recombinant proteins Search Results


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  • 99
    Thermo Fisher recombinant proteins
    Recombinant Proteins, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 1595 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore recombinant proteins
    Recombinant Proteins, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 1870 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    R&D Systems il 17a
    Rhodomyrtone inhibits <t>TNF-α+IL-17A-induced</t> inflammatory responses in skin organ cultures. . Bars indicate mean ± SD, n = 5. Statistical significance determined by one-way ANOVA and Dunn’s multiple comparison test, indicated as* p
    Il 17a, supplied by R&D Systems, used in various techniques. Bioz Stars score: 99/100, based on 3062 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher basic fibroblast growth factor
    Rhodomyrtone inhibits <t>TNF-α+IL-17A-induced</t> inflammatory responses in skin organ cultures. . Bars indicate mean ± SD, n = 5. Statistical significance determined by one-way ANOVA and Dunn’s multiple comparison test, indicated as* p
    Basic Fibroblast Growth Factor, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 3713 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore recombinant protein
    Rhodomyrtone inhibits <t>TNF-α+IL-17A-induced</t> inflammatory responses in skin organ cultures. . Bars indicate mean ± SD, n = 5. Statistical significance determined by one-way ANOVA and Dunn’s multiple comparison test, indicated as* p
    Recombinant Protein, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 9202 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    R&D Systems activin a
    Follistatin blocks the growth arrest induced by <t>activin</t> A in LE6 cells. (A) LE6 cells were treated with activin A (200 ng/ml) in the presence or absence of the indicated doses of follistatin (25, 50, 100, 200, 400, 600 ng/ml). Cell proliferation was detected by CCK-8 assay. (B) LE6 cells were grown in LE media in the presence or absence of activin A (200 ng/ml), follistatin (400 ng/ml) or activin A (200 ng/ml) plus follistatin (400 ng/ml). DNA synthesis was detected by BrdU incorporation assay using FACS. (C) LE6 cells were treated with or without activin A (200 ng/ml), follistatin (400 ng/ml) or activin A (200 ng/ml) plus follistatin (400 ng/ml) for 30 min. Phosphorylated SMAD2/3 was detected by western-blot. (D) LE6 cells were treated with either media alone, activin A (200 ng/ml), follistatin (400 ng/ml) or activin(200 ng/ml) plus follistatin (400 ng/ml). Then phosphorylated Rb, cyclinD1, cyclinE, p21 WAF1/Cip1 and p15 INK4B were analyzed by western-blot.
    Activin A, supplied by R&D Systems, used in various techniques. Bioz Stars score: 99/100, based on 2597 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    R&D Systems egf
    Follistatin blocks the growth arrest induced by <t>activin</t> A in LE6 cells. (A) LE6 cells were treated with activin A (200 ng/ml) in the presence or absence of the indicated doses of follistatin (25, 50, 100, 200, 400, 600 ng/ml). Cell proliferation was detected by CCK-8 assay. (B) LE6 cells were grown in LE media in the presence or absence of activin A (200 ng/ml), follistatin (400 ng/ml) or activin A (200 ng/ml) plus follistatin (400 ng/ml). DNA synthesis was detected by BrdU incorporation assay using FACS. (C) LE6 cells were treated with or without activin A (200 ng/ml), follistatin (400 ng/ml) or activin A (200 ng/ml) plus follistatin (400 ng/ml) for 30 min. Phosphorylated SMAD2/3 was detected by western-blot. (D) LE6 cells were treated with either media alone, activin A (200 ng/ml), follistatin (400 ng/ml) or activin(200 ng/ml) plus follistatin (400 ng/ml). Then phosphorylated Rb, cyclinD1, cyclinE, p21 WAF1/Cip1 and p15 INK4B were analyzed by western-blot.
    Egf, supplied by R&D Systems, used in various techniques. Bioz Stars score: 99/100, based on 3130 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    R&D Systems netrin 1
    <t>Netrin-1</t> promotes inflammation resolution in diabetic corneal wound healing. Corneal epithelium was scraped in control and diabetic mice with or without subconjunctival injection of netrin-1 (50 ng per eye). Mouse cornea was collected and immunofluorescent stained with neutrophil marker Ly6G at 48 h and M2 macrophage markers CD68 (green fluorescence) and CD206 (red fluorescence) at 72 h after epithelial scrape ( A , 3 mice per group). Flow cytometry was performed to quantify the number and percentage of neutrophils (CD45 + CD11b + Ly6G+) ( B , 12 mice per group) at 48-hour post-wounded, M1 macrophages (CD45 + F4/80 + CD86+) and M2 macrophages (CD45 + F4/80 + CD206+) at 72-hour post-wounded. ( C , 12 mice per group at each detection) mRNA expression levels of iNOS and IL-12 (48 h after epithelial scrape), arginase-1 and IL-10 (72 h after epithelial scrape) were analyzed by RT-qPCR from the control and diabetic mouse corneas ( D , 6 mice per group); *p
    Netrin 1, supplied by R&D Systems, used in various techniques. Bioz Stars score: 95/100, based on 927 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    R&D Systems hmgb1
    High-mobility group protein B1 <t>(HMGB1)</t> and heat shock protein 70 (HSP70) stimulate Toll-like receptor 4 (TLR4; +/+) but not TLR4 (−/−) bone marrow (BM)-derived macrophages: BM-derived macrophages stimulated in vitro with media, HSP70 (3 μg/ml), and HMGB1 (3 μg/ml) Interleukin 6 (IL6) was measured by quantitative reverse transcriptase-PCR. Each sample normalized to glyceraldehyde 3-phosphate dehydrogenase (GAPDH). The calibrator gene is the media-treated BMDM. n = 4. Means and standard deviation are shown. * P
    Hmgb1, supplied by R&D Systems, used in various techniques. Bioz Stars score: 99/100, based on 1177 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher tnf α
    Quantification of inflammatory indicators in mice infected with recombinant Mycobacterium smegmatis . The spleen was excised from the infected mice ( n = 3 per group), and the spleen lymphocytes were separated and incubated overnight with PBS, pMVp262/MS and Rv0888NS/MS, D438A/MS, and H481N/MS. The cytokines were determined using the cultured supernatant. (A) Analysis of the IL-1β. (B) Analysis of IL-6. (C) Analysis of <t>TNF-α.</t> The error bars show the SEM of three independent experiments with three wells per group. (A–C) One-way ANOVA followed by Bonferroni’s multiple comparison post hoc test, * P
    Tnf α, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 21879 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    R&D Systems timp 1
    Quantification of inflammatory indicators in mice infected with recombinant Mycobacterium smegmatis . The spleen was excised from the infected mice ( n = 3 per group), and the spleen lymphocytes were separated and incubated overnight with PBS, pMVp262/MS and Rv0888NS/MS, D438A/MS, and H481N/MS. The cytokines were determined using the cultured supernatant. (A) Analysis of the IL-1β. (B) Analysis of IL-6. (C) Analysis of <t>TNF-α.</t> The error bars show the SEM of three independent experiments with three wells per group. (A–C) One-way ANOVA followed by Bonferroni’s multiple comparison post hoc test, * P
    Timp 1, supplied by R&D Systems, used in various techniques. Bioz Stars score: 93/100, based on 1606 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher il 10
    Quantification of inflammatory indicators in mice infected with recombinant Mycobacterium smegmatis . The spleen was excised from the infected mice ( n = 3 per group), and the spleen lymphocytes were separated and incubated overnight with PBS, pMVp262/MS and Rv0888NS/MS, D438A/MS, and H481N/MS. The cytokines were determined using the cultured supernatant. (A) Analysis of the IL-1β. (B) Analysis of IL-6. (C) Analysis of <t>TNF-α.</t> The error bars show the SEM of three independent experiments with three wells per group. (A–C) One-way ANOVA followed by Bonferroni’s multiple comparison post hoc test, * P
    Il 10, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 2937 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher interleukin 6
    VLP effectively stimulates BMDCs in vitro to secrete cytokines. BMDCs were stimulated with M2e5x, M1 VLP, or protein (0.3, 1, 5 μg/mL) for 24 h. ( a – c ) The levels of cytokines ( a ) TNF-α, ( b ) <t>IL-6,</t> ( c ) IFN-γ were determined in the culture supernatants by ELISA assay. Un: Medium only, P: proteins, V: VLP. The statistical significance was confirmed by one-way ANOVA and Dunnett’s multiple comparison test. Error bars indicate the means ± SEM of concentrations from individual animals. *; p
    Interleukin 6, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 1698 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher cd34
    VLP effectively stimulates BMDCs in vitro to secrete cytokines. BMDCs were stimulated with M2e5x, M1 VLP, or protein (0.3, 1, 5 μg/mL) for 24 h. ( a – c ) The levels of cytokines ( a ) TNF-α, ( b ) <t>IL-6,</t> ( c ) IFN-γ were determined in the culture supernatants by ELISA assay. Un: Medium only, P: proteins, V: VLP. The statistical significance was confirmed by one-way ANOVA and Dunnett’s multiple comparison test. Error bars indicate the means ± SEM of concentrations from individual animals. *; p
    Cd34, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 3850 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    R&D Systems wnt3a
    VLP effectively stimulates BMDCs in vitro to secrete cytokines. BMDCs were stimulated with M2e5x, M1 VLP, or protein (0.3, 1, 5 μg/mL) for 24 h. ( a – c ) The levels of cytokines ( a ) TNF-α, ( b ) <t>IL-6,</t> ( c ) IFN-γ were determined in the culture supernatants by ELISA assay. Un: Medium only, P: proteins, V: VLP. The statistical significance was confirmed by one-way ANOVA and Dunnett’s multiple comparison test. Error bars indicate the means ± SEM of concentrations from individual animals. *; p
    Wnt3a, supplied by R&D Systems, used in various techniques. Bioz Stars score: 99/100, based on 2816 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    R&D Systems gdf 15
    Hepcidin and <t>GDF-15</t> are increased and their expression is correlated in breast tumors (A and B) Box plot with Tukey whisker of (A) GDF15 and (B) HAMP mRNA expression (log2 transformed) in normal adjacent tissue (n=61) compared to primary tumor tissue (n=526) in the TCGA breast cancer dataset. (C) GDF15 transcripts in TCGA samples from breast cancer patients divided by HAMP expression (below and above the mean) shown as box and whisker plot. (D) HAMP transcripts in TCGA samples from breast cancer patients divided by GDF15 expression (below and above the mean) shown as box and whisker. (E) Representative images of immunohistochemical staining of tumor tissue from patients with invasive ductal carcinoma (IDC). Proteins stained are Hepcidin, GDF-15, Pan-Cytokeratin and IgG control. (F) Scatter plot displays quantification of staining of epithelial cells from tissues from 56 BRCA patients. A regression analysis was performed to examine correlation of staining intensities (R 2 =0.4434 p
    Gdf 15, supplied by R&D Systems, used in various techniques. Bioz Stars score: 99/100, based on 643 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher human recombinant epidermal growth factor
    Hepcidin and <t>GDF-15</t> are increased and their expression is correlated in breast tumors (A and B) Box plot with Tukey whisker of (A) GDF15 and (B) HAMP mRNA expression (log2 transformed) in normal adjacent tissue (n=61) compared to primary tumor tissue (n=526) in the TCGA breast cancer dataset. (C) GDF15 transcripts in TCGA samples from breast cancer patients divided by HAMP expression (below and above the mean) shown as box and whisker plot. (D) HAMP transcripts in TCGA samples from breast cancer patients divided by GDF15 expression (below and above the mean) shown as box and whisker. (E) Representative images of immunohistochemical staining of tumor tissue from patients with invasive ductal carcinoma (IDC). Proteins stained are Hepcidin, GDF-15, Pan-Cytokeratin and IgG control. (F) Scatter plot displays quantification of staining of epithelial cells from tissues from 56 BRCA patients. A regression analysis was performed to examine correlation of staining intensities (R 2 =0.4434 p
    Human Recombinant Epidermal Growth Factor, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 1496 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    R&D Systems recombinant human tnf α
    The level of <t>TNF-α</t> protein increased in TOLF. A) Fold change of TNF-α protein in the ossified ligamentum flavum of TOLF. Proteins were extracted from the ossified ligamentum flavum (OS-Lig) and normal ligamentum flavum (Lig). The samples were then labeled with iTRAQ following the manufacture’s protocol. *: A star indicates statistical significance compared to control group with p
    Recombinant Human Tnf α, supplied by R&D Systems, used in various techniques. Bioz Stars score: 99/100, based on 1509 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher granulocyte macrophage colony stimulating factor
    The level of <t>TNF-α</t> protein increased in TOLF. A) Fold change of TNF-α protein in the ossified ligamentum flavum of TOLF. Proteins were extracted from the ossified ligamentum flavum (OS-Lig) and normal ligamentum flavum (Lig). The samples were then labeled with iTRAQ following the manufacture’s protocol. *: A star indicates statistical significance compared to control group with p
    Granulocyte Macrophage Colony Stimulating Factor, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 98/100, based on 390 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    R&D Systems pdgf bb
    Incubation of <t>TGF-β1</t> NAB in VSMCs increased the expression of lamin A in cocultured ECs, but <t>PDGF-BB</t> had no significant effect ( A ). NABs of both PDGF-BB and TGF-β1 incubation in VSMCs decreased expressions of LOX and phospho-ERK1/2 ( A
    Pdgf Bb, supplied by R&D Systems, used in various techniques. Bioz Stars score: 99/100, based on 2186 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    R&D Systems ang 1
    Enzyme linked immunosorbent assay (ELISA). a, b Representative graph showing expression of vascular endothelial growth factor A (VEGF) ( a ) and angiopoietin 1 <t>(Ang-1)</t> ( b ). Data presented as mean ± standard deviation. * p ≤ 0.05, *** p ≤ 0.001, n = 4. d days
    Ang 1, supplied by R&D Systems, used in various techniques. Bioz Stars score: 99/100, based on 637 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    R&D Systems human il 6
    3.2. Association of extracellular histones, HMGB1, sTM, and <t>IL-6</t> with ALI
    Human Il 6, supplied by R&D Systems, used in various techniques. Bioz Stars score: 99/100, based on 1161 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Rhodomyrtone inhibits TNF-α+IL-17A-induced inflammatory responses in skin organ cultures. . Bars indicate mean ± SD, n = 5. Statistical significance determined by one-way ANOVA and Dunn’s multiple comparison test, indicated as* p

