anti β actin antibody  (Abcam)

 
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    Name:
    Anti Histone H3 citrulline R2 R8 R17 antibody ChIP Grade
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    ab5103
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    Structured Review

    Abcam anti β actin antibody
    LPS stimulates NETs formation in a time and concentration-dependent manner. (A, B, and C) PMNs were treated with or without LPS (1 μg/mL) for 2 h. Optical micrographs and their higher-magnification views; Scale bar: 50 μm (A). High-resolution SEM micrographs; Scale bar: 10 μm (B). Immunofluorescence staining of the cytomembrane rupture and DNA release in LPS-stimulated PMNs co-stained by membrane lipid dye (DiI, red), extracellular DNA dye (SYTOX Green, green) and another DNA dye (Hochest 33258, blue); Scale bar: 50 μm (C). (D and E) Immunofluorescence staining of LPS-stimulated PMNs characterized by extrusion of DNA (DAPI, blue) and NE (green); Scale bar: 50 μm. PMNs were treated with the same concentration (1 μg/mL) of LPS for 0, 2, and 4 h (D); or treated with different concentrations (0, 0.1, 1.0, and 10 μg/mL) of LPS for 2 h (E). (F, G, and H) PMNs were treated with sequential concentrations (0. 0.1, 1.0, 10 μg/mL) of LPS for a series of times (0, 1, 2, 3, and 4 h). Fluorescent values of extracellular DNA stained with SYTOX Green (F). Western blotting analysis of HiCt3 in PMNs, and <t>β-Actin</t> used as a protein loading control (G and H). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

    https://www.bioz.com/result/anti β actin antibody/product/Abcam
    Average 98 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    anti β actin antibody - by Bioz Stars, 2021-03
    98/100 stars

    Images

    1) Product Images from "A novel cell-based assay for dynamically detecting neutrophil extracellular traps-induced lung epithelial injuries"

    Article Title: A novel cell-based assay for dynamically detecting neutrophil extracellular traps-induced lung epithelial injuries

    Journal: Experimental Cell Research

    doi: 10.1016/j.yexcr.2020.112101

    LPS stimulates NETs formation in a time and concentration-dependent manner. (A, B, and C) PMNs were treated with or without LPS (1 μg/mL) for 2 h. Optical micrographs and their higher-magnification views; Scale bar: 50 μm (A). High-resolution SEM micrographs; Scale bar: 10 μm (B). Immunofluorescence staining of the cytomembrane rupture and DNA release in LPS-stimulated PMNs co-stained by membrane lipid dye (DiI, red), extracellular DNA dye (SYTOX Green, green) and another DNA dye (Hochest 33258, blue); Scale bar: 50 μm (C). (D and E) Immunofluorescence staining of LPS-stimulated PMNs characterized by extrusion of DNA (DAPI, blue) and NE (green); Scale bar: 50 μm. PMNs were treated with the same concentration (1 μg/mL) of LPS for 0, 2, and 4 h (D); or treated with different concentrations (0, 0.1, 1.0, and 10 μg/mL) of LPS for 2 h (E). (F, G, and H) PMNs were treated with sequential concentrations (0. 0.1, 1.0, 10 μg/mL) of LPS for a series of times (0, 1, 2, 3, and 4 h). Fluorescent values of extracellular DNA stained with SYTOX Green (F). Western blotting analysis of HiCt3 in PMNs, and β-Actin used as a protein loading control (G and H). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)
    Figure Legend Snippet: LPS stimulates NETs formation in a time and concentration-dependent manner. (A, B, and C) PMNs were treated with or without LPS (1 μg/mL) for 2 h. Optical micrographs and their higher-magnification views; Scale bar: 50 μm (A). High-resolution SEM micrographs; Scale bar: 10 μm (B). Immunofluorescence staining of the cytomembrane rupture and DNA release in LPS-stimulated PMNs co-stained by membrane lipid dye (DiI, red), extracellular DNA dye (SYTOX Green, green) and another DNA dye (Hochest 33258, blue); Scale bar: 50 μm (C). (D and E) Immunofluorescence staining of LPS-stimulated PMNs characterized by extrusion of DNA (DAPI, blue) and NE (green); Scale bar: 50 μm. PMNs were treated with the same concentration (1 μg/mL) of LPS for 0, 2, and 4 h (D); or treated with different concentrations (0, 0.1, 1.0, and 10 μg/mL) of LPS for 2 h (E). (F, G, and H) PMNs were treated with sequential concentrations (0. 0.1, 1.0, 10 μg/mL) of LPS for a series of times (0, 1, 2, 3, and 4 h). Fluorescent values of extracellular DNA stained with SYTOX Green (F). Western blotting analysis of HiCt3 in PMNs, and β-Actin used as a protein loading control (G and H). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

    Techniques Used: Concentration Assay, Immunofluorescence, Staining, Western Blot

    Related Articles

    Chromatin Immunoprecipitation:

    Article Title: Two-in-one: UV radiation simultaneously induces apoptosis and NETosis
    Article Snippet: .. The antibodies used were anti-phospho-ERK1/2 (AW39R; EMD Millipore) at 1:500; anti-phospho-p38 MAPK (D3F9) XP (Cell Signaling) at 1:500; anti-cleaved caspase 3 (ASP175, Cell Signalling); and anti-histone H3 (citrulline R2+R8+R17) ChIP Grade (ab5103, Abcam). ..

