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  • 87
    Thermo Fisher gene exp cygb hs00370478 m1
    Quantitative real-time RT-PCR analysis of <t>HO-1</t> (a), AQP-1 (b), and Bcl-2 (c) expressions in mixed hTECs under basal conditions and after GO-induced oxidative stress (1 IU/mL) either or not in the presence of epoetin delta (100 IU/mL). GAPDH was used as endogenous control housekeeping gene. Preconditioning the cells with EPO resulted in a significant upregulation of HO-1 and AQP-1 mRNA. GO-induced oxidative stress further increased HO-1, AQP-1, and Bcl-2 mRNA expressions with maximum levels 60 minutes after induction of oxidative stress. Data are presented as the mean ± SD of triplicate determinations of 2 runs (i.e., 6 values each). * P
    Gene Exp Cygb Hs00370478 M1, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 87/100, based on 17 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    97
    Millipore 3t3 f442a cygb cells
    <t>Cygb</t> overexpression increases adipogenic differentiation. a Structure of Cygb gene construct, b relative mRNA expression level of Cygb in transduced cells, c oil red staining of differentiated <t>3T3-F442A–GFP</t> and 3T3-F442A–Cygb cells, d gene expression levels of PPARγ, CEBPα and FABP4 in differentiated 3T3-F442A–GFP and 3T3-F442A–Cygb cells at the end of 8 day differentiation period. * P
    3t3 F442a Cygb Cells, supplied by Millipore, used in various techniques. Bioz Stars score: 97/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Santa Cruz Biotechnology cygb
    Western blot analysis of <t>Cygb,</t> PPARγ, CEBPα and <t>FABP4.</t> during differentiation process. a Western blot results of Cygb, PPARγ, CEBPα and FABP4 during differentiation process, b quantitative results of western blot band intensities. * P
    Cygb, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 88/100, based on 23 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Abcam anti cygb
    Inhibition of liver fibrosis development in long-term TAA-treated <t>Cygb-TG</t> mice. WT and Cygb-TG (TG) mice were subjected to TAA treatment for 10 weeks (TAA-10w). ( a ) Representative liver images of H E, Sirius Red and Fast Green (SiR-FG) staining for collagen deposition and IF staining for the detection of αSMA (red) and CYGB (green); inset, 6x from the original pictures. P, portal vein; C, central vein. ( b ) Percentages of positive SiR-FG staining area per total areas of whole lobe liver sections were quantified (n = 5). ( c ) Hydroxyproline content in the liver tissue (µg/mg total protein) (n = 9 to 10). ( d ) Percentages of CYGB- and αSMA- positive areas per total areas of whole- lobe liver sections were quantified. ( e ) Immunoblot analysis shows COL1α1, αSMA, CYGB, and <t>mCherry</t> protein expression in the liver tissues of WT and TG mice in images and according to densitometric intensity quantification. GAPDH was used as a loading control and for normalization. Full-length Western blots of one gel are presented in Supplementary Fig. S8 . ( f ) mRNA expression levels of Cygb , mCherry , Col1α1 , αSma , Tgf-β1 Tgf-β3 , and Ppar-γ in the liver were determined by RT-qPCR ( n = 9 to 10). Levels were normalized to Gapdh . ( g ) Liver tissues from WT and TG mice were examined by immunoblotting for phosphorylated- and total SMAD3. The densitometric intensity of phosphorylated-SMAD3 was quantified. GAPDH was used as a loading control and for normalization. Full-length Western blots of one gel are presented in Supplementary Fig. S9 . Data are expressed as the mean ± SD (n = 5). Values are given as the mean ± SD of all experiments. *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001.
    Anti Cygb, supplied by Abcam, used in various techniques. Bioz Stars score: 92/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    90
    Santa Cruz Biotechnology anti cygb
    The expression of <t>Cygb</t> in HI animals injected with shRNA or cDNA. The expression of Cygb mRNA at 18 h post-HI was confirmed by RT-PCR ( A ). The expression of CYGB protein at 24 h post-HI was confirmed by Western blotting using anti-CYGB antibody ( C ). <t>β-Actin</t>
    Anti Cygb, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 90/100, based on 14 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Genoway cygb knockout ko mouse model
    The expression of <t>Cygb</t> in HI animals injected with shRNA or cDNA. The expression of Cygb mRNA at 18 h post-HI was confirmed by RT-PCR ( A ). The expression of CYGB protein at 24 h post-HI was confirmed by Western blotting using anti-CYGB antibody ( C ). <t>β-Actin</t>
    Cygb Knockout Ko Mouse Model, supplied by Genoway, used in various techniques. Bioz Stars score: 92/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    cygb  (Abnova)
    88
    Abnova cygb
    5F 203 confers anticancer activity, induces <t>CYGB</t> and pro-apoptotic protein expression and produces caspase 3 cleavage in vitro. (A) Colony formation analysis in MDA-MB-468 cells untransfected, transfected with empty vector or transfected to overexpress CYGB. Lower panel: quantification of colony numbers. (B) Assessment of migration in untransfected, shGFP and shCYGB MDA-MB-468 cells following 5F 203 treatment. Lower panel: wound closure, normalized to control. (C,D) Determination of CYGB, <t>BAK-1,</t> LTA and GADD45A protein expression as well as caspase 3 cleavage in untransfected, shGFP and shCYGB MDA-MB-468 cells following treatment with 5F 203 (1 μM, 72h). Scale bar = 100 μm. Bars, SEM. Statistically significant where indicated as designated * P
    Cygb, supplied by Abnova, used in various techniques. Bioz Stars score: 88/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Thermo Fisher rabbit anti cygb polyclonal ab
    5F 203 confers anticancer activity, induces <t>CYGB</t> and pro-apoptotic protein expression and produces caspase 3 cleavage in vitro. (A) Colony formation analysis in MDA-MB-468 cells untransfected, transfected with empty vector or transfected to overexpress CYGB. Lower panel: quantification of colony numbers. (B) Assessment of migration in untransfected, shGFP and shCYGB MDA-MB-468 cells following 5F 203 treatment. Lower panel: wound closure, normalized to control. (C,D) Determination of CYGB, <t>BAK-1,</t> LTA and GADD45A protein expression as well as caspase 3 cleavage in untransfected, shGFP and shCYGB MDA-MB-468 cells following treatment with 5F 203 (1 μM, 72h). Scale bar = 100 μm. Bars, SEM. Statistically significant where indicated as designated * P
    Rabbit Anti Cygb Polyclonal Ab, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 92/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Genoway project number genoway jco hsa2 cygb 010611
    5F 203 confers anticancer activity, induces <t>CYGB</t> and pro-apoptotic protein expression and produces caspase 3 cleavage in vitro. (A) Colony formation analysis in MDA-MB-468 cells untransfected, transfected with empty vector or transfected to overexpress CYGB. Lower panel: quantification of colony numbers. (B) Assessment of migration in untransfected, shGFP and shCYGB MDA-MB-468 cells following 5F 203 treatment. Lower panel: wound closure, normalized to control. (C,D) Determination of CYGB, <t>BAK-1,</t> LTA and GADD45A protein expression as well as caspase 3 cleavage in untransfected, shGFP and shCYGB MDA-MB-468 cells following treatment with 5F 203 (1 μM, 72h). Scale bar = 100 μm. Bars, SEM. Statistically significant where indicated as designated * P
    Project Number Genoway Jco Hsa2 Cygb 010611, supplied by Genoway, used in various techniques. Bioz Stars score: 92/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Abnova anti cygb
    Expression analysis of LEDGF/p75 and <t>CYGB</t> in docetaxel resistant DU 145 cells. LEDGF/p75 and CYGB transcript and protein overexpression in a docetaxel resistant DU 145 cell line (DU 145-DR) were verified using qPCR ( A ) and immunoblotting ( B ), respectively. Transient knockdown of LEDGF/p75 and its effect on CYGB expression were evaluated in DU145-DR cells by qPCR ( C ) and immunoblotting ( D ). Protein loading was assessed with antibody to <t>β-actin.</t> * P
    Anti Cygb, supplied by Abnova, used in various techniques. Bioz Stars score: 85/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Abcam anti cytoglobin cygb
    Expression analysis of LEDGF/p75 and <t>CYGB</t> in docetaxel resistant DU 145 cells. LEDGF/p75 and CYGB transcript and protein overexpression in a docetaxel resistant DU 145 cell line (DU 145-DR) were verified using qPCR ( A ) and immunoblotting ( B ), respectively. Transient knockdown of LEDGF/p75 and its effect on CYGB expression were evaluated in DU145-DR cells by qPCR ( C ) and immunoblotting ( D ). Protein loading was assessed with antibody to <t>β-actin.</t> * P
    Anti Cytoglobin Cygb, supplied by Abcam, used in various techniques. Bioz Stars score: 94/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Sigma-Genosys cygb amplicon
    Expression analysis of LEDGF/p75 and <t>CYGB</t> in docetaxel resistant DU 145 cells. LEDGF/p75 and CYGB transcript and protein overexpression in a docetaxel resistant DU 145 cell line (DU 145-DR) were verified using qPCR ( A ) and immunoblotting ( B ), respectively. Transient knockdown of LEDGF/p75 and its effect on CYGB expression were evaluated in DU145-DR cells by qPCR ( C ) and immunoblotting ( D ). Protein loading was assessed with antibody to <t>β-actin.</t> * P
    Cygb Amplicon, supplied by Sigma-Genosys, used in various techniques. Bioz Stars score: 85/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Marinus cygb genes
    Expression analysis of LEDGF/p75 and <t>CYGB</t> in docetaxel resistant DU 145 cells. LEDGF/p75 and CYGB transcript and protein overexpression in a docetaxel resistant DU 145 cell line (DU 145-DR) were verified using qPCR ( A ) and immunoblotting ( B ), respectively. Transient knockdown of LEDGF/p75 and its effect on CYGB expression were evaluated in DU145-DR cells by qPCR ( C ) and immunoblotting ( D ). Protein loading was assessed with antibody to <t>β-actin.</t> * P
    Cygb Genes, supplied by Marinus, used in various techniques. Bioz Stars score: 88/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    OriGene cygb cdna clones
    Expression analysis of LEDGF/p75 and <t>CYGB</t> in docetaxel resistant DU 145 cells. LEDGF/p75 and CYGB transcript and protein overexpression in a docetaxel resistant DU 145 cell line (DU 145-DR) were verified using qPCR ( A ) and immunoblotting ( B ), respectively. Transient knockdown of LEDGF/p75 and its effect on CYGB expression were evaluated in DU145-DR cells by qPCR ( C ) and immunoblotting ( D ). Protein loading was assessed with antibody to <t>β-actin.</t> * P
    Cygb Cdna Clones, supplied by OriGene, used in various techniques. Bioz Stars score: 92/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Qiagen cygb sirna
    Quantification of <t>CYGB</t> expression in oesophageal cells subjected to transient transfection or <t>siRNA</t> knockdown. Levels of CYGB expression, compared to ACTB expression, were determined by real time RT-PCR in a parallel sample for each comet assay performed. These expression levels therefore relate directly to the comet assay data presented in Figure 3 . The geometric mean of all experiments is shown for both TE-8 cells (orange bars) and NE1 cells (pink bars).
    Cygb Sirna, supplied by Qiagen, used in various techniques. Bioz Stars score: 99/100, based on 27 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    86
    Thermo Fisher cygb sirna
    Quantification of <t>CYGB</t> expression in oesophageal cells subjected to transient transfection or <t>siRNA</t> knockdown. Levels of CYGB expression, compared to ACTB expression, were determined by real time RT-PCR in a parallel sample for each comet assay performed. These expression levels therefore relate directly to the comet assay data presented in Figure 3 . The geometric mean of all experiments is shown for both TE-8 cells (orange bars) and NE1 cells (pink bars).
    Cygb Sirna, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 86/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Dojindo Labs cygb
    Effect of <t>Cygb</t> deficiency in the expression of bile transporters and CD10 in acute <t>BDL</t> mice. Hepatic mRNA level of ( A ) sinusoidal (Mrp2, Mdr2, Bsep) and ( B ) canalicular (Mrp3, Ntcp, Oatp1) transporters of bile components in sham (S) and acute BDL (24–48 h) mice. ( C ) Immunofluorescence (top panels) and immunoblot (bottom panels) of CD10 in sham and BDL-24 h mice. GAPDH was used as loading control. All gels were run under the same experimental conditions. The cropped gels are used and full-length gels are presented in Supplementary Fig. S9 . ( D ) Quantitative densitometry of CD10 (top panel) and hepatic mRNA level (bottom panel) of CD10 in sham (S) and acute BDL (24–72 h) and chronic BDL (1–3 W) mice. Open bars, WT; close bars, Cygb −/− . Data represent the mean ± SD. Sham (n = 3), BDL (n = 4–8). *p
    Cygb, supplied by Dojindo Labs, used in various techniques. Bioz Stars score: 91/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    SABiosciences cytoglobin cygb genes
    Effect of <t>Cygb</t> deficiency in the expression of bile transporters and CD10 in acute <t>BDL</t> mice. Hepatic mRNA level of ( A ) sinusoidal (Mrp2, Mdr2, Bsep) and ( B ) canalicular (Mrp3, Ntcp, Oatp1) transporters of bile components in sham (S) and acute BDL (24–48 h) mice. ( C ) Immunofluorescence (top panels) and immunoblot (bottom panels) of CD10 in sham and BDL-24 h mice. GAPDH was used as loading control. All gels were run under the same experimental conditions. The cropped gels are used and full-length gels are presented in Supplementary Fig. S9 . ( D ) Quantitative densitometry of CD10 (top panel) and hepatic mRNA level (bottom panel) of CD10 in sham (S) and acute BDL (24–72 h) and chronic BDL (1–3 W) mice. Open bars, WT; close bars, Cygb −/− . Data represent the mean ± SD. Sham (n = 3), BDL (n = 4–8). *p
    Cytoglobin Cygb Genes, supplied by SABiosciences, used in various techniques. Bioz Stars score: 88/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Thermo Fisher gene exp cygb hs00964894 g1
    Effect of <t>Cygb</t> deficiency in the expression of bile transporters and CD10 in acute <t>BDL</t> mice. Hepatic mRNA level of ( A ) sinusoidal (Mrp2, Mdr2, Bsep) and ( B ) canalicular (Mrp3, Ntcp, Oatp1) transporters of bile components in sham (S) and acute BDL (24–48 h) mice. ( C ) Immunofluorescence (top panels) and immunoblot (bottom panels) of CD10 in sham and BDL-24 h mice. GAPDH was used as loading control. All gels were run under the same experimental conditions. The cropped gels are used and full-length gels are presented in Supplementary Fig. S9 . ( D ) Quantitative densitometry of CD10 (top panel) and hepatic mRNA level (bottom panel) of CD10 in sham (S) and acute BDL (24–72 h) and chronic BDL (1–3 W) mice. Open bars, WT; close bars, Cygb −/− . Data represent the mean ± SD. Sham (n = 3), BDL (n = 4–8). *p
    Gene Exp Cygb Hs00964894 G1, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Thermo Fisher gene exp cygb mm00446071 m1
    Effect of <t>Cygb</t> deficiency in the expression of bile transporters and CD10 in acute <t>BDL</t> mice. Hepatic mRNA level of ( A ) sinusoidal (Mrp2, Mdr2, Bsep) and ( B ) canalicular (Mrp3, Ntcp, Oatp1) transporters of bile components in sham (S) and acute BDL (24–48 h) mice. ( C ) Immunofluorescence (top panels) and immunoblot (bottom panels) of CD10 in sham and BDL-24 h mice. GAPDH was used as loading control. All gels were run under the same experimental conditions. The cropped gels are used and full-length gels are presented in Supplementary Fig. S9 . ( D ) Quantitative densitometry of CD10 (top panel) and hepatic mRNA level (bottom panel) of CD10 in sham (S) and acute BDL (24–72 h) and chronic BDL (1–3 W) mice. Open bars, WT; close bars, Cygb −/− . Data represent the mean ± SD. Sham (n = 3), BDL (n = 4–8). *p
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    89
    OriGene human cygb gene
    Effect of <t>Cygb</t> deficiency in the expression of bile transporters and CD10 in acute <t>BDL</t> mice. Hepatic mRNA level of ( A ) sinusoidal (Mrp2, Mdr2, Bsep) and ( B ) canalicular (Mrp3, Ntcp, Oatp1) transporters of bile components in sham (S) and acute BDL (24–48 h) mice. ( C ) Immunofluorescence (top panels) and immunoblot (bottom panels) of CD10 in sham and BDL-24 h mice. GAPDH was used as loading control. All gels were run under the same experimental conditions. The cropped gels are used and full-length gels are presented in Supplementary Fig. S9 . ( D ) Quantitative densitometry of CD10 (top panel) and hepatic mRNA level (bottom panel) of CD10 in sham (S) and acute BDL (24–72 h) and chronic BDL (1–3 W) mice. Open bars, WT; close bars, Cygb −/− . Data represent the mean ± SD. Sham (n = 3), BDL (n = 4–8). *p
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    OriGene human cygb protein
    Effect of <t>Cygb</t> deficiency in the expression of bile transporters and CD10 in acute <t>BDL</t> mice. Hepatic mRNA level of ( A ) sinusoidal (Mrp2, Mdr2, Bsep) and ( B ) canalicular (Mrp3, Ntcp, Oatp1) transporters of bile components in sham (S) and acute BDL (24–48 h) mice. ( C ) Immunofluorescence (top panels) and immunoblot (bottom panels) of CD10 in sham and BDL-24 h mice. GAPDH was used as loading control. All gels were run under the same experimental conditions. The cropped gels are used and full-length gels are presented in Supplementary Fig. S9 . ( D ) Quantitative densitometry of CD10 (top panel) and hepatic mRNA level (bottom panel) of CD10 in sham (S) and acute BDL (24–72 h) and chronic BDL (1–3 W) mice. Open bars, WT; close bars, Cygb −/− . Data represent the mean ± SD. Sham (n = 3), BDL (n = 4–8). *p
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    Image Search Results


