Disintegration of cytoskeletal structure of actin filaments in energy-depleted endothelial cells

1993 ◽  
Vol 264 (5) ◽  
pp. H1599-H1608 ◽  
Author(s):  
W. Kuhne ◽  
M. Besselmann ◽  
T. Noll ◽  
A. Muhs ◽  
H. Watanabe ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Actin Filaments ◽  
Fold Increase ◽  
Metabolic Inhibition ◽  
Ionophore A23187 ◽  
Aortic Endothelial Cells ◽  
Monolayer Cultures ◽  
Cell Retraction ◽  
Porcine Aortic Endothelial Cells ◽  
Reversible Phase

In a previous study [Watanabe, H., W. Kuhne, R. Spahr, P. Schwartz, and H. M. Piper. Am. J. Physiol. 260 (Heart Circ. Physiol. 29): H1344-H1352, 1991] metabolic inhibition (5 mM KCN + 5 mM 2-deoxy-D-glucose, for 2 h) was found to cause disintegration of F-actin filaments, cell retraction, and augmented paracellular macromolecule permeability in monolayer cultures of porcine aortic endothelial cells after a rapid depletion of ATP stores (90% in 5 min). These changes were reversible. In the present study, the nature of this cytoskeletal disintegration was investigated. 1) Disintegration of F-actin filaments within 2-h incubation under metabolic inhibition was accompanied by appearance of F-actin clumps in the cells, but total contents of F-actin remained unaltered. 2) Cytosolic Ca2+ levels rapidly rose in metabolically inhibited cells; after 2 h a 10-fold increase was observed. 3) Presence of the Ca2+ ionophore A23187 (10 microM) mimicked the reversible effect of metabolic inhibition on F-actin filaments and monolayer permeability but not the extensive depletion of ATP stores. 4) Existence of the Ca(2+)-activatable actin-severing protein gelsolin in endothelial cells was demonstrated. The results show that during the reversible phase of endothelial energy depletion disintegration of F-actin filaments is only partial, since it is based on their fragmentation and not depolymerization. Increase in cytosolic Ca2+ levels seems to be the primary cause for the fragmentation, possibly through the activation of gelsolin.

scholarly journals Saccharide expression on wounded endothelial cell monolayers in vitro

1989 ◽  
Vol 93 (1) ◽  
pp. 163-172
Author(s):  
R.Y. Ball ◽  
R.W. Stoddart ◽  
C.J. Jones ◽  
M.J. Mitchinson
Keyword(s):  
Cell Proliferation ◽  
Endothelial Cells ◽  
Cellular Response ◽  
Cell Spreading ◽  
Aortic Endothelial Cells ◽  
Cell Monolayers ◽  
Sequence Of Events ◽  
Monolayer Cultures ◽  
Porcine Aortic Endothelial Cells

Monolayer cultures of porcine aortic endothelial cells were used as a model of the endothelium of large arteries. Linear wounds were produced in such cultures by scraping and the subsequent sequence of events in nearby cells was analysed. The earliest detectable event was cellular spreading at the margins of the wounds (2 h) followed by cell migration (starting at 6–8 h) and cell proliferation in regions adjacent to the wound (16 h and later). Cell spreading was associated with the appearance of saccharides selectively at the spreading margins of the cells, which bound the lectins, ConA, LCA and PSA, and were sensitive to alpha-mannosidase. Terminal alpha-mannosyl residues were therefore present. The appearance of these saccharides suggests a mechanism by which monocytes might adhere to and/or migrate through the endothelium of vessels at sites of cellular response to injury.

Calcium, calmodulin, and the production of prostacyclin by cultured vascular endothelial cells

1983 ◽  
Vol 3 (11) ◽  
pp. 1007-1015 ◽  
Author(s):  
J. M. Seid ◽  
S. Macneil ◽  
S. Tomlinson
Keyword(s):  
Endothelial Cells ◽  
Vascular Endothelial Cells ◽  
Calcium Ionophore ◽  
Calcium Ionophore A23187 ◽  
Ionophore A23187 ◽  
Vascular Endothelial ◽  
Calmodulin Antagonist ◽  
Aortic Endothelial Cells ◽  
Intracellular Calcium Mobilization ◽  
Porcine Aortic Endothelial Cells

