scholarly journals Aldosterone Induces Superoxide Generation via Rac1 Activation in Endothelial Cells

Endocrinology ◽  
2007 ◽  
Vol 149 (3) ◽  
pp. 1009-1014 ◽  
Author(s):  
Fumiko Iwashima ◽  
Takanobu Yoshimoto ◽  
Isao Minami ◽  
Maya Sakurada ◽  
Yuki Hirono ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
G Protein ◽  
Mineralocorticoid Receptor ◽  
Nicotinamide Adenine Dinucleotide Phosphate ◽  
Oxidase Inhibitor ◽  
Dominant Negative ◽  
Superoxide Generation ◽  
Cellular Mechanisms ◽  
Mineralocorticoid Receptor Antagonist ◽  
Aortic Endothelial Cells

Currently, aldosterone is believed to be involved in the development of cardiovascular injury as a potential cardiovascular risk hormone. However, its exact cellular mechanisms remain obscure. This study was undertaken to examine the effect of aldosterone on superoxide production in cultured rat aortic endothelial cells with possible involvement of the small GTP-binding (G) protein Rac1. The aldosterone levels showed a time-dependent (6–24 h) and dose-dependent (10−8 to 10−6m) increase in superoxide generation, whose effect was abolished by mineralocorticoid receptor antagonist (eplerenone), Src inhibitor (PP2), and reduced nicotinamide adenine dinucleotide phosphate [NAD(P)H] oxidase inhibitor (apocynin). Aldosterone activated NADP(H) oxidase and Rac1, whose effects were abolished by eplerenone. The aldosterone-induced superoxide generation was abolished either by nonselective small G protein inhibitor (Clostridium difficile toxin A) or dominant-negative Rac1. Dominant-negative Rac1 also inhibited aldosterone-induced ACE gene expression. Thus, the present study is the first to demonstrate that aldosterone induces superoxide generation via mineralocorticoid receptor-mediated activation of NAD(P)H-oxidase and Rac1 in endothelial cells, thereby contributing to the development of aldosterone-induced vascular injury.

Functional expression of Kir2.x in human aortic endothelial cells: the dominant role of Kir2.2

2005 ◽  
Vol 289 (5) ◽  
pp. C1134-C1144 ◽  
Author(s):  
Yun Fang ◽  
Gernot Schram ◽  
Victor G. Romanenko ◽  
Congzhu Shi ◽  
Lisa Conti ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Dominant Role ◽  
Functional Expression ◽  
Inward Rectifier ◽  
Dominant Negative ◽  
Current Sensitivity ◽  
Aortic Endothelial Cells ◽  
Human Aortic Endothelial Cells ◽  
Aortic Endothelial

Inward rectifier K+ channels (Kir) are a significant determinant of endothelial cell (EC) membrane potential, which plays an important role in endothelium-dependent vasodilatation. In the present study, several complementary strategies were applied to determine the Kir2 subunit composition of human aortic endothelial cells (HAECs). Expression levels of Kir2.1, Kir2.2, and Kir2.4 mRNA were similar, whereas Kir2.3 mRNA expression was significantly weaker. Western blot analysis showed clear Kir2.1 and Kir2.2 protein expression, but Kir2.3 protein was undetectable. Functional analysis of endothelial inward rectifier K+ current ( IK) demonstrated that 1) IK current sensitivity to Ba2+ and pH were consistent with currents determined using Kir2.1 and Kir2.2 but not Kir2.3 and Kir2.4, and 2) unitary conductance distributions showed two prominent peaks corresponding to known unitary conductances of Kir2.1 and Kir2.2 channels with a ratio of ∼4:6. When HAECs were transfected with dominant-negative (dn)Kir2.x mutants, endogenous current was reduced ∼50% by dnKir2.1 and ∼85% by dnKir2.2, whereas no significant effect was observed with dnKir2.3 or dnKir2.4. These studies suggest that Kir2.2 and Kir2.1 are primary determinants of endogenous K+ conductance in HAECs under resting conditions and that Kir2.2 provides the dominant conductance in these cells.

scholarly journals PKC-α mediates flow-stimulated superoxide production in thick ascending limbs

2010 ◽  
Vol 298 (4) ◽  
pp. F885-F891 ◽  
Author(s):  
Nancy J. Hong ◽  
Guillermo B. Silva ◽  
Jeffrey L. Garvin
Keyword(s):  
Nadph Oxidase ◽  
Oxidase Activity ◽  
Mesangial Cells ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Oxidase Inhibitor ◽  
Dominant Negative ◽  
Aortic Endothelial Cells ◽  
Nadph Oxidase Activity ◽  
Measured Flow