    Quantitative real-time RT-PCR analysis of HO-1 (a), AQP-1 (b), and Bcl-2 (c) expressions in mixed hTECs under basal conditions and after GO-induced oxidative stress (1 IU/mL) either or not in the presence of epoetin delta (100 IU/mL). GAPDH was used as endogenous control housekeeping gene. Preconditioning the cells with EPO resulted in a significant upregulation of HO-1 and AQP-1 mRNA. GO-induced oxidative stress further increased HO-1, AQP-1, and Bcl-2 mRNA expressions with maximum levels 60 minutes after induction of oxidative stress. Data are presented as the mean ± SD of triplicate determinations of 2 runs (i.e., 6 values each). * P

    Journal: Journal of Biomedicine and Biotechnology

    Article Title: Epoetin Delta Reduces Oxidative Stress in Primary Human Renal Tubular Cells

    doi: 10.1155/2010/395785

    Figure Lengend Snippet: Quantitative real-time RT-PCR analysis of HO-1 (a), AQP-1 (b), and Bcl-2 (c) expressions in mixed hTECs under basal conditions and after GO-induced oxidative stress (1 IU/mL) either or not in the presence of epoetin delta (100 IU/mL). GAPDH was used as endogenous control housekeeping gene. Preconditioning the cells with EPO resulted in a significant upregulation of HO-1 and AQP-1 mRNA. GO-induced oxidative stress further increased HO-1, AQP-1, and Bcl-2 mRNA expressions with maximum levels 60 minutes after induction of oxidative stress. Data are presented as the mean ± SD of triplicate determinations of 2 runs (i.e., 6 values each). * P

    Article Snippet: Ready to use, predesigned, primer and probe sets (Applied Biosystems) for human genes of interest (Hs00157965_m1 for HO-1, Hs00166067_m1 for AQP-1, Hs00153350_m1 for Bcl-2, Hs00266395_m1 for CPM, Hs00175210_m1 for DPPIV, Hs00370478_m1 for Cygb) and the housekeeping gene GAPDH (Hs99999905_m1) were used according to the manufacturer's guidelines.

