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primary antibodies against mouse il 36 α  (R&D Systems)


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    R&D Systems primary antibodies against mouse il 36 α
    IL-36α was induced in UUO kidney and renal TECs. (A) Real-time PCR showing relative renal levels of <t>IL-36</t> mRNAs in sham control and UUO mice. (B) IHC staining showing IL-36α protein expression in sham control and UUO mice; the arrows indicate IL-36α–positive renal TECs of atrophic tubules. Original magnification, ×400. (C) Quantitative analysis for the IL-36α–positive tubules. Data are means±SEM of nine mice per group. Western blot analysis for IL-36α protein expression in mouse tubular epithelial cells (mTECs) M-1 under (D) mechanically induced pressure, (E) H2O2, or (F) HMGB-1. (G) Semiquantitative IL-36α levels in mTECs M-1 lysates. Western blot analysis for IL-36α protein expression in house tubular epithelial cells (hTECs) HK-2 under (H) mechanically induced pressure, (I) H2O2, or (J) HMGB-1. (K) Semiquantitative IL-36α levels in hTECs HK-2 lysates. Data are means±SEM of three replicative results obtained from in vitro experiments. *P<0.05; **P<0.01; ***P<0.01; #not detectable.
    Primary Antibodies Against Mouse Il 36 α, supplied by R&D Systems, used in various techniques. Bioz Stars score: 93/100, based on 35 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/primary antibodies against mouse il 36 α/product/R&D Systems
    Average 93 stars, based on 35 article reviews
    primary antibodies against mouse il 36 α - by Bioz Stars, 2026-03
    93/100 stars

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    1) Product Images from "IL-36 Signaling Facilitates Activation of the NLRP3 Inflammasome and IL-23/IL-17 Axis in Renal Inflammation and Fibrosis"

    Article Title: IL-36 Signaling Facilitates Activation of the NLRP3 Inflammasome and IL-23/IL-17 Axis in Renal Inflammation and Fibrosis

    Journal: Journal of the American Society of Nephrology : JASN

    doi: 10.1681/ASN.2016080840

    IL-36α was induced in UUO kidney and renal TECs. (A) Real-time PCR showing relative renal levels of IL-36 mRNAs in sham control and UUO mice. (B) IHC staining showing IL-36α protein expression in sham control and UUO mice; the arrows indicate IL-36α–positive renal TECs of atrophic tubules. Original magnification, ×400. (C) Quantitative analysis for the IL-36α–positive tubules. Data are means±SEM of nine mice per group. Western blot analysis for IL-36α protein expression in mouse tubular epithelial cells (mTECs) M-1 under (D) mechanically induced pressure, (E) H2O2, or (F) HMGB-1. (G) Semiquantitative IL-36α levels in mTECs M-1 lysates. Western blot analysis for IL-36α protein expression in house tubular epithelial cells (hTECs) HK-2 under (H) mechanically induced pressure, (I) H2O2, or (J) HMGB-1. (K) Semiquantitative IL-36α levels in hTECs HK-2 lysates. Data are means±SEM of three replicative results obtained from in vitro experiments. *P<0.05; **P<0.01; ***P<0.01; #not detectable.
    Figure Legend Snippet: IL-36α was induced in UUO kidney and renal TECs. (A) Real-time PCR showing relative renal levels of IL-36 mRNAs in sham control and UUO mice. (B) IHC staining showing IL-36α protein expression in sham control and UUO mice; the arrows indicate IL-36α–positive renal TECs of atrophic tubules. Original magnification, ×400. (C) Quantitative analysis for the IL-36α–positive tubules. Data are means±SEM of nine mice per group. Western blot analysis for IL-36α protein expression in mouse tubular epithelial cells (mTECs) M-1 under (D) mechanically induced pressure, (E) H2O2, or (F) HMGB-1. (G) Semiquantitative IL-36α levels in mTECs M-1 lysates. Western blot analysis for IL-36α protein expression in house tubular epithelial cells (hTECs) HK-2 under (H) mechanically induced pressure, (I) H2O2, or (J) HMGB-1. (K) Semiquantitative IL-36α levels in hTECs HK-2 lysates. Data are means±SEM of three replicative results obtained from in vitro experiments. *P<0.05; **P<0.01; ***P<0.01; #not detectable.