We showed that luminal flow increases net superoxide (O2−) production via NADPH oxidase in thick ascending limbs. Protein kinase C (PKC) activates NADPH oxidase activity in phagocytes, cardiomyocytes, aortic endothelial cells, vascular smooth muscle cells, and renal mesangial cells. However, the flow-activated pathway that induces NADPH oxidase activity in thick ascending limbs is unclear. We hypothesized that PKC mediates flow-stimulated net O2− production by thick ascending limbs. Initiation of flow (20 nl/min) increased net O2− production from 4 ± 1 to 61 ± 12 AU/s ( P < 0.007; n = 5). The NADPH oxidase inhibitor apocynin completely blocked the flow-induced increase in net O2− production (2 ± 1 vs. 1 ± 1 AU/s; P > 0.05; n = 5). Flow-stimulated O2− was also blocked in p47phox-deficient mice. We measured flow-stimulated PKC activity with a fluorescence resonance energy transfer (FRET)-based membrane-targeted PKC activity reporter and found that the FRET ratio increased from 0.87 ± 0.02 to 0.96 ± 0.04 AU ( P < 0.05; n = 6). In the absence of flow, the PKC activator phorbol 12-myristate 13-acetate (200 nM) enhanced net O2− production from 5 ± 2 to 92 ± 6 AU/s ( P < 0.001; n = 6). The PKC-α- and βI-selective inhibitor Gö 6976 (100 nM) decreased flow-stimulated net O2− production from 54 ± 15 to 2 ± 1 AU/s ( P < 0.04; n = 5). Flow-induced net O2− production was inhibited in thick ascending limbs transduced with dominant-negative (dn)PKC-α but not dnPKCβI or LacZ (Δ = 11 ± 3 AU/s for dnPKCα, 55 ± 7 AU/s for dnPKCβI, and 63 ± 7 AU/s for LacZ; P < 0.001; n = 6). We concluded that flow stimulates net O2− production in thick ascending limbs via PKC-α-mediated activation of NADPH oxidase.

scholarly journals G Protein-Coupled Estrogen Receptor 1 (GPER1) Mediates Aldosterone-Induced Endothelial Inflammation in a Mineralocorticoid Receptor-Independent Manner

2021 ◽  
Vol 2021 ◽  
pp. 1-8
Author(s):  
Ziwei Tang ◽  
Qifu Li ◽  
Qingfeng Cheng ◽  
Mei Mei ◽  
Ying Song ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Estrogen Receptor ◽  
Inflammatory Response ◽  
G Protein ◽  
Mineralocorticoid Receptor ◽  
Umbilical Vein ◽  
Vascular Inflammation ◽  
Independent Manner ◽  
G Protein Coupled ◽  
Estrogen Receptor 1

Objective. It has been increasingly appreciated that G protein-coupled estrogen receptor 1 (GPER1) mediates both proinflammatory and anti-inflammatory response of estrogen. It is also involved in some rapid vascular effects of aldosterone in a mineralocorticoid receptor (MR) independent manner. However, whether GPER1 mediates aldosterone-induced inflammation response in endothelial cells and its relationship with MR are yet undetermined and therefore require further explanation. Method. Based on the hypothesis that GPER1 plays a role in the aldosterone-related vascular inflammation, the present study utilized a model of human umbilical vein endothelial cells transfected with MR siRNA and induced for inflammatory response with increasing concentration of aldosterone. Results. It was discovered that induction of aldosterone had no effect on the expression of GPER1 but promoted the expression of MR. Suppression of MR did not influence GPER1 expression, and GPER1 was capable of mediating part of aldosterone-induced endothelial inflammatory response. This effect may involve phosphoinositide 3-kinases (PI3K) pathway signaling. Conclusion. These findings not only demonstrated the role of GPER1 in aldosterone-induced vascular inflammation but also suggested an alternative for pharmaceutical treatment of hyperaldosteronism considering the unsatisfying effect on cardiovascular risks with MR antagonists.