    Techniques: Quantitative RT-PCR

    Cygb overexpression increases adipogenic differentiation. a Structure of Cygb gene construct, b relative mRNA expression level of Cygb in transduced cells, c oil red staining of differentiated 3T3-F442A–GFP and 3T3-F442A–Cygb cells, d gene expression levels of PPARγ, CEBPα and FABP4 in differentiated 3T3-F442A–GFP and 3T3-F442A–Cygb cells at the end of 8 day differentiation period. * P

    Journal: Cytotechnology

    Article Title: Cytoglobin: a potential marker for adipogenic differentiation in preadipocytes in vitro

    doi: 10.1007/s10616-016-0047-2

    Figure Lengend Snippet: Cygb overexpression increases adipogenic differentiation. a Structure of Cygb gene construct, b relative mRNA expression level of Cygb in transduced cells, c oil red staining of differentiated 3T3-F442A–GFP and 3T3-F442A–Cygb cells, d gene expression levels of PPARγ, CEBPα and FABP4 in differentiated 3T3-F442A–GFP and 3T3-F442A–Cygb cells at the end of 8 day differentiation period. * P

    Article Snippet: Differentiated 3T3-F442A–GFP and 3T3-F442A–Cygb cells were stained with Oil red O (ORO) (Sigma, St. Louis, MO, USA) to observe lipid vesicles.

    Techniques: Over Expression, Construct, Expressing, Staining

    Diagram showing the differentiation procedure of 3T3-F442A cells. Cygb expression increased at the end of the adipogenesis process

    Journal: Cytotechnology

    Article Title: Cytoglobin: a potential marker for adipogenic differentiation in preadipocytes in vitro

    doi: 10.1007/s10616-016-0047-2

    Figure Lengend Snippet: Diagram showing the differentiation procedure of 3T3-F442A cells. Cygb expression increased at the end of the adipogenesis process

    Article Snippet: Differentiated 3T3-F442A–GFP and 3T3-F442A–Cygb cells were stained with Oil red O (ORO) (Sigma, St. Louis, MO, USA) to observe lipid vesicles.

    Techniques: Expressing

    Western blot analysis of Cygb, PPARγ, CEBPα and FABP4. during differentiation process. a Western blot results of Cygb, PPARγ, CEBPα and FABP4 during differentiation process, b quantitative results of western blot band intensities. * P

    Journal: Cytotechnology

    Article Title: Cytoglobin: a potential marker for adipogenic differentiation in preadipocytes in vitro

    doi: 10.1007/s10616-016-0047-2

    Figure Lengend Snippet: Western blot analysis of Cygb, PPARγ, CEBPα and FABP4. during differentiation process. a Western blot results of Cygb, PPARγ, CEBPα and FABP4 during differentiation process, b quantitative results of western blot band intensities. * P

    Article Snippet: Primary antibodies against PPARγ (1:1000 dilution; MA5-14889; Sigma), CEBPα (1:1000 dilution; 8178; Cell Signaling Technology), FABP4 (1:1000 dilution; sc-271529; Santa Cruz Biotechnology), and Cygb (1:1000 dilution; sc-271529; Santa Cruz Biotechnology) were used for western blot analysis.

    Techniques: Western Blot

    Cygb overexpression increases adipogenic differentiation. a Structure of Cygb gene construct, b relative mRNA expression level of Cygb in transduced cells, c oil red staining of differentiated 3T3-F442A–GFP and 3T3-F442A–Cygb cells, d gene expression levels of PPARγ, CEBPα and FABP4 in differentiated 3T3-F442A–GFP and 3T3-F442A–Cygb cells at the end of 8 day differentiation period. * P

    Journal: Cytotechnology

    Article Title: Cytoglobin: a potential marker for adipogenic differentiation in preadipocytes in vitro

    doi: 10.1007/s10616-016-0047-2

    Figure Lengend Snippet: Cygb overexpression increases adipogenic differentiation. a Structure of Cygb gene construct, b relative mRNA expression level of Cygb in transduced cells, c oil red staining of differentiated 3T3-F442A–GFP and 3T3-F442A–Cygb cells, d gene expression levels of PPARγ, CEBPα and FABP4 in differentiated 3T3-F442A–GFP and 3T3-F442A–Cygb cells at the end of 8 day differentiation period. * P

    Article Snippet: Primary antibodies against PPARγ (1:1000 dilution; MA5-14889; Sigma), CEBPα (1:1000 dilution; 8178; Cell Signaling Technology), FABP4 (1:1000 dilution; sc-271529; Santa Cruz Biotechnology), and Cygb (1:1000 dilution; sc-271529; Santa Cruz Biotechnology) were used for western blot analysis.

    Techniques: Over Expression, Construct, Expressing, Staining

    Gene expression analysis of Cygb, PPARγ, CEBPα and FABP4 during differentiation process. Indicated genes showed an increased in relative mRNA expression in a time dependent manner. * P

    Journal: Cytotechnology

    Article Title: Cytoglobin: a potential marker for adipogenic differentiation in preadipocytes in vitro

    doi: 10.1007/s10616-016-0047-2

    Figure Lengend Snippet: Gene expression analysis of Cygb, PPARγ, CEBPα and FABP4 during differentiation process. Indicated genes showed an increased in relative mRNA expression in a time dependent manner. * P

    Article Snippet: Primary antibodies against PPARγ (1:1000 dilution; MA5-14889; Sigma), CEBPα (1:1000 dilution; 8178; Cell Signaling Technology), FABP4 (1:1000 dilution; sc-271529; Santa Cruz Biotechnology), and Cygb (1:1000 dilution; sc-271529; Santa Cruz Biotechnology) were used for western blot analysis.

    Techniques: Expressing

    Immunocytochemistry analysis of PPARγ, CEBPα and FABP4 during differentiation process. PPARγ, CEBPα and FABP4 staining at day 0 and day 8 is shown. Expression of PPARγ, CEBPα and FABP4 increased during differentiation process. DAPI: Nuclei staining, FITC: Cygb staining. Scale bar 10 µm

    Journal: Cytotechnology

    Article Title: Cytoglobin: a potential marker for adipogenic differentiation in preadipocytes in vitro

    doi: 10.1007/s10616-016-0047-2

    Figure Lengend Snippet: Immunocytochemistry analysis of PPARγ, CEBPα and FABP4 during differentiation process. PPARγ, CEBPα and FABP4 staining at day 0 and day 8 is shown. Expression of PPARγ, CEBPα and FABP4 increased during differentiation process. DAPI: Nuclei staining, FITC: Cygb staining. Scale bar 10 µm

    Article Snippet: Primary antibodies against PPARγ (1:1000 dilution; MA5-14889; Sigma), CEBPα (1:1000 dilution; 8178; Cell Signaling Technology), FABP4 (1:1000 dilution; sc-271529; Santa Cruz Biotechnology), and Cygb (1:1000 dilution; sc-271529; Santa Cruz Biotechnology) were used for western blot analysis.

    Techniques: Immunocytochemistry, Staining, Expressing

    Inhibition of liver fibrosis development in long-term TAA-treated Cygb-TG mice. WT and Cygb-TG (TG) mice were subjected to TAA treatment for 10 weeks (TAA-10w). ( a ) Representative liver images of H E, Sirius Red and Fast Green (SiR-FG) staining for collagen deposition and IF staining for the detection of αSMA (red) and CYGB (green); inset, 6x from the original pictures. P, portal vein; C, central vein. ( b ) Percentages of positive SiR-FG staining area per total areas of whole lobe liver sections were quantified (n = 5). ( c ) Hydroxyproline content in the liver tissue (µg/mg total protein) (n = 9 to 10). ( d ) Percentages of CYGB- and αSMA- positive areas per total areas of whole- lobe liver sections were quantified. ( e ) Immunoblot analysis shows COL1α1, αSMA, CYGB, and mCherry protein expression in the liver tissues of WT and TG mice in images and according to densitometric intensity quantification. GAPDH was used as a loading control and for normalization. Full-length Western blots of one gel are presented in Supplementary Fig. S8 . ( f ) mRNA expression levels of Cygb , mCherry , Col1α1 , αSma , Tgf-β1 Tgf-β3 , and Ppar-γ in the liver were determined by RT-qPCR ( n = 9 to 10). Levels were normalized to Gapdh . ( g ) Liver tissues from WT and TG mice were examined by immunoblotting for phosphorylated- and total SMAD3. The densitometric intensity of phosphorylated-SMAD3 was quantified. GAPDH was used as a loading control and for normalization. Full-length Western blots of one gel are presented in Supplementary Fig. S9 . Data are expressed as the mean ± SD (n = 5). Values are given as the mean ± SD of all experiments. *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001.

