Purine metabolism in cultured aortic and coronary endothelial cells

1989 ◽  
Vol 67 (1) ◽  
pp. 8-15 ◽  
Author(s):  
Christine Des Rosiers ◽  
Stephan Nees ◽  
Eckehart Gerlach
Keyword(s):  
Endothelial Cells ◽  
Adrenergic Receptor ◽  
De Novo ◽  
De Novo Synthesis ◽  
Purine Nucleotides ◽  
Aortic Endothelial Cells ◽  
Purine Bases ◽  
Coronary Endothelial Cells ◽  
Salvage Pathways ◽  
Aortic Endothelial

Purine salvage pathways in cultured endothelial cells of macrovascular (pig aorta) and microvascular (guinea pig coronary system) origin were investigated by measuring the incorporation of radioactive purine bases (adenine or hypoxanthine) or nucleosides (adenosine or inosine) into purine nucleotides. These precursors were used at initial extracellular concentrations of 0.1, 5, and 500 μM. In both types of endothelial cells, purine nucleotide synthesis occurred with all four substrates. Aortic endothelial cells salvaged adenine best among purines and nucleosides when applied at 0.1 μM. At 5 and 500 μM, adenosine was the best precursor. In contrast, microvascular endothelial cells from the coronary system used adenosine most efficiently at all concentrations studied. The synthetic capacity of salvage pathways was greater than that of the de novo pathway. As measured using radioactive formate or glycine, de novo synthesis of purine nucleotides was barely detectable in aortic endothelial cells, whereas it readily occurred in coronary endothelial cells. Purine de novo synthesis in coronary endothelial cells was inhibited by physiological concentrations of purine bases and nucleosides, and by ribose or isoproterenol. The isoproterenol-induced inhibition was prevented by the β-adrenergic receptor antagonist propranolol. The end product of purine catabolism in aortic endothelial cells was found to be hypoxanthine, whereas coronary endothelial cells degraded hypoxanthine further to xanthine and uric acid, a reaction catalyzed by the enzyme xanthine dehydrogenase.Key words: purine metabolism, aortic endothelial cells, coronary endothelial cells, β-adrenergic receptor.

Regulation of purine nucleotide biosynthesis: in yeast and beyond

2006 ◽  
Vol 34 (5) ◽  
pp. 786-790 ◽  
Author(s):  
R.J. Rolfes
Keyword(s):  
Saccharomyces Cerevisiae ◽  
De Novo ◽  
Purine Nucleotide ◽  
De Novo Synthesis ◽  
Normal Functioning ◽  
Purine Nucleotides ◽  
Yeast Saccharomyces Cerevisiae ◽  
Nucleotide Biosynthesis ◽  
Salvage Pathways ◽  
Pool Sizes

Purine nucleotides are critically important for the normal functioning of cells due to their myriad of activities. It is important for cells to maintain a balance in the pool sizes of the adenine-containing and guanine-containing nucleotides, which occurs by a combination of de novo synthesis and salvage pathways that interconvert the purine nucleotides. This review describes the mechanism for regulation of the biosynthetic genes in the yeast Saccharomyces cerevisiae and compares this mechanism with that described in several microbial species.

Abstract 611: AP-1 and ETS Family Transcription Factors Co-localize at Enhancers in Human Aortic Endothelial Cells

2015 ◽  
Vol 35 (suppl_1) ◽  
Author(s):  
Nicholas T Hogan ◽  
Casey E Romanoski ◽  
Michael T Lam ◽  
Christopher K Glass
Keyword(s):  
Endothelial Cells ◽  
Transcription Factor ◽  
Transcription Factors ◽  
De Novo ◽  
Regulatory Elements ◽  
Specific Gene ◽  
Motif Analysis ◽  
Aortic Endothelial Cells ◽  
Human Aortic Endothelial Cells ◽  
Aortic Endothelial

