mouse monoclonal anti fhl2  (Hycult Biotech)

Journal: Cells

Article Title: Four-And-A-Half LIM-Domain Protein 2 (FHL2) Deficiency Aggravates Cholestatic Liver Injury

doi: 10.3390/cells9010248

Figure Lengend Snippet: Primer sequences for quantitative real-time PCR.

Article Snippet: Protein extraction from liver tissues, protein extraction from cells, and analysis by Western blotting were performed as described in [ ] by applying mouse monoclonal anti-FHL2 (HycultBiotech, Uden, The Netherlands; HM2136, 1:300), rabbit monoclonal anti-alpha-smooth muscle actin (α-SMA) (Abcam, Cambridge, United Kingdom; ab32575, 1:1000), rabbit monoclonal anti-B-cell lymphoma 2 (BCL2) (Epitomics, Burlingame, CA, USA; #1017, 1:1000), rabbit polyclonal anti-metalloproteinase 13 (anti-MMP13) (Abcam, Cambridge, United Kingdom; ab39012, 1:1000), rabbit polyclonal anti-MMP14 (Chemicon, Burlington, MA, USA; AB815, 1:1000) and mouse monoclonal anti-actin (ACTB; Merck Millipore, Billerica, MA, USA; MAB1501, 1:10,000) antibodies.

Techniques:

Journal: Cells

Article Title: Four-And-A-Half LIM-Domain Protein 2 (FHL2) Deficiency Aggravates Cholestatic Liver Injury

doi: 10.3390/cells9010248

Figure Lengend Snippet: Fhl2 expression and effect of Fhl2 deficiency on hepatocellular injury and inflammation in the mouse model of bile duct ligation (BDL). Fhl2 -deficient ( Fhl2 -ko) and wild type (wt) mice were either bile duct ligated (BDL) or sham-operated (CTR). ( A ) Fhl2 mRNA levels in wt BDL and CTR mice analyzed by qRT-PCR. ( B ) Representative hematoxylin and eosin stainings of liver tissue samples (20× magnification). ( C ) ALT (alanine aminotransferase) and ( D ) bilirubin serum levels. ( E ) Hmox-1 and ( F ) Mcp-1 mRNA expression levels in liver tissue analyzed by qRT-PCR. ( G ) Immunohistochemical CD3 staining of liver tissue samples (20× magnification). ( H ) Il-1 and ( I ) Tnf mRNA expression levels in liver tissue analyzed by qRT-PCR. (*: p < 0.05).

Article Snippet: Protein extraction from liver tissues, protein extraction from cells, and analysis by Western blotting were performed as described in [ ] by applying mouse monoclonal anti-FHL2 (HycultBiotech, Uden, The Netherlands; HM2136, 1:300), rabbit monoclonal anti-alpha-smooth muscle actin (α-SMA) (Abcam, Cambridge, United Kingdom; ab32575, 1:1000), rabbit monoclonal anti-B-cell lymphoma 2 (BCL2) (Epitomics, Burlingame, CA, USA; #1017, 1:1000), rabbit polyclonal anti-metalloproteinase 13 (anti-MMP13) (Abcam, Cambridge, United Kingdom; ab39012, 1:1000), rabbit polyclonal anti-MMP14 (Chemicon, Burlington, MA, USA; AB815, 1:1000) and mouse monoclonal anti-actin (ACTB; Merck Millipore, Billerica, MA, USA; MAB1501, 1:10,000) antibodies.

Techniques: Expressing, Ligation, Quantitative RT-PCR, Immunohistochemical staining, Staining

Journal: Cells

Article Title: Four-And-A-Half LIM-Domain Protein 2 (FHL2) Deficiency Aggravates Cholestatic Liver Injury

doi: 10.3390/cells9010248

Figure Lengend Snippet: Effect of Fhl2 deficiency on hepatic fibrosis in the mouse model of bile duct ligation (BDL). Fhl2 -deficient ( Fhl2 -ko) and wild type (wt) mice were either subjected to BDL or were sham-operated (CTR). ( A ) Hepatic α-sma mRNA expression levels. ( B ) Immunohistochemical α-sma staining of liver tissue sections (20× magnification). Hepatic ( C ) Tgf-β and ( D ) Col1a1 mRNA expression levels. (E) Sirius Red/Fast Green staining of liver tissue sections (20× magnification). Hepatic ( F ) Mmp1 , ( G ) Mmp2 , ( H ) Mmp9 , and ( I ) Pai-1 mRNA expression levels in liver tissue analyzed by qRT-PCR. (*: p < 0.05).