    Journal: Scientific Reports

    Article Title: Selective overexpression of cytoglobin in stellate cells attenuates thioacetamide-induced liver fibrosis in mice

    doi: 10.1038/s41598-018-36215-4

    Figure Lengend Snippet: Inhibition of liver fibrosis development in long-term TAA-treated Cygb-TG mice. WT and Cygb-TG (TG) mice were subjected to TAA treatment for 10 weeks (TAA-10w). ( a ) Representative liver images of H E, Sirius Red and Fast Green (SiR-FG) staining for collagen deposition and IF staining for the detection of αSMA (red) and CYGB (green); inset, 6x from the original pictures. P, portal vein; C, central vein. ( b ) Percentages of positive SiR-FG staining area per total areas of whole lobe liver sections were quantified (n = 5). ( c ) Hydroxyproline content in the liver tissue (µg/mg total protein) (n = 9 to 10). ( d ) Percentages of CYGB- and αSMA- positive areas per total areas of whole- lobe liver sections were quantified. ( e ) Immunoblot analysis shows COL1α1, αSMA, CYGB, and mCherry protein expression in the liver tissues of WT and TG mice in images and according to densitometric intensity quantification. GAPDH was used as a loading control and for normalization. Full-length Western blots of one gel are presented in Supplementary Fig. S8 . ( f ) mRNA expression levels of Cygb , mCherry , Col1α1 , αSma , Tgf-β1 Tgf-β3 , and Ppar-γ in the liver were determined by RT-qPCR ( n = 9 to 10). Levels were normalized to Gapdh . ( g ) Liver tissues from WT and TG mice were examined by immunoblotting for phosphorylated- and total SMAD3. The densitometric intensity of phosphorylated-SMAD3 was quantified. GAPDH was used as a loading control and for normalization. Full-length Western blots of one gel are presented in Supplementary Fig. S9 . Data are expressed as the mean ± SD (n = 5). Values are given as the mean ± SD of all experiments. *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001.

    Article Snippet: After the membranes were blocked by 5% skim milk, they were probed with the following primary antibodies: anti-CYGB (1:2000; our laboratory), anti-mCherry (1:1000; Abcam, Japan), anti-αSMA (1:2000; DAKO, UK), anti-phosphorylated- and total-SMAD3 (1:1000; Abcam, Japan), anti-phosphorylated- and total-NF-κB (1:1000; Cell Signaling, Japan), or anti-GAPDH (1:2000; Santa Cruz Biotechnology, Santa Cruz, CA).

    Techniques: Inhibition, Mouse Assay, Staining, Expressing, Western Blot, Quantitative RT-PCR

    Overexpression of Cygb attenuates HSC activation. ( a ) Morphology of primary hepatic stellate cells from WT (HSCs Cygb- WT ) and TG mice (HSCs Cygb-TG ) at days 1, 4 and 7. ( b ) Image of lipid droplets and fluorescent mCherry protein in primary HSCs Cygb-TG at day 1. Inset, 3x from original pictures. ( c ) Immunoblot analysis of CYGB, mCherry and αSMA protein expression in primary HSCs Cygb- WT and HSCs Cygb-TG under normal conditions. Right panels show the quantification of the densitometric intensity. Levels were normalized to GAPDH. Full-length Western blots in one gel are presented in Supplementary Fig. S7 . ( d ) mRNA expression levels of Cygb , αSma , Col1α1 and Tgf-β3 in primary HSCs Cygb- WT (white bars) and HSCs Cygb-TG (red bars) at days 1 and 4 were determined by RT-qPCR (n = 5 to 10). Levels were normalized to Gapdh . Values are given as the mean ± SD of all experiments. *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001.

    Journal: Scientific Reports

    Article Title: Selective overexpression of cytoglobin in stellate cells attenuates thioacetamide-induced liver fibrosis in mice

    doi: 10.1038/s41598-018-36215-4

    Figure Lengend Snippet: Overexpression of Cygb attenuates HSC activation. ( a ) Morphology of primary hepatic stellate cells from WT (HSCs Cygb- WT ) and TG mice (HSCs Cygb-TG ) at days 1, 4 and 7. ( b ) Image of lipid droplets and fluorescent mCherry protein in primary HSCs Cygb-TG at day 1. Inset, 3x from original pictures. ( c ) Immunoblot analysis of CYGB, mCherry and αSMA protein expression in primary HSCs Cygb- WT and HSCs Cygb-TG under normal conditions. Right panels show the quantification of the densitometric intensity. Levels were normalized to GAPDH. Full-length Western blots in one gel are presented in Supplementary Fig. S7 . ( d ) mRNA expression levels of Cygb , αSma , Col1α1 and Tgf-β3 in primary HSCs Cygb- WT (white bars) and HSCs Cygb-TG (red bars) at days 1 and 4 were determined by RT-qPCR (n = 5 to 10). Levels were normalized to Gapdh . Values are given as the mean ± SD of all experiments. *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001.

    Article Snippet: After the membranes were blocked by 5% skim milk, they were probed with the following primary antibodies: anti-CYGB (1:2000; our laboratory), anti-mCherry (1:1000; Abcam, Japan), anti-αSMA (1:2000; DAKO, UK), anti-phosphorylated- and total-SMAD3 (1:1000; Abcam, Japan), anti-phosphorylated- and total-NF-κB (1:1000; Cell Signaling, Japan), or anti-GAPDH (1:2000; Santa Cruz Biotechnology, Santa Cruz, CA).

    Techniques: Over Expression, Activation Assay, Mouse Assay, Expressing, Western Blot, Quantitative RT-PCR

    Generation of hepatic stellate cell-specific Cygb-transgenic mice. ( a ) DNA construct map for Cygb-mCherry. The Cygb-2A-mCherry reporter was generated with the BAC clone RP23-330N7A, and a partial genomic map of the Cygb gene with coding exons (black boxes), noncoding regions including the promoter of the Cygb gene (light grey boxes), and flanking introns (solid lines) is shown. The 2A-mCherry reporter gene, which was flanked by 110 bp of the upstream sequence of the Cygb gene stop codon and 87 bp of the downstream sequence of its stop codon, was precisely transferred to the Cygb gene. ( b ) Genotyping of Cygb-transgenic ( TG) mice in the F1 generation by Southern blots showed TG mice (lanes 9, 11, 14, and 15) bearing 10 copies of the Cygb transgenes. NC, DNA negative control; PC, DNA positive controls with 1, 3, 10, and 30 copies. ( c ) Genotyping of offspring by real-time qRT-PCR. DNA isolated from tail biopsies of Cygb 10 copies-transgenic founder (TgFD10c) mice was used as a positive control, and DNA from the 1-copy founder (TgFD1c) was used as a reference sample. Relative quantification of Cygb (green bars) and mCherry (red bars) DNA is shown. Mouse numbers BAC 687, 690–694, and 696 were clarified as Cygb-TG, and the remaining mice were WT. The relative number of DNA copies was normalized to Gapdh levels. ( d ) Macroscopic view of multiple organs of WT and Cygb-TG mice under a contrast photo (right panel) and fluorescence images of Cherry (left panel). ( e ) Real-time qRT-PCR analysis shows the Cygb expression levels in multiple organs of WT (white bars) and TG mice (green bars). Transcriptional levels of only mCherry were examined in TG mice (red bars). Gapdh was used as an endogenous control. ( f ) Immunoblot analysis showed the CYGB and mCherry protein levels in multiple organs of WT and TG mice. GAPDH was used as the loading control and for normalization. Full-length Western blots in one gel are presented in Supplementary Fig. S6 . ( g ) Representative imaging of CYGB (green) and mCherry (red) immunofluorescence staining from WT and TG livers. Br, Brain; Li, Liver; Panc, Pancreas; He, Heart; Lg, Lung; Int, Intestine; Sp, Spleen; Kid, Kidney. P, portal vein; C, central vein.

    Journal: Scientific Reports

    Article Title: Selective overexpression of cytoglobin in stellate cells attenuates thioacetamide-induced liver fibrosis in mice

    doi: 10.1038/s41598-018-36215-4

    Figure Lengend Snippet: Generation of hepatic stellate cell-specific Cygb-transgenic mice. ( a ) DNA construct map for Cygb-mCherry. The Cygb-2A-mCherry reporter was generated with the BAC clone RP23-330N7A, and a partial genomic map of the Cygb gene with coding exons (black boxes), noncoding regions including the promoter of the Cygb gene (light grey boxes), and flanking introns (solid lines) is shown. The 2A-mCherry reporter gene, which was flanked by 110 bp of the upstream sequence of the Cygb gene stop codon and 87 bp of the downstream sequence of its stop codon, was precisely transferred to the Cygb gene. ( b ) Genotyping of Cygb-transgenic ( TG) mice in the F1 generation by Southern blots showed TG mice (lanes 9, 11, 14, and 15) bearing 10 copies of the Cygb transgenes. NC, DNA negative control; PC, DNA positive controls with 1, 3, 10, and 30 copies. ( c ) Genotyping of offspring by real-time qRT-PCR. DNA isolated from tail biopsies of Cygb 10 copies-transgenic founder (TgFD10c) mice was used as a positive control, and DNA from the 1-copy founder (TgFD1c) was used as a reference sample. Relative quantification of Cygb (green bars) and mCherry (red bars) DNA is shown. Mouse numbers BAC 687, 690–694, and 696 were clarified as Cygb-TG, and the remaining mice were WT. The relative number of DNA copies was normalized to Gapdh levels. ( d ) Macroscopic view of multiple organs of WT and Cygb-TG mice under a contrast photo (right panel) and fluorescence images of Cherry (left panel). ( e ) Real-time qRT-PCR analysis shows the Cygb expression levels in multiple organs of WT (white bars) and TG mice (green bars). Transcriptional levels of only mCherry were examined in TG mice (red bars). Gapdh was used as an endogenous control. ( f ) Immunoblot analysis showed the CYGB and mCherry protein levels in multiple organs of WT and TG mice. GAPDH was used as the loading control and for normalization. Full-length Western blots in one gel are presented in Supplementary Fig. S6 . ( g ) Representative imaging of CYGB (green) and mCherry (red) immunofluorescence staining from WT and TG livers. Br, Brain; Li, Liver; Panc, Pancreas; He, Heart; Lg, Lung; Int, Intestine; Sp, Spleen; Kid, Kidney. P, portal vein; C, central vein.