    Journal: PLoS ONE

    Article Title: The small molecule rhodomyrtone suppresses TNF-α and IL-17A-induced keratinocyte inflammatory responses: A potential new therapeutic for psoriasis

    doi: 10.1371/journal.pone.0205340

    Figure Lengend Snippet: Rhodomyrtone inhibits TNF-α+IL-17A-induced inflammatory responses in skin organ cultures. . Bars indicate mean ± SD, n = 5. Statistical significance determined by one-way ANOVA and Dunn’s multiple comparison test, indicated as* p

    Article Snippet: NHK were pretreated with rhodomyrtone (50–600 ng/ml) for 1 h before stimulation with recombinant human 20ng/ml IL-17A and 10 ng/ml TNF-α (both R & D Systems) for 24 h. Cells were harvested for RNA isolation in complete RLT buffer (Qiagen) or protein isolation in complete RIPA buffer.

    Techniques:

    Rhodomyrtone preferentially down-regulates transcripts involved in epidermal responses, myeloid leukocyte chemotaxis and inflammatory responses. Global transcriptome sequencing (RNA-seq) was performed on NHK treated with IL-17A+TNF for 24 h with or without 1 h pre-treatment with 400 ng/ml rhodomyrtone. Rhodomyrtone inhibited the expression of 579 of 1066 IL-17A/TNF-induced genes by NHK (2-fold change, TREAT p ] showed a decrease in the expression of IL-17/TNF induced transcripts area under curve = 0.46 (f) vs. 0.30 (h) and a decrease in genes overlapping with a plaque psoriasis gene signature, area under curve = 0.21 (g) vs. 0.07 (i).

    Journal: PLoS ONE

    Article Title: The small molecule rhodomyrtone suppresses TNF-α and IL-17A-induced keratinocyte inflammatory responses: A potential new therapeutic for psoriasis

    doi: 10.1371/journal.pone.0205340

    Figure Lengend Snippet: Rhodomyrtone preferentially down-regulates transcripts involved in epidermal responses, myeloid leukocyte chemotaxis and inflammatory responses. Global transcriptome sequencing (RNA-seq) was performed on NHK treated with IL-17A+TNF for 24 h with or without 1 h pre-treatment with 400 ng/ml rhodomyrtone. Rhodomyrtone inhibited the expression of 579 of 1066 IL-17A/TNF-induced genes by NHK (2-fold change, TREAT p ] showed a decrease in the expression of IL-17/TNF induced transcripts area under curve = 0.46 (f) vs. 0.30 (h) and a decrease in genes overlapping with a plaque psoriasis gene signature, area under curve = 0.21 (g) vs. 0.07 (i).

    Article Snippet: NHK were pretreated with rhodomyrtone (50–600 ng/ml) for 1 h before stimulation with recombinant human 20ng/ml IL-17A and 10 ng/ml TNF-α (both R & D Systems) for 24 h. Cells were harvested for RNA isolation in complete RLT buffer (Qiagen) or protein isolation in complete RIPA buffer.

    Techniques: Chemotaxis Assay, Sequencing, RNA Sequencing Assay, Expressing

    Rhodomyrtone decreases expression of inflammatory transcripts by TNF-α+IL-17A stimulated in skin organ cultures. Skin biopsies were pre-treated with 400 ng/ml rhodomyrtone 6 h before stimulation with 10 ng/ml TNF-α and 20 ng/ml IL-17A. Rhodomyrtone treatment decreases production of DEFB4 , IL1B , IL17C , IL36G , LCN2 , PI3 , S100A7 , and S100A8 transcripts as measured by qRT-PCR at 72 h. Statistical significance determined by one-way ANOVA and Dunn’s multiple comparison test, indicated as * p

    Journal: PLoS ONE

    Article Title: The small molecule rhodomyrtone suppresses TNF-α and IL-17A-induced keratinocyte inflammatory responses: A potential new therapeutic for psoriasis

    doi: 10.1371/journal.pone.0205340

    Figure Lengend Snippet: Rhodomyrtone decreases expression of inflammatory transcripts by TNF-α+IL-17A stimulated in skin organ cultures. Skin biopsies were pre-treated with 400 ng/ml rhodomyrtone 6 h before stimulation with 10 ng/ml TNF-α and 20 ng/ml IL-17A. Rhodomyrtone treatment decreases production of DEFB4 , IL1B , IL17C , IL36G , LCN2 , PI3 , S100A7 , and S100A8 transcripts as measured by qRT-PCR at 72 h. Statistical significance determined by one-way ANOVA and Dunn’s multiple comparison test, indicated as * p

    Article Snippet: NHK were pretreated with rhodomyrtone (50–600 ng/ml) for 1 h before stimulation with recombinant human 20ng/ml IL-17A and 10 ng/ml TNF-α (both R & D Systems) for 24 h. Cells were harvested for RNA isolation in complete RLT buffer (Qiagen) or protein isolation in complete RIPA buffer.

    Techniques: Expressing, Quantitative RT-PCR

    Rhodomyrtone dose dependently decreases production of CXCL1 , IL-36G , IL-36RN , MMP-13 , S100A7 , S100A8 , and S100A12 transcripts by IL-17A/TNF-α-stimulated primary human keratinocytes. Keratinocytes were pretreated with rhodomyrtone (50, 100, 200, 400, and 600 ng/ml) before stimulation with 20ng/ml IL-17A+10ng/ml TNF-α. The mRNA expression levels were measured at 24 h by qRT-PCR. Values are mean±SD of results from three independent experiments in triplicate. Statistical significance determined using one-way ANOVA and Dunnett’s multiple comparison test and noted **** p

    Journal: PLoS ONE

    Article Title: The small molecule rhodomyrtone suppresses TNF-α and IL-17A-induced keratinocyte inflammatory responses: A potential new therapeutic for psoriasis

    doi: 10.1371/journal.pone.0205340

    Figure Lengend Snippet: Rhodomyrtone dose dependently decreases production of CXCL1 , IL-36G , IL-36RN , MMP-13 , S100A7 , S100A8 , and S100A12 transcripts by IL-17A/TNF-α-stimulated primary human keratinocytes. Keratinocytes were pretreated with rhodomyrtone (50, 100, 200, 400, and 600 ng/ml) before stimulation with 20ng/ml IL-17A+10ng/ml TNF-α. The mRNA expression levels were measured at 24 h by qRT-PCR. Values are mean±SD of results from three independent experiments in triplicate. Statistical significance determined using one-way ANOVA and Dunnett’s multiple comparison test and noted **** p

    Article Snippet: NHK were pretreated with rhodomyrtone (50–600 ng/ml) for 1 h before stimulation with recombinant human 20ng/ml IL-17A and 10 ng/ml TNF-α (both R & D Systems) for 24 h. Cells were harvested for RNA isolation in complete RLT buffer (Qiagen) or protein isolation in complete RIPA buffer.