Introduction: Sequence-specific transcription factors bind DNA regulatory elements and play a key role in establishing cellular identity. Studies comparing macrophages to B cells have revealed that small numbers of such collaborative or lineage-determining transcription factors (LDTF) establish distinct enhancers in each cell type. These factors also allow for the binding of signal dependent transcription factors. Here we present data which suggest members of the AP-1, ETS, and STAT transcription factor families serve as collaborative transcriptional regulators in human aortic endothelial cells (HAEC). Hypothesis: We hypothesize that a set of AP-1 and ETS transcription factors collaborate to establish key endothelial cell enhancers. Methods: Working in HAEC, we measured poised and active enhancers using ChIP-seq for the epigenetic histone modifications H3K4me2 and H3K27Ac, performed motif analysis, and measured transcription factor binding for candidate factors. Knockdowns of JUN, ERG, and STAT3 followed by RNA-seq were used to evaluate altered enhancer function and gene targets of candidate factors. Results: Our de novo motif analysis revealed that motifs for ETS and AP-1 transcription factors are highly enriched at HAEC enhancers. ChIP-seq experiments for JUN, JUNB, ERG, and STAT3 showed between 8,000 and 55,000 intergenic peaks for each factor. Together these peaks bind 50% of poised enhancers, with a subset co-localizing at these sites. Gene ontology analysis showed that gene targets of these enhancers are involved in endothelial-specific functions. Further, knockdown of JUN, ERG, and STAT3 resulted in a twofold or greater change in expression of hundreds of HAEC transcripts. Conclusion: The genome-wide pattern of JUN, JUNB, ERG, and STAT3 co-localization at enhancers in HAEC suggests these factors serve as key regulators that collaboratively modulate endothelial-specific gene expression. Further investigation of candidate lineage-determining transcription factors using pro-atherogenic signals could reveal regulatory mechanisms of disease-relevant endothelial transcriptional programs.

Prostaglandin H synthase: protein synthesis-independent regulation in bovine aortic endothelial cells

1997 ◽  
Vol 273 (5) ◽  
pp. C1749-C1755 ◽  
Author(s):  
Moti Rosenstock ◽  
Abraham Danon ◽  
Gilad Rimon
Keyword(s):  
Endothelial Cells ◽  
Protein Synthesis ◽  
Tyrosine Kinase ◽  
De Novo ◽  
Prostaglandin H Synthase ◽  
Western Blots ◽  
Aortic Endothelial Cells ◽  
Bovine Aortic Endothelial Cells ◽  
Prostaglandin H ◽  
Aortic Endothelial

The objective of the present study was to examine whether prostaglandin H synthase (PGHS) can be regulated by pathways independent of de novo synthesis of PGHS. Incubation of bovine aortic endothelial cells (BAEC) for as short as 5 min with NaF (40 mM) resulted in a 60% increase in PGHS activity. PGHS activity induced by NaF was unaffected by either 10 μM cycloheximide or 1 μM actinomycin D. Aspirin (25 μM) completely inhibited resting PGHS activity, and NaF did not induce further stimulation. NS-398 (500 nM), a specific PGHS-2 inhibitor, was ineffective. Basic fibroblast growth factor (bFGF) induced a significant increase in PGHS activity within 30 min and was insensitive to cycloheximide. The levels of PGHS-1 and PGHS-2 proteins, as measured by Western blots, were not affected by NaF or bFGF. The tyrosine kinase inhibitor genistein attenuated PGHS activity that was induced by NaF and bFGF, whereas the tyrosine phosphatase inhibitor, sodium orthovanadate, augmented these responses. The G protein activators 5′-guanylyl imidodiphosphate and guanosine 5′- O-(3-thiotriphosphate) inhibited both resting and NaF-induced PGHS activities. These results suggest that, in BAEC, PGHS-1 activity can be regulated by tyrosine kinase and/or G proteins, independently of de novo protein synthesis.

scholarly journals Linoleyl hydroperoxide transcriptionally upregulates heme oxygenase-1 gene expression in human renal epithelial and aortic endothelial cells.