The production of prostacyclin (PGI2) by cultured porcine aortic endothelial cells, in response to serum and the calcium ionophore A23187, was inhibited by TMB-8, an antagonist of intracellular calcium mobilization. The calcium-channel blocker methoxyverapamil (D600) inhibited serum-induced PGI2 production in but had little effect on A23187-induced PGI2 production. Calmodulin activity was detected in endothelial-cell Jysates and was inhibited by the calmodulin antagonist W7, which also inhibited PGI2 production in response to both agonists. Calcium and calmodulin appear to play an important role in mediating PGI2 production by the vascular endothelium.

scholarly journals Rho GTPases have diverse effects on the organization of the actin filament system

2004 ◽  
Vol 377 (2) ◽  
pp. 327-337 ◽  
Author(s):  
Pontus ASPENSTRÖM ◽  
Åsa FRANSSON ◽  
Jan SARAS
Keyword(s):  
Endothelial Cells ◽  
Actin Filament ◽  
Focal Adhesion ◽  
Rho Gtpases ◽  
Actin Filaments ◽  
Rho Gtpase ◽  
Major Influence ◽  
Aortic Endothelial Cells ◽  
Filament System ◽  
Porcine Aortic Endothelial Cells

The Rho GTPases are related to the Ras proto-oncogenes and consist of 22 family members. These proteins have important roles in regulating the organization of the actin filament system, and thereby the morphogenesis of vertebrate cells as well as their ability to migrate. In an effort to compare the effects of all members of the Rho GTPase family, active Rho GTPases were transfected into porcine aortic endothelial cells and the effects on the actin filament system were monitored. Cdc42, TCL (TC10-like), Rac1–Rac3 and RhoG induced the formation of lamellipodia, whereas Cdc42, Rac1 and Rac2 also induced the formation of thick bundles of actin filaments. In contrast, transfection with TC10 or Chp resulted in the formation of focal adhesion-like structures, whereas Wrch-1 induced long and thin filopodia. Transfection with RhoA, RhoB or RhoC induced the assembly of stress fibres, whereas Rnd1–Rnd3 resulted in the loss of stress fibres, but this effect was associated with the formation of actin- and ezrin-containing dorsal microvilli. Cells expressing RhoD and Rif had extremely long and flexible filopodia. None of the RhoBTB or Miro GTPases had any major influence on the organization of the actin filament system; instead, RhoBTB1 and RhoBTB2 were present in vesicular structures, and Miro-1 and Miro-2 were present in mitochondria. Collectively, the data obtained in this study to some extent confirm earlier observations, but also allow the identification of previously undetected roles of the different members of the Rho GTPases.

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.

Involvement of Ca2+/calmodulin-dependent protein kinase II in endothelial NO production and endothelium-dependent relaxation

2003 ◽  
Vol 284 (6) ◽  
pp. H2311-H2319 ◽  
Author(s):  
Jean-Christophe Schneider ◽  
Driss El Kebir ◽  
Christiane Chéreau ◽  
Sophie Lanone ◽  
Xiao-Lin Huang ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Protein Kinase ◽  
No Production ◽  
Ionophore A23187 ◽  
Dependent Protein Kinase ◽  
Aortic Endothelial Cells ◽  
Protein Kinase Ii ◽  
Dependent Protein ◽  
Endothelium Dependent Relaxation ◽  
Aortic Endothelial

Nitric oxide (NO) is synthesized froml-arginine by the Ca2+/calmodulin-sensitive endothelial NO synthase (NOS) isoform (eNOS). The present study assesses the role of Ca2+/calmodulin-dependent protein kinase II (CaMK II) in endothelium-dependent relaxation and NO synthesis. The effects of three CaMK II inhibitors were investigated in endothelium-intact aortic rings of normotensive rats. NO synthesis was assessed by a NO sensor and chemiluminescence in culture medium of cultured porcine aortic endothelial cells stimulated with the Ca2+ ionophore A23187 and thapsigargin. Rat aortic endothelial NOS activity was measured by the conversion ofl-[3H]arginine tol-[3H]citrulline. Three CaMK II inhibitors, polypeptide 281–302, KN-93, and lavendustin C, attenuated the endothelium-dependent relaxation of endothelium-intact rat aortic rings in response to acetylcholine, A23187, and thapsigargin. None of the CaMK II inhibitors affected the relaxation induced by NO donors. In a porcine aortic endothelial cell line, KN-93 decreased NO synthesis and caused a rightward shift of the concentration-response curves to A23187 and thapsigargin. In rat aortic endothelial cells, KN-93 significantly decreased bradykinin-induced eNOS activity. These results suggest that CaMK II was involved in NO synthesis as a result of Ca2+-dependent activation of eNOS.

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.

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