    Techniques: Expressing, Quantitative RT-PCR

    TNF-α and IL-17A drive psoriasis transcript expression in ex vivo skin organ cultures. To validate the use of cytokine-stimulated normal human skin biopsies as a model of skin inflammation, we treated biopsies with a combination of 10 ng/ml TNF-α and 20 ng/ml IL-17A for 12, 24, 48, and 72 h and monitored expression of DEFB4 , PI3 , LCN2 , S100A7 , and IL36G transcripts by qRT-PCR. Values are mean ± SD of results in triplicate from 3 donors. Statistical significance indicated by * p

    Journal: PLoS ONE

    Article Title: The small molecule rhodomyrtone suppresses TNF-α and IL-17A-induced keratinocyte inflammatory responses: A potential new therapeutic for psoriasis

    doi: 10.1371/journal.pone.0205340

    Figure Lengend Snippet: TNF-α and IL-17A drive psoriasis transcript expression in ex vivo skin organ cultures. To validate the use of cytokine-stimulated normal human skin biopsies as a model of skin inflammation, we treated biopsies with a combination of 10 ng/ml TNF-α and 20 ng/ml IL-17A for 12, 24, 48, and 72 h and monitored expression of DEFB4 , PI3 , LCN2 , S100A7 , and IL36G transcripts by qRT-PCR. Values are mean ± SD of results in triplicate from 3 donors. Statistical significance indicated by * p

    Article Snippet: NHK were pretreated with rhodomyrtone (50–600 ng/ml) for 1 h before stimulation with recombinant human 20ng/ml IL-17A and 10 ng/ml TNF-α (both R & D Systems) for 24 h. Cells were harvested for RNA isolation in complete RLT buffer (Qiagen) or protein isolation in complete RIPA buffer.

    Techniques: Expressing, Ex Vivo, Quantitative RT-PCR

    Rhodomyrtone inhibits IL-17A/TNF-α-induced inflammatory responses in keratinocytes. Primary human keratinocytes were pretreated with rhodomyrtone (50, 100, 200, 400, and 600 ng/ml) for 1 h before stimulation with 20 ng/ml IL-17A+10 ng/ml TNF-α. HBD-2 (a), and CXCL-1 (b), were assayed in conditioned media at 24 h by ELISA and found to be reduced in a dose-dependent manner in the presence of rhodomyrtone. Values are mean ± SD of results from three independent experiments in triplicate. Statistical significance determined using one-way ANOVA and Dunnett’s multiple comparison test and noted **** p

    Journal: PLoS ONE

    Article Title: The small molecule rhodomyrtone suppresses TNF-α and IL-17A-induced keratinocyte inflammatory responses: A potential new therapeutic for psoriasis

    doi: 10.1371/journal.pone.0205340

    Figure Lengend Snippet: Rhodomyrtone inhibits IL-17A/TNF-α-induced inflammatory responses in keratinocytes. Primary human keratinocytes were pretreated with rhodomyrtone (50, 100, 200, 400, and 600 ng/ml) for 1 h before stimulation with 20 ng/ml IL-17A+10 ng/ml TNF-α. HBD-2 (a), and CXCL-1 (b), were assayed in conditioned media at 24 h by ELISA and found to be reduced in a dose-dependent manner in the presence of rhodomyrtone. Values are mean ± SD of results from three independent experiments in triplicate. Statistical significance determined using one-way ANOVA and Dunnett’s multiple comparison test and noted **** p

    Article Snippet: NHK were pretreated with rhodomyrtone (50–600 ng/ml) for 1 h before stimulation with recombinant human 20ng/ml IL-17A and 10 ng/ml TNF-α (both R & D Systems) for 24 h. Cells were harvested for RNA isolation in complete RLT buffer (Qiagen) or protein isolation in complete RIPA buffer.

    Techniques: Enzyme-linked Immunosorbent Assay

    Follistatin blocks the growth arrest induced by activin A in LE6 cells. (A) LE6 cells were treated with activin A (200 ng/ml) in the presence or absence of the indicated doses of follistatin (25, 50, 100, 200, 400, 600 ng/ml). Cell proliferation was detected by CCK-8 assay. (B) LE6 cells were grown in LE media in the presence or absence of activin A (200 ng/ml), follistatin (400 ng/ml) or activin A (200 ng/ml) plus follistatin (400 ng/ml). DNA synthesis was detected by BrdU incorporation assay using FACS. (C) LE6 cells were treated with or without activin A (200 ng/ml), follistatin (400 ng/ml) or activin A (200 ng/ml) plus follistatin (400 ng/ml) for 30 min. Phosphorylated SMAD2/3 was detected by western-blot. (D) LE6 cells were treated with either media alone, activin A (200 ng/ml), follistatin (400 ng/ml) or activin(200 ng/ml) plus follistatin (400 ng/ml). Then phosphorylated Rb, cyclinD1, cyclinE, p21 WAF1/Cip1 and p15 INK4B were analyzed by western-blot.

    Journal: Cell Communication and Signaling : CCS

    Article Title: Activin A induces growth arrest through a SMAD- dependent pathway in hepatic progenitor cells

    doi: 10.1186/1478-811X-12-18

    Figure Lengend Snippet: Follistatin blocks the growth arrest induced by activin A in LE6 cells. (A) LE6 cells were treated with activin A (200 ng/ml) in the presence or absence of the indicated doses of follistatin (25, 50, 100, 200, 400, 600 ng/ml). Cell proliferation was detected by CCK-8 assay. (B) LE6 cells were grown in LE media in the presence or absence of activin A (200 ng/ml), follistatin (400 ng/ml) or activin A (200 ng/ml) plus follistatin (400 ng/ml). DNA synthesis was detected by BrdU incorporation assay using FACS. (C) LE6 cells were treated with or without activin A (200 ng/ml), follistatin (400 ng/ml) or activin A (200 ng/ml) plus follistatin (400 ng/ml) for 30 min. Phosphorylated SMAD2/3 was detected by western-blot. (D) LE6 cells were treated with either media alone, activin A (200 ng/ml), follistatin (400 ng/ml) or activin(200 ng/ml) plus follistatin (400 ng/ml). Then phosphorylated Rb, cyclinD1, cyclinE, p21 WAF1/Cip1 and p15 INK4B were analyzed by western-blot.

    Article Snippet: Activin A and TGF-β1 enzyme-linked immunosorbent assay Supernatants were collected from confluent LE6 cells treated with or without activin A and tested in triplicate for activin A and TGF-β1 concentrations using rat activin A ELISA kit and rat TGF-β1 ELISA kit (R & D system, MN USA).

    Techniques: CCK-8 Assay, DNA Synthesis, BrdU Incorporation Assay, FACS, Western Blot

    Activin A inhibited proliferation of LE6 cells. (A) 2 × 10 5 LE6 cells/ were seed into 6-well plate and treated with serum free LE media in the present of activin A for up to 48 hours. Cell media was harvested and concentration of TGF-β1 was detected by ELISA kit. The figure showed result of one of three independent assays (B) LE6 cells were grown in 8% FBS LE media in the present or absence of activin A (25, 50, 100, 200, 300 ng/ml) for 72 hours. Cell proliferation was assessed by CCK-8 assay. Data showed result of one of three independent assays. (C and D) LE6 cells were grown in 8% FBS LE media in the presence or absence of activin A (200 ng/ml) for 72 hours. Cell DNA synthesis was assessed by BrdU incorporation assay using FACS (C) . R2 = BrdU positive cells. Cell apoptosis was assessed by AnnexinV/PI assay using FACS (D) . LL = Live cells or annexin V and PI negative cells, LR = Early apoptotic cells or annexin V positive cells, UR = Late apoptotic and necrotic cells or annexin V and PI positive cells, UL = Necrotic cells or PI positive cells. Bar chart showed the apoptosis rate (UR + LR) of control and activin treated LE6 cells. The figure showed result of one of three independent assays.

    Journal: Cell Communication and Signaling : CCS

    Article Title: Activin A induces growth arrest through a SMAD- dependent pathway in hepatic progenitor cells

    doi: 10.1186/1478-811X-12-18

    Figure Lengend Snippet: Activin A inhibited proliferation of LE6 cells. (A) 2 × 10 5 LE6 cells/ were seed into 6-well plate and treated with serum free LE media in the present of activin A for up to 48 hours. Cell media was harvested and concentration of TGF-β1 was detected by ELISA kit. The figure showed result of one of three independent assays (B) LE6 cells were grown in 8% FBS LE media in the present or absence of activin A (25, 50, 100, 200, 300 ng/ml) for 72 hours. Cell proliferation was assessed by CCK-8 assay. Data showed result of one of three independent assays. (C and D) LE6 cells were grown in 8% FBS LE media in the presence or absence of activin A (200 ng/ml) for 72 hours. Cell DNA synthesis was assessed by BrdU incorporation assay using FACS (C) . R2 = BrdU positive cells. Cell apoptosis was assessed by AnnexinV/PI assay using FACS (D) . LL = Live cells or annexin V and PI negative cells, LR = Early apoptotic cells or annexin V positive cells, UR = Late apoptotic and necrotic cells or annexin V and PI positive cells, UL = Necrotic cells or PI positive cells. Bar chart showed the apoptosis rate (UR + LR) of control and activin treated LE6 cells. The figure showed result of one of three independent assays.

    Article Snippet: Activin A and TGF-β1 enzyme-linked immunosorbent assay Supernatants were collected from confluent LE6 cells treated with or without activin A and tested in triplicate for activin A and TGF-β1 concentrations using rat activin A ELISA kit and rat TGF-β1 ELISA kit (R & D system, MN USA).

