recombinant human rh il 17a  (R&D Systems)

 
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    Name:
    Recombinant Human IL 17A Protein CF
    Description:
    The Recombinant Human IL 17A Protein from R D Systems is derived from E coli The Recombinant Human IL 17A Protein has been validated for the following applications Bioactivity
    Catalog Number:
    317-ILB-050
    Price:
    329
    Category:
    Proteins and Enzymes
    Source:
    E. coli-derived Recombinant Human IL-17A Protein
    Applications:
    Bioactivity
    Purity:
    >97%, by SDS-PAGE under reducing conditions and visualized by silver stain.
    Conjugate:
    Unconjugated
    Size:
    50 ug
    Buy from Supplier


    Structured Review

    R&D Systems recombinant human rh il 17a
    JNK1-dependent IL-17 and TGF-β signaling. The binding of <t>IL-17A/F</t> to the IL-17RA/IL-17RC receptor facilitates the recruitment of ACT1 to the receptor, which mediates the activation of JNK1, ERK, p38, and NF-κB (p65/p50) signaling, leading to the production of pro-inflammatory cytokines and chemokines (e.g. CXCL1 , IL6 ). Similarly, TGF-β binds to its receptor (TGFBR1/TGFBR2), leading to the activation of JNK1, ERK, p38, and SMAD (SMAD2/3/4) signaling. This pathway ultimately results in the production of extracellular matrix proteins and regulators (e.g. FN1 , IL11 ). The mutation (yellow star) in JNK1 impairs the JNK1-dependent activation of downstream AP-1 (c-Jun/ATF-2), thereby reducing the JNK1-dependent cellular responses to IL-17 and TGF-β.
    The Recombinant Human IL 17A Protein from R D Systems is derived from E coli The Recombinant Human IL 17A Protein has been validated for the following applications Bioactivity
    https://www.bioz.com/result/recombinant human rh il 17a/product/R&D Systems
    Average 92 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    recombinant human rh il 17a - by Bioz Stars, 2021-09
    92/100 stars

    Images

    1) Product Images from "Chronic mucocutaneous candidiasis and connective tissue disorder in humans with impaired JNK1-dependent responses to IL-17A/F and TGF-β"

    Article Title: Chronic mucocutaneous candidiasis and connective tissue disorder in humans with impaired JNK1-dependent responses to IL-17A/F and TGF-β

    Journal: Science immunology

    doi: 10.1126/sciimmunol.aax7965

    JNK1-dependent IL-17 and TGF-β signaling. The binding of IL-17A/F to the IL-17RA/IL-17RC receptor facilitates the recruitment of ACT1 to the receptor, which mediates the activation of JNK1, ERK, p38, and NF-κB (p65/p50) signaling, leading to the production of pro-inflammatory cytokines and chemokines (e.g. CXCL1 , IL6 ). Similarly, TGF-β binds to its receptor (TGFBR1/TGFBR2), leading to the activation of JNK1, ERK, p38, and SMAD (SMAD2/3/4) signaling. This pathway ultimately results in the production of extracellular matrix proteins and regulators (e.g. FN1 , IL11 ). The mutation (yellow star) in JNK1 impairs the JNK1-dependent activation of downstream AP-1 (c-Jun/ATF-2), thereby reducing the JNK1-dependent cellular responses to IL-17 and TGF-β.
    Figure Legend Snippet: JNK1-dependent IL-17 and TGF-β signaling. The binding of IL-17A/F to the IL-17RA/IL-17RC receptor facilitates the recruitment of ACT1 to the receptor, which mediates the activation of JNK1, ERK, p38, and NF-κB (p65/p50) signaling, leading to the production of pro-inflammatory cytokines and chemokines (e.g. CXCL1 , IL6 ). Similarly, TGF-β binds to its receptor (TGFBR1/TGFBR2), leading to the activation of JNK1, ERK, p38, and SMAD (SMAD2/3/4) signaling. This pathway ultimately results in the production of extracellular matrix proteins and regulators (e.g. FN1 , IL11 ). The mutation (yellow star) in JNK1 impairs the JNK1-dependent activation of downstream AP-1 (c-Jun/ATF-2), thereby reducing the JNK1-dependent cellular responses to IL-17 and TGF-β.

    Techniques Used: Binding Assay, Activation Assay, Mutagenesis

    The MAPK8 variant impairs fibroblast responses to IL-17A/F. ( A ) Production of GRO-α (top panel) and IL-6 (bottom panel) by SV40-fibroblasts from healthy controls (C1 and C2), patients (P2 and P3), and an IL-17RA-deficient ( IL17RA −/− ) stimulated with IL-17A, IL-17F, or IL-17A/F (10, 100, or 1000 ng/mL) for 24 h. ( B ) Production of GRO-α (top panel) and IL-6 (bottom panel) by SV40-fibroblasts from healthy controls (C1 and C2), patients (P2 and P3), and a NEMO-deficient ( NEMO −/− ) stimulated with TNF-α (20 ng/mL) or IL-1β (10 ng/mL) for 24 h. ( C ) Production of GRO-α (top panel) and IL-6 (bottom panel) by SV40-fibroblasts from healthy controls (C1 and C2) and patients (P2 and P3) transfected with empty vector (EV) or plasmids encoding WT JNK1α1 (α1), JNK1α2 (α2), JNK1β1 (β1), JNK1β2 (β2), all four isoforms (α1/α2/β1/β2), JNK1ES (ES), or JNK1IR (IR), in the presence of IL-17A (100 ng/mL), for 24 h. ( D ) Production of GRO-α (left panel) and IL-6 (right panel) by primary fibroblasts from healthy controls (C1, C2, and C3) transfected with control siRNA (50 nM) or MAPK8 siRNA (50 nM) for 24 h and then stimulated with IL-17A (100 ng/mL) for an additional 24 h. The values shown are the means ± SEM of three independent experiments ( A-D ). *, P
    Figure Legend Snippet: The MAPK8 variant impairs fibroblast responses to IL-17A/F. ( A ) Production of GRO-α (top panel) and IL-6 (bottom panel) by SV40-fibroblasts from healthy controls (C1 and C2), patients (P2 and P3), and an IL-17RA-deficient ( IL17RA −/− ) stimulated with IL-17A, IL-17F, or IL-17A/F (10, 100, or 1000 ng/mL) for 24 h. ( B ) Production of GRO-α (top panel) and IL-6 (bottom panel) by SV40-fibroblasts from healthy controls (C1 and C2), patients (P2 and P3), and a NEMO-deficient ( NEMO −/− ) stimulated with TNF-α (20 ng/mL) or IL-1β (10 ng/mL) for 24 h. ( C ) Production of GRO-α (top panel) and IL-6 (bottom panel) by SV40-fibroblasts from healthy controls (C1 and C2) and patients (P2 and P3) transfected with empty vector (EV) or plasmids encoding WT JNK1α1 (α1), JNK1α2 (α2), JNK1β1 (β1), JNK1β2 (β2), all four isoforms (α1/α2/β1/β2), JNK1ES (ES), or JNK1IR (IR), in the presence of IL-17A (100 ng/mL), for 24 h. ( D ) Production of GRO-α (left panel) and IL-6 (right panel) by primary fibroblasts from healthy controls (C1, C2, and C3) transfected with control siRNA (50 nM) or MAPK8 siRNA (50 nM) for 24 h and then stimulated with IL-17A (100 ng/mL) for an additional 24 h. The values shown are the means ± SEM of three independent experiments ( A-D ). *, P

    Techniques Used: Variant Assay, Transfection, Plasmid Preparation

    Compromised T-cell differentiation in the patients. ( A ) Percentage of total, naïve (CCR7 + CD45RA + ), central memory (CM; CCR7 + CD45RA − ), effector memory (EM; CCR7 − CD45RA − ), or CD45RA + effector memory (EMRA; CCR7 − CD45RA + ) CD4 + and CD8 + T cells from healthy controls ( n =40) and patients (P2 and P3). ( B ) Frequency of T H 1 (CXCR5 − CXCR3 + CCR6 − ), T H 2 (CXCR5 − CXCR3 − CCR6 − CCR4 + ), T H 17 (CXCR5 − CXCR3 − CCR6 + CCR4 + ), T H 1* (CXCR5 − CXCR3 + CCR6 + CCR4 + ), T FH (CXCR5 + ), and T reg (CD25 + FOXP3 + ) subsets among CD4 + T cells from healthy controls (T H 1, T H 2, T H 17, T H 1*, and T FH , n =34; T reg , n =17) and patients (P2 and P3). ( C ) Production of IL-17A and IL-22 by whole blood from healthy controls ( n =33) and patients (P2 and P3) after stimulation with PMA plus ionomycin for 24 h. ( D ) Percentage of IL-17A + , IL-17F + , and IFN-γ + cells among memory CD4 + T cells from healthy controls ( n =36) and patients (P2 and P3) activated by T-cell activation and expansion (TAE) beads or PMA plus ionomycin (P/I) for 12 h. ( E ) Cytokine production by naïve CD4 + T cells from healthy controls ( n =8) and patients (P2 and P3) cultured under T H 0-, T H 17- or T H 1-polarizing conditions. ( F and G ) Frequency of IL-17A + and IFN-γ + cells among naïve ( F ) and memory ( G ) CD4 + T cells from healthy controls ( n =10) and patients (P2 and P3) cultured under T H 0-, T H 17- or T H 1-polarizing conditions. C, Healthy controls; P, P2 and P3. Horizontal bars represent median values ( A-G) . *, P
    Figure Legend Snippet: Compromised T-cell differentiation in the patients. ( A ) Percentage of total, naïve (CCR7 + CD45RA + ), central memory (CM; CCR7 + CD45RA − ), effector memory (EM; CCR7 − CD45RA − ), or CD45RA + effector memory (EMRA; CCR7 − CD45RA + ) CD4 + and CD8 + T cells from healthy controls ( n =40) and patients (P2 and P3). ( B ) Frequency of T H 1 (CXCR5 − CXCR3 + CCR6 − ), T H 2 (CXCR5 − CXCR3 − CCR6 − CCR4 + ), T H 17 (CXCR5 − CXCR3 − CCR6 + CCR4 + ), T H 1* (CXCR5 − CXCR3 + CCR6 + CCR4 + ), T FH (CXCR5 + ), and T reg (CD25 + FOXP3 + ) subsets among CD4 + T cells from healthy controls (T H 1, T H 2, T H 17, T H 1*, and T FH , n =34; T reg , n =17) and patients (P2 and P3). ( C ) Production of IL-17A and IL-22 by whole blood from healthy controls ( n =33) and patients (P2 and P3) after stimulation with PMA plus ionomycin for 24 h. ( D ) Percentage of IL-17A + , IL-17F + , and IFN-γ + cells among memory CD4 + T cells from healthy controls ( n =36) and patients (P2 and P3) activated by T-cell activation and expansion (TAE) beads or PMA plus ionomycin (P/I) for 12 h. ( E ) Cytokine production by naïve CD4 + T cells from healthy controls ( n =8) and patients (P2 and P3) cultured under T H 0-, T H 17- or T H 1-polarizing conditions. ( F and G ) Frequency of IL-17A + and IFN-γ + cells among naïve ( F ) and memory ( G ) CD4 + T cells from healthy controls ( n =10) and patients (P2 and P3) cultured under T H 0-, T H 17- or T H 1-polarizing conditions. C, Healthy controls; P, P2 and P3. Horizontal bars represent median values ( A-G) . *, P