Article Snippet: Protein extraction from liver tissues, protein extraction from cells, and analysis by Western blotting were performed as described in [ ] by applying mouse monoclonal anti-FHL2 (HycultBiotech, Uden, The Netherlands; HM2136, 1:300), rabbit monoclonal anti-alpha-smooth muscle actin (α-SMA) (Abcam, Cambridge, United Kingdom; ab32575, 1:1000), rabbit monoclonal anti-B-cell lymphoma 2 (BCL2) (Epitomics, Burlingame, CA, USA; #1017, 1:1000), rabbit polyclonal anti-metalloproteinase 13 (anti-MMP13) (Abcam, Cambridge, United Kingdom; ab39012, 1:1000), rabbit polyclonal anti-MMP14 (Chemicon, Burlington, MA, USA; AB815, 1:1000) and mouse monoclonal anti-actin (ACTB; Merck Millipore, Billerica, MA, USA; MAB1501, 1:10,000) antibodies.

Techniques: Ligation, Expressing, Immunohistochemical staining, Staining, Quantitative RT-PCR

Journal: Cells

Article Title: Four-And-A-Half LIM-Domain Protein 2 (FHL2) Deficiency Aggravates Cholestatic Liver Injury

doi: 10.3390/cells9010248

Figure Lengend Snippet: Effect of FHL2 depletion on bile acid-induced hepatocellular injury in vitro. ( A , B ) FHL2 mRNA and protein expression in HepG2 cells transfected with si-pools against FHL2 (siFHL2) and si-control-pools (siCtr). ( C ) Representative microscopical images 72 h after transfection (10X magnification). ( D ) Quantification of lactate dehydrogenase (LDH) release into the supernatant. ( E ) FHL2 , ( F ) p47phox , and ( H ) BAX mRNA, and ( G ) B-cell lymphoma 2 (BCL2) protein expression after treatment with deoxycholic acid (DCA) for 24 h and control cells (CTR). (*: p < 0.05).

Article Snippet: Protein extraction from liver tissues, protein extraction from cells, and analysis by Western blotting were performed as described in [ ] by applying mouse monoclonal anti-FHL2 (HycultBiotech, Uden, The Netherlands; HM2136, 1:300), rabbit monoclonal anti-alpha-smooth muscle actin (α-SMA) (Abcam, Cambridge, United Kingdom; ab32575, 1:1000), rabbit monoclonal anti-B-cell lymphoma 2 (BCL2) (Epitomics, Burlingame, CA, USA; #1017, 1:1000), rabbit polyclonal anti-metalloproteinase 13 (anti-MMP13) (Abcam, Cambridge, United Kingdom; ab39012, 1:1000), rabbit polyclonal anti-MMP14 (Chemicon, Burlington, MA, USA; AB815, 1:1000) and mouse monoclonal anti-actin (ACTB; Merck Millipore, Billerica, MA, USA; MAB1501, 1:10,000) antibodies.

Techniques: In Vitro, Expressing, Transfection

Journal: Cells

Article Title: Four-And-A-Half LIM-Domain Protein 2 (FHL2) Deficiency Aggravates Cholestatic Liver Injury

doi: 10.3390/cells9010248

Figure Lengend Snippet: Effect of FHL2 depletion on bile acid metabolism. HepG2 cells were transfected with si-pools against FHL2 (siFHL2) and si-control-pools (siCtr). Analysis of ( A ) CYP7A1 , ( B ) NTCP , ( C ) BSEP , and ( D ) MRP2 mRNA expression after treatment with deoxycholic acid (DCA) for 24 h and control cells (CTR). (*: p < 0.05).

Article Snippet: Protein extraction from liver tissues, protein extraction from cells, and analysis by Western blotting were performed as described in [ ] by applying mouse monoclonal anti-FHL2 (HycultBiotech, Uden, The Netherlands; HM2136, 1:300), rabbit monoclonal anti-alpha-smooth muscle actin (α-SMA) (Abcam, Cambridge, United Kingdom; ab32575, 1:1000), rabbit monoclonal anti-B-cell lymphoma 2 (BCL2) (Epitomics, Burlingame, CA, USA; #1017, 1:1000), rabbit polyclonal anti-metalloproteinase 13 (anti-MMP13) (Abcam, Cambridge, United Kingdom; ab39012, 1:1000), rabbit polyclonal anti-MMP14 (Chemicon, Burlington, MA, USA; AB815, 1:1000) and mouse monoclonal anti-actin (ACTB; Merck Millipore, Billerica, MA, USA; MAB1501, 1:10,000) antibodies.

Techniques: Transfection, Expressing

Journal: Cells

Article Title: Four-And-A-Half LIM-Domain Protein 2 (FHL2) Deficiency Aggravates Cholestatic Liver Injury

doi: 10.3390/cells9010248

Figure Lengend Snippet: Effect of FHL2 depletion on pro-fibrogenic gene expression in hepatic stellate cells in vitro. ( A ) FHL2 mRNA expression during activation of primary human hepatic stellate cells (HSCs). ( B , C ) FHL2 mRNA and protein expression in LX-2 cells transfected with si-pools against FHL2 (siFHL2) and si-control-pools (siCtr). ( D ) Representative microscopical images 72 h after transfection (10× magnification). ( E , F ) α-SMA mRNA and protein expression. ( G ) COL1A1 , ( H ) TGF-β , ( I ) MMP1 , ( J ) MMP3 , ( K ) MMP2 , ( L ) MMP9 and ( M ) MMP10 mRNA expression and ( N , O ) MMP13 and ( P , Q ) MMP14 mRNA and protein expression 72 h after transfection. (*: p < 0.05).

Article Snippet: Protein extraction from liver tissues, protein extraction from cells, and analysis by Western blotting were performed as described in [ ] by applying mouse monoclonal anti-FHL2 (HycultBiotech, Uden, The Netherlands; HM2136, 1:300), rabbit monoclonal anti-alpha-smooth muscle actin (α-SMA) (Abcam, Cambridge, United Kingdom; ab32575, 1:1000), rabbit monoclonal anti-B-cell lymphoma 2 (BCL2) (Epitomics, Burlingame, CA, USA; #1017, 1:1000), rabbit polyclonal anti-metalloproteinase 13 (anti-MMP13) (Abcam, Cambridge, United Kingdom; ab39012, 1:1000), rabbit polyclonal anti-MMP14 (Chemicon, Burlington, MA, USA; AB815, 1:1000) and mouse monoclonal anti-actin (ACTB; Merck Millipore, Billerica, MA, USA; MAB1501, 1:10,000) antibodies.

Techniques: Expressing, In Vitro, Activation Assay, Transfection

Journal: PLoS ONE

Article Title: Transcriptional Regulation of the Tumor Suppressor FHL2 by p53 in Human Kidney and Liver Cells

doi: 10.1371/journal.pone.0099359

Figure Lengend Snippet: pcDNA3.1 served as empty vector control of TP53 and the untreated HEK 293 cells lysate showed the endogenous levels of p53 and FHL2. β-actin was used as internal control.

Article Snippet: The following primary antibodies were used in the Western blot analysis: mouse monoclonal anti-FHL2 (1∶1000), mouse monoclonal anti-TP53 (1∶1000) and mouse monoclonal anti-β-actin (1∶5000; all from Santa Cruz Biotechnology).