    Techniques: Concentration Assay, Enzyme-linked Immunosorbent Assay, CCK-8 Assay, DNA Synthesis, BrdU Incorporation Assay, FACS

    Activin A activates SMAD signaling in LE6 cells. (A) LE6 cells were treated with activin A (200 ng/ml) for 6 h, 12 h, 24 h 48 h and 72 h, cell lysis was analyzed by western-blot using specific antibody to phosphorylated Rb, cyclinD1, cyclinE , p21 WAF1/Cip1 , p15 INK4B p27 Kip1 , CDK2 and CDK4. β-actin was used as a loading control. (B) LE6 cells were treated with activin A (200 ng/ml) for 1 h, 6 h, 12 h and 24 h, cell lysis was analyzed by western-blot using specific antibody to phosphorylated JNK1/2, ERK1/2 and p38. (C) LE6 cells were treated with activin A (200 ng/ml) for 15 min, 30 min, 60 min and 120 min, cell lysis was analyzed by western-blot using specific antibody to phosphorylated SMAD2, SMAD3. (D) LE6 cells were treated with activin A (200 ng/ml) for 60 min, cell lysis was incubated with anti-SMAD2/3 antibody and protein A/G agarose overnight, then analyzed by western-blot using specific antibody to SMAD4. (E) LE6 cells were treated with activin A (200 ng/ml) for 15 min and 30 min, Nuclear and cytosolic fractions of cells were analyzed by western-blot using specific antibody to SMAD2/3.GAPDH or laminB was used as loading control for cytosolic or nuclear fraction.

    Journal: Cell Communication and Signaling : CCS

    Article Title: Activin A induces growth arrest through a SMAD- dependent pathway in hepatic progenitor cells

    doi: 10.1186/1478-811X-12-18

    Figure Lengend Snippet: Activin A activates SMAD signaling in LE6 cells. (A) LE6 cells were treated with activin A (200 ng/ml) for 6 h, 12 h, 24 h 48 h and 72 h, cell lysis was analyzed by western-blot using specific antibody to phosphorylated Rb, cyclinD1, cyclinE , p21 WAF1/Cip1 , p15 INK4B p27 Kip1 , CDK2 and CDK4. β-actin was used as a loading control. (B) LE6 cells were treated with activin A (200 ng/ml) for 1 h, 6 h, 12 h and 24 h, cell lysis was analyzed by western-blot using specific antibody to phosphorylated JNK1/2, ERK1/2 and p38. (C) LE6 cells were treated with activin A (200 ng/ml) for 15 min, 30 min, 60 min and 120 min, cell lysis was analyzed by western-blot using specific antibody to phosphorylated SMAD2, SMAD3. (D) LE6 cells were treated with activin A (200 ng/ml) for 60 min, cell lysis was incubated with anti-SMAD2/3 antibody and protein A/G agarose overnight, then analyzed by western-blot using specific antibody to SMAD4. (E) LE6 cells were treated with activin A (200 ng/ml) for 15 min and 30 min, Nuclear and cytosolic fractions of cells were analyzed by western-blot using specific antibody to SMAD2/3.GAPDH or laminB was used as loading control for cytosolic or nuclear fraction.

    Article Snippet: Activin A and TGF-β1 enzyme-linked immunosorbent assay Supernatants were collected from confluent LE6 cells treated with or without activin A and tested in triplicate for activin A and TGF-β1 concentrations using rat activin A ELISA kit and rat TGF-β1 ELISA kit (R & D system, MN USA).

    Techniques: Lysis, Western Blot, Incubation

    Knockdown of Smad4 blocks the anti-proliferative effect of activin A in LE6 cells. (A) SMAD4 expression was determined by western-blot in LE6 cells after Smad4 knockdown (sh1, 2, 3, 4), and compared with control cells (wt) and vehicle cells (V). (B and C) Control cells, vehicle and Smad4 knockdown LE6 cells (LE6- Smad4 KD) were treated with or without 200 ng/ml activin A. Phosphorylated SMAD2 was detected by western-blot (B) and SMAD2/3/4 complex formation was detected by co-immuno precipitation (C) . (D) sh Smad4 -LE6 cells were treated with or without 200 ng/ml activin A and their proliferation assessed by CCK-8 and BrdU incorporation assay. c: control, v: vehicle, sh3: LE6-smad4KD. (E) Indicated cells were treated with or without 200 ng/ml activin A, phosphorylated Rb, cyclinD1, cyclinE , p21 WAF1/Cip1 and p15 INK4B were analyzed by western blotting.

    Journal: Cell Communication and Signaling : CCS

    Article Title: Activin A induces growth arrest through a SMAD- dependent pathway in hepatic progenitor cells

    doi: 10.1186/1478-811X-12-18

    Figure Lengend Snippet: Knockdown of Smad4 blocks the anti-proliferative effect of activin A in LE6 cells. (A) SMAD4 expression was determined by western-blot in LE6 cells after Smad4 knockdown (sh1, 2, 3, 4), and compared with control cells (wt) and vehicle cells (V). (B and C) Control cells, vehicle and Smad4 knockdown LE6 cells (LE6- Smad4 KD) were treated with or without 200 ng/ml activin A. Phosphorylated SMAD2 was detected by western-blot (B) and SMAD2/3/4 complex formation was detected by co-immuno precipitation (C) . (D) sh Smad4 -LE6 cells were treated with or without 200 ng/ml activin A and their proliferation assessed by CCK-8 and BrdU incorporation assay. c: control, v: vehicle, sh3: LE6-smad4KD. (E) Indicated cells were treated with or without 200 ng/ml activin A, phosphorylated Rb, cyclinD1, cyclinE , p21 WAF1/Cip1 and p15 INK4B were analyzed by western blotting.

    Article Snippet: Activin A and TGF-β1 enzyme-linked immunosorbent assay Supernatants were collected from confluent LE6 cells treated with or without activin A and tested in triplicate for activin A and TGF-β1 concentrations using rat activin A ELISA kit and rat TGF-β1 ELISA kit (R & D system, MN USA).

    Techniques: Expressing, Western Blot, Immunoprecipitation, CCK-8 Assay, BrdU Incorporation Assay

    Activin A and follistatin expression in the 2-AAF/PH model correlates with HPC proliferation. (A, E, I) Negative control. (B-D) HPCs and interlobular or terminal duct cells showed positive immuno-reactivity (brown) to pan-CK monoclonal antibody. (×100 magnification). (F-H) Expression of Activin A in 2-AAF/PH model (×100 magnification). (J-L) Expression of Follistatin in 2-AAF/PH model (×100 magnification). (B , F and J) 2-AAF treated rats without PH; (C, G) 9th day after PH; (D, H) 15th day after PH; (K) 2th day after PH; (L) 4th day after PH. (M) The change of HPCs counts in residual liver tissue following PH in 2-AAF/PH model. Expression of activinA (N) and follistatin (O) were detected by IHC and IOD values were analyzed with Image-Pro Plus software (v. 5.0) and recorded in the histograms. (P and Q) Expression of activin βA (P) and follistatin (Q) mRNA in residual liver tissue of partial hepatectomic rats treated with 2-AAF. Data were log-transformed and analyzed by ANOVA for group difference (2-AAF/PH VS shame operation) and time difference (each time point VS 0 day) * P

    Journal: Cell Communication and Signaling : CCS

    Article Title: Activin A induces growth arrest through a SMAD- dependent pathway in hepatic progenitor cells

    doi: 10.1186/1478-811X-12-18

    Figure Lengend Snippet: Activin A and follistatin expression in the 2-AAF/PH model correlates with HPC proliferation. (A, E, I) Negative control. (B-D) HPCs and interlobular or terminal duct cells showed positive immuno-reactivity (brown) to pan-CK monoclonal antibody. (×100 magnification). (F-H) Expression of Activin A in 2-AAF/PH model (×100 magnification). (J-L) Expression of Follistatin in 2-AAF/PH model (×100 magnification). (B , F and J) 2-AAF treated rats without PH; (C, G) 9th day after PH; (D, H) 15th day after PH; (K) 2th day after PH; (L) 4th day after PH. (M) The change of HPCs counts in residual liver tissue following PH in 2-AAF/PH model. Expression of activinA (N) and follistatin (O) were detected by IHC and IOD values were analyzed with Image-Pro Plus software (v. 5.0) and recorded in the histograms. (P and Q) Expression of activin βA (P) and follistatin (Q) mRNA in residual liver tissue of partial hepatectomic rats treated with 2-AAF. Data were log-transformed and analyzed by ANOVA for group difference (2-AAF/PH VS shame operation) and time difference (each time point VS 0 day) * P

    Article Snippet: Activin A and TGF-β1 enzyme-linked immunosorbent assay Supernatants were collected from confluent LE6 cells treated with or without activin A and tested in triplicate for activin A and TGF-β1 concentrations using rat activin A ELISA kit and rat TGF-β1 ELISA kit (R & D system, MN USA).

    Techniques: Expressing, Negative Control, Immunohistochemistry, Software, Transformation Assay

    Netrin-1 promotes inflammation resolution in diabetic corneal wound healing. Corneal epithelium was scraped in control and diabetic mice with or without subconjunctival injection of netrin-1 (50 ng per eye). Mouse cornea was collected and immunofluorescent stained with neutrophil marker Ly6G at 48 h and M2 macrophage markers CD68 (green fluorescence) and CD206 (red fluorescence) at 72 h after epithelial scrape ( A , 3 mice per group). Flow cytometry was performed to quantify the number and percentage of neutrophils (CD45 + CD11b + Ly6G+) ( B , 12 mice per group) at 48-hour post-wounded, M1 macrophages (CD45 + F4/80 + CD86+) and M2 macrophages (CD45 + F4/80 + CD206+) at 72-hour post-wounded. ( C , 12 mice per group at each detection) mRNA expression levels of iNOS and IL-12 (48 h after epithelial scrape), arginase-1 and IL-10 (72 h after epithelial scrape) were analyzed by RT-qPCR from the control and diabetic mouse corneas ( D , 6 mice per group); *p

    Journal: Scientific Reports

    Article Title: Netrin-1 promotes diabetic corneal wound healing through molecular mechanisms mediated via the adenosine 2B receptor

    doi: 10.1038/s41598-018-24506-9

    Figure Lengend Snippet: Netrin-1 promotes inflammation resolution in diabetic corneal wound healing. Corneal epithelium was scraped in control and diabetic mice with or without subconjunctival injection of netrin-1 (50 ng per eye). Mouse cornea was collected and immunofluorescent stained with neutrophil marker Ly6G at 48 h and M2 macrophage markers CD68 (green fluorescence) and CD206 (red fluorescence) at 72 h after epithelial scrape ( A , 3 mice per group). Flow cytometry was performed to quantify the number and percentage of neutrophils (CD45 + CD11b + Ly6G+) ( B , 12 mice per group) at 48-hour post-wounded, M1 macrophages (CD45 + F4/80 + CD86+) and M2 macrophages (CD45 + F4/80 + CD206+) at 72-hour post-wounded. ( C , 12 mice per group at each detection) mRNA expression levels of iNOS and IL-12 (48 h after epithelial scrape), arginase-1 and IL-10 (72 h after epithelial scrape) were analyzed by RT-qPCR from the control and diabetic mouse corneas ( D , 6 mice per group); *p

    Article Snippet: After the corneal epithelium scraped, diabetic mice were injected subconjunctivally with 50 ng netrin-1 (5 μ L/eye, R & D Systems, Minneapolis, MN) or equal phosphate buffer saline (PBS) as the vehicle control.