    Article Snippet: After the membranes were blocked by 5% skim milk, they were probed with the following primary antibodies: anti-CYGB (1:2000; our laboratory), anti-mCherry (1:1000; Abcam, Japan), anti-αSMA (1:2000; DAKO, UK), anti-phosphorylated- and total-SMAD3 (1:1000; Abcam, Japan), anti-phosphorylated- and total-NF-κB (1:1000; Cell Signaling, Japan), or anti-GAPDH (1:2000; Santa Cruz Biotechnology, Santa Cruz, CA).

    Techniques: Transgenic Assay, Mouse Assay, Construct, Generated, BAC Assay, Sequencing, Negative Control, Quantitative RT-PCR, Isolation, Positive Control, Fluorescence, Expressing, Western Blot, Imaging, Immunofluorescence, Staining

    Overview of the potential mechanisms involved in the DpC-mediated effects on neuroblastoma. DpC increases TNFα expression in neuroblastoma cells, which may (1) activate cytotoxic T cells to destroy tumor cells and/or (2) acts on the TNFα receptor ( TNFR ) to activate down-stream signaling pathways. These include the MAPK/p38/JNK and NF-ĸB signaling cascades, which lead to nuclear transcription of numerous genes, including those that induce apoptosis, as well as cytokines such as TNFα. Activation of TNFR also promotes cleavage of caspase 8, leading to caspase 3 cleavage and subsequent apoptosis. Moreover, DpC is also highly redox active, resulting in the production of reactive oxygen species (ROS) [ 13 ]. The generation of ROS triggers the release of cytochrome c from mitochondria [ 8 ], leading to cleavage of caspase 9, which then also cleaves caspase 3, leading to apoptosis. Further, the increased ROS also leads to upregulation of neuroglobin ( Ngb ) and cytoglobin ( Cygb ) expression as both of these proteins respond to oxidative stress. Together, these molecular effects, which promote apoptosis, could contribute to the anti-cancer activity of DpC in neuroblastoma

    Journal: Journal of Hematology & Oncology

    Article Title: The novel thiosemicarbazone, di-2-pyridylketone 4-cyclohexyl-4-methyl-3-thiosemicarbazone (DpC), inhibits neuroblastoma growth in vitro and in vivo via multiple mechanisms

    doi: 10.1186/s13045-016-0330-x

    Figure Lengend Snippet: Overview of the potential mechanisms involved in the DpC-mediated effects on neuroblastoma. DpC increases TNFα expression in neuroblastoma cells, which may (1) activate cytotoxic T cells to destroy tumor cells and/or (2) acts on the TNFα receptor ( TNFR ) to activate down-stream signaling pathways. These include the MAPK/p38/JNK and NF-ĸB signaling cascades, which lead to nuclear transcription of numerous genes, including those that induce apoptosis, as well as cytokines such as TNFα. Activation of TNFR also promotes cleavage of caspase 8, leading to caspase 3 cleavage and subsequent apoptosis. Moreover, DpC is also highly redox active, resulting in the production of reactive oxygen species (ROS) [ 13 ]. The generation of ROS triggers the release of cytochrome c from mitochondria [ 8 ], leading to cleavage of caspase 9, which then also cleaves caspase 3, leading to apoptosis. Further, the increased ROS also leads to upregulation of neuroglobin ( Ngb ) and cytoglobin ( Cygb ) expression as both of these proteins respond to oxidative stress. Together, these molecular effects, which promote apoptosis, could contribute to the anti-cancer activity of DpC in neuroblastoma

    Article Snippet: Antibodies against Ngb and Cygb (Abcam, Cambridge, UK) were kindly provided by Dr. Tan-Un (School of Professional and Continuing Education, The University of Hong Kong, Hong Kong, People’s Republic of China).

    Techniques: Expressing, Activation Assay, Activity Assay

    a Incubation of DpC (25 μM) with SK-N-LP neuroblastoma cells and HK2 non-tumorigenic, immortalized kidney cells induces Cygb and Ngb expression after a 24-h incubation. In these studies, non-tumorigenic, immortalized cell lines (i.e., MSC, H9c2, or HK2), or neoplastic, neuroblastoma (SK-N-LP) cells, were incubated for either 0, 12 or 24 h/37 °C with either control medium ( Con ; no agent added), Dp44mT (25 μM), or DpC (25 μM) and then flow cytometric analysis performed using Flow Jo 8.8.2. Black , red , and blue lines represent the control, or the cells treated with the iron chelators for 12 and 24 h, respectively. Results shown are typical experiments of three performed. b Western blot analysis of Cygb and Ngb expression in SK-N-LP cells following incubation with control media ( Con ), Dp44mT (25 μM), or DpC (25 μM) for 24 h/37 °C. The bands presented in the blots are representative of three repeats and the lanes have been cropped from raw data images containing all three repeats (raw data shown in Additional File 1 ) for clarity (lanes separated by dotted lines ). Densitometry data in ( b ) are presented as the mean ± SEM ( n = 3). * p

    Journal: Journal of Hematology & Oncology

    Article Title: The novel thiosemicarbazone, di-2-pyridylketone 4-cyclohexyl-4-methyl-3-thiosemicarbazone (DpC), inhibits neuroblastoma growth in vitro and in vivo via multiple mechanisms

    doi: 10.1186/s13045-016-0330-x

    Figure Lengend Snippet: a Incubation of DpC (25 μM) with SK-N-LP neuroblastoma cells and HK2 non-tumorigenic, immortalized kidney cells induces Cygb and Ngb expression after a 24-h incubation. In these studies, non-tumorigenic, immortalized cell lines (i.e., MSC, H9c2, or HK2), or neoplastic, neuroblastoma (SK-N-LP) cells, were incubated for either 0, 12 or 24 h/37 °C with either control medium ( Con ; no agent added), Dp44mT (25 μM), or DpC (25 μM) and then flow cytometric analysis performed using Flow Jo 8.8.2. Black , red , and blue lines represent the control, or the cells treated with the iron chelators for 12 and 24 h, respectively. Results shown are typical experiments of three performed. b Western blot analysis of Cygb and Ngb expression in SK-N-LP cells following incubation with control media ( Con ), Dp44mT (25 μM), or DpC (25 μM) for 24 h/37 °C. The bands presented in the blots are representative of three repeats and the lanes have been cropped from raw data images containing all three repeats (raw data shown in Additional File 1 ) for clarity (lanes separated by dotted lines ). Densitometry data in ( b ) are presented as the mean ± SEM ( n = 3). * p

    Article Snippet: Antibodies against Ngb and Cygb (Abcam, Cambridge, UK) were kindly provided by Dr. Tan-Un (School of Professional and Continuing Education, The University of Hong Kong, Hong Kong, People’s Republic of China).

    Techniques: Incubation, Expressing, Flow Cytometry, Western Blot

    The expression of Cygb in HI animals injected with shRNA or cDNA. The expression of Cygb mRNA at 18 h post-HI was confirmed by RT-PCR ( A ). The expression of CYGB protein at 24 h post-HI was confirmed by Western blotting using anti-CYGB antibody ( C ). β-Actin

    Journal: The Journal of Biological Chemistry

    Article Title: Mechanisms of Neuroprotection from Hypoxia-Ischemia (HI) Brain Injury by Up-regulation of Cytoglobin (CYGB) in a Neonatal Rat Model *

    doi: 10.1074/jbc.M112.428789

    Figure Lengend Snippet: The expression of Cygb in HI animals injected with shRNA or cDNA. The expression of Cygb mRNA at 18 h post-HI was confirmed by RT-PCR ( A ). The expression of CYGB protein at 24 h post-HI was confirmed by Western blotting using anti-CYGB antibody ( C ). β-Actin

    Article Snippet: The membrane was blocked by a 1-h incubation at room temperature in a Tris-buffered saline solution (TBS-T; 20 m m Tris, pH 7.6, 135 m m NaCl, and 0.05% Tween) containing 5% nonfat dry milk and then incubated with different primary antibodies, including anti-CYGB (1:400 dilution; Santa Cruz Biotechnology, Inc.), anti-β-actin (1:1000 dilution; CST) overnight at 4 °C.

    Techniques: Expressing, Injection, shRNA, Reverse Transcription Polymerase Chain Reaction, Western Blot

    Infection of neonatal brain with adenovirus carrying Cygb cDNA or shRNA. CYGB expression at various time points was confirmed by Western blotting using anti-CYGB antibody. β-actin was used as an internal control. A and C , representative electrophoretic

    Journal: The Journal of Biological Chemistry

    Article Title: Mechanisms of Neuroprotection from Hypoxia-Ischemia (HI) Brain Injury by Up-regulation of Cytoglobin (CYGB) in a Neonatal Rat Model *

    doi: 10.1074/jbc.M112.428789

    Figure Lengend Snippet: Infection of neonatal brain with adenovirus carrying Cygb cDNA or shRNA. CYGB expression at various time points was confirmed by Western blotting using anti-CYGB antibody. β-actin was used as an internal control. A and C , representative electrophoretic

    Article Snippet: The membrane was blocked by a 1-h incubation at room temperature in a Tris-buffered saline solution (TBS-T; 20 m m Tris, pH 7.6, 135 m m NaCl, and 0.05% Tween) containing 5% nonfat dry milk and then incubated with different primary antibodies, including anti-CYGB (1:400 dilution; Santa Cruz Biotechnology, Inc.), anti-β-actin (1:1000 dilution; CST) overnight at 4 °C.

