scholarly journals Identification of a heparin-binding protein using monoclonal antibodies that block heparin binding to porcine aortic endothelial cells

1995 ◽  
Vol 311 (2) ◽  
pp. 461-469 ◽  
Author(s):  
W A Patton ◽  
C A Granzow ◽  
L A Getts ◽  
S C Thomas ◽  
L M Zotter ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Monoclonal Antibodies ◽  
Endothelial Cell ◽  
Binding Protein ◽  
Heparan Sulphate ◽  
Heparin Binding ◽  
Aortic Endothelial Cells ◽  
Heparin Binding Protein ◽  
Porcine Aortic Endothelial Cells ◽  
Aortic Endothelial

The binding of heparin or heparan sulphate to a variety of cell types results in specific changes in cell function. Endothelial cells treated with heparin alter their synthesis of heparan sulphate proteoglycans and extracellular matrix proteins. In order to identify a putative endothelial cell heparin receptor that could be involved in heparin signalling, anti-(endothelial cell) monoclonal antibodies that significantly inhibit heparin binding to endothelial cells were prepared. Four of these antibodies were employed in affinity-chromatographic isolation of a heparin-binding protein from detergent-solubilized endothelial cells. The heparin-binding protein isolated from porcine aortic endothelial cells using four different monoclonal antibodies has an M(r) of 45,000 assessed by SDS/PAGE. The 45,000-M(r) heparin-binding polypeptide is isolated as a multimer. The antibody-isolated protein binds to heparin-affinity columns as does the pure 45,000-M(r) polypeptide, consistent with its identification as a putative endothelial heparin receptor.

Molecular characterization of the Na-K-Cl cotransporter of bovine aortic endothelial cells

1997 ◽  
Vol 273 (1) ◽  
pp. C188-C197 ◽  
Author(s):  
T. R. Yerby ◽  
C. R. Vibat ◽  
D. Sun ◽  
J. A. Payne ◽  
M. E. O'Donnell
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Amino Acid ◽  
Blot Analysis ◽  
Cdna Probe ◽  
Bovine Aortic Endothelial Cell ◽  
Coding Region ◽  
Cerebral Microvessels ◽  
Aortic Endothelial Cells ◽  
Aortic Endothelial

The Na-K-Cl cotransporter is an important regulator of endothelial cell volume and may also contribute to flux of Na and Cl across the endothelium of the blood-brain barrier. To date, two Na-K-Cl cotransport isoforms have been identified, the cotransporter in secretory epithelia, NKCC1, and that in absorptive renal epithelia, NKCC2. Our previous studies showed that a monoclonal antibody to the cotransporter of human colonic T84 epithelial cells, an NKCC1 isoform, recognizes a 170-kDa glycoprotein from endothelial cells. The molecular identity of the Na-K-Cl cotransporter present in endothelial cells, however, has been unknown. In addition, although evidence has been provided that phosphorylation of the endothelial cotransporter plays a role in regulating its activity, little is known about potential sites for protein kinase interaction with the cotransporter. The present study was conducted to determine the molecular structure of the endothelial Na-K-Cl cotransporter. Using a 1.0-kilobase (kb) cDNA fragment from a conserved region of the T84 cell cotransporter, we screened a bovine aortic endothelial cell cDNA library and subsequently identified and sequenced two overlapping clones that together spanned the entire coding region. The endothelial cotransporter is a 1,201-amino acid protein with 12 putative transmembrane segments and large amino and carboxy termini, each containing several consensus sites for phosphorylation by protein kinases. Comparison of the endothelial cotransporter amino acid sequence with known NKCC1 and NKCC2 sequences revealed a 96% identity with NKCC1. Northern blot analysis using a cDNA probe from the endothelial cotransporter revealed high expression of approximately 7.5-kb transcripts in a number of bovine tissues. Finally, a prominent expression of Na-K-Cl cotransporter was found by Western blot analysis in both cultured and freshly isolated endothelial cells of bovine aorta and cerebral microvessels.

scholarly journals Clinopodium tomentosum (Kunth) Govaerts Leaf Extract Influences in vitro Cell Proliferation and Angiogenesis on Primary Cultures of Porcine Aortic Endothelial Cells

2020 ◽  
Vol 2020 ◽  
pp. 1-11
Author(s):  
Irvin Tubon ◽  
Chiara Bernardini ◽  
Fabiana Antognoni ◽  
Roberto Mandrioli ◽  
Giulia Potente ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Primary Cultures ◽  
Ethanolic Extract ◽  
Total Phenolic ◽  
Dependent Manner ◽  
Aortic Endothelial Cells ◽  
Phenolic Components ◽  
Porcine Aortic Endothelial Cells ◽  
Aortic Endothelial