1998 ◽  
Vol 9 (11) ◽  
pp. 1990-1997
Author(s):  
A Agarwal ◽  
F Shiraishi ◽  
G A Visner ◽  
H S Nick
Keyword(s):  
Endothelial Cells ◽  
Renal Disease ◽  
Heme Oxygenase ◽  
Molecular Mechanisms ◽  
De Novo ◽  
Oxidized Ldl ◽  
Heme Oxygenase 1 ◽  
Aortic Endothelial Cells ◽  
Mrna Induction ◽  
Aortic Endothelial

Atherogenic lipoproteins such as oxidized LDL are implicated in the pathogenesis of atherosclerosis and renal disease. Fatty acid hydroperoxides and phospholipids such as linoleyl hydroperoxide (LAox or 13-HPODE) and lysophosphatidylcholine (lyso-PC), abundant components of oxidized LDL, mediate the effects of atherogenic lipids. Oxidized LDL has been shown to induce heme oxygenase-1 (HO-1), a microsomal enzyme that is involved in heme detoxification and is a major endogenous source of carbon monoxide. HO-1 is also induced by many other stimuli that shift cellular redox. To identify the constituents and molecular mechanisms of oxidized LDL-mediated HO-1 induction, human renal epithelial cells and aortic endothelial cells were exposed to LAox and lyso-PC. Exposure to LAox (25 microM) showed an approximately 16-fold induction of HO-1 mRNA, whereas exposure to lyso-PC (25 microM) showed only an approximate 2.6-fold increase. Treatment with actinomycin-D (4 microM), a transcriptional inhibitor, as well as nuclear run-on assays, demonstrated that LAox-mediated HO-1 gene induction is dependent on de novo transcription. Cycloheximide did not affect LAox-mediated HO-1 mRNA induction, suggesting that new protein synthesis is not required for transcriptional induction. Transfection of a human HO-1 promoter-reporter gene construct showed that LAox upregulation of HO-1 occurs via mechanisms different from those of known inducers, heme and cadmium. These studies are the first demonstration that LAox induces HO-1 by transcriptional mechanisms and may have implications in the pathogenesis of cell injury in atherosclerosis and progressive renal disease.

scholarly journals Matrix metalloproteinases cleave the β2-adrenergic receptor in spontaneously hypertensive rats

2010 ◽  
Vol 299 (1) ◽  
pp. H25-H35 ◽  
Author(s):  
Stephen F. Rodrigues ◽  
Edward D. Tran ◽  
Zuleica B. Fortes ◽  
Geert W. Schmid-Schönbein
Keyword(s):  
Endothelial Cells ◽  
Adrenergic Receptor ◽  
Wistar Rats ◽  
Extracellular Domain ◽  
Aortic Endothelial Cells ◽  
Spontaneously Hypertensive ◽  
Wky Rats ◽  
Arteriolar Tone ◽  
Intracellular Domains ◽  
Aortic Endothelial

We recently observed the enhanced serine and matrix metalloproteinase (MMP) activity in the spontaneously hypertensive rat (SHR) compared with its normotensive Wistar-Kyoto (WKY) rat and the cleavage of membrane receptors in the SHR by MMPs. We demonstrate in vivo that MMP-7 and MMP-9 injection leads to a vasoconstrictor response in microvessels of rats that is blocked by a specific MMP inhibitor (GM-6001, 1 μM). Multiple pathways may be responsible. Since the β2-adrenergic receptor (β2-AR) is susceptible to the action of endogenous MMPs, we hypothesize that MMPs in the plasma of SHRs are able to cleave the extracellular domain of the β2-AR. SHR arterioles respond in an attenuated fashion to β2-AR agonists and antagonists. Aorta and heart muscle of control Wistar rats were exposed for 24 h (37°C) to fresh plasma of male Wistar and WKY rats and SHRs with and without doxycycline (30 μM) and EDTA (10 mM) to reduce MMP activity. The density of extracellular and intracellular domains of β2-AR was determined by immunohistochemistry. The density of the extracellular domain of β2-AR is reduced in aortic endothelial cells and cardiac microvessels of SHRs compared with that of WKY or Wistar rats. Treatment of the aorta and the heart of control Wistar rats with plasma from SHRs, but not from WKY rats, reduced the number of extracellular domains, but not intracellular domains, of β2-AR in aortic endothelial cells and cardiac microvessels. MMP inhibitors (EDTA and doxycycline) prevented the cleavage of the extracellular domain. Thus MMPs may contribute to the reduced density of the extracellular domain of β2-AR in blood vessels and to the increased arteriolar tone of SHRs compared with normotensive rats.