    Techniques: Infection, shRNA, Expressing, Western Blot

    5F 203 confers anticancer activity, induces CYGB and pro-apoptotic protein expression and produces caspase 3 cleavage in vitro. (A) Colony formation analysis in MDA-MB-468 cells untransfected, transfected with empty vector or transfected to overexpress CYGB. Lower panel: quantification of colony numbers. (B) Assessment of migration in untransfected, shGFP and shCYGB MDA-MB-468 cells following 5F 203 treatment. Lower panel: wound closure, normalized to control. (C,D) Determination of CYGB, BAK-1, LTA and GADD45A protein expression as well as caspase 3 cleavage in untransfected, shGFP and shCYGB MDA-MB-468 cells following treatment with 5F 203 (1 μM, 72h). Scale bar = 100 μm. Bars, SEM. Statistically significant where indicated as designated * P

    Journal: Journal of cellular biochemistry

    Article Title: Putative tumor suppressor cytoglobin promotes aryl hydrocarbon receptor ligand-mediated triple negative breast cancer cell death

    doi: 10.1002/jcb.27887

    Figure Lengend Snippet: 5F 203 confers anticancer activity, induces CYGB and pro-apoptotic protein expression and produces caspase 3 cleavage in vitro. (A) Colony formation analysis in MDA-MB-468 cells untransfected, transfected with empty vector or transfected to overexpress CYGB. Lower panel: quantification of colony numbers. (B) Assessment of migration in untransfected, shGFP and shCYGB MDA-MB-468 cells following 5F 203 treatment. Lower panel: wound closure, normalized to control. (C,D) Determination of CYGB, BAK-1, LTA and GADD45A protein expression as well as caspase 3 cleavage in untransfected, shGFP and shCYGB MDA-MB-468 cells following treatment with 5F 203 (1 μM, 72h). Scale bar = 100 μm. Bars, SEM. Statistically significant where indicated as designated * P

    Article Snippet: Antibodies for CYGB (Abnova Corporation Cat# H00114757-A01, RRID:AB_461584), BAK-1 (Cell Signaling Technology Cat# 3814S, RRID:AB_2290287), GADD45a (Cell Signaling Technology Cat# 4632S, RRID:AB_10544538), caspase 3 (Cell Signaling Technology Cat# 9662P, RRID:AB_10839261) and LTA (Abnova Corporation Cat# PAB8441, RRID:AB_1676074) were used. and antibodies for β-actin were obtained from Sigma-Aldrich (St. Louis, MO).

    Techniques: Activity Assay, Expressing, In Vitro, Multiple Displacement Amplification, Transfection, Plasmid Preparation, Migration

    5F 203 confers anticancer activity, induces CYGB and pro-apoptotic protein expression and produces caspase 3 cleavage in vivo. (A,B) Effect of Phortress on tumor growth (as measured in tumor volume, N =11 and tumor mass, N = 9) for 3 cycles of 3 doses every other day (Q2D X 3) followed by a 10 d rest period as determined in Materials and Methods. (C,D) CYGB, BAK-1, and GADD45A protein expression as well as caspase 3 cleavage determination in tumor xenografts. N = 6. Bars, SEM. Statistically significant where indicated as designated * P

    Journal: Journal of cellular biochemistry

    Article Title: Putative tumor suppressor cytoglobin promotes aryl hydrocarbon receptor ligand-mediated triple negative breast cancer cell death

    doi: 10.1002/jcb.27887

    Figure Lengend Snippet: 5F 203 confers anticancer activity, induces CYGB and pro-apoptotic protein expression and produces caspase 3 cleavage in vivo. (A,B) Effect of Phortress on tumor growth (as measured in tumor volume, N =11 and tumor mass, N = 9) for 3 cycles of 3 doses every other day (Q2D X 3) followed by a 10 d rest period as determined in Materials and Methods. (C,D) CYGB, BAK-1, and GADD45A protein expression as well as caspase 3 cleavage determination in tumor xenografts. N = 6. Bars, SEM. Statistically significant where indicated as designated * P

    Article Snippet: Antibodies for CYGB (Abnova Corporation Cat# H00114757-A01, RRID:AB_461584), BAK-1 (Cell Signaling Technology Cat# 3814S, RRID:AB_2290287), GADD45a (Cell Signaling Technology Cat# 4632S, RRID:AB_10544538), caspase 3 (Cell Signaling Technology Cat# 9662P, RRID:AB_10839261) and LTA (Abnova Corporation Cat# PAB8441, RRID:AB_1676074) were used. and antibodies for β-actin were obtained from Sigma-Aldrich (St. Louis, MO).

    Techniques: Activity Assay, Expressing, In Vivo

    Expression analysis of LEDGF/p75 and CYGB in docetaxel resistant DU 145 cells. LEDGF/p75 and CYGB transcript and protein overexpression in a docetaxel resistant DU 145 cell line (DU 145-DR) were verified using qPCR ( A ) and immunoblotting ( B ), respectively. Transient knockdown of LEDGF/p75 and its effect on CYGB expression were evaluated in DU145-DR cells by qPCR ( C ) and immunoblotting ( D ). Protein loading was assessed with antibody to β-actin. * P

    Journal: The Prostate

    Article Title: Pathway Specific Gene Expression Profiling Reveals Oxidative Stress Genes Potentially Regulated by Transcription Co-Activator LEDGF/p75 in Prostate Cancer Cells

    doi: 10.1002/pros.21463

    Figure Lengend Snippet: Expression analysis of LEDGF/p75 and CYGB in docetaxel resistant DU 145 cells. LEDGF/p75 and CYGB transcript and protein overexpression in a docetaxel resistant DU 145 cell line (DU 145-DR) were verified using qPCR ( A ) and immunoblotting ( B ), respectively. Transient knockdown of LEDGF/p75 and its effect on CYGB expression were evaluated in DU145-DR cells by qPCR ( C ) and immunoblotting ( D ). Protein loading was assessed with antibody to β-actin. * P

    Article Snippet: The following antibodies were used: Mouse monoclonals anti-β-actin (Sigma-Aldrich), and anti-CYGB (1:200, Abnova); rabbit polyclonals anti-PRDX1 (1:1,000, Abcam), anti-PRDX2 (1:10,000, Abcam), anti-PRDX3 (1:5,000, Abcam), anti-PRDX4 (1:1,000, Abcam), anti-PRDX5 (1:1,000, Santa Cruz Biotechnology), anti-PRDX6 (1:3,000, Abcam), anti-LEDGF/p75 (1:1,000, Novus Biologicals); and horseradish peroxidase (HRP)-labeled secondary IgG antibodies (Zymed).

    Techniques: Expressing, Over Expression, Real-time Polymerase Chain Reaction

    Analysis of changes in CYGB protein expression in response to knockdown or overexpression of LEDG/p75. Expression of CYGB protein was evaluated by immunoblotting in PC-3 cells with transient knockdown ( A ) and stable overexpression of LEDGF/p75 ( B ), to further validate the qPCR data. Protein loading was assessed with antibody to β-actin.

    Journal: The Prostate

    Article Title: Pathway Specific Gene Expression Profiling Reveals Oxidative Stress Genes Potentially Regulated by Transcription Co-Activator LEDGF/p75 in Prostate Cancer Cells

    doi: 10.1002/pros.21463

    Figure Lengend Snippet: Analysis of changes in CYGB protein expression in response to knockdown or overexpression of LEDG/p75. Expression of CYGB protein was evaluated by immunoblotting in PC-3 cells with transient knockdown ( A ) and stable overexpression of LEDGF/p75 ( B ), to further validate the qPCR data. Protein loading was assessed with antibody to β-actin.

    Article Snippet: The following antibodies were used: Mouse monoclonals anti-β-actin (Sigma-Aldrich), and anti-CYGB (1:200, Abnova); rabbit polyclonals anti-PRDX1 (1:1,000, Abcam), anti-PRDX2 (1:10,000, Abcam), anti-PRDX3 (1:5,000, Abcam), anti-PRDX4 (1:1,000, Abcam), anti-PRDX5 (1:1,000, Santa Cruz Biotechnology), anti-PRDX6 (1:3,000, Abcam), anti-LEDGF/p75 (1:1,000, Novus Biologicals); and horseradish peroxidase (HRP)-labeled secondary IgG antibodies (Zymed).

    Techniques: Expressing, Over Expression, Real-time Polymerase Chain Reaction

    Quantification of CYGB expression in oesophageal cells subjected to transient transfection or siRNA knockdown. Levels of CYGB expression, compared to ACTB expression, were determined by real time RT-PCR in a parallel sample for each comet assay performed. These expression levels therefore relate directly to the comet assay data presented in Figure 3 . The geometric mean of all experiments is shown for both TE-8 cells (orange bars) and NE1 cells (pink bars).

    Journal: PLoS ONE

    Article Title: Protection from Intracellular Oxidative Stress by Cytoglobin in Normal and Cancerous Oesophageal Cells

    doi: 10.1371/journal.pone.0030587

    Figure Lengend Snippet: Quantification of CYGB expression in oesophageal cells subjected to transient transfection or siRNA knockdown. Levels of CYGB expression, compared to ACTB expression, were determined by real time RT-PCR in a parallel sample for each comet assay performed. These expression levels therefore relate directly to the comet assay data presented in Figure 3 . The geometric mean of all experiments is shown for both TE-8 cells (orange bars) and NE1 cells (pink bars).