    Confocal Microscopy:

    Article Title: DNA threads released by activated CD4+ T lymphocytes provide autocrine costimulation
    Article Snippet: To assess THREDs formation in vivo, we isolated inguinal lymph nodes from naïve and EAE mice at 7 dpi. .. Confocal microscopy analysis of lymph node sections was performed by staining with mAb to CD4 (4SM95; e-Bioscience) and a polyclonal Ab to histone H3 (citrulline R2 + R8 + R17; Abcam, ab5103). ..

    Staining:

    Article Title: DNA threads released by activated CD4+ T lymphocytes provide autocrine costimulation
    Article Snippet: To assess THREDs formation in vivo, we isolated inguinal lymph nodes from naïve and EAE mice at 7 dpi. .. Confocal microscopy analysis of lymph node sections was performed by staining with mAb to CD4 (4SM95; e-Bioscience) and a polyclonal Ab to histone H3 (citrulline R2 + R8 + R17; Abcam, ab5103). ..

    Article Title: Neutrophil extracellular traps form predominantly during the organizing stage of human venous thromboembolism development
    Article Snippet: A cardiovascular pathologist categorized the specimens using histomorphologic features as unorganized (lack of collagen deposition, interspersed regions of fibrin/platelets and red blood cells), organizing (poorly organized collagen and/or neovasculature, and/or granulation tissue-like appearance), or organized (well organized collagen with intermixed hemosiderin-laden macrophages, with or without a well-formed neovasculature) [ ]. .. Following deparaffinization, microwave antigen retrieval used citrate buffer (pH 6.7) for 8 min, followed by the incubation of slides in heated buffer for an additional 20 min. After endogenous peroxidase activity blocking in methanol/H2 O2 (100:1) for 30 min at room temperature, single immunohistochemical staining used an anti-H3Cit antibody (ab5103, 1:250 dilution, Abcam, Cambridge, MA, USA). .. Histofine Simple Stain MAX PO (MULTI) (414152F, ready to use, Nichirei Corporation, Tokyo, Japan) served as a secondary antibody.

    Article Title: Milk fat globule-EGF factor 8 suppresses the aberrant immune response of systemic lupus erythematosus-derived neutrophils and associated tissue damage
    Article Snippet: Briefly, 2 × 105 BMDNs were seeded onto poly-l -lysine-coated coverslips (Sigma-Aldrich). .. In experiments without stimulation or with stimulation with 2% serum (extracted from pristane-treated WT or Mfge8 −/− mice for 6 months), the cells were incubated for 2 h. In experiments with 1 μ g/ml LPS, 100 nM PMA or 100 ng/ml MIP-2 stimulation, the incubation was for 4 h. NETs were stained with rabbit anti-histone H3 (citrulline R2+R8+R17) antibody and mouse anti-MPO antibody, followed by incubation with Alexa Fluor 647-conjugated mouse IgG and Alexa Fluor 488-conjugated rabbit IgG (Abcam). .. DNA was stained with DAPI.

    Article Title: Streptococcus agalactiae Induces Placental Macrophages To Release Extracellular Traps Loaded with Tissue Remodeling Enzymes via an Oxidative Burst-Dependent Mechanism
    Article Snippet: Coverslips were washed once with PBS prior to staining with SYTOX Green (final concentration, 10 μM) (ThermoFisher) for double-stranded DNA (dsDNA), and Hoechst 33342 (final concentration, 5 μM ) (ThermoFisher) for condensed chromatin (nuclei). .. Additional staining for histones and MMPs was accomplished by blocking cells in 1% bovine serum albumin in PBS for 30 min at 37°C followed by a 1-h incubation at 37°C with antibodies for histone H3 (ab5103; Abcam, Cambridge, MA), neutrophil elastase (ab68672; Abcam), myeloperoxidase (ab9535; Abcam), matrix metalloproteinase 1 (MMP-1) (ab551168; Abcam), MMP-7 (ab5706; Abcam), MMP-8 (ab81286; Abcam), MMP-9 (ab38898; Abcam), or MMP-12 (ab137444; Abcam). .. Cells were then washed three times with 1% BSA in PBS, followed by a 30-min incubation with an Alexa Fluor 594-conjugated goat anti-rabbit secondary antibody (ThermoFisher) and two additional washes with 1% BSA in PBS prior to mounting coverslips onto glass microscope slides with Aqua Poly/Mount (Polysciences Inc., Warrington, PA).

    Incubation:

    Article Title: Neutrophil extracellular traps form predominantly during the organizing stage of human venous thromboembolism development
    Article Snippet: A cardiovascular pathologist categorized the specimens using histomorphologic features as unorganized (lack of collagen deposition, interspersed regions of fibrin/platelets and red blood cells), organizing (poorly organized collagen and/or neovasculature, and/or granulation tissue-like appearance), or organized (well organized collagen with intermixed hemosiderin-laden macrophages, with or without a well-formed neovasculature) [ ]. .. Following deparaffinization, microwave antigen retrieval used citrate buffer (pH 6.7) for 8 min, followed by the incubation of slides in heated buffer for an additional 20 min. After endogenous peroxidase activity blocking in methanol/H2 O2 (100:1) for 30 min at room temperature, single immunohistochemical staining used an anti-H3Cit antibody (ab5103, 1:250 dilution, Abcam, Cambridge, MA, USA). .. Histofine Simple Stain MAX PO (MULTI) (414152F, ready to use, Nichirei Corporation, Tokyo, Japan) served as a secondary antibody.