Clinopodium tomentosum (Kunth) Govaerts is an endemic species in Ecuador, where it is used as an anti-inflammatory plant to treat respiratory and digestive affections. In this work, effects of a Clinopodium tomentosum ethanolic extract (CTEE), prepared from aerial parts of the plant, were investigated on vascular endothelium functions. In particularly, angiogenesis activity was evaluated, using primary cultures of porcine aortic endothelial cells (pAECs). Cells were cultured for 24 h in the presence of CTEE different concentrations (10, 25, 50, and 100 μg/ml); no viability alterations were found in the 10-50 μg/ml range, while a slight, but significant, proliferative effect was observed at the highest dose. In addition, treatment with CTEE was able to rescue LPS-induced injury in terms of cell viability. The CTEE ability to affect angiogenesis was evaluated by scratch test analysis and by an in vitro capillary-like network assay. Treatment with 25-50 μg/ml of extract caused a significant increase in pAEC’s migration and tube formation capabilities compared to untreated cells, as results from the increased master junctions’ number. On the other hand, CTEE at 100 μg/ml did not induce the same effects. Quantitative PCR data demonstrated that FLK-1 mRNA expression significantly increased at a CTEE dose of 25 μg/ml. The CTEE phytochemical composition was assessed through HPLC-DAD; rosmarinic acid among phenolic acids and hesperidin among flavonoids were found as major phenolic components. Total phenolic content and total flavonoid content assays showed that flavonoids are the most abundant class of polyphenols. The CTEE antioxidant activity was also showed by means of the DPPH and ORAC assays. Results indicate that CTEE possesses an angiogenic capacity in a dose-dependent manner; this represents an initial step in elucidating the mechanism of the therapeutic use of the plant.

scholarly journals TNF-α, IL-4, and IFN-γ Regulate Differential Expression of P- and E-Selectin Expression by Porcine Aortic Endothelial Cells

2000 ◽  
Vol 164 (6) ◽  
pp. 3309-3315 ◽  
Author(s):  
Claire J. Stocker ◽  
Katharine L. Sugars ◽  
Olivier A. Harari ◽  
R. Clive Landis ◽  
Bernard J. Morley ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Differential Expression ◽  
Aortic Endothelial Cells ◽  
Tnf Α ◽  
Ifn Γ ◽  
Porcine Aortic Endothelial Cells ◽  
Aortic Endothelial

scholarly journals African Swine Fever Virus Infection of Porcine Aortic Endothelial Cells Leads to Inhibition of Inflammatory Responses, Activation of the Thrombotic State, and Apoptosis

2001 ◽  
Vol 75 (21) ◽  
pp. 10372-10382 ◽  
Author(s):  
Isabelle Vallée ◽  
Stephen W. G. Tait ◽  
Penelope P. Powell
Keyword(s):  
Endothelial Cells ◽  
African Swine Fever Virus ◽  
African Swine Fever ◽  
Surface Expression ◽  
Fever Virus ◽  
Parp Cleavage ◽  
Aortic Endothelial Cells ◽  
Porcine Aortic Endothelial Cells ◽  
Aortic Endothelial

ABSTRACT African swine fever (ASF) is an asymptomatic infection of warthogs and bushpigs, which has become an emergent disease of domestic pigs, characterized by hemorrhage, lymphopenia, and disseminated intravascular coagulation. It is caused by a large icosohedral double-stranded DNA virus, African swine fever virus (ASFV), with infection of macrophages well characterized in vitro and in vivo. This study shows that virulent isolates of ASFV also infect primary cultures of porcine aortic endothelial cells and bushpig endothelial cells (BPECs) in vitro. Kinetics of early and late gene expression, viral factory formation, replication, and secretion were similar in endothelial cells and macrophages. However, ASFV-infected endothelial cells died by apoptosis, detected morphologically by terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling and nuclear condensation and biochemically by poly(ADP-ribose) polymerase (PARP) cleavage at 4 h postinfection (hpi). Immediate-early proinflammatory responses were inhibited, characterized by a lack of E-selectin surface expression and interleukin 6 (IL-6) and IL-8 mRNA synthesis. Moreover, ASFV actively downregulated interferon-induced major histocompatibility complex class I surface expression, a strategy by which viruses evade the immune system. Significantly, Western blot analysis showed that the 65-kDa subunit of the transcription factor NF-κB, a central regulator of the early response to viral infection, decreased by 8 hpi and disappeared by 18 hpi. Both disappearance of NF-κB p65 and cleavage of PARP were reversed by the caspase inhibitor z-VAD-fmk. Interestingly, surface expression and mRNA transcription of tissue factor, an important initiator of the coagulation cascade, increased 4 h after ASFV infection. These data suggest a central role for vascular endothelial cells in the hemorrhagic pathogenesis of the disease. Since BPECs infected with ASFV also undergo apoptosis, resistance of the natural host must involve complex pathological factors other than viral tropism.

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