    Techniques Used: Real-time Polymerase Chain Reaction, Immunohistochemistry, Expressing, Western Blot, In Vitro

    IL-36 signaling facilitated NLRP3 inflammasome activation in renal tissues of UUO mice and renal TECs. (A) Representative Western blots analysis and semiquantitation for NLRP3, IL-18, and IL-1β in renal tissues. (B) Caspase-1 activity determined by an activity assay in renal tissues. (C) ELISA for urinary protein levels of IL-1β in normal (sham control) or pelvic (UUO) urine samples. Data are means±SEM of nine mice per group. M-1 TECs treated with (D) rIL-36α, LPS (2 μg/ml), or saline for 24 hours (representative Western blots and semiquantitation for NLRP3, pro–IL-18, and pro–IL-1β in lysates) or (E) rIL-36α, LPS, or saline for 24 hours and an additional 1 hour of ATP incubation (representative Western blots for IL-18, IL-1β, and caspase-1 p10 in supernatants). (F) Caspase-1 activity determined by an activity assay in lysate. Data are means±SEM of three replicative results obtained from in vitro experiments. *P<0.05; **P<0.01; ***P<0.01.
    Figure Legend Snippet: IL-36 signaling facilitated NLRP3 inflammasome activation in renal tissues of UUO mice and renal TECs. (A) Representative Western blots analysis and semiquantitation for NLRP3, IL-18, and IL-1β in renal tissues. (B) Caspase-1 activity determined by an activity assay in renal tissues. (C) ELISA for urinary protein levels of IL-1β in normal (sham control) or pelvic (UUO) urine samples. Data are means±SEM of nine mice per group. M-1 TECs treated with (D) rIL-36α, LPS (2 μg/ml), or saline for 24 hours (representative Western blots and semiquantitation for NLRP3, pro–IL-18, and pro–IL-1β in lysates) or (E) rIL-36α, LPS, or saline for 24 hours and an additional 1 hour of ATP incubation (representative Western blots for IL-18, IL-1β, and caspase-1 p10 in supernatants). (F) Caspase-1 activity determined by an activity assay in lysate. Data are means±SEM of three replicative results obtained from in vitro experiments. *P<0.05; **P<0.01; ***P<0.01.

    Techniques Used: Activation Assay, Western Blot, Activity Assay, Enzyme-linked Immunosorbent Assay, Incubation, In Vitro

    IL-36 signaling facilitated NLRP3 inflammasome activation in cultured macrophages. (A) ELISA analysis of IL-1β protein levels in supernatant of murine J774A.1 macrophages incubated for 30 minutes with or without rIL-36α (150 ng/ml), 5.5 hours with or without 0.5 μg/ml LPS, and 30 minutes with or without 5 mM ATP. (B) Representative Western blots and semiquantitation for NLRP3 and pro–IL-1β in lysates of J774A.1 macrophages incubated with or without rIL-36α (150 ng/ml) for 30 minutes and then treated with or without LPS for 6 hours. (C) Representative Western blots for caspase-1 p10 in supernatants of J774A.1 incubated with or without rIL-36α (150 ng/ml), 5.5 hours with or without 0.5 μg g/ml LPS, and 30 minutes with or without 5 mM ATP. (D) ELISA analysis of IL-1β protein levels in supernatant of peritoneal macrophages from IL-36R KO and WT mice incubated for 5.5 hours with or without 1 μg g/ml LPS and 30 minutes with or without 5 mM ATP. (E) ELISA analysis of IL-1β protein levels in supernatant of BMDMs from WT, TLR4 KO, and MyD88 KO mice treated with similar conditions as J774A.1 above. (F) Representative Western blots and semiquantitation for NLRP3 and pro–IL-1β in lysates of BMDMs from WT, TLR4 KO, and MyD88 KO mice treated with or without rIL-36α (150 ng/ml). Data are means±SEM of three replicative results obtained from the experiments. Mϕ, macrophage *P<0.05; **P<0.01; ***P<0.01.
    Figure Legend Snippet: IL-36 signaling facilitated NLRP3 inflammasome activation in cultured macrophages. (A) ELISA analysis of IL-1β protein levels in supernatant of murine J774A.1 macrophages incubated for 30 minutes with or without rIL-36α (150 ng/ml), 5.5 hours with or without 0.5 μg/ml LPS, and 30 minutes with or without 5 mM ATP. (B) Representative Western blots and semiquantitation for NLRP3 and pro–IL-1β in lysates of J774A.1 macrophages incubated with or without rIL-36α (150 ng/ml) for 30 minutes and then treated with or without LPS for 6 hours. (C) Representative Western blots for caspase-1 p10 in supernatants of J774A.1 incubated with or without rIL-36α (150 ng/ml), 5.5 hours with or without 0.5 μg g/ml LPS, and 30 minutes with or without 5 mM ATP. (D) ELISA analysis of IL-1β protein levels in supernatant of peritoneal macrophages from IL-36R KO and WT mice incubated for 5.5 hours with or without 1 μg g/ml LPS and 30 minutes with or without 5 mM ATP. (E) ELISA analysis of IL-1β protein levels in supernatant of BMDMs from WT, TLR4 KO, and MyD88 KO mice treated with similar conditions as J774A.1 above. (F) Representative Western blots and semiquantitation for NLRP3 and pro–IL-1β in lysates of BMDMs from WT, TLR4 KO, and MyD88 KO mice treated with or without rIL-36α (150 ng/ml). Data are means±SEM of three replicative results obtained from the experiments. Mϕ, macrophage *P<0.05; **P<0.01; ***P<0.01.