    Article Title: Deep vein thrombosis in mice is regulated by platelet HMGB1 through release of neutrophil-extracellular traps and DNA
    Article Snippet: .. The samples were then incubated overnight with a combination of the following primary antibodies: anti-CD41 (5 μg/ml, rat IgG; Abcam, ab33661); citH3 (5 μg/ml, rabbit IgG; Abcam ab5103); Ly6G-APC (1:1000, BD Pharmogen 560599) or HMGB1 (2 μg/ml, rabbit IgG; Abcam 18256). .. Sections were then incubated with Alexa 488-conjugated F-actin phalloidin (1:500, Invitrogen, San Diego, CA, USA) in the presence of the following secondary antibodies depending on the primary antibody pairing: Cy5–conjugated goat anti-rat IgG (1:1000, for anti-CD41 antibody, Jackson Immunoresearch 112-165-167); 488-conjugated goat anti-rat IgG (1:500 for CD41 antibody, Molecular Probes, ); Cy3-conjugated goat anti-rabbit IgG (1:1000, for anti-HMGB1 antibody, Jackson Immunoresearch 111-165-003) for 1 h. A Hoeschst nuclear stain was applied for 30 s and slides were prepared for imaging.

    Article Title: Milk fat globule-EGF factor 8 suppresses the aberrant immune response of systemic lupus erythematosus-derived neutrophils and associated tissue damage
    Article Snippet: Briefly, 2 × 105 BMDNs were seeded onto poly-l -lysine-coated coverslips (Sigma-Aldrich). .. In experiments without stimulation or with stimulation with 2% serum (extracted from pristane-treated WT or Mfge8 −/− mice for 6 months), the cells were incubated for 2 h. In experiments with 1 μ g/ml LPS, 100 nM PMA or 100 ng/ml MIP-2 stimulation, the incubation was for 4 h. NETs were stained with rabbit anti-histone H3 (citrulline R2+R8+R17) antibody and mouse anti-MPO antibody, followed by incubation with Alexa Fluor 647-conjugated mouse IgG and Alexa Fluor 488-conjugated rabbit IgG (Abcam). .. DNA was stained with DAPI.

    Article Title: Streptococcus agalactiae Induces Placental Macrophages To Release Extracellular Traps Loaded with Tissue Remodeling Enzymes via an Oxidative Burst-Dependent Mechanism
    Article Snippet: Coverslips were washed once with PBS prior to staining with SYTOX Green (final concentration, 10 μM) (ThermoFisher) for double-stranded DNA (dsDNA), and Hoechst 33342 (final concentration, 5 μM ) (ThermoFisher) for condensed chromatin (nuclei). .. Additional staining for histones and MMPs was accomplished by blocking cells in 1% bovine serum albumin in PBS for 30 min at 37°C followed by a 1-h incubation at 37°C with antibodies for histone H3 (ab5103; Abcam, Cambridge, MA), neutrophil elastase (ab68672; Abcam), myeloperoxidase (ab9535; Abcam), matrix metalloproteinase 1 (MMP-1) (ab551168; Abcam), MMP-7 (ab5706; Abcam), MMP-8 (ab81286; Abcam), MMP-9 (ab38898; Abcam), or MMP-12 (ab137444; Abcam). .. Cells were then washed three times with 1% BSA in PBS, followed by a 30-min incubation with an Alexa Fluor 594-conjugated goat anti-rabbit secondary antibody (ThermoFisher) and two additional washes with 1% BSA in PBS prior to mounting coverslips onto glass microscope slides with Aqua Poly/Mount (Polysciences Inc., Warrington, PA).

    Activity Assay:

    Article Title: Neutrophil extracellular traps form predominantly during the organizing stage of human venous thromboembolism development
    Article Snippet: A cardiovascular pathologist categorized the specimens using histomorphologic features as unorganized (lack of collagen deposition, interspersed regions of fibrin/platelets and red blood cells), organizing (poorly organized collagen and/or neovasculature, and/or granulation tissue-like appearance), or organized (well organized collagen with intermixed hemosiderin-laden macrophages, with or without a well-formed neovasculature) [ ]. .. Following deparaffinization, microwave antigen retrieval used citrate buffer (pH 6.7) for 8 min, followed by the incubation of slides in heated buffer for an additional 20 min. After endogenous peroxidase activity blocking in methanol/H2 O2 (100:1) for 30 min at room temperature, single immunohistochemical staining used an anti-H3Cit antibody (ab5103, 1:250 dilution, Abcam, Cambridge, MA, USA). .. Histofine Simple Stain MAX PO (MULTI) (414152F, ready to use, Nichirei Corporation, Tokyo, Japan) served as a secondary antibody.

