IL-1β levels in the supernatant were measured by ELISA. (C) Total RNA from cells transfected with IL-10 siRNA and treated with KW3110 (10 μg/mL) for 6 h was extracted and reverse-transcribed. Quantitative RT-PCR was performed to amplify mouse GAPDH and IL-10 . The IL-10 values were normalized to the expression of GAPDH . (D, E) Cells were transfected with IL-10 or IL-10 receptor α siRNAs and treated with KW3110 (10 μg/mL) for 24 h. IL-10 levels in the supernatant were measured by ELISA for (D). The cells were then treated with LPS (10 μg/mL) for 4 h and ATP (2 mM) for 1 h. IL-1β levels in the supernatant were measured by ELISA for (E). siNT ; siRNA for non-targeting, siIL-10 ; siRNA for IL-10 , siIL-10R ; siRNA for IL-10 receptor α. Values indicate means ± SEM (triplicate data). Statistical differences were analyzed by ANOVA followed by Dunnett’s test for (A) and Tukey’s test for (B-E) (**, P

    Article Snippet: The list of purchased items includes: ultrapure lipopolysaccharide from E . coli 0111:B4 strain (LPS) (Invivogen, San Diego, CA, USA), adenosine 5′-triphosphate (ATP) and cytochalasin D (Sigma, St. Louis, MO, USA), recombinant mouse and human IL-10 proteins (R & D Systems, MN, USA), anti-mouse caspase-1 (p20) (Adipogen Life Sciences (San Diego, CA, USA); catalog number: AG-20B-0042-C100; mouse-monoclonal antibody), anti-mouse IL-1β (catalog number: 12507S; rabbit-monoclonal antibody) and anti-mouse β-actin (catalog number: 3700; mouse-monoclonal antibody) (Cell Signaling Technology, Danvers, MA, USA) and secondary antibodies, anti-mouse IgG horseradish peroxidase-linked whole antibody from sheep (catalog number: NA931) and anti-rabbit IgG horseradish peroxidase-linked whole antibody from donkey (catalog number: NA934) (GE Healthcare, Chicago, IL, USA).

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

    A signaling diagram of anti-inflammatory mechanism of KW3110. While LPS/ATP stimulation induces caspase-1 activation and IL-1β maturation in non-treated J774A.1 cells, KW3110 treatment suppresses these inflammatory responses in the cells through promoting IL-10 production.

    Journal: PLoS ONE

    Article Title: Lactic acid bacterium, Lactobacillus paracasei KW3110, suppresses inflammatory stress-induced caspase-1 activation by promoting interleukin-10 production in mouse and human immune cells

    doi: 10.1371/journal.pone.0237754

    Figure Lengend Snippet: A signaling diagram of anti-inflammatory mechanism of KW3110. While LPS/ATP stimulation induces caspase-1 activation and IL-1β maturation in non-treated J774A.1 cells, KW3110 treatment suppresses these inflammatory responses in the cells through promoting IL-10 production.

    Article Snippet: The list of purchased items includes: ultrapure lipopolysaccharide from E . coli 0111:B4 strain (LPS) (Invivogen, San Diego, CA, USA), adenosine 5′-triphosphate (ATP) and cytochalasin D (Sigma, St. Louis, MO, USA), recombinant mouse and human IL-10 proteins (R & D Systems, MN, USA), anti-mouse caspase-1 (p20) (Adipogen Life Sciences (San Diego, CA, USA); catalog number: AG-20B-0042-C100; mouse-monoclonal antibody), anti-mouse IL-1β (catalog number: 12507S; rabbit-monoclonal antibody) and anti-mouse β-actin (catalog number: 3700; mouse-monoclonal antibody) (Cell Signaling Technology, Danvers, MA, USA) and secondary antibodies, anti-mouse IgG horseradish peroxidase-linked whole antibody from sheep (catalog number: NA931) and anti-rabbit IgG horseradish peroxidase-linked whole antibody from donkey (catalog number: NA934) (GE Healthcare, Chicago, IL, USA).