    Techniques: Mouse Assay, Injection, Staining, Marker, Fluorescence, Flow Cytometry, Cytometry, Expressing, Quantitative RT-PCR

    Netrin-1 promotes corneal epithelial wound healing in diabetic mice. Corneal epithelium was scraped in control and diabetic mice with or without subconjunctival injection of netrin-1 (50 ng per eye). The corneal epithelial wound defect was stained with fluorescein sodium at 24, 48, and 72 h after epithelial scrape ( A , 8 mice per group). The histogram of residual epithelial defect is presented as the percentage of the original wound area ( B ). *p

    Journal: Scientific Reports

    Article Title: Netrin-1 promotes diabetic corneal wound healing through molecular mechanisms mediated via the adenosine 2B receptor

    doi: 10.1038/s41598-018-24506-9

    Figure Lengend Snippet: Netrin-1 promotes corneal epithelial wound healing in diabetic mice. Corneal epithelium was scraped in control and diabetic mice with or without subconjunctival injection of netrin-1 (50 ng per eye). The corneal epithelial wound defect was stained with fluorescein sodium at 24, 48, and 72 h after epithelial scrape ( A , 8 mice per group). The histogram of residual epithelial defect is presented as the percentage of the original wound area ( B ). *p

    Article Snippet: After the corneal epithelium scraped, diabetic mice were injected subconjunctivally with 50 ng netrin-1 (5 μ L/eye, R & D Systems, Minneapolis, MN) or equal phosphate buffer saline (PBS) as the vehicle control.

    Techniques: Mouse Assay, Injection, Staining

    Netrin-1 promotes the migration, proliferation, and ERK and EGFR phosphorylation of corneal epithelial cells impaired by hyperglycemia. Confluent mouse corneal epithelial cells were wounded after incubation with 5 mM glucose, 30 mM mannose or 30 mM glucose. Cell migration was observed with or without 50 ng/mL netrin-1 treatment for 24 h ( A ). Migration rate was analyzed and represented as the percentage of primary wounding area ( B , n = 3 per group). Cell proliferation was measured by cell counting after 3 days of netrin-1 treatment ( C , n = 3 per group). The phosphorylated levels of ERK and EGFR were analyzed by Western blotting after 3 days incubation until monolayer formation. We administrated netrin-1 (50ng/ml) and after 8 or 15 minutes we collected the cells and then detected the p-ERK or p-EGFR expression by western blotting. ( D , n = 4 per group). The expression of p-ERK/p-EGFR were also confirmed by immunofluorescence staining in the netrin-1 injected diabetic mouse corneal epithelium 48 h after injury ( E , 3 mice per group). *p

    Journal: Scientific Reports

    Article Title: Netrin-1 promotes diabetic corneal wound healing through molecular mechanisms mediated via the adenosine 2B receptor

    doi: 10.1038/s41598-018-24506-9

    Figure Lengend Snippet: Netrin-1 promotes the migration, proliferation, and ERK and EGFR phosphorylation of corneal epithelial cells impaired by hyperglycemia. Confluent mouse corneal epithelial cells were wounded after incubation with 5 mM glucose, 30 mM mannose or 30 mM glucose. Cell migration was observed with or without 50 ng/mL netrin-1 treatment for 24 h ( A ). Migration rate was analyzed and represented as the percentage of primary wounding area ( B , n = 3 per group). Cell proliferation was measured by cell counting after 3 days of netrin-1 treatment ( C , n = 3 per group). The phosphorylated levels of ERK and EGFR were analyzed by Western blotting after 3 days incubation until monolayer formation. We administrated netrin-1 (50ng/ml) and after 8 or 15 minutes we collected the cells and then detected the p-ERK or p-EGFR expression by western blotting. ( D , n = 4 per group). The expression of p-ERK/p-EGFR were also confirmed by immunofluorescence staining in the netrin-1 injected diabetic mouse corneal epithelium 48 h after injury ( E , 3 mice per group). *p

    Article Snippet: After the corneal epithelium scraped, diabetic mice were injected subconjunctivally with 50 ng netrin-1 (5 μ L/eye, R & D Systems, Minneapolis, MN) or equal phosphate buffer saline (PBS) as the vehicle control.

    Techniques: Migration, Incubation, Cell Counting, Western Blot, Expressing, Immunofluorescence, Staining, Injection, Mouse Assay

    The A2BAR receptor mediates the promotion of netrin-1 on diabetic corneal wound healing. A2BAR receptor-specific antagonist or anti-UNC5B blocking antibody was injected subconjunctivally at 24 h before and 0, 24, and 48 h after the scrape of corneal epithelium in netrin-1 treated diabetic mice. Corneal epithelium defect was stained with fluorescein sodium and presented as the percentage of the original wound at 72 h after epithelial scraped ( A , 3 mice per group). Activation of ERK and EGFR in corneal epithelium were analyzed by western blotting at 48 h after epithelial scraped ( B , 3 mice per group). Neutrophils infiltration to the cornea and the cell numbers were examined by immunofluorescence staining ( C , 3 mice per group) and flow cytometry ( D , 12 mice per group) at 48 h after epithelial scraped. Macrophages infiltration to the cornea and the cell numbers were examined by immunofluorescence staining (C, 3 mice per group) and flow cytometry ( D , 12 mice per group of each detection) at 72 h after epithelial scraped. A2R anta: A2BAR antagonist; anti-U5B: anti-UNC5B blocking antibody. *p

    Journal: Scientific Reports

    Article Title: Netrin-1 promotes diabetic corneal wound healing through molecular mechanisms mediated via the adenosine 2B receptor

    doi: 10.1038/s41598-018-24506-9

    Figure Lengend Snippet: The A2BAR receptor mediates the promotion of netrin-1 on diabetic corneal wound healing. A2BAR receptor-specific antagonist or anti-UNC5B blocking antibody was injected subconjunctivally at 24 h before and 0, 24, and 48 h after the scrape of corneal epithelium in netrin-1 treated diabetic mice. Corneal epithelium defect was stained with fluorescein sodium and presented as the percentage of the original wound at 72 h after epithelial scraped ( A , 3 mice per group). Activation of ERK and EGFR in corneal epithelium were analyzed by western blotting at 48 h after epithelial scraped ( B , 3 mice per group). Neutrophils infiltration to the cornea and the cell numbers were examined by immunofluorescence staining ( C , 3 mice per group) and flow cytometry ( D , 12 mice per group) at 48 h after epithelial scraped. Macrophages infiltration to the cornea and the cell numbers were examined by immunofluorescence staining (C, 3 mice per group) and flow cytometry ( D , 12 mice per group of each detection) at 72 h after epithelial scraped. A2R anta: A2BAR antagonist; anti-U5B: anti-UNC5B blocking antibody. *p

    Article Snippet: After the corneal epithelium scraped, diabetic mice were injected subconjunctivally with 50 ng netrin-1 (5 μ L/eye, R & D Systems, Minneapolis, MN) or equal phosphate buffer saline (PBS) as the vehicle control.

    Techniques: Blocking Assay, Injection, Mouse Assay, Staining, Activation Assay, Western Blot, Immunofluorescence, Flow Cytometry, Cytometry

    Hyperglycemia downregulates netrin-1 expression in corneal epithelium. Normal and diabetic mouse corneal epithelium was collected before (as unwound group) or 48 h after epithelial scrape (as wound group). Netrin-1 expression was examined by using immunofluorescence staining ( A , 3 mice per group), RT-qPCR ( B , 9 mice per group) and ELISA ( C , 9 mice per group) with the age-matched normal mice as control. *p

    Journal: Scientific Reports

    Article Title: Netrin-1 promotes diabetic corneal wound healing through molecular mechanisms mediated via the adenosine 2B receptor

    doi: 10.1038/s41598-018-24506-9

    Figure Lengend Snippet: Hyperglycemia downregulates netrin-1 expression in corneal epithelium. Normal and diabetic mouse corneal epithelium was collected before (as unwound group) or 48 h after epithelial scrape (as wound group). Netrin-1 expression was examined by using immunofluorescence staining ( A , 3 mice per group), RT-qPCR ( B , 9 mice per group) and ELISA ( C , 9 mice per group) with the age-matched normal mice as control. *p

    Article Snippet: After the corneal epithelium scraped, diabetic mice were injected subconjunctivally with 50 ng netrin-1 (5 μ L/eye, R & D Systems, Minneapolis, MN) or equal phosphate buffer saline (PBS) as the vehicle control.