    Techniques Used: Cell Differentiation, Activation Assay, Cell Culture

    2) Product Images from "Transcriptome profiling unveils the role of cholesterol in IL-17A signaling in psoriasis"

    Article Title: Transcriptome profiling unveils the role of cholesterol in IL-17A signaling in psoriasis

    Journal: Scientific Reports

    doi: 10.1038/srep19295

    Validation of Micro-array data in human primary keratinocytes (NHEK) and HaCaT. ( A ) Estimation of total cholesterol content in IL-17A stimulated keratinocytes NHEK and HaCaT cells (normalized to total protein content) as well as in psoriatic lesional and non-lesional skin (normalized to tissue weight). (B) Real Time PCR analysis to determine the expression of SREBP transcriptional factors as well as their downstream genes HMGCR, FASN and ACACA in keratinocytes post IL-17A stimulation. The graph represents fold change in gene expression normalized to 18s rRNA expression. (C) Western blot analysis for the expression of genes involved in cholesterol biosynthesis (SREBP2, HMGCR) and fatty acid synthesis (SREBP1, FASN, ACACA) upon IL-17A treatment in keratinocytes. GAPDH was used as a loading control for densitometric analysis. Graph represents the fold change in protein expression compared to unstimulated cells. The data is expressed as the mean ± S.D. of 3 independent experiments. * indicates p-value (≤0.05) in comparison to unstimulated cells.
    Figure Legend Snippet: Validation of Micro-array data in human primary keratinocytes (NHEK) and HaCaT. ( A ) Estimation of total cholesterol content in IL-17A stimulated keratinocytes NHEK and HaCaT cells (normalized to total protein content) as well as in psoriatic lesional and non-lesional skin (normalized to tissue weight). (B) Real Time PCR analysis to determine the expression of SREBP transcriptional factors as well as their downstream genes HMGCR, FASN and ACACA in keratinocytes post IL-17A stimulation. The graph represents fold change in gene expression normalized to 18s rRNA expression. (C) Western blot analysis for the expression of genes involved in cholesterol biosynthesis (SREBP2, HMGCR) and fatty acid synthesis (SREBP1, FASN, ACACA) upon IL-17A treatment in keratinocytes. GAPDH was used as a loading control for densitometric analysis. Graph represents the fold change in protein expression compared to unstimulated cells. The data is expressed as the mean ± S.D. of 3 independent experiments. * indicates p-value (≤0.05) in comparison to unstimulated cells.

    Techniques Used: Microarray, Real-time Polymerase Chain Reaction, Expressing, Western Blot

    Gene expression profiling in IL-17A stimulated human primary keratinocytes and Ingenuity Pathway Analysis (IPA) summary. ( A ) Schema for Illumina Gene expression profiling on un-stimulated and IL-17A stimulated primary keratinocytes at 6h and data was average normalized and list of 528 differentially expressed genes were obtained with detection p-value (≤0.01) and differential p-value of (≤0.05). (B) List of Top five diseases and canonical pathways along with their respective ratio and p-values obtained from IPA analysis. Two key transcription factors SREBP1 and SREBP2 found to have a negative activation z-score as calculated by IPA.
    Figure Legend Snippet: Gene expression profiling in IL-17A stimulated human primary keratinocytes and Ingenuity Pathway Analysis (IPA) summary. ( A ) Schema for Illumina Gene expression profiling on un-stimulated and IL-17A stimulated primary keratinocytes at 6h and data was average normalized and list of 528 differentially expressed genes were obtained with detection p-value (≤0.01) and differential p-value of (≤0.05). (B) List of Top five diseases and canonical pathways along with their respective ratio and p-values obtained from IPA analysis. Two key transcription factors SREBP1 and SREBP2 found to have a negative activation z-score as calculated by IPA.

    Techniques Used: Expressing, Indirect Immunoperoxidase Assay, Activation Assay

    Cellular cholesterol depletion inhibits IL-17A signaling. ( A ) Analysis of CCL20, IL-8 and S100A7 expression in cholesterol depleted (MBCD) and replenished (MC) cells post IL-17A stimulation by Q-RT PCR. Graph represents the fold change in gene expression compared to unstimulated cells. The data is expressed as the mean ± S.D. of 3 independent experiments with ***p-value (≤0.001), **p-value (≤0.01), *p-value (≤0.05). (B) Localization of NF-κB p65 visualized 1h post IL-17A stimulation in cholesterol depleted and replenished HaCaT cells under fluorescence microscope after immunofluorescence staining with NF-κB p65 antibody (green). In addition, the cells were stained with DAPI to visualize nuclei (blue). Scale bar = 50 um.
    Figure Legend Snippet: Cellular cholesterol depletion inhibits IL-17A signaling. ( A ) Analysis of CCL20, IL-8 and S100A7 expression in cholesterol depleted (MBCD) and replenished (MC) cells post IL-17A stimulation by Q-RT PCR. Graph represents the fold change in gene expression compared to unstimulated cells. The data is expressed as the mean ± S.D. of 3 independent experiments with ***p-value (≤0.001), **p-value (≤0.01), *p-value (≤0.05). (B) Localization of NF-κB p65 visualized 1h post IL-17A stimulation in cholesterol depleted and replenished HaCaT cells under fluorescence microscope after immunofluorescence staining with NF-κB p65 antibody (green). In addition, the cells were stained with DAPI to visualize nuclei (blue). Scale bar = 50 um.

    Techniques Used: Expressing, Reverse Transcription Polymerase Chain Reaction, Fluorescence, Microscopy, Immunofluorescence, Staining

    Central role of cholesterol homeostasis in IL-17A function. IL-17A stimulated keratinocytes showed elevated levels of cellular cholesterol. This increased cellular cholesterol by a feedback mechanism may be involved in inhibiting the cholesterol and fatty acid synthesis machinery via SREBP family of transcriptional factors. Elevated cholesterol also regulates the expression of CCL20, IL-8 and S100A7 in keratinocytes via activation of NF-κB and thus may play a crucial role in IL-17A induced psoriatic inflammation.
    Figure Legend Snippet: Central role of cholesterol homeostasis in IL-17A function. IL-17A stimulated keratinocytes showed elevated levels of cellular cholesterol. This increased cellular cholesterol by a feedback mechanism may be involved in inhibiting the cholesterol and fatty acid synthesis machinery via SREBP family of transcriptional factors. Elevated cholesterol also regulates the expression of CCL20, IL-8 and S100A7 in keratinocytes via activation of NF-κB and thus may play a crucial role in IL-17A induced psoriatic inflammation.

    Techniques Used: Expressing, Activation Assay

    Cytokine profiling in psoriatic lesional and non-lesional epidermis. ( A ) Cytokine PCR array for lesional (1–7) and non-lesional (NL- average of expression values of 4 tissues) skin biopsies. Heat map depicts the relative expression values in psoriatic lesional (n = 7) and non- lesional skin (n = 4) for the 84 cytokines. (B) Expression of Th17 related cytokines including TNF-α, IL-24, IL-21, IL-17A, IL-17B, IL-17C, IL-22, IL-23 and IL-16 were measured by Q-RT PCR and represented as a dot plot of relative expression (Delta Ct) values for both non-lesional (n = 5) and lesional skin (n=10). Significant ***p-value (≤0.001), **p-value (≤0.01), *p-value (≤0.05) was calculated using Student’s two-tailed t test.
    Figure Legend Snippet: Cytokine profiling in psoriatic lesional and non-lesional epidermis. ( A ) Cytokine PCR array for lesional (1–7) and non-lesional (NL- average of expression values of 4 tissues) skin biopsies. Heat map depicts the relative expression values in psoriatic lesional (n = 7) and non- lesional skin (n = 4) for the 84 cytokines. (B) Expression of Th17 related cytokines including TNF-α, IL-24, IL-21, IL-17A, IL-17B, IL-17C, IL-22, IL-23 and IL-16 were measured by Q-RT PCR and represented as a dot plot of relative expression (Delta Ct) values for both non-lesional (n = 5) and lesional skin (n=10). Significant ***p-value (≤0.001), **p-value (≤0.01), *p-value (≤0.05) was calculated using Student’s two-tailed t test.