    Blocking Assay:

    Article Title: Neutrophil extracellular traps form predominantly during the organizing stage of human venous thromboembolism development
    Article Snippet: A cardiovascular pathologist categorized the specimens using histomorphologic features as unorganized (lack of collagen deposition, interspersed regions of fibrin/platelets and red blood cells), organizing (poorly organized collagen and/or neovasculature, and/or granulation tissue-like appearance), or organized (well organized collagen with intermixed hemosiderin-laden macrophages, with or without a well-formed neovasculature) [ ]. .. Following deparaffinization, microwave antigen retrieval used citrate buffer (pH 6.7) for 8 min, followed by the incubation of slides in heated buffer for an additional 20 min. After endogenous peroxidase activity blocking in methanol/H2 O2 (100:1) for 30 min at room temperature, single immunohistochemical staining used an anti-H3Cit antibody (ab5103, 1:250 dilution, Abcam, Cambridge, MA, USA). .. Histofine Simple Stain MAX PO (MULTI) (414152F, ready to use, Nichirei Corporation, Tokyo, Japan) served as a secondary antibody.

    Article Title: Streptococcus agalactiae Induces Placental Macrophages To Release Extracellular Traps Loaded with Tissue Remodeling Enzymes via an Oxidative Burst-Dependent Mechanism
    Article Snippet: Coverslips were washed once with PBS prior to staining with SYTOX Green (final concentration, 10 μM) (ThermoFisher) for double-stranded DNA (dsDNA), and Hoechst 33342 (final concentration, 5 μM ) (ThermoFisher) for condensed chromatin (nuclei). .. Additional staining for histones and MMPs was accomplished by blocking cells in 1% bovine serum albumin in PBS for 30 min at 37°C followed by a 1-h incubation at 37°C with antibodies for histone H3 (ab5103; Abcam, Cambridge, MA), neutrophil elastase (ab68672; Abcam), myeloperoxidase (ab9535; Abcam), matrix metalloproteinase 1 (MMP-1) (ab551168; Abcam), MMP-7 (ab5706; Abcam), MMP-8 (ab81286; Abcam), MMP-9 (ab38898; Abcam), or MMP-12 (ab137444; Abcam). .. Cells were then washed three times with 1% BSA in PBS, followed by a 30-min incubation with an Alexa Fluor 594-conjugated goat anti-rabbit secondary antibody (ThermoFisher) and two additional washes with 1% BSA in PBS prior to mounting coverslips onto glass microscope slides with Aqua Poly/Mount (Polysciences Inc., Warrington, PA).

    Immunohistochemistry:

    Article Title: Neutrophil extracellular traps form predominantly during the organizing stage of human venous thromboembolism development
    Article Snippet: A cardiovascular pathologist categorized the specimens using histomorphologic features as unorganized (lack of collagen deposition, interspersed regions of fibrin/platelets and red blood cells), organizing (poorly organized collagen and/or neovasculature, and/or granulation tissue-like appearance), or organized (well organized collagen with intermixed hemosiderin-laden macrophages, with or without a well-formed neovasculature) [ ]. .. Following deparaffinization, microwave antigen retrieval used citrate buffer (pH 6.7) for 8 min, followed by the incubation of slides in heated buffer for an additional 20 min. After endogenous peroxidase activity blocking in methanol/H2 O2 (100:1) for 30 min at room temperature, single immunohistochemical staining used an anti-H3Cit antibody (ab5103, 1:250 dilution, Abcam, Cambridge, MA, USA). .. Histofine Simple Stain MAX PO (MULTI) (414152F, ready to use, Nichirei Corporation, Tokyo, Japan) served as a secondary antibody.

    Mouse Assay:

    Article Title: Milk fat globule-EGF factor 8 suppresses the aberrant immune response of systemic lupus erythematosus-derived neutrophils and associated tissue damage
    Article Snippet: Briefly, 2 × 105 BMDNs were seeded onto poly-l -lysine-coated coverslips (Sigma-Aldrich). .. In experiments without stimulation or with stimulation with 2% serum (extracted from pristane-treated WT or Mfge8 −/− mice for 6 months), the cells were incubated for 2 h. In experiments with 1 μ g/ml LPS, 100 nM PMA or 100 ng/ml MIP-2 stimulation, the incubation was for 4 h. NETs were stained with rabbit anti-histone H3 (citrulline R2+R8+R17) antibody and mouse anti-MPO antibody, followed by incubation with Alexa Fluor 647-conjugated mouse IgG and Alexa Fluor 488-conjugated rabbit IgG (Abcam). .. DNA was stained with DAPI.

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  • 99
    Abcam anti β actin antibody
    Protein expression levels of PERK, IRE1α and ATF6 in ovaries of aged mice. Representative western blot images (a) and quantification of PERK, IRE1α and ATF6 protein expression levels (b). <t>β-ACTIN</t> served as an internal control. Data are expressed as the mean ± SEM, expressed by error bars.
    Anti β Actin Antibody, supplied by Abcam, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti β actin antibody/product/Abcam
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    anti β actin antibody - by Bioz Stars, 2021-03
    99/100 stars
      Buy from Supplier