    Techniques Used: Activation Assay, Cell Culture, Enzyme-linked Immunosorbent Assay, Incubation, Western Blot

    IL-36 signaling activated NLRP3 inflammasome activation in BMDCs. (A) ELISA analysis of IL-1β protein levels in supernatant of BMDCs from WT, IL-36R KO, and NLRP3 KO mice incubated with or without rIL-36α (150 ng/ml) for 24 hours. (B) Representative Western blots and (C) semiquantitation for NLRP3, pro–IL-1β, and caspase-1 p10 in lysates of BMDCs from WT, IL-36R KO, and NLRP3 KO mice incubated with or without rIL-36α (150 ng/ml) for 24 hours. Data are means±SEM of three replicative results obtained from the experiments. *P<0.01; ***P<0.01; #not detectable.
    Figure Legend Snippet: IL-36 signaling activated NLRP3 inflammasome activation in BMDCs. (A) ELISA analysis of IL-1β protein levels in supernatant of BMDCs from WT, IL-36R KO, and NLRP3 KO mice incubated with or without rIL-36α (150 ng/ml) for 24 hours. (B) Representative Western blots and (C) semiquantitation for NLRP3, pro–IL-1β, and caspase-1 p10 in lysates of BMDCs from WT, IL-36R KO, and NLRP3 KO mice incubated with or without rIL-36α (150 ng/ml) for 24 hours. Data are means±SEM of three replicative results obtained from the experiments. *P<0.01; ***P<0.01; #not detectable.

    Techniques Used: Activation Assay, Enzyme-linked Immunosorbent Assay, Incubation, Western Blot

    IL-36 signaling facilitated DC-induced T cells activation in UUO mice and cell models. (A) Flow cytometric analysis showing the percentages of CD4+/CD69+ and CD8+/CD69+ in renal draining lymphoid cells at day 7 after UUO or sham operation. Data are means±SEM of nine mice per group. (B) BMDCs were incubated with or without rIL-36α (150 ng/ml) for 16 hours, pulsed with OVA257–264 or OVA323–339 peptide, and then, cocultured with OT-I CD8 or OT-II CD4 T cells, respectively. After 3 days, the T cell proliferation was measured by [3H]thymidine incorporation. Data are means±SEM of three replicative results obtained from the experiments. (C) Flow cytometric analysis showing expression levels CD40, CD80, and CD86 (within gated CD11c cells) in BMDCs from WT and IL-36R KO mice, which were incubated with or without rIL-36α (150 ng/ml) for 24 hours. Data are means±SEM of three replicative results obtained from the experiments. (D) Flow cytometric analysis showing the numbers of CD11c+ cells in renal draining lymphoid cells at day 7 after UUO or sham operation; data are means±SEM of five mice per group. (E) Flow cytometric analysis showing the CD86+CD11c+ cells in renal draining lymphoid cells at day 7 after UUO or sham operation. Data are means±SEM of five mice per group. rdLN, renal draining lymph node. *P<0.05; **P<0.01; ***P<0.01.
    Figure Legend Snippet: IL-36 signaling facilitated DC-induced T cells activation in UUO mice and cell models. (A) Flow cytometric analysis showing the percentages of CD4+/CD69+ and CD8+/CD69+ in renal draining lymphoid cells at day 7 after UUO or sham operation. Data are means±SEM of nine mice per group. (B) BMDCs were incubated with or without rIL-36α (150 ng/ml) for 16 hours, pulsed with OVA257–264 or OVA323–339 peptide, and then, cocultured with OT-I CD8 or OT-II CD4 T cells, respectively. After 3 days, the T cell proliferation was measured by [3H]thymidine incorporation. Data are means±SEM of three replicative results obtained from the experiments. (C) Flow cytometric analysis showing expression levels CD40, CD80, and CD86 (within gated CD11c cells) in BMDCs from WT and IL-36R KO mice, which were incubated with or without rIL-36α (150 ng/ml) for 24 hours. Data are means±SEM of three replicative results obtained from the experiments. (D) Flow cytometric analysis showing the numbers of CD11c+ cells in renal draining lymphoid cells at day 7 after UUO or sham operation; data are means±SEM of five mice per group. (E) Flow cytometric analysis showing the CD86+CD11c+ cells in renal draining lymphoid cells at day 7 after UUO or sham operation. Data are means±SEM of five mice per group. rdLN, renal draining lymph node. *P<0.05; **P<0.01; ***P<0.01.