    Techniques: Activation Assay

    Effects of phagocytosis blocker on KW3110-induced IL-10 production and IL-1β suppression in J774A.1 cells. (A) Total RNA from J774A.1 cells treated with cytochalasin D (1 μg/mL) for 30 min and KW3110 (5 μg/mL) for 6 h was extracted and reverse-transcribed. Quantitative RT-PCR was performed to amplify mouse GAPDH and IL-10 . IL-10 values were normalized to the expression of GAPDH . (B, C) The cells were treated with cytochalasin D (1 μg/mL) for 30 min and KW3110 (5 μg/mL) for 24 h. IL-10 levels in the supernatant were measured by ELISA for (B). The cells were then treated with LPS (10 μg/mL) for 4 h and ATP (2 mM) for 1 h. IL-1β levels in the supernatant were measured by ELISA for (C). CytD; cytochalasin D. Values indicate means ± SEM (triplicate data). Statistical differences were analyzed by ANOVA followed by Tukey’s test (**, P

    Journal: PLoS ONE

    Article Title: Lactic acid bacterium, Lactobacillus paracasei KW3110, suppresses inflammatory stress-induced caspase-1 activation by promoting interleukin-10 production in mouse and human immune cells

    doi: 10.1371/journal.pone.0237754

    Figure Lengend Snippet: Effects of phagocytosis blocker on KW3110-induced IL-10 production and IL-1β suppression in J774A.1 cells. (A) Total RNA from J774A.1 cells treated with cytochalasin D (1 μg/mL) for 30 min and KW3110 (5 μg/mL) for 6 h was extracted and reverse-transcribed. Quantitative RT-PCR was performed to amplify mouse GAPDH and IL-10 . IL-10 values were normalized to the expression of GAPDH . (B, C) The cells were treated with cytochalasin D (1 μg/mL) for 30 min and KW3110 (5 μg/mL) for 24 h. IL-10 levels in the supernatant were measured by ELISA for (B). The cells were then treated with LPS (10 μg/mL) for 4 h and ATP (2 mM) for 1 h. IL-1β levels in the supernatant were measured by ELISA for (C). CytD; cytochalasin D. Values indicate means ± SEM (triplicate data). Statistical differences were analyzed by ANOVA followed by Tukey’s test (**, P

    Article Snippet: The list of purchased items includes: ultrapure lipopolysaccharide from E . coli 0111:B4 strain (LPS) (Invivogen, San Diego, CA, USA), adenosine 5′-triphosphate (ATP) and cytochalasin D (Sigma, St. Louis, MO, USA), recombinant mouse and human IL-10 proteins (R & D Systems, MN, USA), anti-mouse caspase-1 (p20) (Adipogen Life Sciences (San Diego, CA, USA); catalog number: AG-20B-0042-C100; mouse-monoclonal antibody), anti-mouse IL-1β (catalog number: 12507S; rabbit-monoclonal antibody) and anti-mouse β-actin (catalog number: 3700; mouse-monoclonal antibody) (Cell Signaling Technology, Danvers, MA, USA) and secondary antibodies, anti-mouse IgG horseradish peroxidase-linked whole antibody from sheep (catalog number: NA931) and anti-rabbit IgG horseradish peroxidase-linked whole antibody from donkey (catalog number: NA934) (GE Healthcare, Chicago, IL, USA).

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

    pmel-1 undergo limited homeostatic proliferation in irradiated wild-type hosts (A) Pmel-1.Rag T cells were isolated from lymph nodes and spleens and further purified by MACS negative selection for CD44 low CD8+. After CFSE labelling, pmel-1.Rag were injected into irradiated B6 hosts. For indicated animals, recombinant IL-12 was administered on days 1, 2 and 3 after adoptive transfer. CFSE dilution of donor T cells (upper panels) and CD44 expression levels on donor and host CD8 T cells (lower panels) was assessed in the spleen at 10 days and 6 weeks after adoptive transfer. (B) Naïve pmel-1 were adoptively transferred into irradiated B6 mice. One day after transfer, mice were given i.v injections of either PBS, IL-15 or, IL-15/Rα. Mice were harvested 3 weeks later and CFSE dilution of donor T cells (upper panels) and CD44 expression levels on donor and host CD8 T cells (lower panels) was assessed. Results are representative of 3 independent experiments. In all cases, donor cells were identified by Thy-1 congenic markers.