    99
    Abcam mouse anti β actin
    Gastrodin (GSTD) increased osteogenic transcription Runx2, decreased adipogenic factor peroxisome proliferator-activated receptor (PPAR)γ isoform 2 and activated nuclear factor-like 2 (NRF2) pathway protein expression levels. Representative western blot analyses for total protein was analyzed by western blotting in dexamethasone (DEX)-treated MC3T3-E1 cells with or without GSTD pretreatment at different doses. The relative protein expression levels were measured using the fold-change in each protein relative to <t>β-actin</t> from the same sample. The data are expressed as the mean ± standard deviation. ## P
    Mouse Anti β Actin, supplied by Abcam, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mouse anti β actin/product/Abcam
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    mouse anti β actin - by Bioz Stars, 2021-03
    99/100 stars
      Buy from Supplier

    99
    Abcam anti β actin
    Hirudin regulated fibrosis-related factors in Ang II-induced myocardial fibroblasts. ( A ) The relative mRNA levels of MMP-2, MMP-9, TIMP-2 was evaluated by qRT-PCR. ( B ) The relative protein levels of MMP-2, MMP-9, TIMP-2 was examined by western blot. ( C ) qRT-PCR was performed to determine the mRNA levels of FN, TGF-β1, COL-I and COL-III. ( D ) Western blot was carried out to detect the protein levels of FN, TGF-β1, COL-I and COL-III, and normalized to <t>β-actin</t> expression. Gray value was detected and counted by quality one. * P
    Anti β Actin, supplied by Abcam, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti β actin/product/Abcam
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    anti β actin - by Bioz Stars, 2021-03
    99/100 stars
      Buy from Supplier

    Image Search Results


    Protein expression levels of PERK, IRE1α and ATF6 in ovaries of aged mice. Representative western blot images (a) and quantification of PERK, IRE1α and ATF6 protein expression levels (b). β-ACTIN served as an internal control. Data are expressed as the mean ± SEM, expressed by error bars.

    Journal: The Journal of Veterinary Medical Science

    Article Title: Intravenous human endothelial progenitor cell administration into aged mice enhances embryo development and oocyte quality by reducing inflammation, endoplasmic reticulum stress and apoptosis

    doi: 10.1292/jvms.18-0242

    Figure Lengend Snippet: Protein expression levels of PERK, IRE1α and ATF6 in ovaries of aged mice. Representative western blot images (a) and quantification of PERK, IRE1α and ATF6 protein expression levels (b). β-ACTIN served as an internal control. Data are expressed as the mean ± SEM, expressed by error bars.

    Article Snippet: The loading and blotting of the amount of protein was verified by reprobing the membrane with anti-β actin antibody (1:5,000; Abcam) followed by Coomassie Blue staining.

    Techniques: Expressing, Mouse Assay, Western Blot

    Gastrodin (GSTD) increased osteogenic transcription Runx2, decreased adipogenic factor peroxisome proliferator-activated receptor (PPAR)γ isoform 2 and activated nuclear factor-like 2 (NRF2) pathway protein expression levels. Representative western blot analyses for total protein was analyzed by western blotting in dexamethasone (DEX)-treated MC3T3-E1 cells with or without GSTD pretreatment at different doses. The relative protein expression levels were measured using the fold-change in each protein relative to β-actin from the same sample. The data are expressed as the mean ± standard deviation. ## P

    Journal: International Journal of Molecular Medicine

    Article Title: Gastrodin protects MC3T3-E1 osteoblasts from dexamethasone-induced cellular dysfunction and promotes bone formation via induction of the NRF2 signaling pathway

    doi: 10.3892/ijmm.2018.3414

    Figure Lengend Snippet: Gastrodin (GSTD) increased osteogenic transcription Runx2, decreased adipogenic factor peroxisome proliferator-activated receptor (PPAR)γ isoform 2 and activated nuclear factor-like 2 (NRF2) pathway protein expression levels. Representative western blot analyses for total protein was analyzed by western blotting in dexamethasone (DEX)-treated MC3T3-E1 cells with or without GSTD pretreatment at different doses. The relative protein expression levels were measured using the fold-change in each protein relative to β-actin from the same sample. The data are expressed as the mean ± standard deviation. ## P

    Article Snippet: Rabbit anti-HO-1 (cat. no. ab68477; 1:1,000), rabbit anti-NQO-1 (cat. no. ab76956; 1:1,000), and mouse anti-β-actin (cat. no. ab8245; 1:1,000), Anti-OCN (cat. no. ab93876; 1:1,000) monoclonal antibodies were purchased from Abcam (Cambridge, MA, USA).

    Techniques: Expressing, Western Blot, Standard Deviation

    Hirudin regulated fibrosis-related factors in Ang II-induced myocardial fibroblasts. ( A ) The relative mRNA levels of MMP-2, MMP-9, TIMP-2 was evaluated by qRT-PCR. ( B ) The relative protein levels of MMP-2, MMP-9, TIMP-2 was examined by western blot. ( C ) qRT-PCR was performed to determine the mRNA levels of FN, TGF-β1, COL-I and COL-III. ( D ) Western blot was carried out to detect the protein levels of FN, TGF-β1, COL-I and COL-III, and normalized to β-actin expression. Gray value was detected and counted by quality one. * P

    Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

    Article Title: Hirudin Protects Ang II-Induced Myocardial Fibroblasts Fibrosis by Inhibiting the Extracellular Signal-Regulated Kinase1/2 (ERK1/2) Pathway

    doi: 10.12659/MSM.909044

    Figure Lengend Snippet: Hirudin regulated fibrosis-related factors in Ang II-induced myocardial fibroblasts. ( A ) The relative mRNA levels of MMP-2, MMP-9, TIMP-2 was evaluated by qRT-PCR. ( B ) The relative protein levels of MMP-2, MMP-9, TIMP-2 was examined by western blot. ( C ) qRT-PCR was performed to determine the mRNA levels of FN, TGF-β1, COL-I and COL-III. ( D ) Western blot was carried out to detect the protein levels of FN, TGF-β1, COL-I and COL-III, and normalized to β-actin expression. Gray value was detected and counted by quality one. * P

    Article Snippet: The membranes were incubated with anti-matrix metalloproteinase-2 (MMP-2) (AmyJet Scientific, 12015-01, 1: 2000), anti-MMP-9 (R & D, AF909-SP, 1: 1000), anti-tissue inhibitor of metalloproteinases-2 (TIMP-2) (R & D, AF971, 1: 1200), anti-fibronectin (FN) (Abcam, ab2413, 1: 800), anti-transforming growth factor beta 1 (TGF-β1) (R & D, FAB766G, 1: 700), anti-collagen-I (COL-I) (Abcam, ab90395, 1: 1000), anti-collagen III (COL-III) (Abcam, ab7778, 1: 800), anti-p-ERK1/2 (CST, 8544, 1: 1000), anti-ERK1/2 (CST, 9102, 1: 700), and anti-β-actin (Abcam, ab8226, 1: 2000) at 4°C for 24 hours.

    Techniques: Quantitative RT-PCR, Western Blot, Expressing

    Hirudin downregulated ERK1/2 pathway in Ang II-induced myocardial fibroblasts. ( A ) The proteins expression levels of ERK1/2 and p-ERK1/2 were examined by western blot. ( B ) Quantification of protein expression was carried out with GraphPad prism 7.0. β-actin was used as internal control. The gray value was evaluated and calculated by quality one. * P

    Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

    Article Title: Hirudin Protects Ang II-Induced Myocardial Fibroblasts Fibrosis by Inhibiting the Extracellular Signal-Regulated Kinase1/2 (ERK1/2) Pathway

    doi: 10.12659/MSM.909044

    Figure Lengend Snippet: Hirudin downregulated ERK1/2 pathway in Ang II-induced myocardial fibroblasts. ( A ) The proteins expression levels of ERK1/2 and p-ERK1/2 were examined by western blot. ( B ) Quantification of protein expression was carried out with GraphPad prism 7.0. β-actin was used as internal control. The gray value was evaluated and calculated by quality one. * P

    Article Snippet: The membranes were incubated with anti-matrix metalloproteinase-2 (MMP-2) (AmyJet Scientific, 12015-01, 1: 2000), anti-MMP-9 (R & D, AF909-SP, 1: 1000), anti-tissue inhibitor of metalloproteinases-2 (TIMP-2) (R & D, AF971, 1: 1200), anti-fibronectin (FN) (Abcam, ab2413, 1: 800), anti-transforming growth factor beta 1 (TGF-β1) (R & D, FAB766G, 1: 700), anti-collagen-I (COL-I) (Abcam, ab90395, 1: 1000), anti-collagen III (COL-III) (Abcam, ab7778, 1: 800), anti-p-ERK1/2 (CST, 8544, 1: 1000), anti-ERK1/2 (CST, 9102, 1: 700), and anti-β-actin (Abcam, ab8226, 1: 2000) at 4°C for 24 hours.

    Techniques: Expressing, Western Blot

    Ado inhibits cell viability and triggers ER stress in HepG2 cells. (A) Time- and dose-dependent cytotoxic effects of Ado on HepG2 cells. Cells were treated with different concentrations of Ado (1.0–4.0 mM) for 12, 24, or 48 h. Cell viability was determined by MTT assay. Results are expressed as percentages of cell growth relative to initial number of viable cells. (B) Cells were treated with 2.0 mM for 6, 12, or 24 h. The cell cycle distribution was evaluated using flow cytometric assay by PI staining. (C) HepG2 cells were treated with 2.0 mM Ado for 24 h. Cells were collected and subjected to total RNA extraction. The mRNA levels of ER stress-related genes GRP78, PERK, ATF4, CHOP, Cyt C and caspase-3 were assessed by RT-PCR. (D) HepG2 cells underwent the aforementioned treatment and were collected and subjected to western blot analyses with specific antibodies directed against GRP78, PERK, ATF4, CHOP, Cyt C and cleaved caspase-3, or β-actin. The density of the corresponding bands was assessed quantitatively using image analysis software and corrected by reference to the value of β-actin. Bar graphs represent the mean fluorescence intensity. *P