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

    High-mobility group protein B1 (HMGB1) and heat shock protein 70 (HSP70) stimulate Toll-like receptor 4 (TLR4; +/+) but not TLR4 (−/−) bone marrow (BM)-derived macrophages: BM-derived macrophages stimulated in vitro with media, HSP70 (3 μg/ml), and HMGB1 (3 μg/ml) Interleukin 6 (IL6) was measured by quantitative reverse transcriptase-PCR. Each sample normalized to glyceraldehyde 3-phosphate dehydrogenase (GAPDH). The calibrator gene is the media-treated BMDM. n = 4. Means and standard deviation are shown. * P

    Journal: Kidney International

    Article Title: Early interleukin 6 production by leukocytes during ischemic acute kidney injury is regulated by TLR4

    doi: 10.1038/ki.2011.140

    Figure Lengend Snippet: High-mobility group protein B1 (HMGB1) and heat shock protein 70 (HSP70) stimulate Toll-like receptor 4 (TLR4; +/+) but not TLR4 (−/−) bone marrow (BM)-derived macrophages: BM-derived macrophages stimulated in vitro with media, HSP70 (3 μg/ml), and HMGB1 (3 μg/ml) Interleukin 6 (IL6) was measured by quantitative reverse transcriptase-PCR. Each sample normalized to glyceraldehyde 3-phosphate dehydrogenase (GAPDH). The calibrator gene is the media-treated BMDM. n = 4. Means and standard deviation are shown. * P

    Article Snippet: Explanations include the release of co-stimulatory cytokines or post-translational modification of HMGB1 by stressed cells so that HMGB1 in the supernatant is more stimulatory than purified HMGB1., Endotoxin was not responsible for the stimulatory effect of our HMGB1 (1690-HM, lot MHZ24 from R & D Systems, Minneapolis, MN) that contained less than 0.00855 EU/μg.

    Techniques: Derivative Assay, In Vitro, Polymerase Chain Reaction, Standard Deviation

    Quantification of inflammatory indicators in mice infected with recombinant Mycobacterium smegmatis . The spleen was excised from the infected mice ( n = 3 per group), and the spleen lymphocytes were separated and incubated overnight with PBS, pMVp262/MS and Rv0888NS/MS, D438A/MS, and H481N/MS. The cytokines were determined using the cultured supernatant. (A) Analysis of the IL-1β. (B) Analysis of IL-6. (C) Analysis of TNF-α. The error bars show the SEM of three independent experiments with three wells per group. (A–C) One-way ANOVA followed by Bonferroni’s multiple comparison post hoc test, * P

    Journal: Frontiers in Immunology

    Article Title: Extracellular Sphingomyelinase Rv0888 of Mycobacterium tuberculosis Contributes to Pathological Lung Injury of Mycobacterium smegmatis in Mice via Inducing Formation of Neutrophil Extracellular Traps

    doi: 10.3389/fimmu.2018.00677

    Figure Lengend Snippet: Quantification of inflammatory indicators in mice infected with recombinant Mycobacterium smegmatis . The spleen was excised from the infected mice ( n = 3 per group), and the spleen lymphocytes were separated and incubated overnight with PBS, pMVp262/MS and Rv0888NS/MS, D438A/MS, and H481N/MS. The cytokines were determined using the cultured supernatant. (A) Analysis of the IL-1β. (B) Analysis of IL-6. (C) Analysis of TNF-α. The error bars show the SEM of three independent experiments with three wells per group. (A–C) One-way ANOVA followed by Bonferroni’s multiple comparison post hoc test, * P

    Article Snippet: IL-6, TNF-α, IL-1β, and IFN-γ levels in the supernatant were detected with a commercially available mouse IL-6, TNF-α, IL-1β, and IFN-γ ELISA Kit (eBioscience, Waltham, MA, USA) according to the manufacturer’s instructions.

    Techniques: Mouse Assay, Infection, Recombinant, Incubation, Mass Spectrometry, Cell Culture

    VLP effectively stimulates BMDCs in vitro to secrete cytokines. BMDCs were stimulated with M2e5x, M1 VLP, or protein (0.3, 1, 5 μg/mL) for 24 h. ( a – c ) The levels of cytokines ( a ) TNF-α, ( b ) IL-6, ( c ) IFN-γ were determined in the culture supernatants by ELISA assay. Un: Medium only, P: proteins, V: VLP. The statistical significance was confirmed by one-way ANOVA and Dunnett’s multiple comparison test. Error bars indicate the means ± SEM of concentrations from individual animals. *; p

    Journal: Vaccines

    Article Title: Virus-Like Particles Are a Superior Platform for Presenting M2e Epitopes to Prime Humoral and Cellular Immunity against Influenza Virus

    doi: 10.3390/vaccines6040066

    Figure Lengend Snippet: VLP effectively stimulates BMDCs in vitro to secrete cytokines. BMDCs were stimulated with M2e5x, M1 VLP, or protein (0.3, 1, 5 μg/mL) for 24 h. ( a – c ) The levels of cytokines ( a ) TNF-α, ( b ) IL-6, ( c ) IFN-γ were determined in the culture supernatants by ELISA assay. Un: Medium only, P: proteins, V: VLP. The statistical significance was confirmed by one-way ANOVA and Dunnett’s multiple comparison test. Error bars indicate the means ± SEM of concentrations from individual animals. *; p

    Article Snippet: Briefly, interleukin-6 (IL-6), IL-10, tumor necrosis factor alpha (TNF-α), and interferon gamma (IFN-γ) kits (eBioscience, San Diego, CA, USA) were used to determine the level of cytokines in the BALF and culture supernatants according to the manufacturer’s procedures.

    Techniques: In Vitro, Enzyme-linked Immunosorbent Assay

    VLP effectively stimulates BMDCs in vitro to secrete cytokines. BMDCs were stimulated with M2e5x, M1 VLP, or protein (0.3, 1, 5 μg/mL) for 24 h. ( a – c ) The levels of cytokines ( a ) TNF-α, ( b ) IL-6, ( c ) IFN-γ were determined in the culture supernatants by ELISA assay. Un: Medium only, P: proteins, V: VLP. The statistical significance was confirmed by one-way ANOVA and Dunnett’s multiple comparison test. Error bars indicate the means ± SEM of concentrations from individual animals. *; p

    Journal: Vaccines

    Article Title: Virus-Like Particles Are a Superior Platform for Presenting M2e Epitopes to Prime Humoral and Cellular Immunity against Influenza Virus

    doi: 10.3390/vaccines6040066

    Figure Lengend Snippet: VLP effectively stimulates BMDCs in vitro to secrete cytokines. BMDCs were stimulated with M2e5x, M1 VLP, or protein (0.3, 1, 5 μg/mL) for 24 h. ( a – c ) The levels of cytokines ( a ) TNF-α, ( b ) IL-6, ( c ) IFN-γ were determined in the culture supernatants by ELISA assay. Un: Medium only, P: proteins, V: VLP. The statistical significance was confirmed by one-way ANOVA and Dunnett’s multiple comparison test. Error bars indicate the means ± SEM of concentrations from individual animals. *; p

    Article Snippet: Briefly, interleukin-6 (IL-6), IL-10, tumor necrosis factor alpha (TNF-α), and interferon gamma (IFN-γ) kits (eBioscience, San Diego, CA, USA) were used to determine the level of cytokines in the BALF and culture supernatants according to the manufacturer’s procedures.

    Techniques: In Vitro, Enzyme-linked Immunosorbent Assay

    VLP is superior to protein in priming acute innate immune responses in vivo at the injection site. Balb/c mice ( n = 3) were i.p. injected with 200 μL of PBS, 10 μg 5xM2e VLP, or 25 μg 5xM2e protein. The levels of cytokine and chemokine were determined in sera and peritoneal exudates collected at the indicated times. The levels of cytokines (TNF-α, IL-6, IFN-γ) and chemokines (MCP-1, RANTES) in serum ( a ) and Peritoneal exudates ( b ). ( c – j ) Different phenotypic innate immune cells were determined in the cells collected from peritoneal exudates at 24 h post injection using flow cytometry. PRO: 5xM2e proteins, VLP: 5xM2e VLP. The statistical significance was confirmed by one-way ANOVA and Dunnett’s multiple comparison test. Error bars indicate the means ± SEM of concentrations from individual animals. *; p

    Journal: Vaccines

    Article Title: Virus-Like Particles Are a Superior Platform for Presenting M2e Epitopes to Prime Humoral and Cellular Immunity against Influenza Virus

    doi: 10.3390/vaccines6040066

    Figure Lengend Snippet: VLP is superior to protein in priming acute innate immune responses in vivo at the injection site. Balb/c mice ( n = 3) were i.p. injected with 200 μL of PBS, 10 μg 5xM2e VLP, or 25 μg 5xM2e protein. The levels of cytokine and chemokine were determined in sera and peritoneal exudates collected at the indicated times. The levels of cytokines (TNF-α, IL-6, IFN-γ) and chemokines (MCP-1, RANTES) in serum ( a ) and Peritoneal exudates ( b ). ( c – j ) Different phenotypic innate immune cells were determined in the cells collected from peritoneal exudates at 24 h post injection using flow cytometry. PRO: 5xM2e proteins, VLP: 5xM2e VLP. The statistical significance was confirmed by one-way ANOVA and Dunnett’s multiple comparison test. Error bars indicate the means ± SEM of concentrations from individual animals. *; p

    Article Snippet: Briefly, interleukin-6 (IL-6), IL-10, tumor necrosis factor alpha (TNF-α), and interferon gamma (IFN-γ) kits (eBioscience, San Diego, CA, USA) were used to determine the level of cytokines in the BALF and culture supernatants according to the manufacturer’s procedures.

    Techniques: In Vivo, Injection, Mouse Assay, Flow Cytometry, Cytometry

    Hepcidin and GDF-15 are increased and their expression is correlated in breast tumors (A and B) Box plot with Tukey whisker of (A) GDF15 and (B) HAMP mRNA expression (log2 transformed) in normal adjacent tissue (n=61) compared to primary tumor tissue (n=526) in the TCGA breast cancer dataset. (C) GDF15 transcripts in TCGA samples from breast cancer patients divided by HAMP expression (below and above the mean) shown as box and whisker plot. (D) HAMP transcripts in TCGA samples from breast cancer patients divided by GDF15 expression (below and above the mean) shown as box and whisker. (E) Representative images of immunohistochemical staining of tumor tissue from patients with invasive ductal carcinoma (IDC). Proteins stained are Hepcidin, GDF-15, Pan-Cytokeratin and IgG control. (F) Scatter plot displays quantification of staining of epithelial cells from tissues from 56 BRCA patients. A regression analysis was performed to examine correlation of staining intensities (R 2 =0.4434 p

    Journal: Oncogene

    Article Title: Contribution of three dimensional architecture and tumor-associated fibroblasts to hepcidin regulation in breast cancer

    doi: 10.1038/s41388-018-0243-y

    Figure Lengend Snippet: Hepcidin and GDF-15 are increased and their expression is correlated in breast tumors (A and B) Box plot with Tukey whisker of (A) GDF15 and (B) HAMP mRNA expression (log2 transformed) in normal adjacent tissue (n=61) compared to primary tumor tissue (n=526) in the TCGA breast cancer dataset. (C) GDF15 transcripts in TCGA samples from breast cancer patients divided by HAMP expression (below and above the mean) shown as box and whisker plot. (D) HAMP transcripts in TCGA samples from breast cancer patients divided by GDF15 expression (below and above the mean) shown as box and whisker. (E) Representative images of immunohistochemical staining of tumor tissue from patients with invasive ductal carcinoma (IDC). Proteins stained are Hepcidin, GDF-15, Pan-Cytokeratin and IgG control. (F) Scatter plot displays quantification of staining of epithelial cells from tissues from 56 BRCA patients. A regression analysis was performed to examine correlation of staining intensities (R 2 =0.4434 p

    Article Snippet: GDF-15 and IL-6 were measured in conditioned growth media using a GDF-15 or IL-6 Human ELISA kit from R & D systems and following manufacturers’ protocol.