Techniques: Plasmid Preparation

Journal: PLoS ONE

Article Title: Transcriptional Regulation of the Tumor Suppressor FHL2 by p53 in Human Kidney and Liver Cells

doi: 10.1371/journal.pone.0099359

Figure Lengend Snippet: TP53 expression plasmid was transiently transfected in HEK293 and Hep3B cells using empty pcDNA3.1 vector as control. Western blotting analysis indicated the successful over-expression of TP53 in both HEK293 and Hep3B cells. The mRNA levels of FHL2-1a and FHL2-1b were determined by real-time PCR. (A) The mRNA levels of FHL2-1a and FHL2-1b in HEK293 cells transfected with pcDNA3.1-TP53 and empty pcDNA3.1 vector. (B) The mRNA levels of FHL2-1a and FHL2-1b in Hep3B cells transfected with pcDNA3.1-TP53 and empty pcDNA3.1 vector.

Article Snippet: The following primary antibodies were used in the Western blot analysis: mouse monoclonal anti-FHL2 (1∶1000), mouse monoclonal anti-TP53 (1∶1000) and mouse monoclonal anti-β-actin (1∶5000; all from Santa Cruz Biotechnology).

Techniques: Expressing, Plasmid Preparation, Transfection, Western Blot, Over Expression, Real-time Polymerase Chain Reaction

Journal: PLoS ONE

Article Title: Transcriptional Regulation of the Tumor Suppressor FHL2 by p53 in Human Kidney and Liver Cells

doi: 10.1371/journal.pone.0099359

Figure Lengend Snippet: ( A ) Relative expression levels of FHL2-1a in Hep3B cells transfected with pcDNA3.1-p53 using pcDNA3.1 empty vector as control. ( B ) Protein expression of p53 and FHL2 were detected by western blotting after transfection with pcDNA3.1 and pcDNA3.1-p53. ( C ) ChIP analysis of p53-enriched DNA fragment for Hep3B cells overexpressed with p53. DNA fragment enriched in the ChIP assay were tested by qPCR. The p53 antibody used for enrichment was indicated in the x axis. Y axis represents fold enrichment of immunoprecipitated DNA (gray bars) against the control (empty bars).

Article Snippet: The following primary antibodies were used in the Western blot analysis: mouse monoclonal anti-FHL2 (1∶1000), mouse monoclonal anti-TP53 (1∶1000) and mouse monoclonal anti-β-actin (1∶5000; all from Santa Cruz Biotechnology).

Techniques: Expressing, Transfection, Plasmid Preparation, Western Blot, Immunoprecipitation

Journal: PLoS ONE

Article Title: Transcriptional Regulation of the Tumor Suppressor FHL2 by p53 in Human Kidney and Liver Cells

doi: 10.1371/journal.pone.0099359

Figure Lengend Snippet: ( A ) Relative expression levels of p53 and FHL2-1a in HepG2 cells treated with p53-siRNA with scRNA as control. ( B ) Protein expression of p53 and FHL2 were detected by western blotting after siRNA treatment. ( C ) ChIP analysis of p53-enriched DNA fragment for HepG2 cells. DNA fragment enriched in the ChIP assay were tested by qPCR. The p53 antibody used for enrichment was indicated in the x axis. Y axis represents fold enrichment of immunoprecipitated DNA (gray bars) against the control (empty bars).

Article Snippet: The following primary antibodies were used in the Western blot analysis: mouse monoclonal anti-FHL2 (1∶1000), mouse monoclonal anti-TP53 (1∶1000) and mouse monoclonal anti-β-actin (1∶5000; all from Santa Cruz Biotechnology).

Techniques: Expressing, Western Blot, Immunoprecipitation

Journal: PLoS ONE

Article Title: Transcriptional Regulation of the Tumor Suppressor FHL2 by p53 in Human Kidney and Liver Cells

doi: 10.1371/journal.pone.0099359

Figure Lengend Snippet: Correlation between the T/N expression level of FHL2 and coding region mutations of TP53 in HCC samples.

Article Snippet: The following primary antibodies were used in the Western blot analysis: mouse monoclonal anti-FHL2 (1∶1000), mouse monoclonal anti-TP53 (1∶1000) and mouse monoclonal anti-β-actin (1∶5000; all from Santa Cruz Biotechnology).

Techniques: Expressing