scholarly journals Shear-induced tyrosine phosphorylation in endothelial cells requires Rac1-dependent production of ROS

1999 ◽  
Vol 276 (4) ◽  
pp. C838-C847 ◽  
Author(s):  
Li-Hong Yeh ◽  
Young J. Park ◽  
Riple J. Hansalia ◽  
Imraan S. Ahmed ◽  
Shailesh S. Deshpande ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Shear Stress ◽  
Map Kinase ◽  
Tyrosine Phosphorylation ◽  
Protein Detection ◽  
Pyrrolidine Dithiocarbamate ◽  
Dependent Manner ◽  
Aortic Endothelial Cells ◽  
Bovine Aortic Endothelial Cells ◽  
Aortic Endothelial

The shear-induced intracellular signal transduction pathway in vascular endothelial cells involves tyrosine phosphorylation and activation of mitogen-activated protein (MAP) kinase, which may be responsible for the sustained release of nitric oxide. MAP kinase is known to be activated by reactive oxygen species (ROS), such as H2O2, in several cell types. ROS production in ligand-stimulated nonphagocytic cells appears to require the participation of a Ras-related small GTP-binding protein, Rac1. We hypothesized that Rac1 might serve as a mediator for the effect of shear stress on MAP kinase activation. Exposure of bovine aortic endothelial cells to laminar shear stress of 20 dyn/cm2for 5–30 min stimulated total cellular and cytosolic tyrosine phosphorylation as well as tyrosine phosphorylation of MAP kinase. Treating endothelial cells with the antioxidants N-acetylcysteine and pyrrolidine dithiocarbamate inhibited in a dose-dependent manner the shear-stimulated increase in total cytosolic and, specifically, MAP kinase tyrosine phosphorylation. Hence, the onset of shear stress caused an enhanced generation of intracellular ROS, as evidenced by an oxidized protein detection kit, which were required for the shear-induced total cellular and MAP kinase tyrosine phosphorylation. Total cellular and MAP kinase tyrosine phosphorylation was completely blocked in sheared bovine aortic endothelial cells expressing a dominant negative Rac1 gene product (N17rac1). We concluded that the GTPase Rac1 mediates the shear-induced tyrosine phosphorylation of MAP kinase via regulation of the flow-dependent redox changes in endothelial cells in physiological and pathological circumstances.

Bovine Caveolin-2 Cloning and Effects of Shear Stress on Its Localization in Bovine Aortic Endothelial Cells

Endothelium ◽  
2004 ◽  
Vol 11 (3-4) ◽  
pp. 189-198 ◽  
Author(s):  
Nolan L. Boyd ◽  
Heonyong Park ◽  
Wen-Ping Sun ◽  
Sarah E. Coleman ◽  
Ramakrishna S. Cherukuri ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Shear Stress ◽  
Aortic Endothelial Cells ◽  
Bovine Aortic Endothelial Cells ◽  
Aortic Endothelial

Sustained JNK activation induces endothelial apoptosis: studies with colchicine and shear stress

1999 ◽  
Vol 277 (4) ◽  
pp. H1593-H1599 ◽  
Author(s):  
Ying-Li Hu ◽  
Song Li ◽  
John Y.-J. Shyy ◽  
Shu Chien
Keyword(s):  
Endothelial Cells ◽  
Shear Stress ◽  
Colchicine Treatment ◽  
Laminar Shear Stress ◽  
Aortic Endothelial Cells ◽  
Bovine Aortic Endothelial Cells ◽  
Jnk Activation ◽  
Laminar Shear ◽  
Sustained Activation ◽  
Aortic Endothelial

The disruption of microtubules by treating bovine aortic endothelial cells with 10−7–10−5M colchicine caused apoptosis, as evidenced by DNA laddering and TdT-mediated dUTP nick end labeling fluorescence staining. Colchicine treatment also induced a sustained activation of c-Jun NH2-terminal kinase (JNK) that lasted for ≥12 h. The blockade of JNK activity by using the negative interfering mutant JNK(K-R) markedly decreased the apoptosis induced by colchicine. Exposure of bovine aortic endothelial cells to laminar shear stress (12 dyn/cm2) caused a transient (<2 h) activation of JNK, and there was no induction of apoptosis. The sustained activation of JNK may play a significant role in the apoptosis induced by colchicine.