    Techniques: Expressing, Whisker Assay, Transformation Assay, Immunohistochemistry, Staining

    Working model of regulation of hepcidin in breast cancer spheroids Hepcidin expression is increased in breast cancer cells relative to non-cancer cells. A. In MCF-7 cells grown as 2D monolayers, BMP6 plays a dominant role in the cancer-dependent increase in hepcidin through activation of a SMAD1-5-8 signaling pathway. B. In breast cancer spheroids, spatial control of hepcidin synthesis is exerted by GDF-15, which augments SMAD 1-5- 8 signaling to further increase hepcidin synthesis. C. The microenvironment is an additional source for increased hepcidin in breast cancer cells, in part due to production of IL-6 by tumor-associated fibroblasts (TAFs).

    Journal: Oncogene

    Article Title: Contribution of three dimensional architecture and tumor-associated fibroblasts to hepcidin regulation in breast cancer

    doi: 10.1038/s41388-018-0243-y

    Figure Lengend Snippet: Working model of regulation of hepcidin in breast cancer spheroids Hepcidin expression is increased in breast cancer cells relative to non-cancer cells. A. In MCF-7 cells grown as 2D monolayers, BMP6 plays a dominant role in the cancer-dependent increase in hepcidin through activation of a SMAD1-5-8 signaling pathway. B. In breast cancer spheroids, spatial control of hepcidin synthesis is exerted by GDF-15, which augments SMAD 1-5- 8 signaling to further increase hepcidin synthesis. C. The microenvironment is an additional source for increased hepcidin in breast cancer cells, in part due to production of IL-6 by tumor-associated fibroblasts (TAFs).

    Article Snippet: GDF-15 and IL-6 were measured in conditioned growth media using a GDF-15 or IL-6 Human ELISA kit from R & D systems and following manufacturers’ protocol.

    Techniques: Expressing, Activation Assay

    GDF-15 is induced in breast cancer spheroids and correlates with hepcidin expression (A–C) RT-qPCR of GDF-15 mRNA (normalized to Cyclophilin A) between (A) MCF-7 and MCF-10A spheroids, (B) patient 107 tumor vs. normal adjacent tumor (normal) spheroids and (C) MCF-7 monolayer vs. MCF-7 spheroids. (D) Secreted GDF-15 from conditioned media of MCF-7 monolayer and spheroids normalized to μg protein. (E–F) RT-qPCR of (E) GDF-15 mRNA (normalized to Cyclophilin A) and (F) hepcidin mRNA (normalized to Cyclophilin A) in MCF-7 monolayer (2D) and spheroids (3D) over time.

    Journal: Oncogene

    Article Title: Contribution of three dimensional architecture and tumor-associated fibroblasts to hepcidin regulation in breast cancer

    doi: 10.1038/s41388-018-0243-y

    Figure Lengend Snippet: GDF-15 is induced in breast cancer spheroids and correlates with hepcidin expression (A–C) RT-qPCR of GDF-15 mRNA (normalized to Cyclophilin A) between (A) MCF-7 and MCF-10A spheroids, (B) patient 107 tumor vs. normal adjacent tumor (normal) spheroids and (C) MCF-7 monolayer vs. MCF-7 spheroids. (D) Secreted GDF-15 from conditioned media of MCF-7 monolayer and spheroids normalized to μg protein. (E–F) RT-qPCR of (E) GDF-15 mRNA (normalized to Cyclophilin A) and (F) hepcidin mRNA (normalized to Cyclophilin A) in MCF-7 monolayer (2D) and spheroids (3D) over time.

    Article Snippet: GDF-15 and IL-6 were measured in conditioned growth media using a GDF-15 or IL-6 Human ELISA kit from R & D systems and following manufacturers’ protocol.

    Techniques: Expressing, Quantitative RT-PCR

    GDF-15 positively regulates hepcidin in breast cancer spheroids via a conserved pSMAD1-5-8 pathway (A) RT-qPCR of Hepcidin mRNA (normalized to Cyclophilin A) and (B) western blot analysis of pro-hepcidin and β-actin following knock-down using non targeting control (NTC) siRNA and GDF-15 siRNA (KD#1) in MCF-7 spheroids. Untreated MCF-7 cells grown in 2D or 3D were used as controls. For statistical analysis and quantification, samples were compared to non-targeting control. (C) Western blot analysis of phosphorylated SMAD2-3, total SMAD2, phosphorylated SMAD1-5-8, total SMAD-1, and β-actin in MCF-7 monolayers and spheroids. (D) Western blot analysis of pSMAD1-5-8, tSMAD-1 and β-actin following siRNA knock-down of NTC and GDF-15 knock-down #1. (E–F) RT-qPCR of (E) hepcidin mRNA (normalized to Cyclophilin A) and (F) GDF-15 mRNA (normalized to Cyclophiin A) following siRNA knock-down of NTC and GDF-15 knock-down #1 for 3 days in MCF-7 monolayer. For statistical analysis, samples were compared to non-targeting control.

    Journal: Oncogene

    Article Title: Contribution of three dimensional architecture and tumor-associated fibroblasts to hepcidin regulation in breast cancer

    doi: 10.1038/s41388-018-0243-y

    Figure Lengend Snippet: GDF-15 positively regulates hepcidin in breast cancer spheroids via a conserved pSMAD1-5-8 pathway (A) RT-qPCR of Hepcidin mRNA (normalized to Cyclophilin A) and (B) western blot analysis of pro-hepcidin and β-actin following knock-down using non targeting control (NTC) siRNA and GDF-15 siRNA (KD#1) in MCF-7 spheroids. Untreated MCF-7 cells grown in 2D or 3D were used as controls. For statistical analysis and quantification, samples were compared to non-targeting control. (C) Western blot analysis of phosphorylated SMAD2-3, total SMAD2, phosphorylated SMAD1-5-8, total SMAD-1, and β-actin in MCF-7 monolayers and spheroids. (D) Western blot analysis of pSMAD1-5-8, tSMAD-1 and β-actin following siRNA knock-down of NTC and GDF-15 knock-down #1. (E–F) RT-qPCR of (E) hepcidin mRNA (normalized to Cyclophilin A) and (F) GDF-15 mRNA (normalized to Cyclophiin A) following siRNA knock-down of NTC and GDF-15 knock-down #1 for 3 days in MCF-7 monolayer. For statistical analysis, samples were compared to non-targeting control.

    Article Snippet: GDF-15 and IL-6 were measured in conditioned growth media using a GDF-15 or IL-6 Human ELISA kit from R & D systems and following manufacturers’ protocol.

    Techniques: Quantitative RT-PCR, Western Blot

    The level of TNF-α protein increased in TOLF. A) Fold change of TNF-α protein in the ossified ligamentum flavum of TOLF. Proteins were extracted from the ossified ligamentum flavum (OS-Lig) and normal ligamentum flavum (Lig). The samples were then labeled with iTRAQ following the manufacture’s protocol. *: A star indicates statistical significance compared to control group with p

    Journal: PLoS ONE

    Article Title: The involvement and possible mechanism of pro-inflammatory tumor necrosis factor alpha (TNF-α) in thoracic ossification of the ligamentum flavum

    doi: 10.1371/journal.pone.0178986

    Figure Lengend Snippet: The level of TNF-α protein increased in TOLF. A) Fold change of TNF-α protein in the ossified ligamentum flavum of TOLF. Proteins were extracted from the ossified ligamentum flavum (OS-Lig) and normal ligamentum flavum (Lig). The samples were then labeled with iTRAQ following the manufacture’s protocol. *: A star indicates statistical significance compared to control group with p

    Article Snippet: Primary ligamentum flavum cells derived from TOLF patients were cultured and treated as indicated with recombinant human TNF-α (100 ng/ml; R & D Systems) for 24 hr.

    Techniques: Labeling

    Proportions of cells in S phase increased after TNF-α stimulation. Primary cells of the ossified ligamentum flavum were cultured in the absence of TNF-α (A) or presence of TNF-α (B). Flow cytometry assay were used to examine the cell cycle. The proportions of cells in S phase of cell cycle increased up to 17.06% after TNF-α stimulation.

    Journal: PLoS ONE

    Article Title: The involvement and possible mechanism of pro-inflammatory tumor necrosis factor alpha (TNF-α) in thoracic ossification of the ligamentum flavum

    doi: 10.1371/journal.pone.0178986

    Figure Lengend Snippet: Proportions of cells in S phase increased after TNF-α stimulation. Primary cells of the ossified ligamentum flavum were cultured in the absence of TNF-α (A) or presence of TNF-α (B). Flow cytometry assay were used to examine the cell cycle. The proportions of cells in S phase of cell cycle increased up to 17.06% after TNF-α stimulation.

    Article Snippet: Primary ligamentum flavum cells derived from TOLF patients were cultured and treated as indicated with recombinant human TNF-α (100 ng/ml; R & D Systems) for 24 hr.