    Techniques Used: Activation Assay, Incubation, Expressing

    IL-36 facilitated Th17 differentiation in UUO mice and in vitro T cell activation. (A) Real-time PCR showing relative renal mRNA levels of IL-23p19 and IL-17A in sham control and day14 UUO mice. Data are means±SEM of nine mice per group. (B) BMDCs were incubated with or without rIL-36α (150 ng/ml) for 16 hours, pulsed with OVA323–339 peptides, and then, cocultured with OT-II CD4 T cells. After 4 days, supernatants were collected, and the production of IL-17A was detected by ELISA. Data are means±SEM of three replicative results obtained from the experiments. (C) Real-time PCR showing mRNA levels of specific transcription factors for different Th cells in OT-II CD4 cocultured with BMDCs (preincubated with or without rIL-36α for 16 hours and then, pulsed with OVA323–339 peptides) for 4 days. Data are means±SEM of three replicative results obtained from the experiments. *P<0.05; **P<0.01; ***P<0.01.
    Figure Legend Snippet: IL-36 facilitated Th17 differentiation in UUO mice and in vitro T cell activation. (A) Real-time PCR showing relative renal mRNA levels of IL-23p19 and IL-17A in sham control and day14 UUO mice. Data are means±SEM of nine mice per group. (B) BMDCs were incubated with or without rIL-36α (150 ng/ml) for 16 hours, pulsed with OVA323–339 peptides, and then, cocultured with OT-II CD4 T cells. After 4 days, supernatants were collected, and the production of IL-17A was detected by ELISA. Data are means±SEM of three replicative results obtained from the experiments. (C) Real-time PCR showing mRNA levels of specific transcription factors for different Th cells in OT-II CD4 cocultured with BMDCs (preincubated with or without rIL-36α for 16 hours and then, pulsed with OVA323–339 peptides) for 4 days. Data are means±SEM of three replicative results obtained from the experiments. *P<0.05; **P<0.01; ***P<0.01.

    Techniques Used: In Vitro, Activation Assay, Real-time Polymerase Chain Reaction, Incubation, Enzyme-linked Immunosorbent Assay

    Proposed mechanistic pathways underlying the pathogenic role of IL-36 in the development of renal TILs. IL-36 may be contributed to the pathogenesis of renal TILs, involving the activation of NLRP3 inflammasome and IL-23/IL-17 axis.
    Figure Legend Snippet: Proposed mechanistic pathways underlying the pathogenic role of IL-36 in the development of renal TILs. IL-36 may be contributed to the pathogenesis of renal TILs, involving the activation of NLRP3 inflammasome and IL-23/IL-17 axis.

    Techniques Used: Activation Assay



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    primary antibodies against mouse il 36 α  (R&D Systems)
    93
    R&D Systems primary antibodies against mouse il 36 α
    IL-36α was induced in UUO kidney and renal TECs. (A) Real-time PCR showing relative renal levels of <t>IL-36</t> mRNAs in sham control and UUO mice. (B) IHC staining showing IL-36α protein expression in sham control and UUO mice; the arrows indicate IL-36α–positive renal TECs of atrophic tubules. Original magnification, ×400. (C) Quantitative analysis for the IL-36α–positive tubules. Data are means±SEM of nine mice per group. Western blot analysis for IL-36α protein expression in mouse tubular epithelial cells (mTECs) M-1 under (D) mechanically induced pressure, (E) H2O2, or (F) HMGB-1. (G) Semiquantitative IL-36α levels in mTECs M-1 lysates. Western blot analysis for IL-36α protein expression in house tubular epithelial cells (hTECs) HK-2 under (H) mechanically induced pressure, (I) H2O2, or (J) HMGB-1. (K) Semiquantitative IL-36α levels in hTECs HK-2 lysates. Data are means±SEM of three replicative results obtained from in vitro experiments. *P<0.05; **P<0.01; ***P<0.01; #not detectable.
    Primary Antibodies Against Mouse Il 36 α, supplied by R&D Systems, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/primary antibodies against mouse il 36 α/product/R&D Systems
    Average 93 stars, based on 1 article reviews
    primary antibodies against mouse il 36 α - by Bioz Stars, 2026-03
    93/100 stars
    		  Buy from Supplier

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    IL-36α was induced in UUO kidney and renal TECs. (A) Real-time PCR showing relative renal levels of IL-36 mRNAs in sham control and UUO mice. (B) IHC staining showing IL-36α protein expression in sham control and UUO mice; the arrows indicate IL-36α–positive renal TECs of atrophic tubules. Original magnification, ×400. (C) Quantitative analysis for the IL-36α–positive tubules. Data are means±SEM of nine mice per group. Western blot analysis for IL-36α protein expression in mouse tubular epithelial cells (mTECs) M-1 under (D) mechanically induced pressure, (E) H2O2, or (F) HMGB-1. (G) Semiquantitative IL-36α levels in mTECs M-1 lysates. Western blot analysis for IL-36α protein expression in house tubular epithelial cells (hTECs) HK-2 under (H) mechanically induced pressure, (I) H2O2, or (J) HMGB-1. (K) Semiquantitative IL-36α levels in hTECs HK-2 lysates. Data are means±SEM of three replicative results obtained from in vitro experiments. *P<0.05; **P<0.01; ***P<0.01; #not detectable.