Abstract 442: FOXO1/Notch Signaling Modulates Ambient Ultrafine Particle Impaired Vascular Repair

2017 ◽  
Vol 37 (suppl_1) ◽  
Author(s):  
Kyung In Baek ◽  
René R Packard ◽  
Arian Saffari ◽  
Zhao Ma ◽  
Anh P Luu ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Notch Signaling ◽  
Ultrafine Particles ◽  
Ultrafine Particle ◽  
Dominant Negative ◽  
Vascular Repair ◽  
Active Components ◽  
Aortic Endothelial Cells ◽  
Notch Activity ◽  
Differentiation And Proliferation

Introduction: Exposure to ultrafine particles (UFP, d < 0.1 μm), redox-active components of particular matter (PM 2.5 ), promotes endothelial dysfunction. Notch signaling in endothelial cells (EC) regulates differentiation and proliferation of vasculature. FOXO1 interacts with Notch signaling by enhancing assembly of activation complex during induction of Notch signaling. Whether UFP impair vascular repair by modulating FOXO1/Notch signaling axis remains elusive. Hypothesis: We hypothesized that UFP impairs vascular repair by attenuating Notch signaling via inhibition on FOXO1. Methods and Results: Control transgenic Tg(fli1:gfp) zebrafish embryos underwent tail amputation at 3 days post fertilization (dpf) developed complete vascular repair at 3 days post amputation (dpa), whereas exposure to UFP, or treatment with ADAM10 inhibitor to prevent Notch activation, or micro-injection of dominant negative(DN) Notch1b mRNA disrupted vascular network and impaired regeneration (* P < 0.05, n=20). By crossing the Notch reporter line Tg(tp1:gfp) with the Tg(flk1:mCherry) line, we demonstrated UFP inhibits endothelial Notch signaling on the amputated site at 3 dpa. Micro-injection of NICD mRNA only partially rescued endothelial Notch activity and impaired vascular repair in the presence of UFP (* P < 0.05, n=20). FOXO1 MO significantly inhibited Notch signaling, mimicking the UFP-impaired vascular repair. Injection of FOXO1 mRNA accentuated Notch activity and rescued UFP-impaired vascular repair. In human aortic endothelial cells, UFP suppressed FOXO1 expression and the co-localization with NICD, but not Master-Mind Like 1(MAML) or active NICD expression (* P < 0.05, n=3). As a corollary, UFP exposure induced dose and time-dependent reduction in Notch reporter activity, FOXO1 mRNA expression and the expression of Notch signaling related genes including the Notch ligand Dll4 and Notch target HES1. (* P < 0.05, n=3). Conclusions: In conclusion, UFP attenuated FOXO1/Notch cooperation to modulate Notch signaling and impaired vascular repair in embryonic zebrafish.

scholarly journals Arsenite stimulates plasma membrane NADPH oxidase in vascular endothelial cells

2001 ◽  
Vol 280 (3) ◽  
pp. L442-L449 ◽  
Author(s):  
Karol R. Smith ◽  
Linda R. Klei ◽  
Aaron Barchowsky
Keyword(s):  
Endothelial Cells ◽  
Angiotensin Ii ◽  
Nadph Oxidase ◽  
Oxidase Activity ◽  
Vascular Endothelial Cells ◽  
Oxidase Inhibitor ◽  
Vascular Endothelial ◽  
Effect Analysis ◽  
Aortic Endothelial Cells ◽  
Necrosis Factor

Low-level arsenite treatment of porcine aortic endothelial cells (PAEC) stimulated superoxide accumulation that was attenuated by inhibitors of NAD(P)H oxidase. To demonstrate whether arsenite stimulated NADPH oxidase, intact PAEC were treated with arsenite for up to 2 h and membrane fractions were prepared to measure NADPH oxidase activity. Arsenite (5 μM) stimulated a twofold increase in activity by 1 h, which was inhibited by the oxidase inhibitor diphenyleneiodonium chloride. Direct treatment of isolated membranes with arsenite had no effect. Analysis of NADPH oxidase components revealed that p67phoxlocalized exclusively to membranes of both control and treated cells. In contrast, cytosolic Rac1 translocated to the membrane fractions of cells treated with arsenite or angiotensin II but not with tumor necrosis factor. Immunodepletion of p67phoxblocked oxidase activity stimulated by all three compounds. However, depleting Rac1 inhibited responses only to arsenite and angiotensin II. These data demonstrate that stimulus-specific activation of NADPH oxidase in endothelial cells was the source of reactive oxygen in endothelial cells after noncytotoxic arsenite exposure.