Effects of Dipyrone on Prostaglandin Production by Human Platelets and Cultured Bovine Aortic Endothelial Cells

1983 ◽  
Vol 49 (02) ◽  
pp. 132-137 ◽  
Author(s):  
A Eldor ◽  
G Polliack ◽  
I Vlodavsky ◽  
M Levy
Keyword(s):  
Endothelial Cells ◽  
Arachidonic Acid ◽  
Competitive Inhibition ◽  
Inhibitory Effect ◽  
Human Platelets ◽  
Ionophore A23187 ◽  
Aortic Endothelial Cells ◽  
Bovine Aortic Endothelial Cells ◽  
Aortic Endothelial

SummaryDipyrone and its metabolites 4-methylaminoantipyrine, 4-aminoantipyrine, 4-acetylaminoantipyrine and 4-formylaminoan- tipyrine inhibited the formation of thromboxane A2 (TXA2) during in vitro platelet aggregation induced by ADP, epinephrine, collagen, ionophore A23187 and arachidonic acid. Inhibition occurred after a short incubation (30–40 sec) and depended on the concentration of the drug or its metabolites and the aggregating agents. The minimal inhibitory concentration of dipyrone needed to completely block aggregation varied between individual donors, and related directly to the inherent capacity of their platelets to synthesize TXA2.Incubation of dipyrone with cultured bovine aortic endothelial cells resulted in a time and dose dependent inhibition of the release of prostacyclin (PGI2) into the culture medium. However, inhibition was abolished when the drug was removed from the culture, or when the cells were stimulated to produce PGI2 with either arachidonic acid or ionophore A23187.These results indicate that dipyrone exerts its inhibitory effect on prostaglandins synthesis by platelets or endothelial cells through a competitive inhibition of the cyclooxygenase system.

Platelets Stimulate Thromboplastin Synthesis in Human Endothelial Cells

1983 ◽  
Vol 49 (02) ◽  
pp. 069-072 ◽  
Author(s):  
U L H Johnsen ◽  
T Lyberg ◽  
K S Galdal ◽  
H Prydz
Keyword(s):  
Endothelial Cells ◽  
De Novo ◽  
Umbilical Vein ◽  
Time Dependent ◽  
De Novo Synthesis ◽  
Human Endothelial Cells ◽  
Tumor Promotor ◽  
Coagulation Assay ◽  
Platelet Aggregates ◽  
Umbilical Vein Endothelial Cells

SummaryHuman umbilical vein endothelial cells in culture synthesize thromboplastin upon stimulation with phytohaemagglutinin (PHA) or the tumor promotor 12-O-tetradecanoyl-phorbol-13-acetate (TPA). The thromboplastin activity is further strongly enhanced in a time dependent reaction by the presence of gel-filtered platelets or platelet aggregates. This effect was demonstrable at platelet concentrations lower than those normally found in plasma, it may thus be of pathophysiological relevance. The thromboplastin activity increased with increasing number of platelets added. Cycloheximide inhibited the increase, suggesting that de novo synthesis of the protein component of thromboplastin, apoprotein III, is necessary.When care was taken to remove monocytes no thromboplastin activity and no apoprotein HI antigen could be demonstrated in suspensions of gel-filtered platelets, platelets aggregated with thrombin or homogenized platelets when studied with a coagulation assay and an antibody neutralization technique.

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