    Journal: Journal of the American Society of Nephrology : JASN

    Article Title: IL-36 Signaling Facilitates Activation of the NLRP3 Inflammasome and IL-23/IL-17 Axis in Renal Inflammation and Fibrosis

    doi: 10.1681/ASN.2016080840

    Figure Lengend Snippet: IL-36α was induced in UUO kidney and renal TECs. (A) Real-time PCR showing relative renal levels of IL-36 mRNAs in sham control and UUO mice. (B) IHC staining showing IL-36α protein expression in sham control and UUO mice; the arrows indicate IL-36α–positive renal TECs of atrophic tubules. Original magnification, ×400. (C) Quantitative analysis for the IL-36α–positive tubules. Data are means±SEM of nine mice per group. Western blot analysis for IL-36α protein expression in mouse tubular epithelial cells (mTECs) M-1 under (D) mechanically induced pressure, (E) H2O2, or (F) HMGB-1. (G) Semiquantitative IL-36α levels in mTECs M-1 lysates. Western blot analysis for IL-36α protein expression in house tubular epithelial cells (hTECs) HK-2 under (H) mechanically induced pressure, (I) H2O2, or (J) HMGB-1. (K) Semiquantitative IL-36α levels in hTECs HK-2 lysates. Data are means±SEM of three replicative results obtained from in vitro experiments. *P<0.05; **P<0.01; ***P<0.01; #not detectable.

    Article Snippet: 50 The target proteins were probed with respective primary antibodies against mouse IL-36 α (AF2297; R&D), β -actin (sc-130656; Santa Cruz Biotech, Santa Cruz, CA), human IL-36 α (AP5234c; Abgent), NLRP3 (AG-20B-0014; Adipogen, San Diego, CA), IL-18 (sc-7954; Santa Cruz Biotech), caspase-1 (AG-20B-0044; Adipogen), or IL-1 β (AF-401; R&D) followed by the addition of HRP-conjugated secondary antibodies (Santa Cruz Biotech).

    Techniques: Real-time Polymerase Chain Reaction, Immunohistochemistry, Expressing, Western Blot, In Vitro

    IL-36 signaling facilitated NLRP3 inflammasome activation in renal tissues of UUO mice and renal TECs. (A) Representative Western blots analysis and semiquantitation for NLRP3, IL-18, and IL-1β in renal tissues. (B) Caspase-1 activity determined by an activity assay in renal tissues. (C) ELISA for urinary protein levels of IL-1β in normal (sham control) or pelvic (UUO) urine samples. Data are means±SEM of nine mice per group. M-1 TECs treated with (D) rIL-36α, LPS (2 μg/ml), or saline for 24 hours (representative Western blots and semiquantitation for NLRP3, pro–IL-18, and pro–IL-1β in lysates) or (E) rIL-36α, LPS, or saline for 24 hours and an additional 1 hour of ATP incubation (representative Western blots for IL-18, IL-1β, and caspase-1 p10 in supernatants). (F) Caspase-1 activity determined by an activity assay in lysate. Data are means±SEM of three replicative results obtained from in vitro experiments. *P<0.05; **P<0.01; ***P<0.01.

    Journal: Journal of the American Society of Nephrology : JASN

    Article Title: IL-36 Signaling Facilitates Activation of the NLRP3 Inflammasome and IL-23/IL-17 Axis in Renal Inflammation and Fibrosis

    doi: 10.1681/ASN.2016080840

    Figure Lengend Snippet: IL-36 signaling facilitated NLRP3 inflammasome activation in renal tissues of UUO mice and renal TECs. (A) Representative Western blots analysis and semiquantitation for NLRP3, IL-18, and IL-1β in renal tissues. (B) Caspase-1 activity determined by an activity assay in renal tissues. (C) ELISA for urinary protein levels of IL-1β in normal (sham control) or pelvic (UUO) urine samples. Data are means±SEM of nine mice per group. M-1 TECs treated with (D) rIL-36α, LPS (2 μg/ml), or saline for 24 hours (representative Western blots and semiquantitation for NLRP3, pro–IL-18, and pro–IL-1β in lysates) or (E) rIL-36α, LPS, or saline for 24 hours and an additional 1 hour of ATP incubation (representative Western blots for IL-18, IL-1β, and caspase-1 p10 in supernatants). (F) Caspase-1 activity determined by an activity assay in lysate. Data are means±SEM of three replicative results obtained from in vitro experiments. *P<0.05; **P<0.01; ***P<0.01.

    Article Snippet: 50 The target proteins were probed with respective primary antibodies against mouse IL-36 α (AF2297; R&D), β -actin (sc-130656; Santa Cruz Biotech, Santa Cruz, CA), human IL-36 α (AP5234c; Abgent), NLRP3 (AG-20B-0014; Adipogen, San Diego, CA), IL-18 (sc-7954; Santa Cruz Biotech), caspase-1 (AG-20B-0044; Adipogen), or IL-1 β (AF-401; R&D) followed by the addition of HRP-conjugated secondary antibodies (Santa Cruz Biotech).