Volatile Anesthetics Affect Calcium Mobilization in Bovine Endothelial Cells

1996 ◽  
Vol 85 (5) ◽  
pp. 1147-1156 ◽  
Author(s):  
Thomas N. Pajewski ◽  
Ning Miao ◽  
Carl III Lynch ◽  
Roger A. Johns
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Adenosine Triphosphate ◽  
Volatile Anesthetic ◽  
Volatile Anesthetics ◽  
Dependent Manner ◽  
Aortic Endothelial Cells ◽  
Inhalational Anesthetics ◽  
Bovine Aortic Endothelial Cells ◽  
Aortic Endothelial

Background The site where volatile anesthetics inhibit endothelium-dependent, nitric oxide-mediated vasodilation is unclear. To determine whether anesthetics could limit endothelium-dependent nitric oxide production by inhibiting receptor-mediated increases in cytosolic Ca2+, experiments were performed to see if the inhalational anesthetics halothane, isoflurane, and enflurane affect intracellular Ca2+ ([Ca2+]i) transients induced by the agonists bradykinin and adenosine triphosphate in cultured bovine aortic endothelial cells. Methods Bovine aortic endothelial cells, which had been loaded with the fluorescent Ca2+ indicator Fura-2, were added to medium preequilibrated with volatile anesthetic (1.25% and 2.5% for isoflurane, 1.755 and 3.5% for enflurane, and 0.75% and 1.5% for halothane). In Ca(2+)-containing medium, intracellular Ca2+ transients were elicited in response to bradykinin (10 nM and 1 microM) or adenosine triphosphate (1 microM and 100 microM). Results Both bradykinin and adenosine triphosphate triggered a rapid rise to peak [Ca2+]i followed by a gradual decline to a plateau above the resting level. Although basal [Ca2+]i was unaltered by the anesthetics, both halothane and enflurane, in a dose-dependent manner, depressed the peak and plateau of the [Ca2+]i transient elicited by 10 nM bradykinin, whereas isoflurane had no effect. When [Ca2+]i transients were elicited by 1 microM bradykinin, halothane (1% and 5%) did not alter peak and plateau levels. Halothane and enflurane also decreased [Ca2+]i transients evoked by 1 microM and 100 microM adenosine triphosphate, whereas isoflurane also had no effect in this setting. Conclusions Halothane and enflurane, but not isoflurane, inhibit bradykinin- and adenosine triphosphate-stimulated Ca2+ transients in endothelial cells. Limitations of Ca2+ availability to activate constitutive endothelial nitric oxide synthase could allow for part, but not all, of the inhibition of endothelium-dependent nitric oxide-mediated vasodilation by inhalational anesthetics.

Vitamin E Binds to Specific Binding Sites and Enhances Prostacyclin Production by Cultured Aortic Endothelial Cells

1992 ◽  
Vol 68 (06) ◽  
pp. 744-751 ◽  
Author(s):  
Makoto Kunisaki ◽  
Fumio Umeda ◽  
Toyoshi Inoguchi ◽  
Hajime Nawata
Keyword(s):  
Endothelial Cells ◽  
Vitamin E ◽  
Binding Sites ◽  
Specific Binding ◽  
Ionophore A23187 ◽  
Dependent Manner ◽  
Aortic Endothelial Cells ◽  
Specific Binding Sites ◽  
Bovine Aortic Endothelial Cells ◽  
Aortic Endothelial

