Role of nitric oxide in autocrine control of growth and apoptosis of endothelial cells

1997 ◽  
Vol 272 (2) ◽  
pp. H760-H768 ◽  
Author(s):  
A. Lopez-Farre ◽  
L. Sanchez de Miguel ◽  
C. Caramelo ◽  
J. Gomez-Macias ◽  
R. Garcia ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Growth Inhibitor ◽  
No Production ◽  
Alternative Mechanism ◽  
Competitive Antagonist ◽  
Autocrine Control ◽  
Bovine Endothelial Cells

Nitric oxide (NO) is a growth inhibitor for diverse cellular types. In the present study, we have found that the inhibition of NO production in bovine endothelial cells by an L-arginine competitive antagonist induces DNA replication and promotes the transition from prereplicative to replicative phases of the endothelial cell cycle and an increase in c-myc and c-fos oncogene-encoded protein expression. The inhibition of NO generation had, however, a markedly different outcome depending on the state of confluence of the cells, i.e., proliferation was found in subconfluent cells, whereas apoptosis occurred in confluent cells. Moreover, Western blot analysis revealed differences in the constitutive NO synthase expression in proliferating compared with growth-arrested cells. In conclusion, these results disclose an alternative mechanism of endothelial cell apoptosis at the confluent state, which is related to NO inhibition. Moreover, the fact that the apoptotic phenomenon occurred in the presence of growth factors indicates the existence of apoptotic mechanisms that do not require growth factor deprivation.

Role of G proteins in shear stress-mediated nitric oxide production by endothelial cells

1994 ◽  
Vol 267 (3) ◽  
pp. C753-C758 ◽  
Author(s):  
M. J. Kuchan ◽  
H. Jo ◽  
J. A. Frangos
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Shear Stress ◽  
G Protein ◽  
G Proteins ◽  
No Production ◽  
Dependent Manner ◽  
Protein Activation ◽  
Synthase Inhibitor

Exposure of cultured endothelial cells to shear stress resulting from well-defined fluid flow stimulates the production of nitric oxide (NO). We have established that an initial burst in production is followed by sustained steady-state NO production. The signal transduction events leading to this stimulation are not well understood. In the present study, we examined the role of regulatory guanine nucleotide binding proteins (G proteins) in shear stress-mediated NO production. In endothelial cells not exposed to shear stress, AIF4-, a general activator of G proteins, markedly elevated the production of guanosine 3',5'-cyclic monophosphate (cGMP). Pretreatment with NO synthase inhibitor N omega-nitro-L-arginine completely blocked this stimulation. Incubation with guanosine 5'-O-(2-thiodiphosphate) (GDP beta S), a general G protein inhibitor, blocked the flow-mediated burst in cGMP production in a dose-dependent manner. Likewise, GDP beta S inhibited NOx (NO2 + NO3) production for the 1st h. However, inhibition was not detectable between 1 and 3 h. Pertussis toxin (PTx) had no effect on the shear response at any time point. The burst in NO production caused by a change in shear stress appears to be dependent on a PTx-refractory G protein. Sustained shear-mediated production is independent of G protein activation.

The role of caveolin-1 in PCB77-induced eNOS phosphorylation in human-derived endothelial cells

2007 ◽  
Vol 293 (6) ◽  
pp. H3340-H3347 ◽  
Author(s):  
Eun Jin Lim ◽  
Eric J. Smart ◽  
Michal Toborek ◽  
Bernhard Hennig
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Dna Binding ◽  
Vascular Endothelial Cell ◽  
Vascular Endothelial ◽  
Human Vascular Endothelial Cell ◽  
Caveolin 1 ◽  
Enos Phosphorylation

Polychlorinated biphenyls (PCBs) may contribute to the pathology of atherosclerosis by activating inflammatory responses in vascular endothelial cells. Endothelial nitric oxide synthase (eNOS) is colocalized with caveolae and is a critical regulator of vascular homeostasis. PCBs may be proatherogenic by causing dysfunctional eNOS signaling. The objective of this study was to investigate the role of caveolin-1 in PCB-induced endothelial dysfunction with a focus on mechanisms associated with eNOS signaling. Cells derived from an immortalized human vascular endothelial cell line were treated with PCB77 to study nitrotyrosine formation through eNOS signaling. Phosphorylation studies of eNOS, caveolin-1, and kinases, such as Src, phosphatidylinositol 3-kinase (PI3K), and Akt, were conducted in cells containing either functional or small-interfering RNA-silenced caveolin-1 protein. We also investigated caveolin-1-regulated mechanisms associated with PCB-induced markers of peroxynitrite formation and DNA binding of NF-κB. Cellular exposure to PCB77 increased eNOS phosphorylation and nitric oxide production, as well as peroxynitrite levels. A subsequent PCB-induced increase in NF-κB DNA binding may have implications in oxidative stress-mediated inflammatory mechanisms. The activation of eNOS by PCB77 treatment was blocked by inhibitors of the Src/PI3K/Akt pathway. PCB77 also increased phosphorylation of caveolin-1, indicating caveolae-dependent endocytosis. Caveolin-1 silencing abolished both the PCB-stimulated Akt and eNOS phosphorylation, suggesting a regulatory role of caveolae in PCB-induced eNOS signaling. These findings suggest that PCB77 induces eNOS phosphorylation in endothelial cells through a Src/PI3K/Akt-dependent mechanism, events regulated by functional caveolin-1. Our data provide evidence that caveolae may play a critical role in regulating vascular endothelial cell activation and toxicity induced by persistent environmental pollutants such as coplanar PCBs.