    Techniques: Activation Assay, Western Blot, Activity Assay, Enzyme-linked Immunosorbent Assay, Incubation, In Vitro

    IL-36 signaling facilitated NLRP3 inflammasome activation in cultured macrophages. (A) ELISA analysis of IL-1β protein levels in supernatant of murine J774A.1 macrophages incubated for 30 minutes with or without rIL-36α (150 ng/ml), 5.5 hours with or without 0.5 μg/ml LPS, and 30 minutes with or without 5 mM ATP. (B) Representative Western blots and semiquantitation for NLRP3 and pro–IL-1β in lysates of J774A.1 macrophages incubated with or without rIL-36α (150 ng/ml) for 30 minutes and then treated with or without LPS for 6 hours. (C) Representative Western blots for caspase-1 p10 in supernatants of J774A.1 incubated with or without rIL-36α (150 ng/ml), 5.5 hours with or without 0.5 μg g/ml LPS, and 30 minutes with or without 5 mM ATP. (D) ELISA analysis of IL-1β protein levels in supernatant of peritoneal macrophages from IL-36R KO and WT mice incubated for 5.5 hours with or without 1 μg g/ml LPS and 30 minutes with or without 5 mM ATP. (E) ELISA analysis of IL-1β protein levels in supernatant of BMDMs from WT, TLR4 KO, and MyD88 KO mice treated with similar conditions as J774A.1 above. (F) Representative Western blots and semiquantitation for NLRP3 and pro–IL-1β in lysates of BMDMs from WT, TLR4 KO, and MyD88 KO mice treated with or without rIL-36α (150 ng/ml). Data are means±SEM of three replicative results obtained from the experiments. Mϕ, macrophage *P<0.05; **P<0.01; ***P<0.01.

    Journal: Journal of the American Society of Nephrology : JASN

    Article Title: IL-36 Signaling Facilitates Activation of the NLRP3 Inflammasome and IL-23/IL-17 Axis in Renal Inflammation and Fibrosis

    doi: 10.1681/ASN.2016080840

    Figure Lengend Snippet: IL-36 signaling facilitated NLRP3 inflammasome activation in cultured macrophages. (A) ELISA analysis of IL-1β protein levels in supernatant of murine J774A.1 macrophages incubated for 30 minutes with or without rIL-36α (150 ng/ml), 5.5 hours with or without 0.5 μg/ml LPS, and 30 minutes with or without 5 mM ATP. (B) Representative Western blots and semiquantitation for NLRP3 and pro–IL-1β in lysates of J774A.1 macrophages incubated with or without rIL-36α (150 ng/ml) for 30 minutes and then treated with or without LPS for 6 hours. (C) Representative Western blots for caspase-1 p10 in supernatants of J774A.1 incubated with or without rIL-36α (150 ng/ml), 5.5 hours with or without 0.5 μg g/ml LPS, and 30 minutes with or without 5 mM ATP. (D) ELISA analysis of IL-1β protein levels in supernatant of peritoneal macrophages from IL-36R KO and WT mice incubated for 5.5 hours with or without 1 μg g/ml LPS and 30 minutes with or without 5 mM ATP. (E) ELISA analysis of IL-1β protein levels in supernatant of BMDMs from WT, TLR4 KO, and MyD88 KO mice treated with similar conditions as J774A.1 above. (F) Representative Western blots and semiquantitation for NLRP3 and pro–IL-1β in lysates of BMDMs from WT, TLR4 KO, and MyD88 KO mice treated with or without rIL-36α (150 ng/ml). Data are means±SEM of three replicative results obtained from the experiments. Mϕ, macrophage *P<0.05; **P<0.01; ***P<0.01.

    Article Snippet: 50 The target proteins were probed with respective primary antibodies against mouse IL-36 α (AF2297; R&D), β -actin (sc-130656; Santa Cruz Biotech, Santa Cruz, CA), human IL-36 α (AP5234c; Abgent), NLRP3 (AG-20B-0014; Adipogen, San Diego, CA), IL-18 (sc-7954; Santa Cruz Biotech), caspase-1 (AG-20B-0044; Adipogen), or IL-1 β (AF-401; R&D) followed by the addition of HRP-conjugated secondary antibodies (Santa Cruz Biotech).

    Techniques: Activation Assay, Cell Culture, Enzyme-linked Immunosorbent Assay, Incubation, Western Blot

    IL-36 signaling activated NLRP3 inflammasome activation in BMDCs. (A) ELISA analysis of IL-1β protein levels in supernatant of BMDCs from WT, IL-36R KO, and NLRP3 KO mice incubated with or without rIL-36α (150 ng/ml) for 24 hours. (B) Representative Western blots and (C) semiquantitation for NLRP3, pro–IL-1β, and caspase-1 p10 in lysates of BMDCs from WT, IL-36R KO, and NLRP3 KO mice incubated with or without rIL-36α (150 ng/ml) for 24 hours. Data are means±SEM of three replicative results obtained from the experiments. *P<0.01; ***P<0.01; #not detectable.

    Journal: Journal of the American Society of Nephrology : JASN

    Article Title: IL-36 Signaling Facilitates Activation of the NLRP3 Inflammasome and IL-23/IL-17 Axis in Renal Inflammation and Fibrosis

    doi: 10.1681/ASN.2016080840

    Figure Lengend Snippet: IL-36 signaling activated NLRP3 inflammasome activation in BMDCs. (A) ELISA analysis of IL-1β protein levels in supernatant of BMDCs from WT, IL-36R KO, and NLRP3 KO mice incubated with or without rIL-36α (150 ng/ml) for 24 hours. (B) Representative Western blots and (C) semiquantitation for NLRP3, pro–IL-1β, and caspase-1 p10 in lysates of BMDCs from WT, IL-36R KO, and NLRP3 KO mice incubated with or without rIL-36α (150 ng/ml) for 24 hours. Data are means±SEM of three replicative results obtained from the experiments. *P<0.01; ***P<0.01; #not detectable.

    Article Snippet: 50 The target proteins were probed with respective primary antibodies against mouse IL-36 α (AF2297; R&D), β -actin (sc-130656; Santa Cruz Biotech, Santa Cruz, CA), human IL-36 α (AP5234c; Abgent), NLRP3 (AG-20B-0014; Adipogen, San Diego, CA), IL-18 (sc-7954; Santa Cruz Biotech), caspase-1 (AG-20B-0044; Adipogen), or IL-1 β (AF-401; R&D) followed by the addition of HRP-conjugated secondary antibodies (Santa Cruz Biotech).

    Techniques: Activation Assay, Enzyme-linked Immunosorbent Assay, Incubation, Western Blot

    IL-36 signaling facilitated DC-induced T cells activation in UUO mice and cell models. (A) Flow cytometric analysis showing the percentages of CD4+/CD69+ and CD8+/CD69+ in renal draining lymphoid cells at day 7 after UUO or sham operation. Data are means±SEM of nine mice per group. (B) BMDCs were incubated with or without rIL-36α (150 ng/ml) for 16 hours, pulsed with OVA257–264 or OVA323–339 peptide, and then, cocultured with OT-I CD8 or OT-II CD4 T cells, respectively. After 3 days, the T cell proliferation was measured by [3H]thymidine incorporation. Data are means±SEM of three replicative results obtained from the experiments. (C) Flow cytometric analysis showing expression levels CD40, CD80, and CD86 (within gated CD11c cells) in BMDCs from WT and IL-36R KO mice, which were incubated with or without rIL-36α (150 ng/ml) for 24 hours. Data are means±SEM of three replicative results obtained from the experiments. (D) Flow cytometric analysis showing the numbers of CD11c+ cells in renal draining lymphoid cells at day 7 after UUO or sham operation; data are means±SEM of five mice per group. (E) Flow cytometric analysis showing the CD86+CD11c+ cells in renal draining lymphoid cells at day 7 after UUO or sham operation. Data are means±SEM of five mice per group. rdLN, renal draining lymph node. *P<0.05; **P<0.01; ***P<0.01.

    Journal: Journal of the American Society of Nephrology : JASN

    Article Title: IL-36 Signaling Facilitates Activation of the NLRP3 Inflammasome and IL-23/IL-17 Axis in Renal Inflammation and Fibrosis

    doi: 10.1681/ASN.2016080840

    Figure Lengend Snippet: IL-36 signaling facilitated DC-induced T cells activation in UUO mice and cell models. (A) Flow cytometric analysis showing the percentages of CD4+/CD69+ and CD8+/CD69+ in renal draining lymphoid cells at day 7 after UUO or sham operation. Data are means±SEM of nine mice per group. (B) BMDCs were incubated with or without rIL-36α (150 ng/ml) for 16 hours, pulsed with OVA257–264 or OVA323–339 peptide, and then, cocultured with OT-I CD8 or OT-II CD4 T cells, respectively. After 3 days, the T cell proliferation was measured by [3H]thymidine incorporation. Data are means±SEM of three replicative results obtained from the experiments. (C) Flow cytometric analysis showing expression levels CD40, CD80, and CD86 (within gated CD11c cells) in BMDCs from WT and IL-36R KO mice, which were incubated with or without rIL-36α (150 ng/ml) for 24 hours. Data are means±SEM of three replicative results obtained from the experiments. (D) Flow cytometric analysis showing the numbers of CD11c+ cells in renal draining lymphoid cells at day 7 after UUO or sham operation; data are means±SEM of five mice per group. (E) Flow cytometric analysis showing the CD86+CD11c+ cells in renal draining lymphoid cells at day 7 after UUO or sham operation. Data are means±SEM of five mice per group. rdLN, renal draining lymph node. *P<0.05; **P<0.01; ***P<0.01.

    Article Snippet: 50 The target proteins were probed with respective primary antibodies against mouse IL-36 α (AF2297; R&D), β -actin (sc-130656; Santa Cruz Biotech, Santa Cruz, CA), human IL-36 α (AP5234c; Abgent), NLRP3 (AG-20B-0014; Adipogen, San Diego, CA), IL-18 (sc-7954; Santa Cruz Biotech), caspase-1 (AG-20B-0044; Adipogen), or IL-1 β (AF-401; R&D) followed by the addition of HRP-conjugated secondary antibodies (Santa Cruz Biotech).

    Techniques: Activation Assay, Incubation, Expressing

    IL-36 facilitated Th17 differentiation in UUO mice and in vitro T cell activation. (A) Real-time PCR showing relative renal mRNA levels of IL-23p19 and IL-17A in sham control and day14 UUO mice. Data are means±SEM of nine mice per group. (B) BMDCs were incubated with or without rIL-36α (150 ng/ml) for 16 hours, pulsed with OVA323–339 peptides, and then, cocultured with OT-II CD4 T cells. After 4 days, supernatants were collected, and the production of IL-17A was detected by ELISA. Data are means±SEM of three replicative results obtained from the experiments. (C) Real-time PCR showing mRNA levels of specific transcription factors for different Th cells in OT-II CD4 cocultured with BMDCs (preincubated with or without rIL-36α for 16 hours and then, pulsed with OVA323–339 peptides) for 4 days. Data are means±SEM of three replicative results obtained from the experiments. *P<0.05; **P<0.01; ***P<0.01.

    Journal: Journal of the American Society of Nephrology : JASN

    Article Title: IL-36 Signaling Facilitates Activation of the NLRP3 Inflammasome and IL-23/IL-17 Axis in Renal Inflammation and Fibrosis

    doi: 10.1681/ASN.2016080840

    Figure Lengend Snippet: IL-36 facilitated Th17 differentiation in UUO mice and in vitro T cell activation. (A) Real-time PCR showing relative renal mRNA levels of IL-23p19 and IL-17A in sham control and day14 UUO mice. Data are means±SEM of nine mice per group. (B) BMDCs were incubated with or without rIL-36α (150 ng/ml) for 16 hours, pulsed with OVA323–339 peptides, and then, cocultured with OT-II CD4 T cells. After 4 days, supernatants were collected, and the production of IL-17A was detected by ELISA. Data are means±SEM of three replicative results obtained from the experiments. (C) Real-time PCR showing mRNA levels of specific transcription factors for different Th cells in OT-II CD4 cocultured with BMDCs (preincubated with or without rIL-36α for 16 hours and then, pulsed with OVA323–339 peptides) for 4 days. Data are means±SEM of three replicative results obtained from the experiments. *P<0.05; **P<0.01; ***P<0.01.

    Article Snippet: 50 The target proteins were probed with respective primary antibodies against mouse IL-36 α (AF2297; R&D), β -actin (sc-130656; Santa Cruz Biotech, Santa Cruz, CA), human IL-36 α (AP5234c; Abgent), NLRP3 (AG-20B-0014; Adipogen, San Diego, CA), IL-18 (sc-7954; Santa Cruz Biotech), caspase-1 (AG-20B-0044; Adipogen), or IL-1 β (AF-401; R&D) followed by the addition of HRP-conjugated secondary antibodies (Santa Cruz Biotech).

    Techniques: In Vitro, Activation Assay, Real-time Polymerase Chain Reaction, Incubation, Enzyme-linked Immunosorbent Assay

    Proposed mechanistic pathways underlying the pathogenic role of IL-36 in the development of renal TILs. IL-36 may be contributed to the pathogenesis of renal TILs, involving the activation of NLRP3 inflammasome and IL-23/IL-17 axis.

    Journal: Journal of the American Society of Nephrology : JASN

    Article Title: IL-36 Signaling Facilitates Activation of the NLRP3 Inflammasome and IL-23/IL-17 Axis in Renal Inflammation and Fibrosis

    doi: 10.1681/ASN.2016080840

    Figure Lengend Snippet: Proposed mechanistic pathways underlying the pathogenic role of IL-36 in the development of renal TILs. IL-36 may be contributed to the pathogenesis of renal TILs, involving the activation of NLRP3 inflammasome and IL-23/IL-17 axis.

    Article Snippet: 50 The target proteins were probed with respective primary antibodies against mouse IL-36 α (AF2297; R&D), β -actin (sc-130656; Santa Cruz Biotech, Santa Cruz, CA), human IL-36 α (AP5234c; Abgent), NLRP3 (AG-20B-0014; Adipogen, San Diego, CA), IL-18 (sc-7954; Santa Cruz Biotech), caspase-1 (AG-20B-0044; Adipogen), or IL-1 β (AF-401; R&D) followed by the addition of HRP-conjugated secondary antibodies (Santa Cruz Biotech).

    Techniques: Activation Assay