    Techniques Used: Polymerase Chain Reaction, Expressing, Reverse Transcription Polymerase Chain Reaction, Two Tailed Test

    3) Product Images from "Chronic mucocutaneous candidiasis and connective tissue disorder in humans with impaired JNK1-dependent responses to IL-17A/F and TGF-β"

    Article Title: Chronic mucocutaneous candidiasis and connective tissue disorder in humans with impaired JNK1-dependent responses to IL-17A/F and TGF-β

    Journal: Science immunology

    doi: 10.1126/sciimmunol.aax7965

    JNK1-dependent IL-17 and TGF-β signaling. The binding of IL-17A/F to the IL-17RA/IL-17RC receptor facilitates the recruitment of ACT1 to the receptor, which mediates the activation of JNK1, ERK, p38, and NF-κB (p65/p50) signaling, leading to the production of pro-inflammatory cytokines and chemokines (e.g. CXCL1 , IL6 ). Similarly, TGF-β binds to its receptor (TGFBR1/TGFBR2), leading to the activation of JNK1, ERK, p38, and SMAD (SMAD2/3/4) signaling. This pathway ultimately results in the production of extracellular matrix proteins and regulators (e.g. FN1 , IL11 ). The mutation (yellow star) in JNK1 impairs the JNK1-dependent activation of downstream AP-1 (c-Jun/ATF-2), thereby reducing the JNK1-dependent cellular responses to IL-17 and TGF-β.
    Figure Legend Snippet: JNK1-dependent IL-17 and TGF-β signaling. The binding of IL-17A/F to the IL-17RA/IL-17RC receptor facilitates the recruitment of ACT1 to the receptor, which mediates the activation of JNK1, ERK, p38, and NF-κB (p65/p50) signaling, leading to the production of pro-inflammatory cytokines and chemokines (e.g. CXCL1 , IL6 ). Similarly, TGF-β binds to its receptor (TGFBR1/TGFBR2), leading to the activation of JNK1, ERK, p38, and SMAD (SMAD2/3/4) signaling. This pathway ultimately results in the production of extracellular matrix proteins and regulators (e.g. FN1 , IL11 ). The mutation (yellow star) in JNK1 impairs the JNK1-dependent activation of downstream AP-1 (c-Jun/ATF-2), thereby reducing the JNK1-dependent cellular responses to IL-17 and TGF-β.

    Techniques Used: Binding Assay, Activation Assay, Mutagenesis

    The MAPK8 variant impairs fibroblast responses to IL-17A/F. ( A ) Production of GRO-α (top panel) and IL-6 (bottom panel) by SV40-fibroblasts from healthy controls (C1 and C2), patients (P2 and P3), and an IL-17RA-deficient ( IL17RA −/− ) stimulated with IL-17A, IL-17F, or IL-17A/F (10, 100, or 1000 ng/mL) for 24 h. ( B ) Production of GRO-α (top panel) and IL-6 (bottom panel) by SV40-fibroblasts from healthy controls (C1 and C2), patients (P2 and P3), and a NEMO-deficient ( NEMO −/− ) stimulated with TNF-α (20 ng/mL) or IL-1β (10 ng/mL) for 24 h. ( C ) Production of GRO-α (top panel) and IL-6 (bottom panel) by SV40-fibroblasts from healthy controls (C1 and C2) and patients (P2 and P3) transfected with empty vector (EV) or plasmids encoding WT JNK1α1 (α1), JNK1α2 (α2), JNK1β1 (β1), JNK1β2 (β2), all four isoforms (α1/α2/β1/β2), JNK1ES (ES), or JNK1IR (IR), in the presence of IL-17A (100 ng/mL), for 24 h. ( D ) Production of GRO-α (left panel) and IL-6 (right panel) by primary fibroblasts from healthy controls (C1, C2, and C3) transfected with control siRNA (50 nM) or MAPK8 siRNA (50 nM) for 24 h and then stimulated with IL-17A (100 ng/mL) for an additional 24 h. The values shown are the means ± SEM of three independent experiments ( A-D ). *, P
    Figure Legend Snippet: The MAPK8 variant impairs fibroblast responses to IL-17A/F. ( A ) Production of GRO-α (top panel) and IL-6 (bottom panel) by SV40-fibroblasts from healthy controls (C1 and C2), patients (P2 and P3), and an IL-17RA-deficient ( IL17RA −/− ) stimulated with IL-17A, IL-17F, or IL-17A/F (10, 100, or 1000 ng/mL) for 24 h. ( B ) Production of GRO-α (top panel) and IL-6 (bottom panel) by SV40-fibroblasts from healthy controls (C1 and C2), patients (P2 and P3), and a NEMO-deficient ( NEMO −/− ) stimulated with TNF-α (20 ng/mL) or IL-1β (10 ng/mL) for 24 h. ( C ) Production of GRO-α (top panel) and IL-6 (bottom panel) by SV40-fibroblasts from healthy controls (C1 and C2) and patients (P2 and P3) transfected with empty vector (EV) or plasmids encoding WT JNK1α1 (α1), JNK1α2 (α2), JNK1β1 (β1), JNK1β2 (β2), all four isoforms (α1/α2/β1/β2), JNK1ES (ES), or JNK1IR (IR), in the presence of IL-17A (100 ng/mL), for 24 h. ( D ) Production of GRO-α (left panel) and IL-6 (right panel) by primary fibroblasts from healthy controls (C1, C2, and C3) transfected with control siRNA (50 nM) or MAPK8 siRNA (50 nM) for 24 h and then stimulated with IL-17A (100 ng/mL) for an additional 24 h. The values shown are the means ± SEM of three independent experiments ( A-D ). *, P

    Techniques Used: Variant Assay, Transfection, Plasmid Preparation

    Compromised T-cell differentiation in the patients. ( A ) Percentage of total, naïve (CCR7 + CD45RA + ), central memory (CM; CCR7 + CD45RA − ), effector memory (EM; CCR7 − CD45RA − ), or CD45RA + effector memory (EMRA; CCR7 − CD45RA + ) CD4 + and CD8 + T cells from healthy controls ( n =40) and patients (P2 and P3). ( B ) Frequency of T H 1 (CXCR5 − CXCR3 + CCR6 − ), T H 2 (CXCR5 − CXCR3 − CCR6 − CCR4 + ), T H 17 (CXCR5 − CXCR3 − CCR6 + CCR4 + ), T H 1* (CXCR5 − CXCR3 + CCR6 + CCR4 + ), T FH (CXCR5 + ), and T reg (CD25 + FOXP3 + ) subsets among CD4 + T cells from healthy controls (T H 1, T H 2, T H 17, T H 1*, and T FH , n =34; T reg , n =17) and patients (P2 and P3). ( C ) Production of IL-17A and IL-22 by whole blood from healthy controls ( n =33) and patients (P2 and P3) after stimulation with PMA plus ionomycin for 24 h. ( D ) Percentage of IL-17A + , IL-17F + , and IFN-γ + cells among memory CD4 + T cells from healthy controls ( n =36) and patients (P2 and P3) activated by T-cell activation and expansion (TAE) beads or PMA plus ionomycin (P/I) for 12 h. ( E ) Cytokine production by naïve CD4 + T cells from healthy controls ( n =8) and patients (P2 and P3) cultured under T H 0-, T H 17- or T H 1-polarizing conditions. ( F and G ) Frequency of IL-17A + and IFN-γ + cells among naïve ( F ) and memory ( G ) CD4 + T cells from healthy controls ( n =10) and patients (P2 and P3) cultured under T H 0-, T H 17- or T H 1-polarizing conditions. C, Healthy controls; P, P2 and P3. Horizontal bars represent median values ( A-G) . *, P
    Figure Legend Snippet: Compromised T-cell differentiation in the patients. ( A ) Percentage of total, naïve (CCR7 + CD45RA + ), central memory (CM; CCR7 + CD45RA − ), effector memory (EM; CCR7 − CD45RA − ), or CD45RA + effector memory (EMRA; CCR7 − CD45RA + ) CD4 + and CD8 + T cells from healthy controls ( n =40) and patients (P2 and P3). ( B ) Frequency of T H 1 (CXCR5 − CXCR3 + CCR6 − ), T H 2 (CXCR5 − CXCR3 − CCR6 − CCR4 + ), T H 17 (CXCR5 − CXCR3 − CCR6 + CCR4 + ), T H 1* (CXCR5 − CXCR3 + CCR6 + CCR4 + ), T FH (CXCR5 + ), and T reg (CD25 + FOXP3 + ) subsets among CD4 + T cells from healthy controls (T H 1, T H 2, T H 17, T H 1*, and T FH , n =34; T reg , n =17) and patients (P2 and P3). ( C ) Production of IL-17A and IL-22 by whole blood from healthy controls ( n =33) and patients (P2 and P3) after stimulation with PMA plus ionomycin for 24 h. ( D ) Percentage of IL-17A + , IL-17F + , and IFN-γ + cells among memory CD4 + T cells from healthy controls ( n =36) and patients (P2 and P3) activated by T-cell activation and expansion (TAE) beads or PMA plus ionomycin (P/I) for 12 h. ( E ) Cytokine production by naïve CD4 + T cells from healthy controls ( n =8) and patients (P2 and P3) cultured under T H 0-, T H 17- or T H 1-polarizing conditions. ( F and G ) Frequency of IL-17A + and IFN-γ + cells among naïve ( F ) and memory ( G ) CD4 + T cells from healthy controls ( n =10) and patients (P2 and P3) cultured under T H 0-, T H 17- or T H 1-polarizing conditions. C, Healthy controls; P, P2 and P3. Horizontal bars represent median values ( A-G) . *, P

    Techniques Used: Cell Differentiation, Activation Assay, Cell Culture

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    other:

    Article Title: Contribution of IL-17 in Steroid Hyporesponsiveness in Obese Asthmatics Through Dysregulation of Glucocorticoid Receptors α and β
    Article Snippet: Mature adipocytes were starved with DMEM/Ham's F-12 (1:1, v/v) media supplemented with: 0.01 M HEPES pH 7.4, 0.5% fetal bovine serum and 100 U/ml penicillin/streptomycin overnight.

    Incubation:

    Article Title: Immunomodulatory role of Parkinson’s disease 7 in inflammatory bowel disease
    Article Snippet: .. Cells were incubated in transfection reagent for 24 h. After transfection, cells were treated with vehicle or with recombinant human IL-17A (100 ng/ml) for 24 h. ..