    Techniques: Cell Culture, Flow Cytometry, Cytometry

    TNF-α activated the expressions of cyclin D1 and c-Myc. Primary ligamentum flavum cells derived from TOLF patients were cultured and treated with TNF-α (100 ng/ml) for 24 hr. Total RNA was isolated and measured by real time RT-PCR. The RNA level from the control group was normalized to a value of 1. Values were presented as the mean ±S.D. A paired t -test was performed comparing control and TNF-α group. *: A star indicates statistical significance compared to control group with p

    Journal: PLoS ONE

    Article Title: The involvement and possible mechanism of pro-inflammatory tumor necrosis factor alpha (TNF-α) in thoracic ossification of the ligamentum flavum

    doi: 10.1371/journal.pone.0178986

    Figure Lengend Snippet: TNF-α activated the expressions of cyclin D1 and c-Myc. Primary ligamentum flavum cells derived from TOLF patients were cultured and treated with TNF-α (100 ng/ml) for 24 hr. Total RNA was isolated and measured by real time RT-PCR. The RNA level from the control group was normalized to a value of 1. Values were presented as the mean ±S.D. A paired t -test was performed comparing control and TNF-α group. *: A star indicates statistical significance compared to control group with p

    Article Snippet: Primary ligamentum flavum cells derived from TOLF patients were cultured and treated as indicated with recombinant human TNF-α (100 ng/ml; R & D Systems) for 24 hr.

    Techniques: Derivative Assay, Cell Culture, Isolation, Quantitative RT-PCR

    TNF-α induced the expression of osteoblast genes in primary cells from TOLF. Primary ligamentum flavum cells were derived from TOLF patients. Primary cells were treated with 100 ng/ml TNF-α or as indicated for 24 hr. Total RNA was isolated and measured by real time RT-PCR. The RNA level from the control group was normalized to a value of 1. Values were presented as the mean ±S.D. A paired t -test was performed comparing control and TNF-α group. *: A star indicates statistical significance compared to control group with p

    Journal: PLoS ONE

    Article Title: The involvement and possible mechanism of pro-inflammatory tumor necrosis factor alpha (TNF-α) in thoracic ossification of the ligamentum flavum

    doi: 10.1371/journal.pone.0178986

    Figure Lengend Snippet: TNF-α induced the expression of osteoblast genes in primary cells from TOLF. Primary ligamentum flavum cells were derived from TOLF patients. Primary cells were treated with 100 ng/ml TNF-α or as indicated for 24 hr. Total RNA was isolated and measured by real time RT-PCR. The RNA level from the control group was normalized to a value of 1. Values were presented as the mean ±S.D. A paired t -test was performed comparing control and TNF-α group. *: A star indicates statistical significance compared to control group with p

    Article Snippet: Primary ligamentum flavum cells derived from TOLF patients were cultured and treated as indicated with recombinant human TNF-α (100 ng/ml; R & D Systems) for 24 hr.

    Techniques: Expressing, Derivative Assay, Isolation, Quantitative RT-PCR

    Incubation of TGF-β1 NAB in VSMCs increased the expression of lamin A in cocultured ECs, but PDGF-BB had no significant effect ( A ). NABs of both PDGF-BB and TGF-β1 incubation in VSMCs decreased expressions of LOX and phospho-ERK1/2 ( A

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

    Article Title: PDGF-BB and TGF-?1 on cross-talk between endothelial and smooth muscle cells in vascular remodeling induced by low shear stress

    doi: 10.1073/pnas.1019219108

    Figure Lengend Snippet: Incubation of TGF-β1 NAB in VSMCs increased the expression of lamin A in cocultured ECs, but PDGF-BB had no significant effect ( A ). NABs of both PDGF-BB and TGF-β1 incubation in VSMCs decreased expressions of LOX and phospho-ERK1/2 ( A

    Article Snippet: Recombinant proteins of PDGF-BB and TGF-β1, NABs of PDGF-BB and TGF-β1, and Quantikine ELISA Kit for PDGF-BB and TGF-β1 were purchased from R & D Systems.

    Techniques: Incubation, Expressing

    PDGF-BB target siRNA transfection significantly decreased the secretions of PDGF-BB and TGF-β1 from ECs under LowSS conditions ( A ). PDGF-BB knockdown in ECs also markedly repressed the secretion of PDGF-BB and TGF-β1 from cocultured VSMCs

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

    Article Title: PDGF-BB and TGF-?1 on cross-talk between endothelial and smooth muscle cells in vascular remodeling induced by low shear stress

    doi: 10.1073/pnas.1019219108

    Figure Lengend Snippet: PDGF-BB target siRNA transfection significantly decreased the secretions of PDGF-BB and TGF-β1 from ECs under LowSS conditions ( A ). PDGF-BB knockdown in ECs also markedly repressed the secretion of PDGF-BB and TGF-β1 from cocultured VSMCs

    Article Snippet: Recombinant proteins of PDGF-BB and TGF-β1, NABs of PDGF-BB and TGF-β1, and Quantikine ELISA Kit for PDGF-BB and TGF-β1 were purchased from R & D Systems.

    Techniques: Transfection

    In ECs and VSMCs, LowSS induced the secretion of PDGF-BB and TGF-β1 at 12 and 24 h ( A and B ) and decreased the expression of lamin A and LOX at 6, 12, and 24 h ( C and D ). The phosphorylation of ERK1/2 was up-regulated by LowSS at 6 and 12 h in

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

    Article Title: PDGF-BB and TGF-?1 on cross-talk between endothelial and smooth muscle cells in vascular remodeling induced by low shear stress

    doi: 10.1073/pnas.1019219108

    Figure Lengend Snippet: In ECs and VSMCs, LowSS induced the secretion of PDGF-BB and TGF-β1 at 12 and 24 h ( A and B ) and decreased the expression of lamin A and LOX at 6, 12, and 24 h ( C and D ). The phosphorylation of ERK1/2 was up-regulated by LowSS at 6 and 12 h in

    Article Snippet: Recombinant proteins of PDGF-BB and TGF-β1, NABs of PDGF-BB and TGF-β1, and Quantikine ELISA Kit for PDGF-BB and TGF-β1 were purchased from R & D Systems.

    Techniques: Expressing

    Enzyme linked immunosorbent assay (ELISA). a, b Representative graph showing expression of vascular endothelial growth factor A (VEGF) ( a ) and angiopoietin 1 (Ang-1) ( b ). Data presented as mean ± standard deviation. * p ≤ 0.05, *** p ≤ 0.001, n = 4. d days

    Journal: Stem Cell Research & Therapy

    Article Title: Human dental pulp stromal cell conditioned medium alters endothelial cell behavior

    doi: 10.1186/s13287-018-0815-3

    Figure Lengend Snippet: Enzyme linked immunosorbent assay (ELISA). a, b Representative graph showing expression of vascular endothelial growth factor A (VEGF) ( a ) and angiopoietin 1 (Ang-1) ( b ). Data presented as mean ± standard deviation. * p ≤ 0.05, *** p ≤ 0.001, n = 4. d days

    Article Snippet: The expression of human VEGF and Ang-1 in supernatants was then quantified using ELISA kits according to the manufacturer’s protocol (R & D Systems).

    Techniques: Enzyme-linked Immunosorbent Assay, Expressing, Standard Deviation

    Real-time reverse transcription-polymerase chain reaction (RT-PCR). a–g Representative graphs showing expression of the angiogenesis-related genes vascular endothelial growth factor A (VEGF-A) ( a ), angiopoietin 2 (Ang-2) ( b ), platelet endothelial cell adhesion molecule 1 (PECAM-1) ( c ), fibroblast growth factor (FGF) ( d ), angiopoietin 1 (Ang-1) ( e ), von Willebrand factor (vWF) ( f ) and platelet-derived growth factor (PDGF) ( g ) in human umbilical vein cord endothelial cells (HUVECs) grown under different culture conditions at 2, 3 and 7 days. Data normalized to glyceraldehyde-3-phosphate dehydrogenase (GAPDH) as housekeeping gene and presented as mean ± SEM. * p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001, n = 4. CM conditioned medium, EGM-2 Endothelial Growth Medium-2, d days

    Journal: Stem Cell Research & Therapy

    Article Title: Human dental pulp stromal cell conditioned medium alters endothelial cell behavior

    doi: 10.1186/s13287-018-0815-3

    Figure Lengend Snippet: Real-time reverse transcription-polymerase chain reaction (RT-PCR). a–g Representative graphs showing expression of the angiogenesis-related genes vascular endothelial growth factor A (VEGF-A) ( a ), angiopoietin 2 (Ang-2) ( b ), platelet endothelial cell adhesion molecule 1 (PECAM-1) ( c ), fibroblast growth factor (FGF) ( d ), angiopoietin 1 (Ang-1) ( e ), von Willebrand factor (vWF) ( f ) and platelet-derived growth factor (PDGF) ( g ) in human umbilical vein cord endothelial cells (HUVECs) grown under different culture conditions at 2, 3 and 7 days. Data normalized to glyceraldehyde-3-phosphate dehydrogenase (GAPDH) as housekeeping gene and presented as mean ± SEM. * p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001, n = 4. CM conditioned medium, EGM-2 Endothelial Growth Medium-2, d days

    Article Snippet: The expression of human VEGF and Ang-1 in supernatants was then quantified using ELISA kits according to the manufacturer’s protocol (R & D Systems).

    Techniques: Reverse Transcription Polymerase Chain Reaction, Expressing, Derivative Assay

    3.2. Association of extracellular histones, HMGB1, sTM, and IL-6 with ALI

    Journal: Medicine

    Article Title: Damage-associated molecular patterns in intensive care unit patients with acute liver injuries

    doi: 10.1097/MD.0000000000012780

    Figure Lengend Snippet: 3.2. Association of extracellular histones, HMGB1, sTM, and IL-6 with ALI

    Article Snippet: Plasma concentrations of soluble thrombomodulin (sTM) and interleukin-6 (IL-6) were determined using commercially available ELISA kits against human thrombomodulin and human IL-6, respectively (Quantikine; R & D Systems, Minneapolis, MN).

    Techniques:

    3.2. Association of extracellular histones, HMGB1, sTM, and IL-6 with ALI

    Journal: Medicine

    Article Title: Damage-associated molecular patterns in intensive care unit patients with acute liver injuries

    doi: 10.1097/MD.0000000000012780

    Figure Lengend Snippet: 3.2. Association of extracellular histones, HMGB1, sTM, and IL-6 with ALI

    Article Snippet: Plasma concentrations of soluble thrombomodulin (sTM) and interleukin-6 (IL-6) were determined using commercially available ELISA kits against human thrombomodulin and human IL-6, respectively (Quantikine; R & D Systems, Minneapolis, MN).

    Techniques: