ikkα mouse anti human monoclonal 1 1000 cat no 11930 cell signaling technology  (Cell Signaling Technology Inc)


Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 86

    Structured Review

    Cell Signaling Technology Inc ikkα mouse anti human monoclonal 1 1000 cat no 11930 cell signaling technology
    Ikkα Mouse Anti Human Monoclonal 1 1000 Cat No 11930 Cell Signaling Technology, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/ikkα mouse anti human monoclonal 1 1000 cat no 11930 cell signaling technology/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    ikkα mouse anti human monoclonal 1 1000 cat no 11930 cell signaling technology - by Bioz Stars, 2024-06
    86/100 stars

    Images

    mouse monoclonal anti human β actin antibodies  (Cell Signaling Technology Inc)


    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 86

    Structured Review

    Cell Signaling Technology Inc mouse monoclonal anti human β actin antibodies
    A P3 ESC show increased nuclear p21 (green signal, left panel) after IL-1β-treatment. <t>β-Actin</t> (red) staining is more diffuse in IL-1β-treated ESC and the cellular orientation is more parallel. Merged channel images (yellow signal, right panels) are included. Magnification × 200. B SA-β-gal staining was used to detect β-galactosidase-positive signals. There was an intense blue color in the cytoplasm of ESC following 24 h of IL-1β treatment (right panel) compared to the untreated control (left panel). Magnification × 200.
    Mouse Monoclonal Anti Human β Actin Antibodies, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mouse monoclonal anti human β actin antibodies/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    mouse monoclonal anti human β actin antibodies - by Bioz Stars, 2024-06
    86/100 stars

    Images

    1) Product Images from "Interleukin-1β induces and accelerates human endometrial stromal cell senescence and impairs decidualization via the c-Jun N-terminal kinase pathway"

    Article Title: Interleukin-1β induces and accelerates human endometrial stromal cell senescence and impairs decidualization via the c-Jun N-terminal kinase pathway

    Journal: Cell Death Discovery

    doi: 10.1038/s41420-024-02048-6

    A P3 ESC show increased nuclear p21 (green signal, left panel) after IL-1β-treatment. β-Actin (red) staining is more diffuse in IL-1β-treated ESC and the cellular orientation is more parallel. Merged channel images (yellow signal, right panels) are included. Magnification × 200. B SA-β-gal staining was used to detect β-galactosidase-positive signals. There was an intense blue color in the cytoplasm of ESC following 24 h of IL-1β treatment (right panel) compared to the untreated control (left panel). Magnification × 200.
    Figure Legend Snippet: A P3 ESC show increased nuclear p21 (green signal, left panel) after IL-1β-treatment. β-Actin (red) staining is more diffuse in IL-1β-treated ESC and the cellular orientation is more parallel. Merged channel images (yellow signal, right panels) are included. Magnification × 200. B SA-β-gal staining was used to detect β-galactosidase-positive signals. There was an intense blue color in the cytoplasm of ESC following 24 h of IL-1β treatment (right panel) compared to the untreated control (left panel). Magnification × 200.

    Techniques Used: Staining

    A After 48 h incubation, Western blotting shows that IL-1β increased IL-1β, IL-6, TNF-α, MMP3, p16, p21, CCL2 and phospho-histone H2A.X in P3 ESC. B Time-course experiments reveal that IL-1β upregulated IL-1β, IL-6, TNF-α, MMP3, p16, p21, HMGB1, CCL2, CCL5 and phospho-histone H2A.X at 24, 48 and 72 h (lanes 2, 4 and 6). Co-incubation in the presence of the JNK inhibitor SP blocked IL-1β-mediated upregulation at all time points (lanes 3, 5 and 7). Incubation with SP alone (lane 8) suppressed many biomarkers below basal (Control, lane 1) levels. β-Actin levels were not affected. Molecular masses (kDa) are indicated by arrows at right. C Based on the time course experiments, the three key senescence-associated biomarkers indicated that the baseline levels of IL-1β and IL-6 were notably low and undetectable by the western blot method at all three time points. Meanwhile, MMP3 expression levels in the untreated control group remained consistently similar across the three observed time points. β-Actin levels were not affected. Molecular masses (kDa) are indicated by arrows at right.
    Figure Legend Snippet: A After 48 h incubation, Western blotting shows that IL-1β increased IL-1β, IL-6, TNF-α, MMP3, p16, p21, CCL2 and phospho-histone H2A.X in P3 ESC. B Time-course experiments reveal that IL-1β upregulated IL-1β, IL-6, TNF-α, MMP3, p16, p21, HMGB1, CCL2, CCL5 and phospho-histone H2A.X at 24, 48 and 72 h (lanes 2, 4 and 6). Co-incubation in the presence of the JNK inhibitor SP blocked IL-1β-mediated upregulation at all time points (lanes 3, 5 and 7). Incubation with SP alone (lane 8) suppressed many biomarkers below basal (Control, lane 1) levels. β-Actin levels were not affected. Molecular masses (kDa) are indicated by arrows at right. C Based on the time course experiments, the three key senescence-associated biomarkers indicated that the baseline levels of IL-1β and IL-6 were notably low and undetectable by the western blot method at all three time points. Meanwhile, MMP3 expression levels in the untreated control group remained consistently similar across the three observed time points. β-Actin levels were not affected. Molecular masses (kDa) are indicated by arrows at right.

    Techniques Used: Incubation, Western Blot, Expressing

    A ELISA confirmed that CCL5 secretion by P3 ESC from each of four subjects was upregulated after 72 h of IL-1β treatment. This effect was reversed by co-incubation with SP ( p < 0.001, ANOVA). Error bars indicate SD of triplicate determinations for subject S4. B When IL-1β was co-incubated with a 100-fold excess of IL-1ra , complete abrogation of the IL-1β effect on IL-8 and IL-6 upregulation was observed. The overall comparison using two-factor ANOVA analysis revealed significant differences ( p < 0.001). Scheffé Test comparisons showed significant differences between Control vs IL-1β (p < 0.001) and IL-1β vs IL-1ra ( p < 0.001). Error bars indicate SD of triplicate determinations. C Time-course of CCL5 ELISA response to IL-1β ± SP (24–72 h). The overall comparison using one-factor ANOVA revealed significant differences between groups (p < 0.001). Error bars indicate SD of triplicate determinations. D Time-course of TNF-α ELISA response to IL-1β ± SP (24–72 h). The overall comparison using one-factor ANOVA revealed significant differences between groups ( p < 0.001). Error bars indicate SD of triplicate determinations. E Western blot showing that preincubation of P3 ESC with SP, for 1 h before addition of IL-1β (PSP), had the same inhibitory effect as simultaneous co-incubation of IL-1β plus SP (SSP) on IL-1β, IL-6, TNF-α and MMP3 reduction. SP alone reduced basal (control) SASP markers. β-Actin levels were not affected. F Western blot reveals inhibition of IL-1β stimulation of IL-1β, IL - 6, MMP3, p21 and HMGB1 by JNK1 (CJ1) or JNK2 (CJ2) siRNA constructs transfected into P3 ESC. β-Actin levels were not affected.
    Figure Legend Snippet: A ELISA confirmed that CCL5 secretion by P3 ESC from each of four subjects was upregulated after 72 h of IL-1β treatment. This effect was reversed by co-incubation with SP ( p < 0.001, ANOVA). Error bars indicate SD of triplicate determinations for subject S4. B When IL-1β was co-incubated with a 100-fold excess of IL-1ra , complete abrogation of the IL-1β effect on IL-8 and IL-6 upregulation was observed. The overall comparison using two-factor ANOVA analysis revealed significant differences ( p < 0.001). Scheffé Test comparisons showed significant differences between Control vs IL-1β (p < 0.001) and IL-1β vs IL-1ra ( p < 0.001). Error bars indicate SD of triplicate determinations. C Time-course of CCL5 ELISA response to IL-1β ± SP (24–72 h). The overall comparison using one-factor ANOVA revealed significant differences between groups (p < 0.001). Error bars indicate SD of triplicate determinations. D Time-course of TNF-α ELISA response to IL-1β ± SP (24–72 h). The overall comparison using one-factor ANOVA revealed significant differences between groups ( p < 0.001). Error bars indicate SD of triplicate determinations. E Western blot showing that preincubation of P3 ESC with SP, for 1 h before addition of IL-1β (PSP), had the same inhibitory effect as simultaneous co-incubation of IL-1β plus SP (SSP) on IL-1β, IL-6, TNF-α and MMP3 reduction. SP alone reduced basal (control) SASP markers. β-Actin levels were not affected. F Western blot reveals inhibition of IL-1β stimulation of IL-1β, IL - 6, MMP3, p21 and HMGB1 by JNK1 (CJ1) or JNK2 (CJ2) siRNA constructs transfected into P3 ESC. β-Actin levels were not affected.

    Techniques Used: Enzyme-linked Immunosorbent Assay, Incubation, Comparison, Western Blot, Inhibition, Construct, Transfection

    A Immunofluorescence- (IFC, top panels) and immunoperoxidase cytochemistry (IPC, lower panels) were used to localize IL-1β in ESC before and after IL-1β stimulation in P3 cells. B IFC (top panels) and IPC (lower panels) identified more IL-1β after IL-1β stimulation in P21 cells than P3 cells. Cell orientation in P21 cells also appears more parallel. C Western blot shows IL-1β, IL-6, TNF-α, MMP3, p16, p21 all increased with extended cell passage (P3 to P21) after incubation with IL-1β. β-Actin levels were unchanged.
    Figure Legend Snippet: A Immunofluorescence- (IFC, top panels) and immunoperoxidase cytochemistry (IPC, lower panels) were used to localize IL-1β in ESC before and after IL-1β stimulation in P3 cells. B IFC (top panels) and IPC (lower panels) identified more IL-1β after IL-1β stimulation in P21 cells than P3 cells. Cell orientation in P21 cells also appears more parallel. C Western blot shows IL-1β, IL-6, TNF-α, MMP3, p16, p21 all increased with extended cell passage (P3 to P21) after incubation with IL-1β. β-Actin levels were unchanged.

    Techniques Used: Immunofluorescence, Western Blot, Incubation

    anti human mouse fra 1  (Cell Signaling Technology Inc)


    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 86

    Structured Review

    Cell Signaling Technology Inc anti human mouse fra 1
    IL-33 Enhances the Expression of Downstream Target Genes of WNT/β-catenin Pathway. A RNA expression of downstream target genes of WNT/β-catenin ( AXIN2, CCN4, CCND1, CD44, <t>Fra-1,</t> JUN, LGR5, MYC, and PPARD ) were detected via real-time PCR in EECs and 12Z cells. B Fra-1 expression in EECs and 12Z cells was detected by western blotting. C CCN4 secretion in the cell supernatant of EECs and 12Z cells was detected via ELISA. D - F After IL-33 or/and ST2 neutralizing antibody treatment, the mRNA expression and protein levels of Fra-1 and CCN4 in EECs and 12Z cells. G Immunohistochemical staining of Fra-1 and CCN4 in eutopic endometrium and ectopic lesions. Data are presented as mean ± SEM. All data were analyzed using one-way ANOVA followed by Dunnett’s post hoc test and Student’s t-test ; * P < 0.05, ** P < 0.01, *** P < 0.001
    Anti Human Mouse Fra 1, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti human mouse fra 1/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    anti human mouse fra 1 - by Bioz Stars, 2024-06
    86/100 stars

    Images

    1) Product Images from "The IL-33-ST2 axis plays a vital role in endometriosis via promoting epithelial–mesenchymal transition by phosphorylating β-catenin"

    Article Title: The IL-33-ST2 axis plays a vital role in endometriosis via promoting epithelial–mesenchymal transition by phosphorylating β-catenin