    Transfection:

    Article Title: Immunomodulatory role of Parkinson’s disease 7 in inflammatory bowel disease
    Article Snippet: .. Cells were incubated in transfection reagent for 24 h. After transfection, cells were treated with vehicle or with recombinant human IL-17A (100 ng/ml) for 24 h. ..

    Enzyme-linked Immunosorbent Assay:

    Article Title: Circulating IL-17A Levels in Postmenopausal Women with Primary Hyperparathyroidism
    Article Snippet: .. Plasma IL-17A levels were assayed by Human IL-17 Quantikine ELISA kit (R & D System, Cat. D1700; intra-assay CV 4.7%, interassay CV 8.4%, sensitivity < 3.0 pg/ml, by considering an additional point to the low range of the standard curve); plasma OPG was determined by the Human Osteoprotegerin ELISA kit (BioVendor, Cat. RD194003200; intra-assay CV 4.9%, interassay CV 9.0%, sensitivity 0.03 pmol/l) and plasma soluble RANKL by the sRANKL (total) human ELISA kit (BioVendor, Cat. RD193004200R; intra-assay CV 11.5%, inter-assay CV 12.7%, sensitivity 0.4 pmol/l). ..

    Intra Assay:

    Article Title: Circulating IL-17A Levels in Postmenopausal Women with Primary Hyperparathyroidism
    Article Snippet: .. Plasma IL-17A levels were assayed by Human IL-17 Quantikine ELISA kit (R & D System, Cat. D1700; intra-assay CV 4.7%, interassay CV 8.4%, sensitivity < 3.0 pg/ml, by considering an additional point to the low range of the standard curve); plasma OPG was determined by the Human Osteoprotegerin ELISA kit (BioVendor, Cat. RD194003200; intra-assay CV 4.9%, interassay CV 9.0%, sensitivity 0.03 pmol/l) and plasma soluble RANKL by the sRANKL (total) human ELISA kit (BioVendor, Cat. RD193004200R; intra-assay CV 11.5%, inter-assay CV 12.7%, sensitivity 0.4 pmol/l). ..

    Inter Assay:

    Article Title: Circulating IL-17A Levels in Postmenopausal Women with Primary Hyperparathyroidism
    Article Snippet: .. Plasma IL-17A levels were assayed by Human IL-17 Quantikine ELISA kit (R & D System, Cat. D1700; intra-assay CV 4.7%, interassay CV 8.4%, sensitivity < 3.0 pg/ml, by considering an additional point to the low range of the standard curve); plasma OPG was determined by the Human Osteoprotegerin ELISA kit (BioVendor, Cat. RD194003200; intra-assay CV 4.9%, interassay CV 9.0%, sensitivity 0.03 pmol/l) and plasma soluble RANKL by the sRANKL (total) human ELISA kit (BioVendor, Cat. RD193004200R; intra-assay CV 11.5%, inter-assay CV 12.7%, sensitivity 0.4 pmol/l). ..

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  • 92
    R&D Systems recombinant human rh il 17a
    JNK1-dependent IL-17 and TGF-β signaling. The binding of <t>IL-17A/F</t> to the IL-17RA/IL-17RC receptor facilitates the recruitment of ACT1 to the receptor, which mediates the activation of JNK1, ERK, p38, and NF-κB (p65/p50) signaling, leading to the production of pro-inflammatory cytokines and chemokines (e.g. CXCL1 , IL6 ). Similarly, TGF-β binds to its receptor (TGFBR1/TGFBR2), leading to the activation of JNK1, ERK, p38, and SMAD (SMAD2/3/4) signaling. This pathway ultimately results in the production of extracellular matrix proteins and regulators (e.g. FN1 , IL11 ). The mutation (yellow star) in JNK1 impairs the JNK1-dependent activation of downstream AP-1 (c-Jun/ATF-2), thereby reducing the JNK1-dependent cellular responses to IL-17 and TGF-β.
    Recombinant Human Rh Il 17a, supplied by R&D Systems, used in various techniques. Bioz Stars score: 92/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/recombinant human rh il 17a/product/R&D Systems
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    94
    R&D Systems anti il 17 antibody
    Inhibition of SOCE reduces IL-6 production induced by the <t>IL-17/TNFα</t> combination. Myoblasts were stimulated with IL-17 and/or TNFα in presence or not of the SOCE inhibitor 2-APB (10, 25, and 50 μM) or BTP2 (10 and 20 μM) for 48 h. CCL20 and IL-6 secretion by myoblasts was quantified by ELISA (A,B) . Data are the mean of 7 independent experiments ± SEM; ** p
    Anti Il 17 Antibody, supplied by R&D Systems, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti il 17 antibody/product/R&D Systems
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    92
    R&D Systems pe anti human il 17rc
    No induction of IL-17A, IL-17F, and <t>IL-17RC</t> mRNA expression in neutrophils incubated with IL-6 plus IL-23, in combination with inactivated Aspergillus fumigatus hyphae or conidia. Neutrophils (5 × 10 6 /ml) were incubated either with 100 ng/ml rIL-17A for 2 h or with or without 20 µg/ml IL-6 plus 2 µg/ml IL-23 for 1 h, prior to adding, or not, inactivated A. fumigatus conidia (1:5 neutrophils/conidia ratio) and hyphae (1:1 neutrophils/hyphae ratio) for additional 1 h. Neutrophils were then harvested for RNA extraction to evaluate IL-17A (A) , IL-17F (B) , IL-17RC (C) , IL-17RA (D) , and SOCS3 (F) mRNA expression by reverse transcription quantitative real-time PCR. Gene expression data are depicted as mean normalized expression (MNE) units after GAPDH mRNA normalization (mean ± SEM, n = 4). Asterisks stand for significant differences as compared to untreated cells: * P
    Pe Anti Human Il 17rc, supplied by R&D Systems, used in various techniques. Bioz Stars score: 92/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 92 stars, based on 1 article reviews
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    pe anti human il 17rc - by Bioz Stars, 2021-09
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    Image Search Results


    JNK1-dependent IL-17 and TGF-β signaling. The binding of IL-17A/F to the IL-17RA/IL-17RC receptor facilitates the recruitment of ACT1 to the receptor, which mediates the activation of JNK1, ERK, p38, and NF-κB (p65/p50) signaling, leading to the production of pro-inflammatory cytokines and chemokines (e.g. CXCL1 , IL6 ). Similarly, TGF-β binds to its receptor (TGFBR1/TGFBR2), leading to the activation of JNK1, ERK, p38, and SMAD (SMAD2/3/4) signaling. This pathway ultimately results in the production of extracellular matrix proteins and regulators (e.g. FN1 , IL11 ). The mutation (yellow star) in JNK1 impairs the JNK1-dependent activation of downstream AP-1 (c-Jun/ATF-2), thereby reducing the JNK1-dependent cellular responses to IL-17 and TGF-β.

    Journal: Science immunology

    Article Title: Chronic mucocutaneous candidiasis and connective tissue disorder in humans with impaired JNK1-dependent responses to IL-17A/F and TGF-β

    doi: 10.1126/sciimmunol.aax7965

    Figure Lengend Snippet: JNK1-dependent IL-17 and TGF-β signaling. The binding of IL-17A/F to the IL-17RA/IL-17RC receptor facilitates the recruitment of ACT1 to the receptor, which mediates the activation of JNK1, ERK, p38, and NF-κB (p65/p50) signaling, leading to the production of pro-inflammatory cytokines and chemokines (e.g. CXCL1 , IL6 ). Similarly, TGF-β binds to its receptor (TGFBR1/TGFBR2), leading to the activation of JNK1, ERK, p38, and SMAD (SMAD2/3/4) signaling. This pathway ultimately results in the production of extracellular matrix proteins and regulators (e.g. FN1 , IL11 ). The mutation (yellow star) in JNK1 impairs the JNK1-dependent activation of downstream AP-1 (c-Jun/ATF-2), thereby reducing the JNK1-dependent cellular responses to IL-17 and TGF-β.

    Article Snippet: After 24 h, cells were left unstimulated or were stimulated with recombinant human (rh) IL-17A (317-ILB; R & D Systems), rh IL-17F (1335-IL; R & D Systems), rh IL-17A/F (5194-IL; R & D Systems), rh TNF-α (210-TA; R & D Systems), rh IL-1β (201-LB; R & D Systems), rh lymphotoxin α1/β2 (8884-LY; R & D Systems), LTA-SA (tlrl-slta; InvivoGen), Pam3 CSK4 (tlrl-pms; InvivoGen), FSL-1 (tlrl-fsl; InvivoGen), Pam2 CSK4 (tlrl-pm2s-1; InvivoGen), and lipopolysaccharide (LPS) (L9764; Sigma-Aldrich) for a further 24 h. ELISA kits were used to determine the levels of GRO-α (DY275; R & D Systems), IL-6 (88-7066; Invitrogen), and IL-8 (M9318; Sanquin) in the supernatants.