SummaryWe evaluated the effect of d-α-tocopherol (vitamin E) on the production of prostacyclin (PGI2) by cultured bovine aortic endothelial cells. Vitamin E at physiological doses significantly enhanced the production of PGI2 by aortic endothelial cells when added to the culture simultaneously with histamine, the Ca2+ ionophore A23187 (A23187), plasma-derived serum (PDS), or arachidonic acid. This effect was found to occur in a time- and dose-dependent manner, and the maximal enhancement was produced by 9.28 µM of vitamin E for 1 h incubations. Significantly lower amounts of lipid peroxides were measured in endothelial cells stimulated by 10% PDS with 9.28 µM of vitamin E than in those stimulated without vitamin E for over 24 h, although the stimulation during the initial 1 to 12 h period did not have a significant effect on lipid peroxide formation in cultured aortic endothelial cells.We also demonstrated that bovine aortic endothelial cells have specific binding sites for [3H]vitamin E that exhibited time- and temperature-dependent saturability. At 4° C, the nonspecific binding was 8–12% of the total binding, and the specific binding reached equilibrium by 2 h. Specific binding increased with the concentration of [3H]vitamin E and became saturated at concentrations between 1.5 µM and 2.0 µM per 2.0 × 105 cells. Raising the unlabeled vitamin E concentration from 97.7 nM to 1,000 µM reduced the specific binding of 2.0 µM [3H]vitamin E. The Scatchard plot of [3H]vitamin E binding to the endothelial cells shows two classes of binding sites: one with a high affinity {K a1 2.48 ± 0.32 × 107 NT-1, n = 6} and a low capacity {n 1 1.20 ± 0.34 × 107 sites/cell} and the other with a low affinity {K a2 1.18 ± 0.32 × 105 M–1} and a high capacity {n 2 3.39 ± 0.53 × 109 sites/cell}.Our results suggest that the endothelial cells binding sites for vitamin E may play some roles in vascular homeostasis in vivo, and that vitamin E may prevent the development of atherosclerotic changes due in part to the enhancement of PGI2 production by the vascular wall and its action as an antioxidant in vascular endothelial cell.

scholarly journals Modulation of sulfated proteoglycan synthesis by bovine aortic endothelial cells during migration.

1986 ◽  
Vol 102 (3) ◽  
pp. 679-687 ◽  
Author(s):  
M G Kinsella ◽  
T N Wight
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell Migration ◽  
Proteoglycan Synthesis ◽  
Dependent Manner ◽  
Sulfate Proteoglycan ◽  
Wound Edge ◽  
Aortic Endothelial Cells ◽  
Bovine Aortic Endothelial Cells ◽  
Sulfate Incorporation ◽  
Aortic Endothelial

The rates of 35S-sulfate incorporation into proteoglycan were compared in multi-scratch wounded and confluent cultures of bovine aortic endothelial cells to determine whether proteoglycan synthesis is altered as cells are stimulated to migrate and proliferate. Incorporation was found to be stimulated in a time-dependent manner, reaching maximal levels 44-50 h after wounding, as cells migrated into wounded areas of the culture dish. Quantitative autoradiography of 35S-sulfate-labeled single-scratch wounded cultures demonstrated a 2-4-fold increase in the number of silver grains over migrating cells near the wound edge when compared to cells remote from the wound edge. Furthermore, when cell proliferation was blocked by inhibition of DNA synthesis, the increase in 35S-sulfate incorporation into proteoglycan after wounding was unaffected. These data indicate that cell division is not required for the modulation of proteoglycan synthesis to occur after wounding. Characterization of the newly synthesized proteoglycan by ion-exchange and molecular sieve chromatography demonstrated that heparan sulfate proteoglycan constitutes approximately 80% of the labeled proteoglycan in postconfluent cultures, while after wounding, chondroitin sulfate proteoglycan and/or dermatan sulfate proteoglycan (CS/DSPG) increases to as much as 60% of the total labeled proteoglycan. These results suggest that CS/DSPG synthesis is stimulated concomitant with the stimulation of endothelial cell migration after wounding.

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