    Article Snippet: Cells were transfected with 25 nM final concentration of either a negative control scrambled siRNA (Ambion; AM4611) or CYGB siRNA (Qiagen; SI03228323 or Ambion; s41571) using 15 µg ml−1 TransIT siQuest reagent (Mirus Bio) according to the manufacturer's instructions.

    Techniques: Expressing, Transfection, Quantitative RT-PCR, Single Cell Gel Electrophoresis

    Impact of BSO treatment and CYGB expression on oxidative stress response. Oxidative stress was measured in a: TE-8 and b: NE1 cells. Median DCF fluorescence is the mean of 5 repeats. All values were normalised to the level seen in the control cells with no BSO. Error bars represent SEM.+2020: transfection reagent only; +CYGB: cells transfected with full length CYGB; −ve siRNA: scrambled control; CYGB k/d: CYGB siRNA. * Statistically significant concentration-dependent response (p

    Journal: PLoS ONE

    Article Title: Protection from Intracellular Oxidative Stress by Cytoglobin in Normal and Cancerous Oesophageal Cells

    doi: 10.1371/journal.pone.0030587

    Figure Lengend Snippet: Impact of BSO treatment and CYGB expression on oxidative stress response. Oxidative stress was measured in a: TE-8 and b: NE1 cells. Median DCF fluorescence is the mean of 5 repeats. All values were normalised to the level seen in the control cells with no BSO. Error bars represent SEM.+2020: transfection reagent only; +CYGB: cells transfected with full length CYGB; −ve siRNA: scrambled control; CYGB k/d: CYGB siRNA. * Statistically significant concentration-dependent response (p

    Article Snippet: Cells were transfected with 25 nM final concentration of either a negative control scrambled siRNA (Ambion; AM4611) or CYGB siRNA (Qiagen; SI03228323 or Ambion; s41571) using 15 µg ml−1 TransIT siQuest reagent (Mirus Bio) according to the manufacturer's instructions.

    Techniques: Expressing, Fluorescence, Transfection, Concentration Assay

    Effect of Cygb deficiency in the expression of bile transporters and CD10 in acute BDL mice. Hepatic mRNA level of ( A ) sinusoidal (Mrp2, Mdr2, Bsep) and ( B ) canalicular (Mrp3, Ntcp, Oatp1) transporters of bile components in sham (S) and acute BDL (24–48 h) mice. ( C ) Immunofluorescence (top panels) and immunoblot (bottom panels) of CD10 in sham and BDL-24 h mice. GAPDH was used as loading control. All gels were run under the same experimental conditions. The cropped gels are used and full-length gels are presented in Supplementary Fig. S9 . ( D ) Quantitative densitometry of CD10 (top panel) and hepatic mRNA level (bottom panel) of CD10 in sham (S) and acute BDL (24–72 h) and chronic BDL (1–3 W) mice. Open bars, WT; close bars, Cygb −/− . Data represent the mean ± SD. Sham (n = 3), BDL (n = 4–8). *p

    Journal: Scientific Reports

    Article Title: Possible Involvement of Nitric Oxide in Enhanced Liver Injury and Fibrogenesis during Cholestasis in Cytoglobin-deficient Mice

    doi: 10.1038/srep41888

    Figure Lengend Snippet: Effect of Cygb deficiency in the expression of bile transporters and CD10 in acute BDL mice. Hepatic mRNA level of ( A ) sinusoidal (Mrp2, Mdr2, Bsep) and ( B ) canalicular (Mrp3, Ntcp, Oatp1) transporters of bile components in sham (S) and acute BDL (24–48 h) mice. ( C ) Immunofluorescence (top panels) and immunoblot (bottom panels) of CD10 in sham and BDL-24 h mice. GAPDH was used as loading control. All gels were run under the same experimental conditions. The cropped gels are used and full-length gels are presented in Supplementary Fig. S9 . ( D ) Quantitative densitometry of CD10 (top panel) and hepatic mRNA level (bottom panel) of CD10 in sham (S) and acute BDL (24–72 h) and chronic BDL (1–3 W) mice. Open bars, WT; close bars, Cygb −/− . Data represent the mean ± SD. Sham (n = 3), BDL (n = 4–8). *p

    Article Snippet: Treatment of nitric oxide inhibitor and donor To investigate the role of NO in BDL model, WT or Cygb−/− mice were administered L-NAME (Dojindo, MD, USA) in drinking water at the dose of 0.5 mg/ml for 9 days (n = 5 per each group).

    Techniques: Expressing, Mouse Assay, Immunofluorescence

    Effect of Cygb deficiency on inflammation and cell death in acute BDL. ( A ) Hepatic mRNA level of chemokine Cxcl-1, Cxcl-2, Cxcl-5, and Ccl-2 in sham (S) and acute BDL (24–48 h). ( B ) Immunohistochemistry of neutrophil- (top panels) and CD68- positive cells (bottom panels) in sham and BDL-24 h mice. ( C ) Quantification of neutrophil- (top panel) and CD68-positive cells (bottom panel) per field. ( D ) TUNEL staining in sham and acute BDL-24 (top panels) and number of TUNEL positive cells (bottom panel) per field. ( E ) Immunoblots of phosphorylated (p) and total NF-κB p65, active and pro caspase 3 (CASP 3), and cytochrome c (CYT C) in sham and BDL-24 h mice. GAPDH was used as loading control. All gels were run under the same experimental conditions. The cropped gels are used and full-length gels are presented in Supplementary Fig. S8 . ( E ) Quantitative densitometry of p-NF-κB and active CASP 3 in sham and BDL-24 h. Open bars, WT; close bars, Cygb −/− . Data represent the mean ± SD. Sham (n = 3), BDL (n = 4–8). *p

    Journal: Scientific Reports

    Article Title: Possible Involvement of Nitric Oxide in Enhanced Liver Injury and Fibrogenesis during Cholestasis in Cytoglobin-deficient Mice

    doi: 10.1038/srep41888

    Figure Lengend Snippet: Effect of Cygb deficiency on inflammation and cell death in acute BDL. ( A ) Hepatic mRNA level of chemokine Cxcl-1, Cxcl-2, Cxcl-5, and Ccl-2 in sham (S) and acute BDL (24–48 h). ( B ) Immunohistochemistry of neutrophil- (top panels) and CD68- positive cells (bottom panels) in sham and BDL-24 h mice. ( C ) Quantification of neutrophil- (top panel) and CD68-positive cells (bottom panel) per field. ( D ) TUNEL staining in sham and acute BDL-24 (top panels) and number of TUNEL positive cells (bottom panel) per field. ( E ) Immunoblots of phosphorylated (p) and total NF-κB p65, active and pro caspase 3 (CASP 3), and cytochrome c (CYT C) in sham and BDL-24 h mice. GAPDH was used as loading control. All gels were run under the same experimental conditions. The cropped gels are used and full-length gels are presented in Supplementary Fig. S8 . ( E ) Quantitative densitometry of p-NF-κB and active CASP 3 in sham and BDL-24 h. Open bars, WT; close bars, Cygb −/− . Data represent the mean ± SD. Sham (n = 3), BDL (n = 4–8). *p

    Article Snippet: Treatment of nitric oxide inhibitor and donor To investigate the role of NO in BDL model, WT or Cygb−/− mice were administered L-NAME (Dojindo, MD, USA) in drinking water at the dose of 0.5 mg/ml for 9 days (n = 5 per each group).

    Techniques: Immunohistochemistry, Mouse Assay, TUNEL Assay, Staining, Western Blot

    Effect of NO inhibitor in Cygb −/− mice after BDL. WT and KO mice were subjected to BDL-48 h together with L-NG-nitroarginine methyl ester (L-NAME) treatment. Control mice received drinking water (DW). ( A ) Concentration of nitrite + nitrate and cGMP in serum. ( B ) Representative macroscopic images and microscopic liver sections stained with H E. ( C ) Serum levels of AST, ALT, total bilirubin, and total bile acid (TBA). ( D ) Immunofluorescence, immunoblots and hepatic mRNA level of CD10. GAPDH was used as loading control. All gels were run under the same experimental conditions. The cropped gels are used and full-length gels are presented in Supplementary Fig. S11 . ( E ) Hepatic mRNA level of Bsep, Mdr2, Oatp1, Ntcp. ( F ) Immunohistochemistry of neutrophils and CD68 and its quantitative analyses (right insets). Open bars, WT; close bars, KO. Data represent the mean ± SD. n = 5. *p

    Journal: Scientific Reports

    Article Title: Possible Involvement of Nitric Oxide in Enhanced Liver Injury and Fibrogenesis during Cholestasis in Cytoglobin-deficient Mice

    doi: 10.1038/srep41888

    Figure Lengend Snippet: Effect of NO inhibitor in Cygb −/− mice after BDL. WT and KO mice were subjected to BDL-48 h together with L-NG-nitroarginine methyl ester (L-NAME) treatment. Control mice received drinking water (DW). ( A ) Concentration of nitrite + nitrate and cGMP in serum. ( B ) Representative macroscopic images and microscopic liver sections stained with H E. ( C ) Serum levels of AST, ALT, total bilirubin, and total bile acid (TBA). ( D ) Immunofluorescence, immunoblots and hepatic mRNA level of CD10. GAPDH was used as loading control. All gels were run under the same experimental conditions. The cropped gels are used and full-length gels are presented in Supplementary Fig. S11 . ( E ) Hepatic mRNA level of Bsep, Mdr2, Oatp1, Ntcp. ( F ) Immunohistochemistry of neutrophils and CD68 and its quantitative analyses (right insets). Open bars, WT; close bars, KO. Data represent the mean ± SD. n = 5. *p

    Article Snippet: Treatment of nitric oxide inhibitor and donor To investigate the role of NO in BDL model, WT or Cygb−/− mice were administered L-NAME (Dojindo, MD, USA) in drinking water at the dose of 0.5 mg/ml for 9 days (n = 5 per each group).

    Techniques: Mouse Assay, Concentration Assay, Staining, AST Assay, Immunofluorescence, Western Blot, Immunohistochemistry

    NO metabolites and oxidative stress condition in Cygb −/− mice under acute BDL. Concentration of nitrite + nitrate ( A ) and cGMP ( B ) in serum, liver lysate and urine in sham (S) and acute BDL-48 h mice. ( C ) Hepatic immunofluorescence, immunoblot and mRNA level of iNOS in sham and acute BDL (24–48 h) mice. ( D ) Malondialdehyde (MDA) content of sham (S) and acute BDL (24–48 h) mice in serum and liver. ( E ) Hepatic immunofluorescence, immunoblot and mRNA of HO-1 in sham or acute BDL mice (24–48 h). GAPDH was used as loading control. All gels were run under the same experimental conditions. The cropped gels are used and full-length gels are presented in Supplementary Fig. S10 . Open bars, WT; close bars, Cygb −/− . Data represent the mean ± SD. Sham (n = 3), BDL (n = 4–8). *p

    Journal: Scientific Reports

    Article Title: Possible Involvement of Nitric Oxide in Enhanced Liver Injury and Fibrogenesis during Cholestasis in Cytoglobin-deficient Mice

    doi: 10.1038/srep41888

    Figure Lengend Snippet: NO metabolites and oxidative stress condition in Cygb −/− mice under acute BDL. Concentration of nitrite + nitrate ( A ) and cGMP ( B ) in serum, liver lysate and urine in sham (S) and acute BDL-48 h mice. ( C ) Hepatic immunofluorescence, immunoblot and mRNA level of iNOS in sham and acute BDL (24–48 h) mice. ( D ) Malondialdehyde (MDA) content of sham (S) and acute BDL (24–48 h) mice in serum and liver. ( E ) Hepatic immunofluorescence, immunoblot and mRNA of HO-1 in sham or acute BDL mice (24–48 h). GAPDH was used as loading control. All gels were run under the same experimental conditions. The cropped gels are used and full-length gels are presented in Supplementary Fig. S10 . Open bars, WT; close bars, Cygb −/− . Data represent the mean ± SD. Sham (n = 3), BDL (n = 4–8). *p

    Article Snippet: Treatment of nitric oxide inhibitor and donor To investigate the role of NO in BDL model, WT or Cygb−/− mice were administered L-NAME (Dojindo, MD, USA) in drinking water at the dose of 0.5 mg/ml for 9 days (n = 5 per each group).

    Techniques: Mouse Assay, Concentration Assay, Immunofluorescence, Multiple Displacement Amplification

    Effect of NO donor on BDL-induced liver injury in WT and Cygb −/− mice after BDL. WT and KO were subjected to BDL-48 h together with saline or sodium nitroprusside (SNP) treatment. ( A ) Concentration of nitrite + nitrate and cGMP in serum. ( B ) Representative macroscopic images, microscopic liver sections stained with H E. ( C ) Serum AST, ALT, total bilirubin, and total bile acid (TBA). ( D ) Immunofluorescent staining, immunoblots and hepatic mRNA level of CD10. GAPDH was used as loading control. All gels were run under the same experimental conditions. The cropped gels are used and full-length gels are presented in Supplementary Fig. S12 . ( E ) Hepatic mRNA level of Bsep, Mdr2, Oatp1, Ntcp. ( F ) Immunohistochemistry of neutrophils and CD68 and its quantitative analyses (right insets). Open bars, WT; close bars, KO. Data represent the mean ± SD. n = 5. *p

    Journal: Scientific Reports

    Article Title: Possible Involvement of Nitric Oxide in Enhanced Liver Injury and Fibrogenesis during Cholestasis in Cytoglobin-deficient Mice

    doi: 10.1038/srep41888

    Figure Lengend Snippet: Effect of NO donor on BDL-induced liver injury in WT and Cygb −/− mice after BDL. WT and KO were subjected to BDL-48 h together with saline or sodium nitroprusside (SNP) treatment. ( A ) Concentration of nitrite + nitrate and cGMP in serum. ( B ) Representative macroscopic images, microscopic liver sections stained with H E. ( C ) Serum AST, ALT, total bilirubin, and total bile acid (TBA). ( D ) Immunofluorescent staining, immunoblots and hepatic mRNA level of CD10. GAPDH was used as loading control. All gels were run under the same experimental conditions. The cropped gels are used and full-length gels are presented in Supplementary Fig. S12 . ( E ) Hepatic mRNA level of Bsep, Mdr2, Oatp1, Ntcp. ( F ) Immunohistochemistry of neutrophils and CD68 and its quantitative analyses (right insets). Open bars, WT; close bars, KO. Data represent the mean ± SD. n = 5. *p

    Article Snippet: Treatment of nitric oxide inhibitor and donor To investigate the role of NO in BDL model, WT or Cygb−/− mice were administered L-NAME (Dojindo, MD, USA) in drinking water at the dose of 0.5 mg/ml for 9 days (n = 5 per each group).

    Techniques: Mouse Assay, Concentration Assay, Staining, AST Assay, Western Blot, Immunohistochemistry

    Severe liver injury in Cygb −/− mice under BDL. ( A ) Kaplan-Meier curve, n = 19 per group. Representative macroscopic images ( B ) and microscopic liver sections stained with H E ( C ) in sham, acute BDL (24–72 h) and chronic BDL (1–3 W). Original magnification, x40. Yellow and black arrows indicate bile infarcts. ( D ) Quantification of area of bile infarcts. ( E ) Levels of serum AST, ALT, and total bilirubin, and hepatic total bile acid (TBA). Data represent the mean ± SD. Sham (n = 3), BDL (n = 4–8). *p

    Journal: Scientific Reports

    Article Title: Possible Involvement of Nitric Oxide in Enhanced Liver Injury and Fibrogenesis during Cholestasis in Cytoglobin-deficient Mice

    doi: 10.1038/srep41888

    Figure Lengend Snippet: Severe liver injury in Cygb −/− mice under BDL. ( A ) Kaplan-Meier curve, n = 19 per group. Representative macroscopic images ( B ) and microscopic liver sections stained with H E ( C ) in sham, acute BDL (24–72 h) and chronic BDL (1–3 W). Original magnification, x40. Yellow and black arrows indicate bile infarcts. ( D ) Quantification of area of bile infarcts. ( E ) Levels of serum AST, ALT, and total bilirubin, and hepatic total bile acid (TBA). Data represent the mean ± SD. Sham (n = 3), BDL (n = 4–8). *p

    Article Snippet: Treatment of nitric oxide inhibitor and donor To investigate the role of NO in BDL model, WT or Cygb−/− mice were administered L-NAME (Dojindo, MD, USA) in drinking water at the dose of 0.5 mg/ml for 9 days (n = 5 per each group).

    Techniques: Mouse Assay, Staining, AST Assay

    Promotion of fibrosis in Cygb −/− mice after chronic BDL. ( A ) Liver sections from Sirius Red and Fast Green (SiR-FG) staining in BDL 2 W. ( B ) Sirius Red positive area (left panel) and hydroxyproline (HP) content of liver (right panel) in sham (S) and chronic BDL (1–3 W) mice. ( C ) Immunohistochemistry for α-SMA (top panels), immunoblot analysis (bottom panels), and its quantitative densitometry (right inset) of the α-SMA expression. GAPDH was used as loading control. All gels were run under the same experimental conditions. The cropped gels are used and full-length gels are presented in Supplementary Fig. S13 . ( D ) Hepatic mRNA level of α-Sma expression. ( E ) Hepatic mRNA level of collagen1a1 and Timp-1. Open bars, WT; close bars, KO. Data represent the mean ± SD. Sham (n = 3), BDL (n = 4–8). *p

    Journal: Scientific Reports

    Article Title: Possible Involvement of Nitric Oxide in Enhanced Liver Injury and Fibrogenesis during Cholestasis in Cytoglobin-deficient Mice

    doi: 10.1038/srep41888

    Figure Lengend Snippet: Promotion of fibrosis in Cygb −/− mice after chronic BDL. ( A ) Liver sections from Sirius Red and Fast Green (SiR-FG) staining in BDL 2 W. ( B ) Sirius Red positive area (left panel) and hydroxyproline (HP) content of liver (right panel) in sham (S) and chronic BDL (1–3 W) mice. ( C ) Immunohistochemistry for α-SMA (top panels), immunoblot analysis (bottom panels), and its quantitative densitometry (right inset) of the α-SMA expression. GAPDH was used as loading control. All gels were run under the same experimental conditions. The cropped gels are used and full-length gels are presented in Supplementary Fig. S13 . ( D ) Hepatic mRNA level of α-Sma expression. ( E ) Hepatic mRNA level of collagen1a1 and Timp-1. Open bars, WT; close bars, KO. Data represent the mean ± SD. Sham (n = 3), BDL (n = 4–8). *p

    Article Snippet: Treatment of nitric oxide inhibitor and donor To investigate the role of NO in BDL model, WT or Cygb−/− mice were administered L-NAME (Dojindo, MD, USA) in drinking water at the dose of 0.5 mg/ml for 9 days (n = 5 per each group).

    Techniques: Mouse Assay, Staining, Immunohistochemistry, Expressing