    Techniques: Binding Assay, Activation Assay, Mutagenesis

    The MAPK8 variant impairs fibroblast responses to IL-17A/F. ( A ) Production of GRO-α (top panel) and IL-6 (bottom panel) by SV40-fibroblasts from healthy controls (C1 and C2), patients (P2 and P3), and an IL-17RA-deficient ( IL17RA −/− ) stimulated with IL-17A, IL-17F, or IL-17A/F (10, 100, or 1000 ng/mL) for 24 h. ( B ) Production of GRO-α (top panel) and IL-6 (bottom panel) by SV40-fibroblasts from healthy controls (C1 and C2), patients (P2 and P3), and a NEMO-deficient ( NEMO −/− ) stimulated with TNF-α (20 ng/mL) or IL-1β (10 ng/mL) for 24 h. ( C ) Production of GRO-α (top panel) and IL-6 (bottom panel) by SV40-fibroblasts from healthy controls (C1 and C2) and patients (P2 and P3) transfected with empty vector (EV) or plasmids encoding WT JNK1α1 (α1), JNK1α2 (α2), JNK1β1 (β1), JNK1β2 (β2), all four isoforms (α1/α2/β1/β2), JNK1ES (ES), or JNK1IR (IR), in the presence of IL-17A (100 ng/mL), for 24 h. ( D ) Production of GRO-α (left panel) and IL-6 (right panel) by primary fibroblasts from healthy controls (C1, C2, and C3) transfected with control siRNA (50 nM) or MAPK8 siRNA (50 nM) for 24 h and then stimulated with IL-17A (100 ng/mL) for an additional 24 h. The values shown are the means ± SEM of three independent experiments ( A-D ). *, P

    Journal: Science immunology

    Article Title: Chronic mucocutaneous candidiasis and connective tissue disorder in humans with impaired JNK1-dependent responses to IL-17A/F and TGF-β

    doi: 10.1126/sciimmunol.aax7965

    Figure Lengend Snippet: The MAPK8 variant impairs fibroblast responses to IL-17A/F. ( A ) Production of GRO-α (top panel) and IL-6 (bottom panel) by SV40-fibroblasts from healthy controls (C1 and C2), patients (P2 and P3), and an IL-17RA-deficient ( IL17RA −/− ) stimulated with IL-17A, IL-17F, or IL-17A/F (10, 100, or 1000 ng/mL) for 24 h. ( B ) Production of GRO-α (top panel) and IL-6 (bottom panel) by SV40-fibroblasts from healthy controls (C1 and C2), patients (P2 and P3), and a NEMO-deficient ( NEMO −/− ) stimulated with TNF-α (20 ng/mL) or IL-1β (10 ng/mL) for 24 h. ( C ) Production of GRO-α (top panel) and IL-6 (bottom panel) by SV40-fibroblasts from healthy controls (C1 and C2) and patients (P2 and P3) transfected with empty vector (EV) or plasmids encoding WT JNK1α1 (α1), JNK1α2 (α2), JNK1β1 (β1), JNK1β2 (β2), all four isoforms (α1/α2/β1/β2), JNK1ES (ES), or JNK1IR (IR), in the presence of IL-17A (100 ng/mL), for 24 h. ( D ) Production of GRO-α (left panel) and IL-6 (right panel) by primary fibroblasts from healthy controls (C1, C2, and C3) transfected with control siRNA (50 nM) or MAPK8 siRNA (50 nM) for 24 h and then stimulated with IL-17A (100 ng/mL) for an additional 24 h. The values shown are the means ± SEM of three independent experiments ( A-D ). *, P

    Article Snippet: After 24 h, cells were left unstimulated or were stimulated with recombinant human (rh) IL-17A (317-ILB; R & D Systems), rh IL-17F (1335-IL; R & D Systems), rh IL-17A/F (5194-IL; R & D Systems), rh TNF-α (210-TA; R & D Systems), rh IL-1β (201-LB; R & D Systems), rh lymphotoxin α1/β2 (8884-LY; R & D Systems), LTA-SA (tlrl-slta; InvivoGen), Pam3 CSK4 (tlrl-pms; InvivoGen), FSL-1 (tlrl-fsl; InvivoGen), Pam2 CSK4 (tlrl-pm2s-1; InvivoGen), and lipopolysaccharide (LPS) (L9764; Sigma-Aldrich) for a further 24 h. ELISA kits were used to determine the levels of GRO-α (DY275; R & D Systems), IL-6 (88-7066; Invitrogen), and IL-8 (M9318; Sanquin) in the supernatants.

    Techniques: Variant Assay, Transfection, Plasmid Preparation

    Compromised T-cell differentiation in the patients. ( A ) Percentage of total, naïve (CCR7 + CD45RA + ), central memory (CM; CCR7 + CD45RA − ), effector memory (EM; CCR7 − CD45RA − ), or CD45RA + effector memory (EMRA; CCR7 − CD45RA + ) CD4 + and CD8 + T cells from healthy controls ( n =40) and patients (P2 and P3). ( B ) Frequency of T H 1 (CXCR5 − CXCR3 + CCR6 − ), T H 2 (CXCR5 − CXCR3 − CCR6 − CCR4 + ), T H 17 (CXCR5 − CXCR3 − CCR6 + CCR4 + ), T H 1* (CXCR5 − CXCR3 + CCR6 + CCR4 + ), T FH (CXCR5 + ), and T reg (CD25 + FOXP3 + ) subsets among CD4 + T cells from healthy controls (T H 1, T H 2, T H 17, T H 1*, and T FH , n =34; T reg , n =17) and patients (P2 and P3). ( C ) Production of IL-17A and IL-22 by whole blood from healthy controls ( n =33) and patients (P2 and P3) after stimulation with PMA plus ionomycin for 24 h. ( D ) Percentage of IL-17A + , IL-17F + , and IFN-γ + cells among memory CD4 + T cells from healthy controls ( n =36) and patients (P2 and P3) activated by T-cell activation and expansion (TAE) beads or PMA plus ionomycin (P/I) for 12 h. ( E ) Cytokine production by naïve CD4 + T cells from healthy controls ( n =8) and patients (P2 and P3) cultured under T H 0-, T H 17- or T H 1-polarizing conditions. ( F and G ) Frequency of IL-17A + and IFN-γ + cells among naïve ( F ) and memory ( G ) CD4 + T cells from healthy controls ( n =10) and patients (P2 and P3) cultured under T H 0-, T H 17- or T H 1-polarizing conditions. C, Healthy controls; P, P2 and P3. Horizontal bars represent median values ( A-G) . *, P

    Journal: Science immunology

    Article Title: Chronic mucocutaneous candidiasis and connective tissue disorder in humans with impaired JNK1-dependent responses to IL-17A/F and TGF-β

    doi: 10.1126/sciimmunol.aax7965

    Figure Lengend Snippet: Compromised T-cell differentiation in the patients. ( A ) Percentage of total, naïve (CCR7 + CD45RA + ), central memory (CM; CCR7 + CD45RA − ), effector memory (EM; CCR7 − CD45RA − ), or CD45RA + effector memory (EMRA; CCR7 − CD45RA + ) CD4 + and CD8 + T cells from healthy controls ( n =40) and patients (P2 and P3). ( B ) Frequency of T H 1 (CXCR5 − CXCR3 + CCR6 − ), T H 2 (CXCR5 − CXCR3 − CCR6 − CCR4 + ), T H 17 (CXCR5 − CXCR3 − CCR6 + CCR4 + ), T H 1* (CXCR5 − CXCR3 + CCR6 + CCR4 + ), T FH (CXCR5 + ), and T reg (CD25 + FOXP3 + ) subsets among CD4 + T cells from healthy controls (T H 1, T H 2, T H 17, T H 1*, and T FH , n =34; T reg , n =17) and patients (P2 and P3). ( C ) Production of IL-17A and IL-22 by whole blood from healthy controls ( n =33) and patients (P2 and P3) after stimulation with PMA plus ionomycin for 24 h. ( D ) Percentage of IL-17A + , IL-17F + , and IFN-γ + cells among memory CD4 + T cells from healthy controls ( n =36) and patients (P2 and P3) activated by T-cell activation and expansion (TAE) beads or PMA plus ionomycin (P/I) for 12 h. ( E ) Cytokine production by naïve CD4 + T cells from healthy controls ( n =8) and patients (P2 and P3) cultured under T H 0-, T H 17- or T H 1-polarizing conditions. ( F and G ) Frequency of IL-17A + and IFN-γ + cells among naïve ( F ) and memory ( G ) CD4 + T cells from healthy controls ( n =10) and patients (P2 and P3) cultured under T H 0-, T H 17- or T H 1-polarizing conditions. C, Healthy controls; P, P2 and P3. Horizontal bars represent median values ( A-G) . *, P

    Article Snippet: After 24 h, cells were left unstimulated or were stimulated with recombinant human (rh) IL-17A (317-ILB; R & D Systems), rh IL-17F (1335-IL; R & D Systems), rh IL-17A/F (5194-IL; R & D Systems), rh TNF-α (210-TA; R & D Systems), rh IL-1β (201-LB; R & D Systems), rh lymphotoxin α1/β2 (8884-LY; R & D Systems), LTA-SA (tlrl-slta; InvivoGen), Pam3 CSK4 (tlrl-pms; InvivoGen), FSL-1 (tlrl-fsl; InvivoGen), Pam2 CSK4 (tlrl-pm2s-1; InvivoGen), and lipopolysaccharide (LPS) (L9764; Sigma-Aldrich) for a further 24 h. ELISA kits were used to determine the levels of GRO-α (DY275; R & D Systems), IL-6 (88-7066; Invitrogen), and IL-8 (M9318; Sanquin) in the supernatants.

    Techniques: Cell Differentiation, Activation Assay, Cell Culture

    Validation of Micro-array data in human primary keratinocytes (NHEK) and HaCaT. ( A ) Estimation of total cholesterol content in IL-17A stimulated keratinocytes NHEK and HaCaT cells (normalized to total protein content) as well as in psoriatic lesional and non-lesional skin (normalized to tissue weight). (B) Real Time PCR analysis to determine the expression of SREBP transcriptional factors as well as their downstream genes HMGCR, FASN and ACACA in keratinocytes post IL-17A stimulation. The graph represents fold change in gene expression normalized to 18s rRNA expression. (C) Western blot analysis for the expression of genes involved in cholesterol biosynthesis (SREBP2, HMGCR) and fatty acid synthesis (SREBP1, FASN, ACACA) upon IL-17A treatment in keratinocytes. GAPDH was used as a loading control for densitometric analysis. Graph represents the fold change in protein expression compared to unstimulated cells. The data is expressed as the mean ± S.D. of 3 independent experiments. * indicates p-value (≤0.05) in comparison to unstimulated cells.