    Journal: Journal of immunology (Baltimore, Md. : 1950)

    Article Title: Self-specific CD8+ T cells maintain a semi-na?ve state following lymphopenia-induced proliferation

    doi: 10.4049/jimmunol.1000109

    Figure Lengend Snippet: pmel-1 undergo limited homeostatic proliferation in irradiated wild-type hosts (A) Pmel-1.Rag T cells were isolated from lymph nodes and spleens and further purified by MACS negative selection for CD44 low CD8+. After CFSE labelling, pmel-1.Rag were injected into irradiated B6 hosts. For indicated animals, recombinant IL-12 was administered on days 1, 2 and 3 after adoptive transfer. CFSE dilution of donor T cells (upper panels) and CD44 expression levels on donor and host CD8 T cells (lower panels) was assessed in the spleen at 10 days and 6 weeks after adoptive transfer. (B) Naïve pmel-1 were adoptively transferred into irradiated B6 mice. One day after transfer, mice were given i.v injections of either PBS, IL-15 or, IL-15/Rα. Mice were harvested 3 weeks later and CFSE dilution of donor T cells (upper panels) and CD44 expression levels on donor and host CD8 T cells (lower panels) was assessed. Results are representative of 3 independent experiments. In all cases, donor cells were identified by Thy-1 congenic markers.

    Article Snippet: In other experiments, mice were treated with recombinant IL-12 (1μg in a total volume of 200μl PBS, administered via tail vein injection) (R & D Systems) on days 1, 2 and 3 after adoptive transfer of pmel-1 into sub-lethally irradiated hosts.

    Techniques: Irradiation, Isolation, Purification, Magnetic Cell Separation, Selection, Injection, Recombinant, Adoptive Transfer Assay, Expressing, Mouse Assay

    Co-treatment with neutralizing anti-IL-12 antibody reverses both rIL-12 and silibinin-mediated protection against UVB-induced apoptosis in JB6 cells. (A–D) JB6 cells were irradiated with 50 mJ/cm 2 UVB and treated with rIL-12 in presence or absence

    Journal: Molecular carcinogenesis

    Article Title: Silibinin Inhibits Ultraviolet B Radiation-Induced DNA-Damage and Apoptosis by Enhancing Interleukin-12 Expression in JB6 Cells and SKH-1 Hairless Mouse Skin

    doi: 10.1002/mc.22000

    Figure Lengend Snippet: Co-treatment with neutralizing anti-IL-12 antibody reverses both rIL-12 and silibinin-mediated protection against UVB-induced apoptosis in JB6 cells. (A–D) JB6 cells were irradiated with 50 mJ/cm 2 UVB and treated with rIL-12 in presence or absence

    Article Snippet: The recombinant mouse IL-12 and neutralizing anti-mouse IL-12 antibody were from R & D Systems (Minneapolis, MN).

    Techniques: Irradiation

    Silibinin enhances the production of IL-12 in UVB-irradiated mouse epidermal keratinocytes in vitro and in vivo . (A) JB6 cells were cultured overnight on 96 well plates, sham irradiated or exposed to 50 mJ/cm 2 UVB, and incubated with silibinin and/or

    Journal: Molecular carcinogenesis

    Article Title: Silibinin Inhibits Ultraviolet B Radiation-Induced DNA-Damage and Apoptosis by Enhancing Interleukin-12 Expression in JB6 Cells and SKH-1 Hairless Mouse Skin

    doi: 10.1002/mc.22000

    Figure Lengend Snippet: Silibinin enhances the production of IL-12 in UVB-irradiated mouse epidermal keratinocytes in vitro and in vivo . (A) JB6 cells were cultured overnight on 96 well plates, sham irradiated or exposed to 50 mJ/cm 2 UVB, and incubated with silibinin and/or

    Article Snippet: The recombinant mouse IL-12 and neutralizing anti-mouse IL-12 antibody were from R & D Systems (Minneapolis, MN).

    Techniques: Irradiation, In Vitro, In Vivo, Cell Culture, Incubation