    Journal: Oncology Reports

    Article Title: Inhibition of autophagy enhances adenosine-induced apoptosis in human hepatoblastoma HepG2 cells

    doi: 10.3892/or.2018.6899

    Figure Lengend Snippet: Ado inhibits cell viability and triggers ER stress in HepG2 cells. (A) Time- and dose-dependent cytotoxic effects of Ado on HepG2 cells. Cells were treated with different concentrations of Ado (1.0–4.0 mM) for 12, 24, or 48 h. Cell viability was determined by MTT assay. Results are expressed as percentages of cell growth relative to initial number of viable cells. (B) Cells were treated with 2.0 mM for 6, 12, or 24 h. The cell cycle distribution was evaluated using flow cytometric assay by PI staining. (C) HepG2 cells were treated with 2.0 mM Ado for 24 h. Cells were collected and subjected to total RNA extraction. The mRNA levels of ER stress-related genes GRP78, PERK, ATF4, CHOP, Cyt C and caspase-3 were assessed by RT-PCR. (D) HepG2 cells underwent the aforementioned treatment and were collected and subjected to western blot analyses with specific antibodies directed against GRP78, PERK, ATF4, CHOP, Cyt C and cleaved caspase-3, or β-actin. The density of the corresponding bands was assessed quantitatively using image analysis software and corrected by reference to the value of β-actin. Bar graphs represent the mean fluorescence intensity. *P

    Article Snippet: ATF4 (cat. no. ab23760), GRP78 (cat. no. ab21685), PERK (cat. no. ab65142), p-AMPK (cat. no. ab133448), AMPK (cat. no. ab80039), p-ULK1 (cat. no. ab229540), ULK1 (cat. no. ab167139), p-mTOR (cat. no. ab84400), mTOR (cat. no. ab32028), CHOP (cat. no. ab11419), cytochrome c (cat. no. ab110325) and β-actin (cat. no. ab8226) were purchased from Abcam (Cambridge, MA, USA).

    Techniques: MTT Assay, Flow Cytometry, Staining, RNA Extraction, Reverse Transcription Polymerase Chain Reaction, Western Blot, Software, Fluorescence

    Inhibition of autophagy enhances ER stress-mediated apoptotic cell death. (A) HepG2 cells were treated with 2.0 mM Ado or 10 µM LY alone. In the combination treatment group, the cells were pre-treated with 10 µM LY for 2 h, followed by 2.0 mM Ado treatment. Cell viability was determined by MTT assay at the indicated time-points. (B) Cell apoptosis was detected by Hoechst staining. Images were captured under a fluorescence microscope (original magnification, ×200). Arrows indicate apoptotic cells. (C) The mRNA levels of Cyt C, AMPK and p62 were assessed by RT-PCR. (D) HepG2 cells underwent the aforementioned treatment and were collected and subjected to western blot analyses with specific antibodies directed against Cyt C, p-AMPK, AMPK, p62, LC3-I and LC3-II, or β-actin. Bar graphs represent the mean protein band intensity. Relative quantity of the aforementioned proteins was computed. (E) Mitochondrial membrane potential (ΔΨm) indicated by Rhodamine-123 was detected by flow cytometry. Bar graphs represent the mean fluorescence intensity. *P

    Journal: Oncology Reports

    Article Title: Inhibition of autophagy enhances adenosine-induced apoptosis in human hepatoblastoma HepG2 cells

    doi: 10.3892/or.2018.6899

    Figure Lengend Snippet: Inhibition of autophagy enhances ER stress-mediated apoptotic cell death. (A) HepG2 cells were treated with 2.0 mM Ado or 10 µM LY alone. In the combination treatment group, the cells were pre-treated with 10 µM LY for 2 h, followed by 2.0 mM Ado treatment. Cell viability was determined by MTT assay at the indicated time-points. (B) Cell apoptosis was detected by Hoechst staining. Images were captured under a fluorescence microscope (original magnification, ×200). Arrows indicate apoptotic cells. (C) The mRNA levels of Cyt C, AMPK and p62 were assessed by RT-PCR. (D) HepG2 cells underwent the aforementioned treatment and were collected and subjected to western blot analyses with specific antibodies directed against Cyt C, p-AMPK, AMPK, p62, LC3-I and LC3-II, or β-actin. Bar graphs represent the mean protein band intensity. Relative quantity of the aforementioned proteins was computed. (E) Mitochondrial membrane potential (ΔΨm) indicated by Rhodamine-123 was detected by flow cytometry. Bar graphs represent the mean fluorescence intensity. *P

    Article Snippet: ATF4 (cat. no. ab23760), GRP78 (cat. no. ab21685), PERK (cat. no. ab65142), p-AMPK (cat. no. ab133448), AMPK (cat. no. ab80039), p-ULK1 (cat. no. ab229540), ULK1 (cat. no. ab167139), p-mTOR (cat. no. ab84400), mTOR (cat. no. ab32028), CHOP (cat. no. ab11419), cytochrome c (cat. no. ab110325) and β-actin (cat. no. ab8226) were purchased from Abcam (Cambridge, MA, USA).