    Journal: Scientific Reports

    Article Title: Transcriptome profiling unveils the role of cholesterol in IL-17A signaling in psoriasis

    doi: 10.1038/srep19295

    Figure Lengend Snippet: Validation of Micro-array data in human primary keratinocytes (NHEK) and HaCaT. ( A ) Estimation of total cholesterol content in IL-17A stimulated keratinocytes NHEK and HaCaT cells (normalized to total protein content) as well as in psoriatic lesional and non-lesional skin (normalized to tissue weight). (B) Real Time PCR analysis to determine the expression of SREBP transcriptional factors as well as their downstream genes HMGCR, FASN and ACACA in keratinocytes post IL-17A stimulation. The graph represents fold change in gene expression normalized to 18s rRNA expression. (C) Western blot analysis for the expression of genes involved in cholesterol biosynthesis (SREBP2, HMGCR) and fatty acid synthesis (SREBP1, FASN, ACACA) upon IL-17A treatment in keratinocytes. GAPDH was used as a loading control for densitometric analysis. Graph represents the fold change in protein expression compared to unstimulated cells. The data is expressed as the mean ± S.D. of 3 independent experiments. * indicates p-value (≤0.05) in comparison to unstimulated cells.

    Article Snippet: Keratinocytes were cultured in keratinocyte serum free media supplemented with epidermal growth factor and bovine pituitary extract and once 80% confluent, cells were stimulated with recombinant human (rh) IL-17A (R & D System, Minneapolis, MN) at 100 ng/ml for 6 hours before harvesting for further experiments.

    Techniques: Microarray, Real-time Polymerase Chain Reaction, Expressing, Western Blot

    Gene expression profiling in IL-17A stimulated human primary keratinocytes and Ingenuity Pathway Analysis (IPA) summary. ( A ) Schema for Illumina Gene expression profiling on un-stimulated and IL-17A stimulated primary keratinocytes at 6h and data was average normalized and list of 528 differentially expressed genes were obtained with detection p-value (≤0.01) and differential p-value of (≤0.05). (B) List of Top five diseases and canonical pathways along with their respective ratio and p-values obtained from IPA analysis. Two key transcription factors SREBP1 and SREBP2 found to have a negative activation z-score as calculated by IPA.

    Journal: Scientific Reports

    Article Title: Transcriptome profiling unveils the role of cholesterol in IL-17A signaling in psoriasis

    doi: 10.1038/srep19295

    Figure Lengend Snippet: Gene expression profiling in IL-17A stimulated human primary keratinocytes and Ingenuity Pathway Analysis (IPA) summary. ( A ) Schema for Illumina Gene expression profiling on un-stimulated and IL-17A stimulated primary keratinocytes at 6h and data was average normalized and list of 528 differentially expressed genes were obtained with detection p-value (≤0.01) and differential p-value of (≤0.05). (B) List of Top five diseases and canonical pathways along with their respective ratio and p-values obtained from IPA analysis. Two key transcription factors SREBP1 and SREBP2 found to have a negative activation z-score as calculated by IPA.

    Article Snippet: Keratinocytes were cultured in keratinocyte serum free media supplemented with epidermal growth factor and bovine pituitary extract and once 80% confluent, cells were stimulated with recombinant human (rh) IL-17A (R & D System, Minneapolis, MN) at 100 ng/ml for 6 hours before harvesting for further experiments.

    Techniques: Expressing, Indirect Immunoperoxidase Assay, Activation Assay

    Cellular cholesterol depletion inhibits IL-17A signaling. ( A ) Analysis of CCL20, IL-8 and S100A7 expression in cholesterol depleted (MBCD) and replenished (MC) cells post IL-17A stimulation by Q-RT PCR. Graph represents the fold change in gene expression compared to unstimulated cells. The data is expressed as the mean ± S.D. of 3 independent experiments with ***p-value (≤0.001), **p-value (≤0.01), *p-value (≤0.05). (B) Localization of NF-κB p65 visualized 1h post IL-17A stimulation in cholesterol depleted and replenished HaCaT cells under fluorescence microscope after immunofluorescence staining with NF-κB p65 antibody (green). In addition, the cells were stained with DAPI to visualize nuclei (blue). Scale bar = 50 um.

    Journal: Scientific Reports

    Article Title: Transcriptome profiling unveils the role of cholesterol in IL-17A signaling in psoriasis

    doi: 10.1038/srep19295

    Figure Lengend Snippet: Cellular cholesterol depletion inhibits IL-17A signaling. ( A ) Analysis of CCL20, IL-8 and S100A7 expression in cholesterol depleted (MBCD) and replenished (MC) cells post IL-17A stimulation by Q-RT PCR. Graph represents the fold change in gene expression compared to unstimulated cells. The data is expressed as the mean ± S.D. of 3 independent experiments with ***p-value (≤0.001), **p-value (≤0.01), *p-value (≤0.05). (B) Localization of NF-κB p65 visualized 1h post IL-17A stimulation in cholesterol depleted and replenished HaCaT cells under fluorescence microscope after immunofluorescence staining with NF-κB p65 antibody (green). In addition, the cells were stained with DAPI to visualize nuclei (blue). Scale bar = 50 um.

    Article Snippet: Keratinocytes were cultured in keratinocyte serum free media supplemented with epidermal growth factor and bovine pituitary extract and once 80% confluent, cells were stimulated with recombinant human (rh) IL-17A (R & D System, Minneapolis, MN) at 100 ng/ml for 6 hours before harvesting for further experiments.

    Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Fluorescence, Microscopy, Immunofluorescence, Staining

    Central role of cholesterol homeostasis in IL-17A function. IL-17A stimulated keratinocytes showed elevated levels of cellular cholesterol. This increased cellular cholesterol by a feedback mechanism may be involved in inhibiting the cholesterol and fatty acid synthesis machinery via SREBP family of transcriptional factors. Elevated cholesterol also regulates the expression of CCL20, IL-8 and S100A7 in keratinocytes via activation of NF-κB and thus may play a crucial role in IL-17A induced psoriatic inflammation.

    Journal: Scientific Reports

    Article Title: Transcriptome profiling unveils the role of cholesterol in IL-17A signaling in psoriasis

    doi: 10.1038/srep19295

    Figure Lengend Snippet: Central role of cholesterol homeostasis in IL-17A function. IL-17A stimulated keratinocytes showed elevated levels of cellular cholesterol. This increased cellular cholesterol by a feedback mechanism may be involved in inhibiting the cholesterol and fatty acid synthesis machinery via SREBP family of transcriptional factors. Elevated cholesterol also regulates the expression of CCL20, IL-8 and S100A7 in keratinocytes via activation of NF-κB and thus may play a crucial role in IL-17A induced psoriatic inflammation.

    Article Snippet: Keratinocytes were cultured in keratinocyte serum free media supplemented with epidermal growth factor and bovine pituitary extract and once 80% confluent, cells were stimulated with recombinant human (rh) IL-17A (R & D System, Minneapolis, MN) at 100 ng/ml for 6 hours before harvesting for further experiments.

    Techniques: Expressing, Activation Assay

    Cytokine profiling in psoriatic lesional and non-lesional epidermis. ( A ) Cytokine PCR array for lesional (1–7) and non-lesional (NL- average of expression values of 4 tissues) skin biopsies. Heat map depicts the relative expression values in psoriatic lesional (n = 7) and non- lesional skin (n = 4) for the 84 cytokines. (B) Expression of Th17 related cytokines including TNF-α, IL-24, IL-21, IL-17A, IL-17B, IL-17C, IL-22, IL-23 and IL-16 were measured by Q-RT PCR and represented as a dot plot of relative expression (Delta Ct) values for both non-lesional (n = 5) and lesional skin (n=10). Significant ***p-value (≤0.001), **p-value (≤0.01), *p-value (≤0.05) was calculated using Student’s two-tailed t test.

    Journal: Scientific Reports

    Article Title: Transcriptome profiling unveils the role of cholesterol in IL-17A signaling in psoriasis

    doi: 10.1038/srep19295

    Figure Lengend Snippet: Cytokine profiling in psoriatic lesional and non-lesional epidermis. ( A ) Cytokine PCR array for lesional (1–7) and non-lesional (NL- average of expression values of 4 tissues) skin biopsies. Heat map depicts the relative expression values in psoriatic lesional (n = 7) and non- lesional skin (n = 4) for the 84 cytokines. (B) Expression of Th17 related cytokines including TNF-α, IL-24, IL-21, IL-17A, IL-17B, IL-17C, IL-22, IL-23 and IL-16 were measured by Q-RT PCR and represented as a dot plot of relative expression (Delta Ct) values for both non-lesional (n = 5) and lesional skin (n=10). Significant ***p-value (≤0.001), **p-value (≤0.01), *p-value (≤0.05) was calculated using Student’s two-tailed t test.

    Article Snippet: Keratinocytes were cultured in keratinocyte serum free media supplemented with epidermal growth factor and bovine pituitary extract and once 80% confluent, cells were stimulated with recombinant human (rh) IL-17A (R & D System, Minneapolis, MN) at 100 ng/ml for 6 hours before harvesting for further experiments.

    Techniques: Polymerase Chain Reaction, Expressing, Reverse Transcription Polymerase Chain Reaction, Two Tailed Test

    Inhibition of SOCE reduces IL-6 production induced by the IL-17/TNFα combination. Myoblasts were stimulated with IL-17 and/or TNFα in presence or not of the SOCE inhibitor 2-APB (10, 25, and 50 μM) or BTP2 (10 and 20 μM) for 48 h. CCL20 and IL-6 secretion by myoblasts was quantified by ELISA (A,B) . Data are the mean of 7 independent experiments ± SEM; ** p

    Journal: Frontiers in Immunology

    Article Title: Blockade of Store-Operated Calcium Entry Reduces IL-17/TNF Cytokine-Induced Inflammatory Response in Human Myoblasts

    doi: 10.3389/fimmu.2018.03170

    Figure Lengend Snippet: Inhibition of SOCE reduces IL-6 production induced by the IL-17/TNFα combination. Myoblasts were stimulated with IL-17 and/or TNFα in presence or not of the SOCE inhibitor 2-APB (10, 25, and 50 μM) or BTP2 (10 and 20 μM) for 48 h. CCL20 and IL-6 secretion by myoblasts was quantified by ELISA (A,B) . Data are the mean of 7 independent experiments ± SEM; ** p

    Article Snippet: For the IL-17 and TNFα neutralization assays, PBMCs activated or not with PHA for 24 h were exposed to an anti-IL-17 antibody (R & D Systems) and/or the anti-TNFα antibody infliximab (Merck, Kenilworth, USA) at 10 μg/mL for 3 h before being added to the HepaRG cells.

