fluorescence protein ad egfp  (Obio Technology Corp Ltd)

Journal: American Journal of Physiology - Cell Physiology

Article Title: Nuclear receptor subfamily 4 group A member 2 inhibits activation of ERK signaling and cell growth in response to β-adrenergic stimulation in adult rat cardiomyocytes

doi: 10.1152/ajpcell.00526.2018

Figure Lengend Snippet: RNA-Seq analysis identifies nuclear receptor subfamily 4 group A (NR4A2) as a transcriptional modulator of growth regulatory pathways in adult cardiomyocytes. A : comparison of NR4A2 total mRNA levels in adenovirus encoding green fluorescent protein (Ad-GFP) control and Ad-h-NR4A2 transduced cells. B : comparison of NR4A2 total protein levels in Ad-GFP control and Ad-h-NR4A2-transduced cells. Representative immunoblot is presented. C : immunolocalization of NR4A2 in nontransduced control and Ad-h-NR4A2-transduced cells. Troponin C was used as marker for cardiac myocytes. Pairwise comparison was performed with unpaired Student’s t -test. Data are means ± SE of 4–5 independent experiments using adult rat ventricular myocytes (ARVMs) isolated from different rats. * P < 0.05. D : heat map representation of 3,021 genes that passed filtering criteria. The 2 clusters of genes differentially regulated (down- and upregulated, respectively) in NR4A2-overexpressing cells compared with the GFP-expressing control cells are indicated. E : selective representation of cell growth and hypertrophy signaling networks containing effectors regulated at gene level in response to NR4A2 overexpression. Upward green and downward red arrows represent increased and decreased expression levels, respectively. Circular arrows indicate changes in isoform expression. Data are based on analysis of 6 independent experiments using ARVMs isolated from different rats. LVDCC, L-type voltage-dependent Ca 2+ channel; β-arr, β-arrestin; GRK, G protein-coupled receptor kinase; AKAP, A-kinase anchoring protein; RCAN1, regulator of calcineurin 1; mTORC1, mammalian target of rapamycin complex 1; PDE, phosphodiesterase; CREB, cAMP response element-binding protein; GSK-3β, glycogen synthase kinase-3β; DUSP, dual-specificity phosphatase; PI3K, phosphoinositide 3-kinase; 4E-BP1, eukaryotic initiation factor 4E-binding protein-1.

Article Snippet: The human NR4A2 adenovirus (Ad-h-NR4A2; cat. no. ADV-217057) and enhanced green fluorescent protein (eGFP) adenovirus (Ad-GFP; cat. no. 1060) were generated, amplified, purified, and titrated by Vector Biolabs.

Techniques: RNA Sequencing, Comparison, Control, Western Blot, Marker, Isolation, Expressing, Over Expression, Binding Assay

Journal: The Journal of Physiology

Article Title: Differential regulation of blood flow‐induced neovascularization and mural cell recruitment by vascular endothelial growth factor and angiopoietin signalling

doi: 10.1113/JP273430

Figure Lengend Snippet: A , images of mesenteries after treatment with Ad. eGFP , and Ad. eNOS /Ad. Ang1 . B , angiogenesis index. C , confocal stack images from mesenteric panels stained with isolectin B4 and Hoechst 33324 or Ki67 and Hoechst 33324. D – I , analysis of vessel density ( D ), branch point density ( E ), proliferating endothelial cell density ( F ), exchange vessel density ( G ), conduit vessel density ( H ) and sprout point density ( I ). * P < 0.05, ** P < 0.01, *** P < 0.001 compared with GFP; # P < 0.05 compared with eNOS. Scale bar: 40 μm.

Article Snippet: Ang1 and Ad. eGFP (enhanced green fluorescent protein) were a gift from Regeneron Inc., Tarrytown, NY, USA (Benest et al . ) and Ad. eNOS (endothelial nitric oxide synthase) from Prof. Keith Channon, University of Oxford (Benest et al . ); Ad. sFlt1 (soluble Fms‐like tyrosine kinase receptor‐1/soluble VEGFR1) was generated by Dr Ewa Paleolog, Imperial College London (Afuwape et al . ) and Ad. sTie2 (soluble Tie2) (Lin et al . ) by Prof. Charles Lin, Vanderbilt University, Nashville, TN, USA.

Techniques: Staining

Journal: The Journal of Physiology

Article Title: Differential regulation of blood flow‐induced neovascularization and mural cell recruitment by vascular endothelial growth factor and angiopoietin signalling

doi: 10.1113/JP273430

Figure Lengend Snippet: A , sample confocal stack images from rat mesentery 6 days post‐treatment stained with isolectin‐B 4 and antibodies for NG2 and αSMA. B , treatment with Ad. eNOS +Ad. Ang1 significantly increased mural cell coverage. C , Ad. eNOS was unable to stimulate arteriolargenesis by itself but did so when together with Ad. Ang1 . Scale bar: 40 μm. *** P < 0.001 vs . eGFP; ### P < 0.001 vs . eNOS.

Article Snippet: Ang1 and Ad. eGFP (enhanced green fluorescent protein) were a gift from Regeneron Inc., Tarrytown, NY, USA (Benest et al . ) and Ad. eNOS (endothelial nitric oxide synthase) from Prof. Keith Channon, University of Oxford (Benest et al . ); Ad. sFlt1 (soluble Fms‐like tyrosine kinase receptor‐1/soluble VEGFR1) was generated by Dr Ewa Paleolog, Imperial College London (Afuwape et al . ) and Ad. sTie2 (soluble Tie2) (Lin et al . ) by Prof. Charles Lin, Vanderbilt University, Nashville, TN, USA.

Techniques: Staining

Journal: The Journal of Physiology

Article Title: Differential regulation of blood flow‐induced neovascularization and mural cell recruitment by vascular endothelial growth factor and angiopoietin signalling

doi: 10.1113/JP273430

Figure Lengend Snippet: A , images of mesenteries after treatment with eNOS with dual VEGF–Ang1 inhibition or eNOS and Ang1 (NO–Tie) with VEGF blockade. B , angiogenesis index. C , sample confocal stack images from rat mesentery 6 days post‐treatment stained with isolectin‐B 4 , Hoechst 33324 and antibodies for Ki67. D – F , quantification of vessel density ( D ), branch point density ( E ) and sprout point density ( F ). G , images of mesenteries after treatment with eNOS with VEGF inhibition, angiopoietin inhibition, dual VEGF–Ang1 or eNOS and Ang1 (NO–Tie) with VEGF blockade stained with isolectin‐B 4 and antibodies for NG2 and αSMA. No arteriolargenesis was seen in any of these conditions. # P < 0.05, ### P < 0.001 vs . eGFP; * P < 0.05, ** P < 0.01 vs . eNOS; + P < 0.05 vs . eNOS–sFlt1–sTie2.

Article Snippet: Ang1 and Ad. eGFP (enhanced green fluorescent protein) were a gift from Regeneron Inc., Tarrytown, NY, USA (Benest et al . ) and Ad. eNOS (endothelial nitric oxide synthase) from Prof. Keith Channon, University of Oxford (Benest et al . ); Ad. sFlt1 (soluble Fms‐like tyrosine kinase receptor‐1/soluble VEGFR1) was generated by Dr Ewa Paleolog, Imperial College London (Afuwape et al . ) and Ad. sTie2 (soluble Tie2) (Lin et al . ) by Prof. Charles Lin, Vanderbilt University, Nashville, TN, USA.

Techniques: Inhibition, Staining