Calcium signalling by G protein-coupled sphingolipid receptors in bovine aortic endothelial cells

1996 ◽  
Vol 354 (4) ◽  
pp. 397-403 ◽  
Author(s):  
Dagmar zu Heringdorf Meyer ◽  
Chris J. van Koppen ◽  
Bernd Windorfer ◽  
Herbert M. Himmel ◽  
Karl H. Jakobs
Keyword(s):  
Endothelial Cells ◽  
G Protein ◽  
Calcium Signalling ◽  
Aortic Endothelial Cells ◽  
Bovine Aortic Endothelial Cells ◽  
G Protein Coupled ◽  
Aortic Endothelial

Systematic study of dominant negative Gα C-terminal minigenes delineate G protein pathways in thrombin-mediated responses of endothelial cells

2000 ◽  
Vol 28 (5) ◽  
pp. A470-A470
Author(s):  
T. O. Thomas ◽  
A. Gilchrist ◽  
A. Li ◽  
J. F. Vanhauwe ◽  
T. A. Voyno-Yasenetskaya ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
G Protein ◽  
Systematic Study ◽  
Dominant Negative

scholarly journals Adaptor Protein Crk Is Required for Ephrin-B1-induced Membrane Ruffling and Focal Complex Assembly of Human Aortic Endothelial Cells

2002 ◽  
Vol 13 (12) ◽  
pp. 4231-4242 ◽  
Author(s):  
Ken-Ichiro Nagashima ◽  
Akira Endo ◽  
Hisakazu Ogita ◽  
Akiko Kawana ◽  
Akiko Yamagishi ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Adaptor Protein ◽  
Dominant Negative ◽  
Aortic Endothelial Cells ◽  
Induced Membrane ◽  
Membrane Ruffling ◽  
Focal Complex ◽  
Human Aortic Endothelial Cells ◽  
Aortic Endothelial

Endothelial cell migration is an essential step in vasculogenesis and angiogenesis, in which receptor tyrosine kinases play a pivotal role. We investigated the mechanism by which ephrin-B1 promotes membrane ruffling in human aortic endothelial cells, because membrane ruffling heralds cell body migration. We especially focused on the role of Crk adaptor protein in EphB-mediated signaling. Using DsRed-tagged Crk and a fluorescent time-lapse microscope, we showed that Crk was recruited to the nascent focal complex after ephrin-B1 stimulation. Furthermore, we found that p130Cas, but not paxillin, recruited Crk to the nascent focal complex. The necessity of Crk in ephrin-B1–induced membrane ruffling was shown both by the overexpression of dominant negative Crk mutants and by the depletion of Crk by using RNA interference. Then, we examined the role of two major downstream molecules of Crk, Rac1 and Rap1. The dominant negative mutant of Rac1 completely inhibited ephrin-B1–induced membrane ruffling and focal complex assembly. In contrast, rap1GAPII, a negative regulator of Rap1, did not inhibit ephrin-B1–induced membrane ruffling. However, in rap1GAPII-expressing cells, ephrin-B1 did not induce membrane spreading, probably due to instability of the focal complex. These results indicated that Crk plays a critical role in Rac1-induced membrane ruffling and Rap1-mediated nascent focal complex stabilization contributing to ephrin-B1–induced human aortic endothelial cells migration.

Ornithine Decarboxylase Activity in Cultured Endothelial Cells Stimulated by Serum, Thrombin and Serotonin

1978 ◽  
Vol 39 (02) ◽  
pp. 496-503 ◽  
Author(s):  
P A D’Amore ◽  
H B Hechtman ◽  
D Shepro
Keyword(s):  
Endothelial Cells ◽  
Ornithine Decarboxylase ◽  
Decarboxylase Activity ◽  
Aortic Endothelial Cells ◽  
Ornithine Decarboxylase Activity ◽  
Rate Limiting Step ◽  
Bovine Aortic Endothelial Cells ◽  
Limiting Step ◽  
Rate Limiting ◽  
Serum Serotonin

SummaryOrnithine decarboxylase (ODC) activity, the rate-limiting step in the synthesis of polyamines, can be demonstrated in cultured, bovine, aortic endothelial cells (EC). Serum, serotonin and thrombin produce a rise in ODC activity. The serotonin-induced ODC activity is significantly blocked by imipramine (10-5 M) or Lilly 11 0140 (10-6M). Preincubation of EC with these blockers together almost completely depresses the 5-HT-stimulated ODC activity. These observations suggest a manner by which platelets may maintain EC structural and metabolic soundness.

Sign in / Sign up

Export Citation Format

Share Document