    Techniques: Inhibition, Enzyme-linked Immunosorbent Assay

    IL-17 and TNFα increase ER stress and mitochondrial ROS in myoblasts. Myoblasts were treated with IL-17 (50 ng/mL) and/or TNFα (1 ng/mL) for 24 h. Expression of BiP/Grp78 protein was measured by western-blot and the band density was normalized with tubulin expression (A,B) . Mitochondrial oxidative stress measurements (ROS) of human myoblasts was measured with the fluorescence intensity of CellRox Dye, using 40x objective of a confocal microscope Nikon A1r, scale bar 70 μm (C,D) . Data are the mean of 4 to 7 independent experiments ± SEM, *** p

    Journal: Frontiers in Immunology

    Article Title: Blockade of Store-Operated Calcium Entry Reduces IL-17/TNF Cytokine-Induced Inflammatory Response in Human Myoblasts

    doi: 10.3389/fimmu.2018.03170

    Figure Lengend Snippet: IL-17 and TNFα increase ER stress and mitochondrial ROS in myoblasts. Myoblasts were treated with IL-17 (50 ng/mL) and/or TNFα (1 ng/mL) for 24 h. Expression of BiP/Grp78 protein was measured by western-blot and the band density was normalized with tubulin expression (A,B) . Mitochondrial oxidative stress measurements (ROS) of human myoblasts was measured with the fluorescence intensity of CellRox Dye, using 40x objective of a confocal microscope Nikon A1r, scale bar 70 μm (C,D) . Data are the mean of 4 to 7 independent experiments ± SEM, *** p

    Article Snippet: For the IL-17 and TNFα neutralization assays, PBMCs activated or not with PHA for 24 h were exposed to an anti-IL-17 antibody (R & D Systems) and/or the anti-TNFα antibody infliximab (Merck, Kenilworth, USA) at 10 μg/mL for 3 h before being added to the HepaRG cells.

    Techniques: Expressing, Western Blot, Fluorescence, Microscopy

    PBMC-myoblast interaction induces a strong production CCL20 and IL-6. PBMC and myoblasts were cultured alone or in co-culture at a ratio of 5 PBMCs for 1 myoblast for 48 h in the presence or not of PHA (5 μg/mL). CCL20 and IL-6 secretion by myoblasts was quantified by ELISA (A–F) . The contribution of direct cell-cell contact was investigated with a cell culture permeable insert (C,D) . PBMCs were pre-incubated for 24 h in presence or not of PHA and then exposed or not to an anti-IL-17 antibody and/or an anti-TNFα antibody for 3 h before being added to the myoblast cultures. Data are expressed as CCL20 and IL-6 supernatant level percentages compared to the non-activated pre-incubated PBMC—myoblast co-cultures (E,F) . Data are the mean of 6–14 independent experiments ± SEM; * p

    Journal: Frontiers in Immunology

    Article Title: Blockade of Store-Operated Calcium Entry Reduces IL-17/TNF Cytokine-Induced Inflammatory Response in Human Myoblasts

    doi: 10.3389/fimmu.2018.03170

    Figure Lengend Snippet: PBMC-myoblast interaction induces a strong production CCL20 and IL-6. PBMC and myoblasts were cultured alone or in co-culture at a ratio of 5 PBMCs for 1 myoblast for 48 h in the presence or not of PHA (5 μg/mL). CCL20 and IL-6 secretion by myoblasts was quantified by ELISA (A–F) . The contribution of direct cell-cell contact was investigated with a cell culture permeable insert (C,D) . PBMCs were pre-incubated for 24 h in presence or not of PHA and then exposed or not to an anti-IL-17 antibody and/or an anti-TNFα antibody for 3 h before being added to the myoblast cultures. Data are expressed as CCL20 and IL-6 supernatant level percentages compared to the non-activated pre-incubated PBMC—myoblast co-cultures (E,F) . Data are the mean of 6–14 independent experiments ± SEM; * p

    Article Snippet: For the IL-17 and TNFα neutralization assays, PBMCs activated or not with PHA for 24 h were exposed to an anti-IL-17 antibody (R & D Systems) and/or the anti-TNFα antibody infliximab (Merck, Kenilworth, USA) at 10 μg/mL for 3 h before being added to the HepaRG cells.

    Techniques: Cell Culture, Co-Culture Assay, Enzyme-linked Immunosorbent Assay, Incubation

    IL-17 and TNFα increase store-operated calcium entry. Myoblasts were treated with IL-17 (50 ng/mL) and/or TNFα (1 ng/mL). mRNA levels of STIM1 and ORAI1 at 12 h was expressed as fold changes compared to control (A,B) . ORAI1 and STIM1 protein was measured by western-blot and the band density was normalized with the tubulin expression. (C–F) Representative image of STIM1 puncta in human myoblast treated with IL-17 (50 ng/mL) and TNFα (1 ng/mL) for 24 h. Image Correlation Spectroscopy (ICS) analysis of STIM1 puncta (left inset) mean density of puncta (μm2). (right inset) mean surface of puncta (puncta/μm2). Data are the mean of 3 independent experiments with cells from 3 different donors ± SEM; * p

    Journal: Frontiers in Immunology

    Article Title: Blockade of Store-Operated Calcium Entry Reduces IL-17/TNF Cytokine-Induced Inflammatory Response in Human Myoblasts

    doi: 10.3389/fimmu.2018.03170

    Figure Lengend Snippet: IL-17 and TNFα increase store-operated calcium entry. Myoblasts were treated with IL-17 (50 ng/mL) and/or TNFα (1 ng/mL). mRNA levels of STIM1 and ORAI1 at 12 h was expressed as fold changes compared to control (A,B) . ORAI1 and STIM1 protein was measured by western-blot and the band density was normalized with the tubulin expression. (C–F) Representative image of STIM1 puncta in human myoblast treated with IL-17 (50 ng/mL) and TNFα (1 ng/mL) for 24 h. Image Correlation Spectroscopy (ICS) analysis of STIM1 puncta (left inset) mean density of puncta (μm2). (right inset) mean surface of puncta (puncta/μm2). Data are the mean of 3 independent experiments with cells from 3 different donors ± SEM; * p

    Article Snippet: For the IL-17 and TNFα neutralization assays, PBMCs activated or not with PHA for 24 h were exposed to an anti-IL-17 antibody (R & D Systems) and/or the anti-TNFα antibody infliximab (Merck, Kenilworth, USA) at 10 μg/mL for 3 h before being added to the HepaRG cells.

    Techniques: Western Blot, Expressing, Spectroscopy

    IL-17 and TNFα mediate muscle damage and weakness through immune and non-immune pathways in myoblasts. The immune cell infiltration in IIM constitutes a local source of cytokines and promotes the cell-cell interactions. IL-17 mainly produced by Th17 cells, and TNFα act in synergy on myoblasts to increase IL-6 and CCL20 secretion. Because IL-6 is involved in the Th17 cell differentiation and CCL20 in dendritic and Th17 cell recruitment, IL-6 and CCL20 mediate a positive feedback loop promoting local IL-17 production. IL-17 and TNFα induce also non-immune pathways with ROS production, ER stress and SOCE activation. The IL-17/TNFα effect of mitochondrial dysfunction, ER stress, and SOCE activation are probably closely linked. SOCE and calcium dysregulation contribute to the IL-6 release induced by IL-17/TNFα. CCL20, chemokine (C-C motif) ligand 20; DC, dendritic cells; ER, endoplasmic reticulum; IL, interleukin; ROS, reactive oxygen species; STIM, stromal interacting molecule; TNFα, tumor necrosis factor-α.

    Journal: Frontiers in Immunology

    Article Title: Blockade of Store-Operated Calcium Entry Reduces IL-17/TNF Cytokine-Induced Inflammatory Response in Human Myoblasts

    doi: 10.3389/fimmu.2018.03170

    Figure Lengend Snippet: IL-17 and TNFα mediate muscle damage and weakness through immune and non-immune pathways in myoblasts. The immune cell infiltration in IIM constitutes a local source of cytokines and promotes the cell-cell interactions. IL-17 mainly produced by Th17 cells, and TNFα act in synergy on myoblasts to increase IL-6 and CCL20 secretion. Because IL-6 is involved in the Th17 cell differentiation and CCL20 in dendritic and Th17 cell recruitment, IL-6 and CCL20 mediate a positive feedback loop promoting local IL-17 production. IL-17 and TNFα induce also non-immune pathways with ROS production, ER stress and SOCE activation. The IL-17/TNFα effect of mitochondrial dysfunction, ER stress, and SOCE activation are probably closely linked. SOCE and calcium dysregulation contribute to the IL-6 release induced by IL-17/TNFα. CCL20, chemokine (C-C motif) ligand 20; DC, dendritic cells; ER, endoplasmic reticulum; IL, interleukin; ROS, reactive oxygen species; STIM, stromal interacting molecule; TNFα, tumor necrosis factor-α.

    Article Snippet: For the IL-17 and TNFα neutralization assays, PBMCs activated or not with PHA for 24 h were exposed to an anti-IL-17 antibody (R & D Systems) and/or the anti-TNFα antibody infliximab (Merck, Kenilworth, USA) at 10 μg/mL for 3 h before being added to the HepaRG cells.