    Techniques: Inhibition, MTT Assay, Staining, Fluorescence, Microscopy, Reverse Transcription Polymerase Chain Reaction, Western Blot, Flow Cytometry, Cytometry

    Ado induces autophagy in HepG2 cells. (A) HepG2 cells were treated with 2.0 mM for 12, or 24 h and cells were stained with MDC. The autophagosomes were detected under fluorescence microscopy (magnification, ×200). (B) The mRNA levels of autophagy-related genes Beclin-1 and p62 were assessed by RT-PCR. (C) HepG2 cells underwent the aforementioned treatment and were collected and subjected to western blot analyses with specific antibodies directed against LC3-I, LC3-II, Beclin-1, p62 or β-actin. Bar graphs represent the mean protein band intensity. Relative quantity of the aforementioned proteins was computed after normalization with β-actin. *P

    Journal: Oncology Reports

    Article Title: Inhibition of autophagy enhances adenosine-induced apoptosis in human hepatoblastoma HepG2 cells

    doi: 10.3892/or.2018.6899

    Figure Lengend Snippet: Ado induces autophagy in HepG2 cells. (A) HepG2 cells were treated with 2.0 mM for 12, or 24 h and cells were stained with MDC. The autophagosomes were detected under fluorescence microscopy (magnification, ×200). (B) The mRNA levels of autophagy-related genes Beclin-1 and p62 were assessed by RT-PCR. (C) HepG2 cells underwent the aforementioned treatment and were collected and subjected to western blot analyses with specific antibodies directed against LC3-I, LC3-II, Beclin-1, p62 or β-actin. Bar graphs represent the mean protein band intensity. Relative quantity of the aforementioned proteins was computed after normalization with β-actin. *P

    Article Snippet: ATF4 (cat. no. ab23760), GRP78 (cat. no. ab21685), PERK (cat. no. ab65142), p-AMPK (cat. no. ab133448), AMPK (cat. no. ab80039), p-ULK1 (cat. no. ab229540), ULK1 (cat. no. ab167139), p-mTOR (cat. no. ab84400), mTOR (cat. no. ab32028), CHOP (cat. no. ab11419), cytochrome c (cat. no. ab110325) and β-actin (cat. no. ab8226) were purchased from Abcam (Cambridge, MA, USA).

    Techniques: Staining, Fluorescence, Microscopy, Reverse Transcription Polymerase Chain Reaction, Western Blot

    Inhibition of autophagy by AMPK siRNA enhances Ado-induced apoptosis and inhibits the AMPK/mTOR/ULK1 pathway. (A) HepG2 cells were transfected with AMPK siRNA (20 µM) or control siRNA (transfected with empty vector, 20 µM), then exposed to 2.0 mM Ado or not for 24 h and then the cell apoptotic ratio was determined by flow cytometry (FACS) with Annexin V-FITC and PI double staining. (B) HepG2 cells underwent the aforementioned treatment and were collected and the protein levels of the AMPK/mTOR/ULK1 pathway were assessed by western blotting. The phosphorylation ratios of p-AMPK/AMPK, p-ULK1/ULK1 and p-mTOR/mTOR were computed (C) The mRNA and protein levels of ER stress-related genes were assessed by RT-PCR and western blotting. HepG2 cells underwent the aforementioned treatment and were collected and subjected to western blot analyses with specific antibodies directed against CHOP, cleaved caspase-3, p62 and Cyt C or β-actin. Bar graphs represent the mean protein band intensity. (D) Mitochondrial membrane potential (ΔΨm) indicated by Rhodamine-123 was detected by flow cytometry. Bar graphs represent the mean fluorescence intensity. *P

    Journal: Oncology Reports

    Article Title: Inhibition of autophagy enhances adenosine-induced apoptosis in human hepatoblastoma HepG2 cells

    doi: 10.3892/or.2018.6899

    Figure Lengend Snippet: Inhibition of autophagy by AMPK siRNA enhances Ado-induced apoptosis and inhibits the AMPK/mTOR/ULK1 pathway. (A) HepG2 cells were transfected with AMPK siRNA (20 µM) or control siRNA (transfected with empty vector, 20 µM), then exposed to 2.0 mM Ado or not for 24 h and then the cell apoptotic ratio was determined by flow cytometry (FACS) with Annexin V-FITC and PI double staining. (B) HepG2 cells underwent the aforementioned treatment and were collected and the protein levels of the AMPK/mTOR/ULK1 pathway were assessed by western blotting. The phosphorylation ratios of p-AMPK/AMPK, p-ULK1/ULK1 and p-mTOR/mTOR were computed (C) The mRNA and protein levels of ER stress-related genes were assessed by RT-PCR and western blotting. HepG2 cells underwent the aforementioned treatment and were collected and subjected to western blot analyses with specific antibodies directed against CHOP, cleaved caspase-3, p62 and Cyt C or β-actin. Bar graphs represent the mean protein band intensity. (D) Mitochondrial membrane potential (ΔΨm) indicated by Rhodamine-123 was detected by flow cytometry. Bar graphs represent the mean fluorescence intensity. *P

    Article Snippet: ATF4 (cat. no. ab23760), GRP78 (cat. no. ab21685), PERK (cat. no. ab65142), p-AMPK (cat. no. ab133448), AMPK (cat. no. ab80039), p-ULK1 (cat. no. ab229540), ULK1 (cat. no. ab167139), p-mTOR (cat. no. ab84400), mTOR (cat. no. ab32028), CHOP (cat. no. ab11419), cytochrome c (cat. no. ab110325) and β-actin (cat. no. ab8226) were purchased from Abcam (Cambridge, MA, USA).

    Techniques: Inhibition, Transfection, Plasmid Preparation, Flow Cytometry, Cytometry, FACS, Double Staining, Western Blot, Reverse Transcription Polymerase Chain Reaction, Fluorescence