    Journal: Cell Communication and Signaling : CCS

    doi: 10.1186/s12964-024-01683-x

    IL-33 Enhances the Expression of Downstream Target Genes of WNT/β-catenin Pathway. A RNA expression of downstream target genes of WNT/β-catenin ( AXIN2, CCN4, CCND1, CD44, Fra-1, JUN, LGR5, MYC, and PPARD ) were detected via real-time PCR in EECs and 12Z cells. B Fra-1 expression in EECs and 12Z cells was detected by western blotting. C CCN4 secretion in the cell supernatant of EECs and 12Z cells was detected via ELISA. D - F After IL-33 or/and ST2 neutralizing antibody treatment, the mRNA expression and protein levels of Fra-1 and CCN4 in EECs and 12Z cells. G Immunohistochemical staining of Fra-1 and CCN4 in eutopic endometrium and ectopic lesions. Data are presented as mean ± SEM. All data were analyzed using one-way ANOVA followed by Dunnett’s post hoc test and Student’s t-test ; * P < 0.05, ** P < 0.01, *** P < 0.001
    Figure Legend Snippet: IL-33 Enhances the Expression of Downstream Target Genes of WNT/β-catenin Pathway. A RNA expression of downstream target genes of WNT/β-catenin ( AXIN2, CCN4, CCND1, CD44, Fra-1, JUN, LGR5, MYC, and PPARD ) were detected via real-time PCR in EECs and 12Z cells. B Fra-1 expression in EECs and 12Z cells was detected by western blotting. C CCN4 secretion in the cell supernatant of EECs and 12Z cells was detected via ELISA. D - F After IL-33 or/and ST2 neutralizing antibody treatment, the mRNA expression and protein levels of Fra-1 and CCN4 in EECs and 12Z cells. G Immunohistochemical staining of Fra-1 and CCN4 in eutopic endometrium and ectopic lesions. Data are presented as mean ± SEM. All data were analyzed using one-way ANOVA followed by Dunnett’s post hoc test and Student’s t-test ; * P < 0.05, ** P < 0.01, *** P < 0.001

    Techniques Used: Expressing, RNA Expression, Real-time Polymerase Chain Reaction, Western Blot, Enzyme-linked Immunosorbent Assay, Immunohistochemical staining, Staining

    IL-33 obviously Induced β-catenin Phosphorylation. A Protein expression of phospho-β-catenin (Ser675 and Ser552), phospho-CREB (Ser133), CREB, phospho-AKT(Ser473) and AKT in EECs and 12Z cells treated by IL-33 or/and ST2 neutralizing antibody. B Protein expression of phospho-β-catenin (Ser675 and Ser552), phospho-CREB (Ser133), CREB, phospho-AKT (Ser473), AKT, and Fra-1 in EECs and 12Z cells treated by IL-33 or/and knocking down ST2 by siRNA. C Expression of CCN4 in supernatant of EEC and 12Z treated by IL-33 or/and knocking down ST2 by siRNA. Data are presented as mean ± SEM. All data were analyzed using one-way ANOVA followed by Dunnett’s post hoc test and Student’s t-test ; * p < 0.05, ** p < 0.01, *** p < 0.001
    Figure Legend Snippet: IL-33 obviously Induced β-catenin Phosphorylation. A Protein expression of phospho-β-catenin (Ser675 and Ser552), phospho-CREB (Ser133), CREB, phospho-AKT(Ser473) and AKT in EECs and 12Z cells treated by IL-33 or/and ST2 neutralizing antibody. B Protein expression of phospho-β-catenin (Ser675 and Ser552), phospho-CREB (Ser133), CREB, phospho-AKT (Ser473), AKT, and Fra-1 in EECs and 12Z cells treated by IL-33 or/and knocking down ST2 by siRNA. C Expression of CCN4 in supernatant of EEC and 12Z treated by IL-33 or/and knocking down ST2 by siRNA. Data are presented as mean ± SEM. All data were analyzed using one-way ANOVA followed by Dunnett’s post hoc test and Student’s t-test ; * p < 0.05, ** p < 0.01, *** p < 0.001

    Techniques Used: Expressing

    IL-33 Mediates β-catenin Phosphorylation at Ser675 and Ser552 by PKA. A , B Protein expression of EMT-related proteins (vimentin, E-cadherin, N-cadherin, and β-catenin), Fra-1,phospho-β-catenin (Ser675 and Ser552), phospho-CREB (Ser133), CREB, phospho-AKT (Ser473) and AKT in EEC and 12Z treated by IL-33, PKA inhibitor-H89 or/and AKT inhibitor- MK2206. C Expression of CCN4 in supernatant of EEC and 12Z treated by IL-33, PKA inhibitor- H89 or/and AKT inhibitor- MK2206. D , F Cyclohexane chase assay of β-catenin in EECs and 12Z cells treated by IL-33. Data are presented as mean ± SEM. All data were analyzed using one-way ANOVA followed by Dunnett’s post hoc test and Student’s t-test ; * P < 0.05, ** P < 0.01, *** P < 0.001
    Figure Legend Snippet: IL-33 Mediates β-catenin Phosphorylation at Ser675 and Ser552 by PKA. A , B Protein expression of EMT-related proteins (vimentin, E-cadherin, N-cadherin, and β-catenin), Fra-1,phospho-β-catenin (Ser675 and Ser552), phospho-CREB (Ser133), CREB, phospho-AKT (Ser473) and AKT in EEC and 12Z treated by IL-33, PKA inhibitor-H89 or/and AKT inhibitor- MK2206. C Expression of CCN4 in supernatant of EEC and 12Z treated by IL-33, PKA inhibitor- H89 or/and AKT inhibitor- MK2206. D , F Cyclohexane chase assay of β-catenin in EECs and 12Z cells treated by IL-33. Data are presented as mean ± SEM. All data were analyzed using one-way ANOVA followed by Dunnett’s post hoc test and Student’s t-test ; * P < 0.05, ** P < 0.01, *** P < 0.001

    Techniques Used: Expressing

    IL-33-ST2 Signal Blocking or β-catenin Knockout Weakens EMT in EEC and 12Z. A . Immunofluorescence staining of phospho-β-catenin at Ser675 and Ser552 (Scale bars, 20 μm). B , C Protein expression of EMT-related proteins (Vimentin, E-cadherin, N-cadherin, and β-catenin) and Fra-1 in EECs and 12Z cells treated by IL-33 or/and knocking β-catenin by siRNA. D Expression of CCN4 in supernatant of EEC and 12Z treated by IL-33 or/and knocking β-catenin by siRNA. Data are presented as mean ± SEM. All data were analyzed using one-way ANOVA followed by Dunnett’s post hoc test and Student’s t-test ; * P < 0.05, ** P < 0.01, *** P < 0.001
    Figure Legend Snippet: IL-33-ST2 Signal Blocking or β-catenin Knockout Weakens EMT in EEC and 12Z. A . Immunofluorescence staining of phospho-β-catenin at Ser675 and Ser552 (Scale bars, 20 μm). B , C Protein expression of EMT-related proteins (Vimentin, E-cadherin, N-cadherin, and β-catenin) and Fra-1 in EECs and 12Z cells treated by IL-33 or/and knocking β-catenin by siRNA. D Expression of CCN4 in supernatant of EEC and 12Z treated by IL-33 or/and knocking β-catenin by siRNA. Data are presented as mean ± SEM. All data were analyzed using one-way ANOVA followed by Dunnett’s post hoc test and Student’s t-test ; * P < 0.05, ** P < 0.01, *** P < 0.001

    Techniques Used: Blocking Assay, Knock-Out, Immunofluorescence, Staining, Expressing

    Schematic illustration of IL-33 Promoting EMT Process in Endometriotic Milieu. Interleukin 33 (IL-33) is highly expressed in ectopic ESCs, acting via the receptor ST2. Ectopic milieu, characterized by ROS, TGF-β1, and high level of estrogen, triggers secretion of IL-33 in ESCs, which in turn, enhanced the aggressive implantation and survival of ESCs. Meanwhile, elevated IL-33 in ectopic milieu also activated WNT/β-catenin pathway in EECs by phosphorylating β-catenin (Ser675 and Ser552), which primes exresssion of ECM related genes (CCN4 and Fra-1) and mesenchymal markers, enhanceing the EMT process and extracellular matrix produnction. Thus, IL-33/ST2 axis plays a pivotal role in endometriosis progress by promoting EMT. This figure was created with biorender.com
    Figure Legend Snippet: Schematic illustration of IL-33 Promoting EMT Process in Endometriotic Milieu. Interleukin 33 (IL-33) is highly expressed in ectopic ESCs, acting via the receptor ST2. Ectopic milieu, characterized by ROS, TGF-β1, and high level of estrogen, triggers secretion of IL-33 in ESCs, which in turn, enhanced the aggressive implantation and survival of ESCs. Meanwhile, elevated IL-33 in ectopic milieu also activated WNT/β-catenin pathway in EECs by phosphorylating β-catenin (Ser675 and Ser552), which primes exresssion of ECM related genes (CCN4 and Fra-1) and mesenchymal markers, enhanceing the EMT process and extracellular matrix produnction. Thus, IL-33/ST2 axis plays a pivotal role in endometriosis progress by promoting EMT. This figure was created with biorender.com

    Techniques Used:

    anti human mouse β actin  (Cell Signaling Technology Inc)


    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 86

    Structured Review

    Cell Signaling Technology Inc anti human mouse β actin
    GSDMD deletion exacerbates lupus-like phenotype in cGVH and NTS models. A Immunochemical images of GSDMD-FL and GSDMD-N pattern in renal section from normal control (NC) and LN patients. Scale bars: 50 μm. B Schematic of murine lupus models. NZB/W F1 mice were sacrificed at 14 weeks and 36 weeks. For cGVH model, C57BL/6 mice received an intraperitoneal injection of 1 × 10^ 8 bm12-derived splenocytes and were sacrificed 10 weeks post-induction. For NTS model, C57BL/6 mice were preimmunized with 0.2 mg sheep IgG intraperitoneally, followed by an intravenous injection of 50 μl sheep NTS on Day 4, and sacrificed on Day 11. C-E Immunoblotting of GSDMD in renal protein extracts of NZB/W F1 mice, cGVH or NTS mice with control (CON) mice. <t>β-actin</t> is loading control. F ELISA determining the level of autoantibodies (anti-dsDNA, anti-ssDNA and anti-histone) in serum of cGVH mice ( n = 4 or 5 per group). G Spleen to body weight ratio in cGVH mice ( n = 8 per group). H Representative PAS staining of kidney sections with quantitative analysis of glomerular cellularity and tubular injury scores in cGVH mice ( n = 5 or 10 per group). Scale bars: 50 μm. I Urine albumin to creatinine ratio in cGVH mice ( n = 8 per group). J Representative images and quantified immunofluorescence density of IgG and C3 deposition in the kidneys from cGVH mice ( n = 5 per group). Scale bars: 50 μm. K Quantitative analysis of glomerular PAS score and tubular injury scores in kidney sections from NTS mice (n = 5 per group). L Urine albumin to creatinine ratio and BUN in NTS mice (n = 5 per group). Data are shown as mean ± SEM. Two-way ANOVA test, ANOVA or Student’s t test was used for statistical analysis. **** P < 0.0001; *** P < 0.001; ** P < 0.01; * P < 0.05; ns, not significant
    Anti Human Mouse β Actin, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti human mouse β actin/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    anti human mouse β actin - by Bioz Stars, 2024-06
    86/100 stars

    Images

    1) Product Images from "Gasdermin D deficiency aborts myeloid calcium influx to drive granulopoiesis in lupus nephritis"

    Article Title: Gasdermin D deficiency aborts myeloid calcium influx to drive granulopoiesis in lupus nephritis

    Journal: Cell Communication and Signaling : CCS

    doi: 10.1186/s12964-024-01681-z

    GSDMD deletion exacerbates lupus-like phenotype in cGVH and NTS models. A Immunochemical images of GSDMD-FL and GSDMD-N pattern in renal section from normal control (NC) and LN patients. Scale bars: 50 μm. B Schematic of murine lupus models. NZB/W F1 mice were sacrificed at 14 weeks and 36 weeks. For cGVH model, C57BL/6 mice received an intraperitoneal injection of 1 × 10^ 8 bm12-derived splenocytes and were sacrificed 10 weeks post-induction. For NTS model, C57BL/6 mice were preimmunized with 0.2 mg sheep IgG intraperitoneally, followed by an intravenous injection of 50 μl sheep NTS on Day 4, and sacrificed on Day 11. C-E Immunoblotting of GSDMD in renal protein extracts of NZB/W F1 mice, cGVH or NTS mice with control (CON) mice. β-actin is loading control. F ELISA determining the level of autoantibodies (anti-dsDNA, anti-ssDNA and anti-histone) in serum of cGVH mice ( n = 4 or 5 per group). G Spleen to body weight ratio in cGVH mice ( n = 8 per group). H Representative PAS staining of kidney sections with quantitative analysis of glomerular cellularity and tubular injury scores in cGVH mice ( n = 5 or 10 per group). Scale bars: 50 μm. I Urine albumin to creatinine ratio in cGVH mice ( n = 8 per group). J Representative images and quantified immunofluorescence density of IgG and C3 deposition in the kidneys from cGVH mice ( n = 5 per group). Scale bars: 50 μm. K Quantitative analysis of glomerular PAS score and tubular injury scores in kidney sections from NTS mice (n = 5 per group). L Urine albumin to creatinine ratio and BUN in NTS mice (n = 5 per group). Data are shown as mean ± SEM. Two-way ANOVA test, ANOVA or Student’s t test was used for statistical analysis. **** P < 0.0001; *** P < 0.001; ** P < 0.01; * P < 0.05; ns, not significant
    Figure Legend Snippet: GSDMD deletion exacerbates lupus-like phenotype in cGVH and NTS models. A Immunochemical images of GSDMD-FL and GSDMD-N pattern in renal section from normal control (NC) and LN patients. Scale bars: 50 μm. B Schematic of murine lupus models. NZB/W F1 mice were sacrificed at 14 weeks and 36 weeks. For cGVH model, C57BL/6 mice received an intraperitoneal injection of 1 × 10^ 8 bm12-derived splenocytes and were sacrificed 10 weeks post-induction. For NTS model, C57BL/6 mice were preimmunized with 0.2 mg sheep IgG intraperitoneally, followed by an intravenous injection of 50 μl sheep NTS on Day 4, and sacrificed on Day 11. C-E Immunoblotting of GSDMD in renal protein extracts of NZB/W F1 mice, cGVH or NTS mice with control (CON) mice. β-actin is loading control. F ELISA determining the level of autoantibodies (anti-dsDNA, anti-ssDNA and anti-histone) in serum of cGVH mice ( n = 4 or 5 per group). G Spleen to body weight ratio in cGVH mice ( n = 8 per group). H Representative PAS staining of kidney sections with quantitative analysis of glomerular cellularity and tubular injury scores in cGVH mice ( n = 5 or 10 per group). Scale bars: 50 μm. I Urine albumin to creatinine ratio in cGVH mice ( n = 8 per group). J Representative images and quantified immunofluorescence density of IgG and C3 deposition in the kidneys from cGVH mice ( n = 5 per group). Scale bars: 50 μm. K Quantitative analysis of glomerular PAS score and tubular injury scores in kidney sections from NTS mice (n = 5 per group). L Urine albumin to creatinine ratio and BUN in NTS mice (n = 5 per group). Data are shown as mean ± SEM. Two-way ANOVA test, ANOVA or Student’s t test was used for statistical analysis. **** P < 0.0001; *** P < 0.001; ** P < 0.01; * P < 0.05; ns, not significant

    Techniques Used: Injection, Derivative Assay, Western Blot, Enzyme-linked Immunosorbent Assay, Staining, Immunofluorescence

    mouse anti human iκβα antibody  (Cell Signaling Technology Inc)


    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 86

    Structured Review

    Cell Signaling Technology Inc mouse anti human iκβα antibody
    Mouse Anti Human Iκβα Antibody, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mouse anti human iκβα antibody/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    mouse anti human iκβα antibody - by Bioz Stars, 2024-06
    86/100 stars

    Images

    human s292 mouse s293 antibody  (Cell Signaling Technology Inc)


    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 86

    Structured Review

    Cell Signaling Technology Inc human s292 mouse s293 antibody
    Human S292 Mouse S293 Antibody, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/human s292 mouse s293 antibody/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    human s292 mouse s293 antibody - by Bioz Stars, 2024-06
    86/100 stars

    Images

    mouse anti human phospho s536 nf κb antibody  (Cell Signaling Technology Inc)


    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 86

    Structured Review

    Cell Signaling Technology Inc mouse anti human phospho s536 nf κb antibody
    LOXL4-mediated regulation of the MMP9 through the activation <t>of</t> <t>NF-κB.</t> (A) The prepared conditioned media from one-day serum-free cultivation of the indicated cells were condensed 10-fold. The media specimens were subjected to SDS-PAGE and then applied to WB analysis to detect MMP or CBB staining as a sample control of proper loading. (B) The prepared whole cell extracts from the indicated cells under the standard culture were also subjected to SDS-PAGE and analyzed by WB for the proteins of interest. CBB visualized the loaded proteins and displayed the stained image as a control of proper sample loading. On the other hand, to correctly detect the phosphorylation levels of Iκβα, the indicated cells were all treated with MG-132 proteasome inhibitor to restore the Iκβα protein levels from their lowered levels caused by proteasome-mediated degradation. (C) The prepared nuclear extracts from the indicated cells were subjected to EMSA analysis to detect NF-κB activity. (D, E) WB analysis was done for the 10-fold condensed cell conditioned media and cell extracts from the LOXL4 wt-overexpressing clones to detect MMP9 (D) and indicated protein molecules (E) in a similar manner as described for panels (A) and (B) , except for the treatment with siRNAs (control siRNA: siCont; ITGB1 siRNA: siITGB1) or NF-κB inhibitor. (F) EMSA analysis of NF-κB was also performed for the nuclear extracts from the same series of cells, as shown in (E) .
    Mouse Anti Human Phospho S536 Nf κb Antibody, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mouse anti human phospho s536 nf κb antibody/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    mouse anti human phospho s536 nf κb antibody - by Bioz Stars, 2024-06
    86/100 stars

    Images

    1) Product Images from "Dissection of the signal transduction machinery responsible for the lysyl oxidase-like 4-mediated increase in invasive motility in triple-negative breast cancer cells: mechanistic insight into the integrin-β1-NF-κB-MMP9 axis"

    Article Title: Dissection of the signal transduction machinery responsible for the lysyl oxidase-like 4-mediated increase in invasive motility in triple-negative breast cancer cells: mechanistic insight into the integrin-β1-NF-κB-MMP9 axis

    Journal: Frontiers in Oncology

    doi: 10.3389/fonc.2024.1371307

    LOXL4-mediated regulation of the MMP9 through the activation of NF-κB. (A) The prepared conditioned media from one-day serum-free cultivation of the indicated cells were condensed 10-fold. The media specimens were subjected to SDS-PAGE and then applied to WB analysis to detect MMP or CBB staining as a sample control of proper loading. (B) The prepared whole cell extracts from the indicated cells under the standard culture were also subjected to SDS-PAGE and analyzed by WB for the proteins of interest. CBB visualized the loaded proteins and displayed the stained image as a control of proper sample loading. On the other hand, to correctly detect the phosphorylation levels of Iκβα, the indicated cells were all treated with MG-132 proteasome inhibitor to restore the Iκβα protein levels from their lowered levels caused by proteasome-mediated degradation. (C) The prepared nuclear extracts from the indicated cells were subjected to EMSA analysis to detect NF-κB activity. (D, E) WB analysis was done for the 10-fold condensed cell conditioned media and cell extracts from the LOXL4 wt-overexpressing clones to detect MMP9 (D) and indicated protein molecules (E) in a similar manner as described for panels (A) and (B) , except for the treatment with siRNAs (control siRNA: siCont; ITGB1 siRNA: siITGB1) or NF-κB inhibitor. (F) EMSA analysis of NF-κB was also performed for the nuclear extracts from the same series of cells, as shown in (E) .
    Figure Legend Snippet: LOXL4-mediated regulation of the MMP9 through the activation of NF-κB. (A) The prepared conditioned media from one-day serum-free cultivation of the indicated cells were condensed 10-fold. The media specimens were subjected to SDS-PAGE and then applied to WB analysis to detect MMP or CBB staining as a sample control of proper loading. (B) The prepared whole cell extracts from the indicated cells under the standard culture were also subjected to SDS-PAGE and analyzed by WB for the proteins of interest. CBB visualized the loaded proteins and displayed the stained image as a control of proper sample loading. On the other hand, to correctly detect the phosphorylation levels of Iκβα, the indicated cells were all treated with MG-132 proteasome inhibitor to restore the Iκβα protein levels from their lowered levels caused by proteasome-mediated degradation. (C) The prepared nuclear extracts from the indicated cells were subjected to EMSA analysis to detect NF-κB activity. (D, E) WB analysis was done for the 10-fold condensed cell conditioned media and cell extracts from the LOXL4 wt-overexpressing clones to detect MMP9 (D) and indicated protein molecules (E) in a similar manner as described for panels (A) and (B) , except for the treatment with siRNAs (control siRNA: siCont; ITGB1 siRNA: siITGB1) or NF-κB inhibitor. (F) EMSA analysis of NF-κB was also performed for the nuclear extracts from the same series of cells, as shown in (E) .

    Techniques Used: Activation Assay, SDS Page, Staining, Activity Assay, Clone Assay

    Identifying TRAF4 and TAK1 on the integrin-β1 signal cascade that links to NF-κB activation. (A) Schematic representation of the NF-κB activation flow via canonical and noncanonical pathways. (B, C) As indicated, the prepared cell lysates of HEK293T transfectants with different combinations of the plasmids were divided into three aliquots. The first was used as the whole cell lysate, i.e., as input (rightmost). The second was used to conduct immunoprecipitations with anti-HA tag antibody-conjugated agarose beads (middle). The remaining third was used to perform similar immunoprecipitations with anti-DYKDDDDK tag antibody-conjugated agarose beads as a negative control for the immunoprecipitation experiment (leftmost). After immunoprecipitation, the expressed products with beads-bound foreign proteins were analyzed by WB with an anti-HA, anti-Myc, or anti-albumin antibody. (D) The indicated cell extracts were incubated with the biotinylated (bio)-integrin-β1 antibody to capture the intrinsic integrin-β1 of the individual cells. On the other hand, bio-control IgG was used as a negative control for the same immunoprecipitation experiment. The captured integrin-β1 was collected by the pull-down method with avidin beads, and the precipitates were further subjected to WB with the indicated antibodies.
    Figure Legend Snippet: Identifying TRAF4 and TAK1 on the integrin-β1 signal cascade that links to NF-κB activation. (A) Schematic representation of the NF-κB activation flow via canonical and noncanonical pathways. (B, C) As indicated, the prepared cell lysates of HEK293T transfectants with different combinations of the plasmids were divided into three aliquots. The first was used as the whole cell lysate, i.e., as input (rightmost). The second was used to conduct immunoprecipitations with anti-HA tag antibody-conjugated agarose beads (middle). The remaining third was used to perform similar immunoprecipitations with anti-DYKDDDDK tag antibody-conjugated agarose beads as a negative control for the immunoprecipitation experiment (leftmost). After immunoprecipitation, the expressed products with beads-bound foreign proteins were analyzed by WB with an anti-HA, anti-Myc, or anti-albumin antibody. (D) The indicated cell extracts were incubated with the biotinylated (bio)-integrin-β1 antibody to capture the intrinsic integrin-β1 of the individual cells. On the other hand, bio-control IgG was used as a negative control for the same immunoprecipitation experiment. The captured integrin-β1 was collected by the pull-down method with avidin beads, and the precipitates were further subjected to WB with the indicated antibodies.

    Techniques Used: Activation Assay, Negative Control, Immunoprecipitation, Incubation, Avidin-Biotin Assay

    The beneficial role of TRAF4 and TAK1 in regulating MMP9 via NF-κB activation. (A, B) The indicated cells with transient transfection of the combination of RFP with the indicated genes (control empty vector EV, dominant negative TRAF4: TRAF4 dn, catalytically unfunctional kinase-dead TAK1: TAK1 KD) were collected by RFP-based cell sorting. None stands for intact non-treated cells with no cell sorting procedure, which were also used as control cells. WB analysis was done for the 10-fold condensed cell conditioned media and whole cell extracts from the LOXL4 wt-overexpressing clones to detect MMP9 (A) and the indicated protein molecules (B) following a procedure similar to that described in <xref ref-type= Figures 2A, B . (C) EMSA analysis of NF-κB was also performed for the whole extracts from the same series of cells, as shown in (B) , because there were too few cells to perform fractionation. (D) Invasion assay set with or without NF-κB inhibitor was carried out to evaluate the effect of NF-κB on the LOXL4-enhanced invasiveness. (E, F) Endogenous LOXL4 in cells and the secreted MMP9 in the conditioned media in the indicated cell lines were detected using the WB procedure (E) . On the other hand, zymography with gelatin substrate was performed to detect secretory MMP activity from the indicated cells. Specification of the MMP9-mediated digestive bands was done by the treatment of the gels with an MMP9-specific inhibitor (F) . Other running gels with no gelatin, also loaded with identical specimens corresponding to WB and zymography, were stained with CBB as sample controls of proper loading (E, F) . (G) The conditioned media prepared from the indicated cells treated or not treated with LOX inhibitor BAPN, MMP inhibitor, or NF-κB inhibitor were subjected to WB (upper) and zymography (lower) to detect their MMP9 proteins and activities. (H) A trans-chamber-based cell invasion assay also evaluated the effects of these inhibitors used in (G) on the invasive activities in the indicated cells. (I) Schematic diagram of the molecular interplay among the indicated vital molecules. (J) Gene expression plots of MMP9 mRNA from breast invasive carcinoma (BRCA) specimens were obtained from a publicly available website ( http://gepia.cancer-pku.cn/ ). (K) Overall survival plots for MMP9 protein expression levels were obtained from a publicly available website ( http://kmplot.com/analysis/ ). Data are mean ± SD. *p<0.05, **p<0.01, ***p<0.001. " title="... role of TRAF4 and TAK1 in regulating MMP9 via NF-κB activation. (A, B) The indicated cells with ..." property="contentUrl" width="100%" height="100%"/>
    Figure Legend Snippet: The beneficial role of TRAF4 and TAK1 in regulating MMP9 via NF-κB activation. (A, B) The indicated cells with transient transfection of the combination of RFP with the indicated genes (control empty vector EV, dominant negative TRAF4: TRAF4 dn, catalytically unfunctional kinase-dead TAK1: TAK1 KD) were collected by RFP-based cell sorting. None stands for intact non-treated cells with no cell sorting procedure, which were also used as control cells. WB analysis was done for the 10-fold condensed cell conditioned media and whole cell extracts from the LOXL4 wt-overexpressing clones to detect MMP9 (A) and the indicated protein molecules (B) following a procedure similar to that described in Figures 2A, B . (C) EMSA analysis of NF-κB was also performed for the whole extracts from the same series of cells, as shown in (B) , because there were too few cells to perform fractionation. (D) Invasion assay set with or without NF-κB inhibitor was carried out to evaluate the effect of NF-κB on the LOXL4-enhanced invasiveness. (E, F) Endogenous LOXL4 in cells and the secreted MMP9 in the conditioned media in the indicated cell lines were detected using the WB procedure (E) . On the other hand, zymography with gelatin substrate was performed to detect secretory MMP activity from the indicated cells. Specification of the MMP9-mediated digestive bands was done by the treatment of the gels with an MMP9-specific inhibitor (F) . Other running gels with no gelatin, also loaded with identical specimens corresponding to WB and zymography, were stained with CBB as sample controls of proper loading (E, F) . (G) The conditioned media prepared from the indicated cells treated or not treated with LOX inhibitor BAPN, MMP inhibitor, or NF-κB inhibitor were subjected to WB (upper) and zymography (lower) to detect their MMP9 proteins and activities. (H) A trans-chamber-based cell invasion assay also evaluated the effects of these inhibitors used in (G) on the invasive activities in the indicated cells. (I) Schematic diagram of the molecular interplay among the indicated vital molecules. (J) Gene expression plots of MMP9 mRNA from breast invasive carcinoma (BRCA) specimens were obtained from a publicly available website ( http://gepia.cancer-pku.cn/ ). (K) Overall survival plots for MMP9 protein expression levels were obtained from a publicly available website ( http://kmplot.com/analysis/ ). Data are mean ± SD. *p<0.05, **p<0.01, ***p<0.001.

    Techniques Used: Activation Assay, Transfection, Plasmid Preparation, Dominant Negative Mutation, FACS, Clone Assay, Fractionation, Invasion Assay, Zymography, Activity Assay, Staining, Expressing

    mouse anti human epcam antibody 5488s  (Cell Signaling Technology Inc)


    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 86

    Structured Review

    Cell Signaling Technology Inc mouse anti human epcam antibody 5488s
    Mouse Anti Human Epcam Antibody 5488s, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mouse anti human epcam antibody 5488s/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    mouse anti human epcam antibody 5488s - by Bioz Stars, 2024-06
    86/100 stars

    Images

    mouse anti human cd68 antibody  (Cell Signaling Technology Inc)


    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 86

    Structured Review

    Cell Signaling Technology Inc mouse anti human cd68 antibody
    Mouse Anti Human Cd68 Antibody, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mouse anti human cd68 antibody/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    mouse anti human cd68 antibody - by Bioz Stars, 2024-06
    86/100 stars

    Images

    primary mouse anti human cd68 antibody  (Cell Signaling Technology Inc)


    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 86

    Structured Review

    Cell Signaling Technology Inc primary mouse anti human cd68 antibody
    Primary Mouse Anti Human Cd68 Antibody, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/primary mouse anti human cd68 antibody/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    primary mouse anti human cd68 antibody - by Bioz Stars, 2024-06
    86/100 stars

    Images

    Similar Products

  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 86
    Cell Signaling Technology Inc ikkα mouse anti human monoclonal 1 1000 cat no 11930 cell signaling technology
    Ikkα Mouse Anti Human Monoclonal 1 1000 Cat No 11930 Cell Signaling Technology, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/ikkα mouse anti human monoclonal 1 1000 cat no 11930 cell signaling technology/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    ikkα mouse anti human monoclonal 1 1000 cat no 11930 cell signaling technology - by Bioz Stars, 2024-06
    86/100 stars
      Buy from Supplier

    86
    Cell Signaling Technology Inc mouse monoclonal anti human β actin antibodies
    A P3 ESC show increased nuclear p21 (green signal, left panel) after IL-1β-treatment. <t>β-Actin</t> (red) staining is more diffuse in IL-1β-treated ESC and the cellular orientation is more parallel. Merged channel images (yellow signal, right panels) are included. Magnification × 200. B SA-β-gal staining was used to detect β-galactosidase-positive signals. There was an intense blue color in the cytoplasm of ESC following 24 h of IL-1β treatment (right panel) compared to the untreated control (left panel). Magnification × 200.
    Mouse Monoclonal Anti Human β Actin Antibodies, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mouse monoclonal anti human β actin antibodies/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    mouse monoclonal anti human β actin antibodies - by Bioz Stars, 2024-06
    86/100 stars
      Buy from Supplier

    86
    Cell Signaling Technology Inc anti human mouse fra 1
    IL-33 Enhances the Expression of Downstream Target Genes of WNT/β-catenin Pathway. A RNA expression of downstream target genes of WNT/β-catenin ( AXIN2, CCN4, CCND1, CD44, <t>Fra-1,</t> JUN, LGR5, MYC, and PPARD ) were detected via real-time PCR in EECs and 12Z cells. B Fra-1 expression in EECs and 12Z cells was detected by western blotting. C CCN4 secretion in the cell supernatant of EECs and 12Z cells was detected via ELISA. D - F After IL-33 or/and ST2 neutralizing antibody treatment, the mRNA expression and protein levels of Fra-1 and CCN4 in EECs and 12Z cells. G Immunohistochemical staining of Fra-1 and CCN4 in eutopic endometrium and ectopic lesions. Data are presented as mean ± SEM. All data were analyzed using one-way ANOVA followed by Dunnett’s post hoc test and Student’s t-test ; * P < 0.05, ** P < 0.01, *** P < 0.001
    Anti Human Mouse Fra 1, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti human mouse fra 1/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    anti human mouse fra 1 - by Bioz Stars, 2024-06
    86/100 stars
      Buy from Supplier

    86
    Cell Signaling Technology Inc anti human mouse β actin
    GSDMD deletion exacerbates lupus-like phenotype in cGVH and NTS models. A Immunochemical images of GSDMD-FL and GSDMD-N pattern in renal section from normal control (NC) and LN patients. Scale bars: 50 μm. B Schematic of murine lupus models. NZB/W F1 mice were sacrificed at 14 weeks and 36 weeks. For cGVH model, C57BL/6 mice received an intraperitoneal injection of 1 × 10^ 8 bm12-derived splenocytes and were sacrificed 10 weeks post-induction. For NTS model, C57BL/6 mice were preimmunized with 0.2 mg sheep IgG intraperitoneally, followed by an intravenous injection of 50 μl sheep NTS on Day 4, and sacrificed on Day 11. C-E Immunoblotting of GSDMD in renal protein extracts of NZB/W F1 mice, cGVH or NTS mice with control (CON) mice. <t>β-actin</t> is loading control. F ELISA determining the level of autoantibodies (anti-dsDNA, anti-ssDNA and anti-histone) in serum of cGVH mice ( n = 4 or 5 per group). G Spleen to body weight ratio in cGVH mice ( n = 8 per group). H Representative PAS staining of kidney sections with quantitative analysis of glomerular cellularity and tubular injury scores in cGVH mice ( n = 5 or 10 per group). Scale bars: 50 μm. I Urine albumin to creatinine ratio in cGVH mice ( n = 8 per group). J Representative images and quantified immunofluorescence density of IgG and C3 deposition in the kidneys from cGVH mice ( n = 5 per group). Scale bars: 50 μm. K Quantitative analysis of glomerular PAS score and tubular injury scores in kidney sections from NTS mice (n = 5 per group). L Urine albumin to creatinine ratio and BUN in NTS mice (n = 5 per group). Data are shown as mean ± SEM. Two-way ANOVA test, ANOVA or Student’s t test was used for statistical analysis. **** P < 0.0001; *** P < 0.001; ** P < 0.01; * P < 0.05; ns, not significant
    Anti Human Mouse β Actin, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti human mouse β actin/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    anti human mouse β actin - by Bioz Stars, 2024-06
    86/100 stars
      Buy from Supplier

    86
    Cell Signaling Technology Inc mouse anti human iκβα antibody
    GSDMD deletion exacerbates lupus-like phenotype in cGVH and NTS models. A Immunochemical images of GSDMD-FL and GSDMD-N pattern in renal section from normal control (NC) and LN patients. Scale bars: 50 μm. B Schematic of murine lupus models. NZB/W F1 mice were sacrificed at 14 weeks and 36 weeks. For cGVH model, C57BL/6 mice received an intraperitoneal injection of 1 × 10^ 8 bm12-derived splenocytes and were sacrificed 10 weeks post-induction. For NTS model, C57BL/6 mice were preimmunized with 0.2 mg sheep IgG intraperitoneally, followed by an intravenous injection of 50 μl sheep NTS on Day 4, and sacrificed on Day 11. C-E Immunoblotting of GSDMD in renal protein extracts of NZB/W F1 mice, cGVH or NTS mice with control (CON) mice. <t>β-actin</t> is loading control. F ELISA determining the level of autoantibodies (anti-dsDNA, anti-ssDNA and anti-histone) in serum of cGVH mice ( n = 4 or 5 per group). G Spleen to body weight ratio in cGVH mice ( n = 8 per group). H Representative PAS staining of kidney sections with quantitative analysis of glomerular cellularity and tubular injury scores in cGVH mice ( n = 5 or 10 per group). Scale bars: 50 μm. I Urine albumin to creatinine ratio in cGVH mice ( n = 8 per group). J Representative images and quantified immunofluorescence density of IgG and C3 deposition in the kidneys from cGVH mice ( n = 5 per group). Scale bars: 50 μm. K Quantitative analysis of glomerular PAS score and tubular injury scores in kidney sections from NTS mice (n = 5 per group). L Urine albumin to creatinine ratio and BUN in NTS mice (n = 5 per group). Data are shown as mean ± SEM. Two-way ANOVA test, ANOVA or Student’s t test was used for statistical analysis. **** P < 0.0001; *** P < 0.001; ** P < 0.01; * P < 0.05; ns, not significant
    Mouse Anti Human Iκβα Antibody, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mouse anti human iκβα antibody/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    mouse anti human iκβα antibody - by Bioz Stars, 2024-06
    86/100 stars
      Buy from Supplier

    86
    Cell Signaling Technology Inc human s292 mouse s293 antibody
    GSDMD deletion exacerbates lupus-like phenotype in cGVH and NTS models. A Immunochemical images of GSDMD-FL and GSDMD-N pattern in renal section from normal control (NC) and LN patients. Scale bars: 50 μm. B Schematic of murine lupus models. NZB/W F1 mice were sacrificed at 14 weeks and 36 weeks. For cGVH model, C57BL/6 mice received an intraperitoneal injection of 1 × 10^ 8 bm12-derived splenocytes and were sacrificed 10 weeks post-induction. For NTS model, C57BL/6 mice were preimmunized with 0.2 mg sheep IgG intraperitoneally, followed by an intravenous injection of 50 μl sheep NTS on Day 4, and sacrificed on Day 11. C-E Immunoblotting of GSDMD in renal protein extracts of NZB/W F1 mice, cGVH or NTS mice with control (CON) mice. <t>β-actin</t> is loading control. F ELISA determining the level of autoantibodies (anti-dsDNA, anti-ssDNA and anti-histone) in serum of cGVH mice ( n = 4 or 5 per group). G Spleen to body weight ratio in cGVH mice ( n = 8 per group). H Representative PAS staining of kidney sections with quantitative analysis of glomerular cellularity and tubular injury scores in cGVH mice ( n = 5 or 10 per group). Scale bars: 50 μm. I Urine albumin to creatinine ratio in cGVH mice ( n = 8 per group). J Representative images and quantified immunofluorescence density of IgG and C3 deposition in the kidneys from cGVH mice ( n = 5 per group). Scale bars: 50 μm. K Quantitative analysis of glomerular PAS score and tubular injury scores in kidney sections from NTS mice (n = 5 per group). L Urine albumin to creatinine ratio and BUN in NTS mice (n = 5 per group). Data are shown as mean ± SEM. Two-way ANOVA test, ANOVA or Student’s t test was used for statistical analysis. **** P < 0.0001; *** P < 0.001; ** P < 0.01; * P < 0.05; ns, not significant
    Human S292 Mouse S293 Antibody, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/human s292 mouse s293 antibody/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    human s292 mouse s293 antibody - by Bioz Stars, 2024-06
    86/100 stars
      Buy from Supplier

    86
    Cell Signaling Technology Inc mouse anti human phospho s536 nf κb antibody
    LOXL4-mediated regulation of the MMP9 through the activation <t>of</t> <t>NF-κB.</t> (A) The prepared conditioned media from one-day serum-free cultivation of the indicated cells were condensed 10-fold. The media specimens were subjected to SDS-PAGE and then applied to WB analysis to detect MMP or CBB staining as a sample control of proper loading. (B) The prepared whole cell extracts from the indicated cells under the standard culture were also subjected to SDS-PAGE and analyzed by WB for the proteins of interest. CBB visualized the loaded proteins and displayed the stained image as a control of proper sample loading. On the other hand, to correctly detect the phosphorylation levels of Iκβα, the indicated cells were all treated with MG-132 proteasome inhibitor to restore the Iκβα protein levels from their lowered levels caused by proteasome-mediated degradation. (C) The prepared nuclear extracts from the indicated cells were subjected to EMSA analysis to detect NF-κB activity. (D, E) WB analysis was done for the 10-fold condensed cell conditioned media and cell extracts from the LOXL4 wt-overexpressing clones to detect MMP9 (D) and indicated protein molecules (E) in a similar manner as described for panels (A) and (B) , except for the treatment with siRNAs (control siRNA: siCont; ITGB1 siRNA: siITGB1) or NF-κB inhibitor. (F) EMSA analysis of NF-κB was also performed for the nuclear extracts from the same series of cells, as shown in (E) .
    Mouse Anti Human Phospho S536 Nf κb Antibody, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mouse anti human phospho s536 nf κb antibody/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    mouse anti human phospho s536 nf κb antibody - by Bioz Stars, 2024-06
    86/100 stars
      Buy from Supplier

    86
    Cell Signaling Technology Inc mouse anti human epcam antibody 5488s
    LOXL4-mediated regulation of the MMP9 through the activation <t>of</t> <t>NF-κB.</t> (A) The prepared conditioned media from one-day serum-free cultivation of the indicated cells were condensed 10-fold. The media specimens were subjected to SDS-PAGE and then applied to WB analysis to detect MMP or CBB staining as a sample control of proper loading. (B) The prepared whole cell extracts from the indicated cells under the standard culture were also subjected to SDS-PAGE and analyzed by WB for the proteins of interest. CBB visualized the loaded proteins and displayed the stained image as a control of proper sample loading. On the other hand, to correctly detect the phosphorylation levels of Iκβα, the indicated cells were all treated with MG-132 proteasome inhibitor to restore the Iκβα protein levels from their lowered levels caused by proteasome-mediated degradation. (C) The prepared nuclear extracts from the indicated cells were subjected to EMSA analysis to detect NF-κB activity. (D, E) WB analysis was done for the 10-fold condensed cell conditioned media and cell extracts from the LOXL4 wt-overexpressing clones to detect MMP9 (D) and indicated protein molecules (E) in a similar manner as described for panels (A) and (B) , except for the treatment with siRNAs (control siRNA: siCont; ITGB1 siRNA: siITGB1) or NF-κB inhibitor. (F) EMSA analysis of NF-κB was also performed for the nuclear extracts from the same series of cells, as shown in (E) .
    Mouse Anti Human Epcam Antibody 5488s, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mouse anti human epcam antibody 5488s/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    mouse anti human epcam antibody 5488s - by Bioz Stars, 2024-06
    86/100 stars
      Buy from Supplier

    86
    Cell Signaling Technology Inc mouse anti human cd68 antibody
    LOXL4-mediated regulation of the MMP9 through the activation <t>of</t> <t>NF-κB.</t> (A) The prepared conditioned media from one-day serum-free cultivation of the indicated cells were condensed 10-fold. The media specimens were subjected to SDS-PAGE and then applied to WB analysis to detect MMP or CBB staining as a sample control of proper loading. (B) The prepared whole cell extracts from the indicated cells under the standard culture were also subjected to SDS-PAGE and analyzed by WB for the proteins of interest. CBB visualized the loaded proteins and displayed the stained image as a control of proper sample loading. On the other hand, to correctly detect the phosphorylation levels of Iκβα, the indicated cells were all treated with MG-132 proteasome inhibitor to restore the Iκβα protein levels from their lowered levels caused by proteasome-mediated degradation. (C) The prepared nuclear extracts from the indicated cells were subjected to EMSA analysis to detect NF-κB activity. (D, E) WB analysis was done for the 10-fold condensed cell conditioned media and cell extracts from the LOXL4 wt-overexpressing clones to detect MMP9 (D) and indicated protein molecules (E) in a similar manner as described for panels (A) and (B) , except for the treatment with siRNAs (control siRNA: siCont; ITGB1 siRNA: siITGB1) or NF-κB inhibitor. (F) EMSA analysis of NF-κB was also performed for the nuclear extracts from the same series of cells, as shown in (E) .
    Mouse Anti Human Cd68 Antibody, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mouse anti human cd68 antibody/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    mouse anti human cd68 antibody - by Bioz Stars, 2024-06
    86/100 stars
      Buy from Supplier

    86
    Cell Signaling Technology Inc primary mouse anti human cd68 antibody
    LOXL4-mediated regulation of the MMP9 through the activation <t>of</t> <t>NF-κB.</t> (A) The prepared conditioned media from one-day serum-free cultivation of the indicated cells were condensed 10-fold. The media specimens were subjected to SDS-PAGE and then applied to WB analysis to detect MMP or CBB staining as a sample control of proper loading. (B) The prepared whole cell extracts from the indicated cells under the standard culture were also subjected to SDS-PAGE and analyzed by WB for the proteins of interest. CBB visualized the loaded proteins and displayed the stained image as a control of proper sample loading. On the other hand, to correctly detect the phosphorylation levels of Iκβα, the indicated cells were all treated with MG-132 proteasome inhibitor to restore the Iκβα protein levels from their lowered levels caused by proteasome-mediated degradation. (C) The prepared nuclear extracts from the indicated cells were subjected to EMSA analysis to detect NF-κB activity. (D, E) WB analysis was done for the 10-fold condensed cell conditioned media and cell extracts from the LOXL4 wt-overexpressing clones to detect MMP9 (D) and indicated protein molecules (E) in a similar manner as described for panels (A) and (B) , except for the treatment with siRNAs (control siRNA: siCont; ITGB1 siRNA: siITGB1) or NF-κB inhibitor. (F) EMSA analysis of NF-κB was also performed for the nuclear extracts from the same series of cells, as shown in (E) .
    Primary Mouse Anti Human Cd68 Antibody, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/primary mouse anti human cd68 antibody/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    primary mouse anti human cd68 antibody - by Bioz Stars, 2024-06
    86/100 stars
      Buy from Supplier

    Image Search Results


    A P3 ESC show increased nuclear p21 (green signal, left panel) after IL-1β-treatment. β-Actin (red) staining is more diffuse in IL-1β-treated ESC and the cellular orientation is more parallel. Merged channel images (yellow signal, right panels) are included. Magnification × 200. B SA-β-gal staining was used to detect β-galactosidase-positive signals. There was an intense blue color in the cytoplasm of ESC following 24 h of IL-1β treatment (right panel) compared to the untreated control (left panel). Magnification × 200.

    Journal: Cell Death Discovery

    Article Title: Interleukin-1β induces and accelerates human endometrial stromal cell senescence and impairs decidualization via the c-Jun N-terminal kinase pathway

    doi: 10.1038/s41420-024-02048-6

    Figure Lengend Snippet: A P3 ESC show increased nuclear p21 (green signal, left panel) after IL-1β-treatment. β-Actin (red) staining is more diffuse in IL-1β-treated ESC and the cellular orientation is more parallel. Merged channel images (yellow signal, right panels) are included. Magnification × 200. B SA-β-gal staining was used to detect β-galactosidase-positive signals. There was an intense blue color in the cytoplasm of ESC following 24 h of IL-1β treatment (right panel) compared to the untreated control (left panel). Magnification × 200.

    Article Snippet: Blots were washed, re-probed with mouse monoclonal anti-human β-actin antibodies (1:1000 dilution, cat# 3700, Cell Signaling) to confirm evenness of loading.

    Techniques: Staining

    A After 48 h incubation, Western blotting shows that IL-1β increased IL-1β, IL-6, TNF-α, MMP3, p16, p21, CCL2 and phospho-histone H2A.X in P3 ESC. B Time-course experiments reveal that IL-1β upregulated IL-1β, IL-6, TNF-α, MMP3, p16, p21, HMGB1, CCL2, CCL5 and phospho-histone H2A.X at 24, 48 and 72 h (lanes 2, 4 and 6). Co-incubation in the presence of the JNK inhibitor SP blocked IL-1β-mediated upregulation at all time points (lanes 3, 5 and 7). Incubation with SP alone (lane 8) suppressed many biomarkers below basal (Control, lane 1) levels. β-Actin levels were not affected. Molecular masses (kDa) are indicated by arrows at right. C Based on the time course experiments, the three key senescence-associated biomarkers indicated that the baseline levels of IL-1β and IL-6 were notably low and undetectable by the western blot method at all three time points. Meanwhile, MMP3 expression levels in the untreated control group remained consistently similar across the three observed time points. β-Actin levels were not affected. Molecular masses (kDa) are indicated by arrows at right.

    Journal: Cell Death Discovery

    Article Title: Interleukin-1β induces and accelerates human endometrial stromal cell senescence and impairs decidualization via the c-Jun N-terminal kinase pathway

    doi: 10.1038/s41420-024-02048-6

    Figure Lengend Snippet: A After 48 h incubation, Western blotting shows that IL-1β increased IL-1β, IL-6, TNF-α, MMP3, p16, p21, CCL2 and phospho-histone H2A.X in P3 ESC. B Time-course experiments reveal that IL-1β upregulated IL-1β, IL-6, TNF-α, MMP3, p16, p21, HMGB1, CCL2, CCL5 and phospho-histone H2A.X at 24, 48 and 72 h (lanes 2, 4 and 6). Co-incubation in the presence of the JNK inhibitor SP blocked IL-1β-mediated upregulation at all time points (lanes 3, 5 and 7). Incubation with SP alone (lane 8) suppressed many biomarkers below basal (Control, lane 1) levels. β-Actin levels were not affected. Molecular masses (kDa) are indicated by arrows at right. C Based on the time course experiments, the three key senescence-associated biomarkers indicated that the baseline levels of IL-1β and IL-6 were notably low and undetectable by the western blot method at all three time points. Meanwhile, MMP3 expression levels in the untreated control group remained consistently similar across the three observed time points. β-Actin levels were not affected. Molecular masses (kDa) are indicated by arrows at right.

    Article Snippet: Blots were washed, re-probed with mouse monoclonal anti-human β-actin antibodies (1:1000 dilution, cat# 3700, Cell Signaling) to confirm evenness of loading.

    Techniques: Incubation, Western Blot, Expressing

    A ELISA confirmed that CCL5 secretion by P3 ESC from each of four subjects was upregulated after 72 h of IL-1β treatment. This effect was reversed by co-incubation with SP ( p < 0.001, ANOVA). Error bars indicate SD of triplicate determinations for subject S4. B When IL-1β was co-incubated with a 100-fold excess of IL-1ra , complete abrogation of the IL-1β effect on IL-8 and IL-6 upregulation was observed. The overall comparison using two-factor ANOVA analysis revealed significant differences ( p < 0.001). Scheffé Test comparisons showed significant differences between Control vs IL-1β (p < 0.001) and IL-1β vs IL-1ra ( p < 0.001). Error bars indicate SD of triplicate determinations. C Time-course of CCL5 ELISA response to IL-1β ± SP (24–72 h). The overall comparison using one-factor ANOVA revealed significant differences between groups (p < 0.001). Error bars indicate SD of triplicate determinations. D Time-course of TNF-α ELISA response to IL-1β ± SP (24–72 h). The overall comparison using one-factor ANOVA revealed significant differences between groups ( p < 0.001). Error bars indicate SD of triplicate determinations. E Western blot showing that preincubation of P3 ESC with SP, for 1 h before addition of IL-1β (PSP), had the same inhibitory effect as simultaneous co-incubation of IL-1β plus SP (SSP) on IL-1β, IL-6, TNF-α and MMP3 reduction. SP alone reduced basal (control) SASP markers. β-Actin levels were not affected. F Western blot reveals inhibition of IL-1β stimulation of IL-1β, IL - 6, MMP3, p21 and HMGB1 by JNK1 (CJ1) or JNK2 (CJ2) siRNA constructs transfected into P3 ESC. β-Actin levels were not affected.

    Journal: Cell Death Discovery

    Article Title: Interleukin-1β induces and accelerates human endometrial stromal cell senescence and impairs decidualization via the c-Jun N-terminal kinase pathway

    doi: 10.1038/s41420-024-02048-6

    Figure Lengend Snippet: A ELISA confirmed that CCL5 secretion by P3 ESC from each of four subjects was upregulated after 72 h of IL-1β treatment. This effect was reversed by co-incubation with SP ( p < 0.001, ANOVA). Error bars indicate SD of triplicate determinations for subject S4. B When IL-1β was co-incubated with a 100-fold excess of IL-1ra , complete abrogation of the IL-1β effect on IL-8 and IL-6 upregulation was observed. The overall comparison using two-factor ANOVA analysis revealed significant differences ( p < 0.001). Scheffé Test comparisons showed significant differences between Control vs IL-1β (p < 0.001) and IL-1β vs IL-1ra ( p < 0.001). Error bars indicate SD of triplicate determinations. C Time-course of CCL5 ELISA response to IL-1β ± SP (24–72 h). The overall comparison using one-factor ANOVA revealed significant differences between groups (p < 0.001). Error bars indicate SD of triplicate determinations. D Time-course of TNF-α ELISA response to IL-1β ± SP (24–72 h). The overall comparison using one-factor ANOVA revealed significant differences between groups ( p < 0.001). Error bars indicate SD of triplicate determinations. E Western blot showing that preincubation of P3 ESC with SP, for 1 h before addition of IL-1β (PSP), had the same inhibitory effect as simultaneous co-incubation of IL-1β plus SP (SSP) on IL-1β, IL-6, TNF-α and MMP3 reduction. SP alone reduced basal (control) SASP markers. β-Actin levels were not affected. F Western blot reveals inhibition of IL-1β stimulation of IL-1β, IL - 6, MMP3, p21 and HMGB1 by JNK1 (CJ1) or JNK2 (CJ2) siRNA constructs transfected into P3 ESC. β-Actin levels were not affected.

    Article Snippet: Blots were washed, re-probed with mouse monoclonal anti-human β-actin antibodies (1:1000 dilution, cat# 3700, Cell Signaling) to confirm evenness of loading.

    Techniques: Enzyme-linked Immunosorbent Assay, Incubation, Comparison, Western Blot, Inhibition, Construct, Transfection

    A Immunofluorescence- (IFC, top panels) and immunoperoxidase cytochemistry (IPC, lower panels) were used to localize IL-1β in ESC before and after IL-1β stimulation in P3 cells. B IFC (top panels) and IPC (lower panels) identified more IL-1β after IL-1β stimulation in P21 cells than P3 cells. Cell orientation in P21 cells also appears more parallel. C Western blot shows IL-1β, IL-6, TNF-α, MMP3, p16, p21 all increased with extended cell passage (P3 to P21) after incubation with IL-1β. β-Actin levels were unchanged.

    Journal: Cell Death Discovery

    Article Title: Interleukin-1β induces and accelerates human endometrial stromal cell senescence and impairs decidualization via the c-Jun N-terminal kinase pathway

    doi: 10.1038/s41420-024-02048-6

    Figure Lengend Snippet: A Immunofluorescence- (IFC, top panels) and immunoperoxidase cytochemistry (IPC, lower panels) were used to localize IL-1β in ESC before and after IL-1β stimulation in P3 cells. B IFC (top panels) and IPC (lower panels) identified more IL-1β after IL-1β stimulation in P21 cells than P3 cells. Cell orientation in P21 cells also appears more parallel. C Western blot shows IL-1β, IL-6, TNF-α, MMP3, p16, p21 all increased with extended cell passage (P3 to P21) after incubation with IL-1β. β-Actin levels were unchanged.

    Article Snippet: Blots were washed, re-probed with mouse monoclonal anti-human β-actin antibodies (1:1000 dilution, cat# 3700, Cell Signaling) to confirm evenness of loading.

    Techniques: Immunofluorescence, Western Blot, Incubation

    IL-33 Enhances the Expression of Downstream Target Genes of WNT/β-catenin Pathway. A RNA expression of downstream target genes of WNT/β-catenin ( AXIN2, CCN4, CCND1, CD44, Fra-1, JUN, LGR5, MYC, and PPARD ) were detected via real-time PCR in EECs and 12Z cells. B Fra-1 expression in EECs and 12Z cells was detected by western blotting. C CCN4 secretion in the cell supernatant of EECs and 12Z cells was detected via ELISA. D - F After IL-33 or/and ST2 neutralizing antibody treatment, the mRNA expression and protein levels of Fra-1 and CCN4 in EECs and 12Z cells. G Immunohistochemical staining of Fra-1 and CCN4 in eutopic endometrium and ectopic lesions. Data are presented as mean ± SEM. All data were analyzed using one-way ANOVA followed by Dunnett’s post hoc test and Student’s t-test ; * P < 0.05, ** P < 0.01, *** P < 0.001

    Journal: Cell Communication and Signaling : CCS

    Article Title: The IL-33-ST2 axis plays a vital role in endometriosis via promoting epithelial–mesenchymal transition by phosphorylating β-catenin

    doi: 10.1186/s12964-024-01683-x

    Figure Lengend Snippet: IL-33 Enhances the Expression of Downstream Target Genes of WNT/β-catenin Pathway. A RNA expression of downstream target genes of WNT/β-catenin ( AXIN2, CCN4, CCND1, CD44, Fra-1, JUN, LGR5, MYC, and PPARD ) were detected via real-time PCR in EECs and 12Z cells. B Fra-1 expression in EECs and 12Z cells was detected by western blotting. C CCN4 secretion in the cell supernatant of EECs and 12Z cells was detected via ELISA. D - F After IL-33 or/and ST2 neutralizing antibody treatment, the mRNA expression and protein levels of Fra-1 and CCN4 in EECs and 12Z cells. G Immunohistochemical staining of Fra-1 and CCN4 in eutopic endometrium and ectopic lesions. Data are presented as mean ± SEM. All data were analyzed using one-way ANOVA followed by Dunnett’s post hoc test and Student’s t-test ; * P < 0.05, ** P < 0.01, *** P < 0.001

    Article Snippet: The following IHC antibodies were used: anti-human ST2 (1:200), anti-human CCN4 (1:200), and anti-human E-cadherin (1:500) (abcam, Cambridge, UK); anti-human vimentin (1:100) (Cell Signaling Technology, Boston, USA); anti-human/mouse Fra-1 (1:200), anti-mouse CCN4 (1:100), and anti-mouse vimentin (1:100) (Affinity Biosciences, China); The primary antibodies used are listed in Supplementary Table 1.

    Techniques: Expressing, RNA Expression, Real-time Polymerase Chain Reaction, Western Blot, Enzyme-linked Immunosorbent Assay, Immunohistochemical staining, Staining

    IL-33 obviously Induced β-catenin Phosphorylation. A Protein expression of phospho-β-catenin (Ser675 and Ser552), phospho-CREB (Ser133), CREB, phospho-AKT(Ser473) and AKT in EECs and 12Z cells treated by IL-33 or/and ST2 neutralizing antibody. B Protein expression of phospho-β-catenin (Ser675 and Ser552), phospho-CREB (Ser133), CREB, phospho-AKT (Ser473), AKT, and Fra-1 in EECs and 12Z cells treated by IL-33 or/and knocking down ST2 by siRNA. C Expression of CCN4 in supernatant of EEC and 12Z treated by IL-33 or/and knocking down ST2 by siRNA. Data are presented as mean ± SEM. All data were analyzed using one-way ANOVA followed by Dunnett’s post hoc test and Student’s t-test ; * p < 0.05, ** p < 0.01, *** p < 0.001

    Journal: Cell Communication and Signaling : CCS

    Article Title: The IL-33-ST2 axis plays a vital role in endometriosis via promoting epithelial–mesenchymal transition by phosphorylating β-catenin

    doi: 10.1186/s12964-024-01683-x

    Figure Lengend Snippet: IL-33 obviously Induced β-catenin Phosphorylation. A Protein expression of phospho-β-catenin (Ser675 and Ser552), phospho-CREB (Ser133), CREB, phospho-AKT(Ser473) and AKT in EECs and 12Z cells treated by IL-33 or/and ST2 neutralizing antibody. B Protein expression of phospho-β-catenin (Ser675 and Ser552), phospho-CREB (Ser133), CREB, phospho-AKT (Ser473), AKT, and Fra-1 in EECs and 12Z cells treated by IL-33 or/and knocking down ST2 by siRNA. C Expression of CCN4 in supernatant of EEC and 12Z treated by IL-33 or/and knocking down ST2 by siRNA. Data are presented as mean ± SEM. All data were analyzed using one-way ANOVA followed by Dunnett’s post hoc test and Student’s t-test ; * p < 0.05, ** p < 0.01, *** p < 0.001

    Article Snippet: The following IHC antibodies were used: anti-human ST2 (1:200), anti-human CCN4 (1:200), and anti-human E-cadherin (1:500) (abcam, Cambridge, UK); anti-human vimentin (1:100) (Cell Signaling Technology, Boston, USA); anti-human/mouse Fra-1 (1:200), anti-mouse CCN4 (1:100), and anti-mouse vimentin (1:100) (Affinity Biosciences, China); The primary antibodies used are listed in Supplementary Table 1.

    Techniques: Expressing

    IL-33 Mediates β-catenin Phosphorylation at Ser675 and Ser552 by PKA. A , B Protein expression of EMT-related proteins (vimentin, E-cadherin, N-cadherin, and β-catenin), Fra-1,phospho-β-catenin (Ser675 and Ser552), phospho-CREB (Ser133), CREB, phospho-AKT (Ser473) and AKT in EEC and 12Z treated by IL-33, PKA inhibitor-H89 or/and AKT inhibitor- MK2206. C Expression of CCN4 in supernatant of EEC and 12Z treated by IL-33, PKA inhibitor- H89 or/and AKT inhibitor- MK2206. D , F Cyclohexane chase assay of β-catenin in EECs and 12Z cells treated by IL-33. Data are presented as mean ± SEM. All data were analyzed using one-way ANOVA followed by Dunnett’s post hoc test and Student’s t-test ; * P < 0.05, ** P < 0.01, *** P < 0.001

    Journal: Cell Communication and Signaling : CCS

    Article Title: The IL-33-ST2 axis plays a vital role in endometriosis via promoting epithelial–mesenchymal transition by phosphorylating β-catenin

    doi: 10.1186/s12964-024-01683-x

    Figure Lengend Snippet: IL-33 Mediates β-catenin Phosphorylation at Ser675 and Ser552 by PKA. A , B Protein expression of EMT-related proteins (vimentin, E-cadherin, N-cadherin, and β-catenin), Fra-1,phospho-β-catenin (Ser675 and Ser552), phospho-CREB (Ser133), CREB, phospho-AKT (Ser473) and AKT in EEC and 12Z treated by IL-33, PKA inhibitor-H89 or/and AKT inhibitor- MK2206. C Expression of CCN4 in supernatant of EEC and 12Z treated by IL-33, PKA inhibitor- H89 or/and AKT inhibitor- MK2206. D , F Cyclohexane chase assay of β-catenin in EECs and 12Z cells treated by IL-33. Data are presented as mean ± SEM. All data were analyzed using one-way ANOVA followed by Dunnett’s post hoc test and Student’s t-test ; * P < 0.05, ** P < 0.01, *** P < 0.001

    Article Snippet: The following IHC antibodies were used: anti-human ST2 (1:200), anti-human CCN4 (1:200), and anti-human E-cadherin (1:500) (abcam, Cambridge, UK); anti-human vimentin (1:100) (Cell Signaling Technology, Boston, USA); anti-human/mouse Fra-1 (1:200), anti-mouse CCN4 (1:100), and anti-mouse vimentin (1:100) (Affinity Biosciences, China); The primary antibodies used are listed in Supplementary Table 1.

    Techniques: Expressing

    IL-33-ST2 Signal Blocking or β-catenin Knockout Weakens EMT in EEC and 12Z. A . Immunofluorescence staining of phospho-β-catenin at Ser675 and Ser552 (Scale bars, 20 μm). B , C Protein expression of EMT-related proteins (Vimentin, E-cadherin, N-cadherin, and β-catenin) and Fra-1 in EECs and 12Z cells treated by IL-33 or/and knocking β-catenin by siRNA. D Expression of CCN4 in supernatant of EEC and 12Z treated by IL-33 or/and knocking β-catenin by siRNA. Data are presented as mean ± SEM. All data were analyzed using one-way ANOVA followed by Dunnett’s post hoc test and Student’s t-test ; * P < 0.05, ** P < 0.01, *** P < 0.001

    Journal: Cell Communication and Signaling : CCS

    Article Title: The IL-33-ST2 axis plays a vital role in endometriosis via promoting epithelial–mesenchymal transition by phosphorylating β-catenin

    doi: 10.1186/s12964-024-01683-x

    Figure Lengend Snippet: IL-33-ST2 Signal Blocking or β-catenin Knockout Weakens EMT in EEC and 12Z. A . Immunofluorescence staining of phospho-β-catenin at Ser675 and Ser552 (Scale bars, 20 μm). B , C Protein expression of EMT-related proteins (Vimentin, E-cadherin, N-cadherin, and β-catenin) and Fra-1 in EECs and 12Z cells treated by IL-33 or/and knocking β-catenin by siRNA. D Expression of CCN4 in supernatant of EEC and 12Z treated by IL-33 or/and knocking β-catenin by siRNA. Data are presented as mean ± SEM. All data were analyzed using one-way ANOVA followed by Dunnett’s post hoc test and Student’s t-test ; * P < 0.05, ** P < 0.01, *** P < 0.001

    Article Snippet: The following IHC antibodies were used: anti-human ST2 (1:200), anti-human CCN4 (1:200), and anti-human E-cadherin (1:500) (abcam, Cambridge, UK); anti-human vimentin (1:100) (Cell Signaling Technology, Boston, USA); anti-human/mouse Fra-1 (1:200), anti-mouse CCN4 (1:100), and anti-mouse vimentin (1:100) (Affinity Biosciences, China); The primary antibodies used are listed in Supplementary Table 1.

    Techniques: Blocking Assay, Knock-Out, Immunofluorescence, Staining, Expressing

    Schematic illustration of IL-33 Promoting EMT Process in Endometriotic Milieu. Interleukin 33 (IL-33) is highly expressed in ectopic ESCs, acting via the receptor ST2. Ectopic milieu, characterized by ROS, TGF-β1, and high level of estrogen, triggers secretion of IL-33 in ESCs, which in turn, enhanced the aggressive implantation and survival of ESCs. Meanwhile, elevated IL-33 in ectopic milieu also activated WNT/β-catenin pathway in EECs by phosphorylating β-catenin (Ser675 and Ser552), which primes exresssion of ECM related genes (CCN4 and Fra-1) and mesenchymal markers, enhanceing the EMT process and extracellular matrix produnction. Thus, IL-33/ST2 axis plays a pivotal role in endometriosis progress by promoting EMT. This figure was created with biorender.com

    Journal: Cell Communication and Signaling : CCS

    Article Title: The IL-33-ST2 axis plays a vital role in endometriosis via promoting epithelial–mesenchymal transition by phosphorylating β-catenin

    doi: 10.1186/s12964-024-01683-x

    Figure Lengend Snippet: Schematic illustration of IL-33 Promoting EMT Process in Endometriotic Milieu. Interleukin 33 (IL-33) is highly expressed in ectopic ESCs, acting via the receptor ST2. Ectopic milieu, characterized by ROS, TGF-β1, and high level of estrogen, triggers secretion of IL-33 in ESCs, which in turn, enhanced the aggressive implantation and survival of ESCs. Meanwhile, elevated IL-33 in ectopic milieu also activated WNT/β-catenin pathway in EECs by phosphorylating β-catenin (Ser675 and Ser552), which primes exresssion of ECM related genes (CCN4 and Fra-1) and mesenchymal markers, enhanceing the EMT process and extracellular matrix produnction. Thus, IL-33/ST2 axis plays a pivotal role in endometriosis progress by promoting EMT. This figure was created with biorender.com

    Article Snippet: The following IHC antibodies were used: anti-human ST2 (1:200), anti-human CCN4 (1:200), and anti-human E-cadherin (1:500) (abcam, Cambridge, UK); anti-human vimentin (1:100) (Cell Signaling Technology, Boston, USA); anti-human/mouse Fra-1 (1:200), anti-mouse CCN4 (1:100), and anti-mouse vimentin (1:100) (Affinity Biosciences, China); The primary antibodies used are listed in Supplementary Table 1.

    Techniques:

    GSDMD deletion exacerbates lupus-like phenotype in cGVH and NTS models. A Immunochemical images of GSDMD-FL and GSDMD-N pattern in renal section from normal control (NC) and LN patients. Scale bars: 50 μm. B Schematic of murine lupus models. NZB/W F1 mice were sacrificed at 14 weeks and 36 weeks. For cGVH model, C57BL/6 mice received an intraperitoneal injection of 1 × 10^ 8 bm12-derived splenocytes and were sacrificed 10 weeks post-induction. For NTS model, C57BL/6 mice were preimmunized with 0.2 mg sheep IgG intraperitoneally, followed by an intravenous injection of 50 μl sheep NTS on Day 4, and sacrificed on Day 11. C-E Immunoblotting of GSDMD in renal protein extracts of NZB/W F1 mice, cGVH or NTS mice with control (CON) mice. β-actin is loading control. F ELISA determining the level of autoantibodies (anti-dsDNA, anti-ssDNA and anti-histone) in serum of cGVH mice ( n = 4 or 5 per group). G Spleen to body weight ratio in cGVH mice ( n = 8 per group). H Representative PAS staining of kidney sections with quantitative analysis of glomerular cellularity and tubular injury scores in cGVH mice ( n = 5 or 10 per group). Scale bars: 50 μm. I Urine albumin to creatinine ratio in cGVH mice ( n = 8 per group). J Representative images and quantified immunofluorescence density of IgG and C3 deposition in the kidneys from cGVH mice ( n = 5 per group). Scale bars: 50 μm. K Quantitative analysis of glomerular PAS score and tubular injury scores in kidney sections from NTS mice (n = 5 per group). L Urine albumin to creatinine ratio and BUN in NTS mice (n = 5 per group). Data are shown as mean ± SEM. Two-way ANOVA test, ANOVA or Student’s t test was used for statistical analysis. **** P < 0.0001; *** P < 0.001; ** P < 0.01; * P < 0.05; ns, not significant

    Journal: Cell Communication and Signaling : CCS

    Article Title: Gasdermin D deficiency aborts myeloid calcium influx to drive granulopoiesis in lupus nephritis

    doi: 10.1186/s12964-024-01681-z

    Figure Lengend Snippet: GSDMD deletion exacerbates lupus-like phenotype in cGVH and NTS models. A Immunochemical images of GSDMD-FL and GSDMD-N pattern in renal section from normal control (NC) and LN patients. Scale bars: 50 μm. B Schematic of murine lupus models. NZB/W F1 mice were sacrificed at 14 weeks and 36 weeks. For cGVH model, C57BL/6 mice received an intraperitoneal injection of 1 × 10^ 8 bm12-derived splenocytes and were sacrificed 10 weeks post-induction. For NTS model, C57BL/6 mice were preimmunized with 0.2 mg sheep IgG intraperitoneally, followed by an intravenous injection of 50 μl sheep NTS on Day 4, and sacrificed on Day 11. C-E Immunoblotting of GSDMD in renal protein extracts of NZB/W F1 mice, cGVH or NTS mice with control (CON) mice. β-actin is loading control. F ELISA determining the level of autoantibodies (anti-dsDNA, anti-ssDNA and anti-histone) in serum of cGVH mice ( n = 4 or 5 per group). G Spleen to body weight ratio in cGVH mice ( n = 8 per group). H Representative PAS staining of kidney sections with quantitative analysis of glomerular cellularity and tubular injury scores in cGVH mice ( n = 5 or 10 per group). Scale bars: 50 μm. I Urine albumin to creatinine ratio in cGVH mice ( n = 8 per group). J Representative images and quantified immunofluorescence density of IgG and C3 deposition in the kidneys from cGVH mice ( n = 5 per group). Scale bars: 50 μm. K Quantitative analysis of glomerular PAS score and tubular injury scores in kidney sections from NTS mice (n = 5 per group). L Urine albumin to creatinine ratio and BUN in NTS mice (n = 5 per group). Data are shown as mean ± SEM. Two-way ANOVA test, ANOVA or Student’s t test was used for statistical analysis. **** P < 0.0001; *** P < 0.001; ** P < 0.01; * P < 0.05; ns, not significant

    Article Snippet: The following primary antibodies were used: anti-human GSDMD (Sigma-Aldrich, Cat# HPA044487; RRID: AB_2678957), anti-mouse GSDMD (Abcam, Cat# ab219800; RRID:AB_2888940), anti-human/mouse β-actin (Cell Signaling Technology, Cat# 3700S), anti-GAPDH (Abcam, Cat# ab8245; RRID:AB_2107448).

    Techniques: Injection, Derivative Assay, Western Blot, Enzyme-linked Immunosorbent Assay, Staining, Immunofluorescence

    LOXL4-mediated regulation of the MMP9 through the activation of NF-κB. (A) The prepared conditioned media from one-day serum-free cultivation of the indicated cells were condensed 10-fold. The media specimens were subjected to SDS-PAGE and then applied to WB analysis to detect MMP or CBB staining as a sample control of proper loading. (B) The prepared whole cell extracts from the indicated cells under the standard culture were also subjected to SDS-PAGE and analyzed by WB for the proteins of interest. CBB visualized the loaded proteins and displayed the stained image as a control of proper sample loading. On the other hand, to correctly detect the phosphorylation levels of Iκβα, the indicated cells were all treated with MG-132 proteasome inhibitor to restore the Iκβα protein levels from their lowered levels caused by proteasome-mediated degradation. (C) The prepared nuclear extracts from the indicated cells were subjected to EMSA analysis to detect NF-κB activity. (D, E) WB analysis was done for the 10-fold condensed cell conditioned media and cell extracts from the LOXL4 wt-overexpressing clones to detect MMP9 (D) and indicated protein molecules (E) in a similar manner as described for panels (A) and (B) , except for the treatment with siRNAs (control siRNA: siCont; ITGB1 siRNA: siITGB1) or NF-κB inhibitor. (F) EMSA analysis of NF-κB was also performed for the nuclear extracts from the same series of cells, as shown in (E) .

    Journal: Frontiers in Oncology

    Article Title: Dissection of the signal transduction machinery responsible for the lysyl oxidase-like 4-mediated increase in invasive motility in triple-negative breast cancer cells: mechanistic insight into the integrin-β1-NF-κB-MMP9 axis

    doi: 10.3389/fonc.2024.1371307

    Figure Lengend Snippet: LOXL4-mediated regulation of the MMP9 through the activation of NF-κB. (A) The prepared conditioned media from one-day serum-free cultivation of the indicated cells were condensed 10-fold. The media specimens were subjected to SDS-PAGE and then applied to WB analysis to detect MMP or CBB staining as a sample control of proper loading. (B) The prepared whole cell extracts from the indicated cells under the standard culture were also subjected to SDS-PAGE and analyzed by WB for the proteins of interest. CBB visualized the loaded proteins and displayed the stained image as a control of proper sample loading. On the other hand, to correctly detect the phosphorylation levels of Iκβα, the indicated cells were all treated with MG-132 proteasome inhibitor to restore the Iκβα protein levels from their lowered levels caused by proteasome-mediated degradation. (C) The prepared nuclear extracts from the indicated cells were subjected to EMSA analysis to detect NF-κB activity. (D, E) WB analysis was done for the 10-fold condensed cell conditioned media and cell extracts from the LOXL4 wt-overexpressing clones to detect MMP9 (D) and indicated protein molecules (E) in a similar manner as described for panels (A) and (B) , except for the treatment with siRNAs (control siRNA: siCont; ITGB1 siRNA: siITGB1) or NF-κB inhibitor. (F) EMSA analysis of NF-κB was also performed for the nuclear extracts from the same series of cells, as shown in (E) .

    Article Snippet: The antibodies used were as follows: mouse anti-HA tag antibody (clone 6E2; Cell Signaling Technology, Danvers, MA), mouse anti-Myc tag antibody (clone 9B11; Cell Signaling Technology), rabbit anti-human MMP9 antibody (Cell Signaling Technology), rabbit anti-human Akt antibody (Cell Signaling Technology), rabbit anti-human phospho (T308)-Akt antibody (Cell Signaling Technology), rabbit anti-human phospho (S473)-Akt antibody (Cell Signaling Technology), rabbit anti-human NF-κB antibody (Cell Signaling Technology), mouse anti-human phospho (S536)-NF-κB antibody (Cell Signaling Technology), rabbit anti-human IKKα antibody (Cell Signaling Technology), rabbit anti-human IKKβ antibody (Cell Signaling Technology), rabbit anti-human phospho (S176/180)-IKKα/β antibody (Cell Signaling Technology), mouse anti-human Iκβα antibody (Cell Signaling Technology), rabbit anti-human phospho (S32)-Iκβα antibody (Cell Signaling Technology), rabbit anti-human integrin-β1 antibody (Proteintech), rabbit anti-human TRAF4 antibody (Proteintech), rabbit anti-human TAK1 antibody (Cell Signaling Technology), rabbit anti-DsRed (RFP) (Takara Bio USA, Mountain View, CA), rabbit anti-GFP (Thermo Fisher Scientific), mouse anti-β-actin (Merk Sigma-Aldrich), and rabbit anti-human albumin (Agilent’s DAKO, Glostrup, Denmark) that cross-reacts with bovine albumin with high affinity in WB.

    Techniques: Activation Assay, SDS Page, Staining, Activity Assay, Clone Assay

    Identifying TRAF4 and TAK1 on the integrin-β1 signal cascade that links to NF-κB activation. (A) Schematic representation of the NF-κB activation flow via canonical and noncanonical pathways. (B, C) As indicated, the prepared cell lysates of HEK293T transfectants with different combinations of the plasmids were divided into three aliquots. The first was used as the whole cell lysate, i.e., as input (rightmost). The second was used to conduct immunoprecipitations with anti-HA tag antibody-conjugated agarose beads (middle). The remaining third was used to perform similar immunoprecipitations with anti-DYKDDDDK tag antibody-conjugated agarose beads as a negative control for the immunoprecipitation experiment (leftmost). After immunoprecipitation, the expressed products with beads-bound foreign proteins were analyzed by WB with an anti-HA, anti-Myc, or anti-albumin antibody. (D) The indicated cell extracts were incubated with the biotinylated (bio)-integrin-β1 antibody to capture the intrinsic integrin-β1 of the individual cells. On the other hand, bio-control IgG was used as a negative control for the same immunoprecipitation experiment. The captured integrin-β1 was collected by the pull-down method with avidin beads, and the precipitates were further subjected to WB with the indicated antibodies.

    Journal: Frontiers in Oncology

    Article Title: Dissection of the signal transduction machinery responsible for the lysyl oxidase-like 4-mediated increase in invasive motility in triple-negative breast cancer cells: mechanistic insight into the integrin-β1-NF-κB-MMP9 axis

    doi: 10.3389/fonc.2024.1371307

    Figure Lengend Snippet: Identifying TRAF4 and TAK1 on the integrin-β1 signal cascade that links to NF-κB activation. (A) Schematic representation of the NF-κB activation flow via canonical and noncanonical pathways. (B, C) As indicated, the prepared cell lysates of HEK293T transfectants with different combinations of the plasmids were divided into three aliquots. The first was used as the whole cell lysate, i.e., as input (rightmost). The second was used to conduct immunoprecipitations with anti-HA tag antibody-conjugated agarose beads (middle). The remaining third was used to perform similar immunoprecipitations with anti-DYKDDDDK tag antibody-conjugated agarose beads as a negative control for the immunoprecipitation experiment (leftmost). After immunoprecipitation, the expressed products with beads-bound foreign proteins were analyzed by WB with an anti-HA, anti-Myc, or anti-albumin antibody. (D) The indicated cell extracts were incubated with the biotinylated (bio)-integrin-β1 antibody to capture the intrinsic integrin-β1 of the individual cells. On the other hand, bio-control IgG was used as a negative control for the same immunoprecipitation experiment. The captured integrin-β1 was collected by the pull-down method with avidin beads, and the precipitates were further subjected to WB with the indicated antibodies.

    Article Snippet: The antibodies used were as follows: mouse anti-HA tag antibody (clone 6E2; Cell Signaling Technology, Danvers, MA), mouse anti-Myc tag antibody (clone 9B11; Cell Signaling Technology), rabbit anti-human MMP9 antibody (Cell Signaling Technology), rabbit anti-human Akt antibody (Cell Signaling Technology), rabbit anti-human phospho (T308)-Akt antibody (Cell Signaling Technology), rabbit anti-human phospho (S473)-Akt antibody (Cell Signaling Technology), rabbit anti-human NF-κB antibody (Cell Signaling Technology), mouse anti-human phospho (S536)-NF-κB antibody (Cell Signaling Technology), rabbit anti-human IKKα antibody (Cell Signaling Technology), rabbit anti-human IKKβ antibody (Cell Signaling Technology), rabbit anti-human phospho (S176/180)-IKKα/β antibody (Cell Signaling Technology), mouse anti-human Iκβα antibody (Cell Signaling Technology), rabbit anti-human phospho (S32)-Iκβα antibody (Cell Signaling Technology), rabbit anti-human integrin-β1 antibody (Proteintech), rabbit anti-human TRAF4 antibody (Proteintech), rabbit anti-human TAK1 antibody (Cell Signaling Technology), rabbit anti-DsRed (RFP) (Takara Bio USA, Mountain View, CA), rabbit anti-GFP (Thermo Fisher Scientific), mouse anti-β-actin (Merk Sigma-Aldrich), and rabbit anti-human albumin (Agilent’s DAKO, Glostrup, Denmark) that cross-reacts with bovine albumin with high affinity in WB.

    Techniques: Activation Assay, Negative Control, Immunoprecipitation, Incubation, Avidin-Biotin Assay

    The beneficial role of TRAF4 and TAK1 in regulating MMP9 via NF-κB activation. (A, B) The indicated cells with transient transfection of the combination of RFP with the indicated genes (control empty vector EV, dominant negative TRAF4: TRAF4 dn, catalytically unfunctional kinase-dead TAK1: TAK1 KD) were collected by RFP-based cell sorting. None stands for intact non-treated cells with no cell sorting procedure, which were also used as control cells. WB analysis was done for the 10-fold condensed cell conditioned media and whole cell extracts from the LOXL4 wt-overexpressing clones to detect MMP9 (A) and the indicated protein molecules (B) following a procedure similar to that described in <xref ref-type= Figures 2A, B . (C) EMSA analysis of NF-κB was also performed for the whole extracts from the same series of cells, as shown in (B) , because there were too few cells to perform fractionation. (D) Invasion assay set with or without NF-κB inhibitor was carried out to evaluate the effect of NF-κB on the LOXL4-enhanced invasiveness. (E, F) Endogenous LOXL4 in cells and the secreted MMP9 in the conditioned media in the indicated cell lines were detected using the WB procedure (E) . On the other hand, zymography with gelatin substrate was performed to detect secretory MMP activity from the indicated cells. Specification of the MMP9-mediated digestive bands was done by the treatment of the gels with an MMP9-specific inhibitor (F) . Other running gels with no gelatin, also loaded with identical specimens corresponding to WB and zymography, were stained with CBB as sample controls of proper loading (E, F) . (G) The conditioned media prepared from the indicated cells treated or not treated with LOX inhibitor BAPN, MMP inhibitor, or NF-κB inhibitor were subjected to WB (upper) and zymography (lower) to detect their MMP9 proteins and activities. (H) A trans-chamber-based cell invasion assay also evaluated the effects of these inhibitors used in (G) on the invasive activities in the indicated cells. (I) Schematic diagram of the molecular interplay among the indicated vital molecules. (J) Gene expression plots of MMP9 mRNA from breast invasive carcinoma (BRCA) specimens were obtained from a publicly available website ( http://gepia.cancer-pku.cn/ ). (K) Overall survival plots for MMP9 protein expression levels were obtained from a publicly available website ( http://kmplot.com/analysis/ ). Data are mean ± SD. *p<0.05, **p<0.01, ***p<0.001. " width="100%" height="100%">

    Journal: Frontiers in Oncology

    Article Title: Dissection of the signal transduction machinery responsible for the lysyl oxidase-like 4-mediated increase in invasive motility in triple-negative breast cancer cells: mechanistic insight into the integrin-β1-NF-κB-MMP9 axis

    doi: 10.3389/fonc.2024.1371307

    Figure Lengend Snippet: The beneficial role of TRAF4 and TAK1 in regulating MMP9 via NF-κB activation. (A, B) The indicated cells with transient transfection of the combination of RFP with the indicated genes (control empty vector EV, dominant negative TRAF4: TRAF4 dn, catalytically unfunctional kinase-dead TAK1: TAK1 KD) were collected by RFP-based cell sorting. None stands for intact non-treated cells with no cell sorting procedure, which were also used as control cells. WB analysis was done for the 10-fold condensed cell conditioned media and whole cell extracts from the LOXL4 wt-overexpressing clones to detect MMP9 (A) and the indicated protein molecules (B) following a procedure similar to that described in Figures 2A, B . (C) EMSA analysis of NF-κB was also performed for the whole extracts from the same series of cells, as shown in (B) , because there were too few cells to perform fractionation. (D) Invasion assay set with or without NF-κB inhibitor was carried out to evaluate the effect of NF-κB on the LOXL4-enhanced invasiveness. (E, F) Endogenous LOXL4 in cells and the secreted MMP9 in the conditioned media in the indicated cell lines were detected using the WB procedure (E) . On the other hand, zymography with gelatin substrate was performed to detect secretory MMP activity from the indicated cells. Specification of the MMP9-mediated digestive bands was done by the treatment of the gels with an MMP9-specific inhibitor (F) . Other running gels with no gelatin, also loaded with identical specimens corresponding to WB and zymography, were stained with CBB as sample controls of proper loading (E, F) . (G) The conditioned media prepared from the indicated cells treated or not treated with LOX inhibitor BAPN, MMP inhibitor, or NF-κB inhibitor were subjected to WB (upper) and zymography (lower) to detect their MMP9 proteins and activities. (H) A trans-chamber-based cell invasion assay also evaluated the effects of these inhibitors used in (G) on the invasive activities in the indicated cells. (I) Schematic diagram of the molecular interplay among the indicated vital molecules. (J) Gene expression plots of MMP9 mRNA from breast invasive carcinoma (BRCA) specimens were obtained from a publicly available website ( http://gepia.cancer-pku.cn/ ). (K) Overall survival plots for MMP9 protein expression levels were obtained from a publicly available website ( http://kmplot.com/analysis/ ). Data are mean ± SD. *p<0.05, **p<0.01, ***p<0.001.

    Article Snippet: The antibodies used were as follows: mouse anti-HA tag antibody (clone 6E2; Cell Signaling Technology, Danvers, MA), mouse anti-Myc tag antibody (clone 9B11; Cell Signaling Technology), rabbit anti-human MMP9 antibody (Cell Signaling Technology), rabbit anti-human Akt antibody (Cell Signaling Technology), rabbit anti-human phospho (T308)-Akt antibody (Cell Signaling Technology), rabbit anti-human phospho (S473)-Akt antibody (Cell Signaling Technology), rabbit anti-human NF-κB antibody (Cell Signaling Technology), mouse anti-human phospho (S536)-NF-κB antibody (Cell Signaling Technology), rabbit anti-human IKKα antibody (Cell Signaling Technology), rabbit anti-human IKKβ antibody (Cell Signaling Technology), rabbit anti-human phospho (S176/180)-IKKα/β antibody (Cell Signaling Technology), mouse anti-human Iκβα antibody (Cell Signaling Technology), rabbit anti-human phospho (S32)-Iκβα antibody (Cell Signaling Technology), rabbit anti-human integrin-β1 antibody (Proteintech), rabbit anti-human TRAF4 antibody (Proteintech), rabbit anti-human TAK1 antibody (Cell Signaling Technology), rabbit anti-DsRed (RFP) (Takara Bio USA, Mountain View, CA), rabbit anti-GFP (Thermo Fisher Scientific), mouse anti-β-actin (Merk Sigma-Aldrich), and rabbit anti-human albumin (Agilent’s DAKO, Glostrup, Denmark) that cross-reacts with bovine albumin with high affinity in WB.

    Techniques: Activation Assay, Transfection, Plasmid Preparation, Dominant Negative Mutation, FACS, Clone Assay, Fractionation, Invasion Assay, Zymography, Activity Assay, Staining, Expressing