    Techniques: Produced, Activated Clotting Time Assay, Cell Differentiation, Activation Assay

    Synergistic effect of IL-17 and TNFα on the CCL20 and IL-6 production by myoblasts. Myoblasts were treated with IL-17 (50 ng/mL) and/or TNFα (1 ng/mL) for 48 h. IL-6 and CCL20 secretion by myoblasts was quantified by ELISA (A,B) . Data are the mean of 5–8 independent experiments ± SEM; * p

    Journal: Frontiers in Immunology

    Article Title: Blockade of Store-Operated Calcium Entry Reduces IL-17/TNF Cytokine-Induced Inflammatory Response in Human Myoblasts

    doi: 10.3389/fimmu.2018.03170

    Figure Lengend Snippet: Synergistic effect of IL-17 and TNFα on the CCL20 and IL-6 production by myoblasts. Myoblasts were treated with IL-17 (50 ng/mL) and/or TNFα (1 ng/mL) for 48 h. IL-6 and CCL20 secretion by myoblasts was quantified by ELISA (A,B) . Data are the mean of 5–8 independent experiments ± SEM; * p

    Article Snippet: For the IL-17 and TNFα neutralization assays, PBMCs activated or not with PHA for 24 h were exposed to an anti-IL-17 antibody (R & D Systems) and/or the anti-TNFα antibody infliximab (Merck, Kenilworth, USA) at 10 μg/mL for 3 h before being added to the HepaRG cells.

    Techniques: Enzyme-linked Immunosorbent Assay

    No induction of IL-17A, IL-17F, and IL-17RC mRNA expression in neutrophils incubated with IL-6 plus IL-23, in combination with inactivated Aspergillus fumigatus hyphae or conidia. Neutrophils (5 × 10 6 /ml) were incubated either with 100 ng/ml rIL-17A for 2 h or with or without 20 µg/ml IL-6 plus 2 µg/ml IL-23 for 1 h, prior to adding, or not, inactivated A. fumigatus conidia (1:5 neutrophils/conidia ratio) and hyphae (1:1 neutrophils/hyphae ratio) for additional 1 h. Neutrophils were then harvested for RNA extraction to evaluate IL-17A (A) , IL-17F (B) , IL-17RC (C) , IL-17RA (D) , and SOCS3 (F) mRNA expression by reverse transcription quantitative real-time PCR. Gene expression data are depicted as mean normalized expression (MNE) units after GAPDH mRNA normalization (mean ± SEM, n = 4). Asterisks stand for significant differences as compared to untreated cells: * P

    Journal: Frontiers in Immunology

    Article Title: A Reappraisal on the Potential Ability of Human Neutrophils to Express and Produce IL-17 Family Members In Vitro: Failure to Reproducibly Detect It

    doi: 10.3389/fimmu.2018.00795

    Figure Lengend Snippet: No induction of IL-17A, IL-17F, and IL-17RC mRNA expression in neutrophils incubated with IL-6 plus IL-23, in combination with inactivated Aspergillus fumigatus hyphae or conidia. Neutrophils (5 × 10 6 /ml) were incubated either with 100 ng/ml rIL-17A for 2 h or with or without 20 µg/ml IL-6 plus 2 µg/ml IL-23 for 1 h, prior to adding, or not, inactivated A. fumigatus conidia (1:5 neutrophils/conidia ratio) and hyphae (1:1 neutrophils/hyphae ratio) for additional 1 h. Neutrophils were then harvested for RNA extraction to evaluate IL-17A (A) , IL-17F (B) , IL-17RC (C) , IL-17RA (D) , and SOCS3 (F) mRNA expression by reverse transcription quantitative real-time PCR. Gene expression data are depicted as mean normalized expression (MNE) units after GAPDH mRNA normalization (mean ± SEM, n = 4). Asterisks stand for significant differences as compared to untreated cells: * P

    Article Snippet: Neutrophils were then stained for 15 min at T room with: APC anti-human IL-17RA/CD217 (clone 424LTS) and APC mouse IgG1к, as isotype control (clone P3.6.2.8.1) from eBioscience (San Diego, CA, USA); PE anti-human IL-17RC (clone 309822) and mouse PE IgG2B isotype control from R & D systems; PE-vio770 anti-human CD11b (clone ICRF44), FITC anti-human CD66b (clone G10F5), and PerCP-Cy5.6 anti-human CD16 (clone 3G8) from BioLegend (San Diego, CA, USA); APC anti-human CD62L (clone 145/15 Miltenyi Biotec), all at working dilutions specified in the corresponding datasheets.

    Techniques: Expressing, Incubation, RNA Extraction, Real-time Polymerase Chain Reaction

    IL-17A, IL-17F, IL-17RA, IL-17RC, CXCL8, TNFα, and SOCS3 mRNA expression, as well as IL-17R surface expression, in neutrophils from patients with psoriasis. (A) Neutrophils isolated from healthy donors (HDs) ( n = 3) or psoriatic patients ( n = 3) were cultured for 20 h with 100 U/ml IFNγ plus 100 ng/ml LPS, 5 µM R848, or 500 ng/ml IL-17A to evaluate IL-17A, IL-17F, IL-17RA, IL-17RC, CXCL8, TNFα, and SOCS3 mRNA expression by reverse transcription quantitative real-time PCR. Gene expression data are depicted as mean normalized expression (MNE) units after GAPDH mRNA normalization. (B) Surface IL-17RA and IL-17RC expression evaluated by flow cytometry in human neutrophils from HDs or psoriatic patients. Values repres ent the mean ± SEM ( n = 3). For the data of panels (A,B) no significant differences between HDs or psoriatic patients were observed by two-way ANOVA followed by Bonferroni’s post-test.

    Journal: Frontiers in Immunology

    Article Title: A Reappraisal on the Potential Ability of Human Neutrophils to Express and Produce IL-17 Family Members In Vitro: Failure to Reproducibly Detect It

    doi: 10.3389/fimmu.2018.00795

    Figure Lengend Snippet: IL-17A, IL-17F, IL-17RA, IL-17RC, CXCL8, TNFα, and SOCS3 mRNA expression, as well as IL-17R surface expression, in neutrophils from patients with psoriasis. (A) Neutrophils isolated from healthy donors (HDs) ( n = 3) or psoriatic patients ( n = 3) were cultured for 20 h with 100 U/ml IFNγ plus 100 ng/ml LPS, 5 µM R848, or 500 ng/ml IL-17A to evaluate IL-17A, IL-17F, IL-17RA, IL-17RC, CXCL8, TNFα, and SOCS3 mRNA expression by reverse transcription quantitative real-time PCR. Gene expression data are depicted as mean normalized expression (MNE) units after GAPDH mRNA normalization. (B) Surface IL-17RA and IL-17RC expression evaluated by flow cytometry in human neutrophils from HDs or psoriatic patients. Values repres ent the mean ± SEM ( n = 3). For the data of panels (A,B) no significant differences between HDs or psoriatic patients were observed by two-way ANOVA followed by Bonferroni’s post-test.

    Article Snippet: Neutrophils were then stained for 15 min at T room with: APC anti-human IL-17RA/CD217 (clone 424LTS) and APC mouse IgG1к, as isotype control (clone P3.6.2.8.1) from eBioscience (San Diego, CA, USA); PE anti-human IL-17RC (clone 309822) and mouse PE IgG2B isotype control from R & D systems; PE-vio770 anti-human CD11b (clone ICRF44), FITC anti-human CD66b (clone G10F5), and PerCP-Cy5.6 anti-human CD16 (clone 3G8) from BioLegend (San Diego, CA, USA); APC anti-human CD62L (clone 145/15 Miltenyi Biotec), all at working dilutions specified in the corresponding datasheets.

    Techniques: Expressing, Isolation, Cell Culture, Real-time Polymerase Chain Reaction, Flow Cytometry, Cytometry

    Expression of surface IL-17RA and IL-17RC in neutrophils activated under various experimental conditions. Expression of surface IL-17RA (left panel) and IL-17RC (right panel) was evaluated by flow cytometry in neutrophils either freshly isolated or cultured for 3 h without or with 100 U/ml IFNγ plus 100 ng/ml LPS, 5 µM R848 (A) , 20 µg/ml IL-6 plus 2 µg/ml IL-23 alone or in the presence of inactivated Aspergillus fumigatus conidia, hyphae or 500 ng/ml rIL-17A (B) . Graphs depict a representative experiment out of three independent ones with similar results. Histograms show staining by specific and isotype control Abs, respectively, for each stimulatory condition.

    Journal: Frontiers in Immunology

    Article Title: A Reappraisal on the Potential Ability of Human Neutrophils to Express and Produce IL-17 Family Members In Vitro: Failure to Reproducibly Detect It

    doi: 10.3389/fimmu.2018.00795

    Figure Lengend Snippet: Expression of surface IL-17RA and IL-17RC in neutrophils activated under various experimental conditions. Expression of surface IL-17RA (left panel) and IL-17RC (right panel) was evaluated by flow cytometry in neutrophils either freshly isolated or cultured for 3 h without or with 100 U/ml IFNγ plus 100 ng/ml LPS, 5 µM R848 (A) , 20 µg/ml IL-6 plus 2 µg/ml IL-23 alone or in the presence of inactivated Aspergillus fumigatus conidia, hyphae or 500 ng/ml rIL-17A (B) . Graphs depict a representative experiment out of three independent ones with similar results. Histograms show staining by specific and isotype control Abs, respectively, for each stimulatory condition.

    Article Snippet: Neutrophils were then stained for 15 min at T room with: APC anti-human IL-17RA/CD217 (clone 424LTS) and APC mouse IgG1к, as isotype control (clone P3.6.2.8.1) from eBioscience (San Diego, CA, USA); PE anti-human IL-17RC (clone 309822) and mouse PE IgG2B isotype control from R & D systems; PE-vio770 anti-human CD11b (clone ICRF44), FITC anti-human CD66b (clone G10F5), and PerCP-Cy5.6 anti-human CD16 (clone 3G8) from BioLegend (San Diego, CA, USA); APC anti-human CD62L (clone 145/15 Miltenyi Biotec), all at working dilutions specified in the corresponding datasheets.

    Techniques: Expressing, Flow Cytometry, Cytometry, Isolation, Cell Culture, Staining