Intercellular calcium signaling and nitric oxide feedback during constriction of rabbit renal afferent arterioles

2007 ◽  
Vol 292 (4) ◽  
pp. F1124-F1131 ◽  
Author(s):  
T. R. Uhrenholt ◽  
J. Schjerning ◽  
P. M. Vanhoutte ◽  
B. L. Jensen ◽  
O. Skøtt
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Smooth Muscle ◽  
Endothelial Cell ◽  
Smooth Muscle Cells ◽  
Muscle Cells ◽  
Calcium Sensitivity ◽  
No Production ◽  
Contractile Apparatus ◽  
Afferent Arterioles

Vasoconstriction and increase in the intracellular calcium concentration ([Ca2+]i) of vascular smooth muscle cells may cause an increase of endothelial cell [Ca2+]i, which, in turn, augments nitric oxide (NO) production and inhibits smooth muscle cell contraction. This hypothesis was tested in microperfused rabbit renal afferent arterioles, using fluorescence imaging microscopy with the calcium-sensitive dye fura-2 and the NO-sensitive dye 4-amino-5-methylamino-2′,7′-difluorescein. Both dyes were loaded into smooth muscle and endothelium. Depolarization with 100 mmol/l KCl led to a transient vasoconstriction which was converted into a sustained response by N-nitro-l-arginine methyl ester (l-NAME). Depolarization increased smooth muscle cell [Ca2+]ifrom 162 ± 15 nmol/l to a peak of 555 ± 70 nmol/l ( n = 7), and this response was inhibited by 80% by the l-type calcium channel blocker calciseptine. After a delay of 10 s, [Ca2+]iincreased in endothelial cells immediately adjacent to reactive smooth muscle cells, and this calcium wave spread in a nonregenerative fashion laterally into the endothelial cell layer with a velocity of 1.2 μm/s. Depolarization with 100 mmol/l KCl led to a significant increase in NO production ([NO]i) which was inhibited by l-NAME ( n = 5). Acetylcholine caused a rapid increase in endothelial [Ca2+]i, which did not transfer to the smooth muscle cells. l-NAME treatment did not affect changes in smooth muscle [Ca2+]iafter depolarization, but it did increase the calcium sensitivity of the contractile apparatus. We conclude that depolarization increases smooth muscle [Ca2+]iwhich is transferred to the endothelial cells and stimulates NO production which curtails vasoconstriction by reducing the calcium sensitivity of the contractile apparatus.

scholarly journals MAGI1 Mediates eNOS Activation and NO Production in Endothelial Cells in Response to Fluid Shear Stress

Cells ◽  
2019 ◽  
Vol 8 (5) ◽  
pp. 388 ◽  
Author(s):  
Kedar Ghimire ◽  
Jelena Zaric ◽  
Begoña Alday-Parejo ◽  
Jochen Seebach ◽  
Mélanie Bousquenaud ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Shear Stress ◽  
Endothelial Cell ◽  
Protein Kinase ◽  
Fluid Shear Stress ◽  
Adaptor Protein ◽  
No Production ◽  
Fluid Shear ◽  
Enos Phosphorylation

Fluid shear stress stimulates endothelial nitric oxide synthase (eNOS) activation and nitric oxide (NO) production through multiple kinases, including protein kinase A (PKA), AMP-activated protein kinase (AMPK), AKT and Ca2+/calmodulin-dependent protein kinase II (CaMKII). Membrane-associated guanylate kinase (MAGUK) with inverted domain structure-1 (MAGI1) is an adaptor protein that stabilizes epithelial and endothelial cell-cell contacts. The aim of this study was to assess the unknown role of endothelial cell MAGI1 in response to fluid shear stress. We show constitutive expression and co-localization of MAGI1 with vascular endothelial cadherin (VE-cadherin) in endothelial cells at cellular junctions under static and laminar flow conditions. Fluid shear stress increases MAGI1 expression. MAGI1 silencing perturbed flow-dependent responses, specifically, Krüppel-like factor 4 (KLF4) expression, endothelial cell alignment, eNOS phosphorylation and NO production. MAGI1 overexpression had opposite effects and induced phosphorylation of PKA, AMPK, and CAMKII. Pharmacological inhibition of PKA and AMPK prevented MAGI1-mediated eNOS phosphorylation. Consistently, MAGI1 silencing and PKA inhibition suppressed the flow-induced NO production. Endothelial cell-specific transgenic expression of MAGI1 induced PKA and eNOS phosphorylation in vivo and increased NO production ex vivo in isolated endothelial cells. In conclusion, we have identified endothelial cell MAGI1 as a previously unrecognized mediator of fluid shear stress-induced and PKA/AMPK dependent eNOS activation and NO production.

scholarly journals Serum Containing Tao-Hong-Si-Wu Decoction Induces Human Endothelial Cell VEGF Production via PI3K/Akt-eNOS Signaling

2013 ◽  
Vol 2013 ◽  
pp. 1-9 ◽  
Author(s):  
DengKe Yin ◽  
ZhuQing Liu ◽  
DaiYin Peng ◽  
Ye Yang ◽  
XiangDong Gao ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Umbilical Vein ◽  
No Production ◽  
Hydroxysafflor Yellow A ◽  
Vegf Expression ◽  
Pi3k Inhibitor Ly294002 ◽  
Ischemic Disease ◽  
Umbilical Vein Endothelial Cells

Tao-Hong-Si-Wu decoction (TSD) is a famous traditional Chinese medicine (TCM) and widely used for ischemic disease in China. TSD medicated serum was prepared after oral administration of TSD (1.6 g/kg) twice a day for 3 days in rats. TSD medicated serum induced human umbilical vein endothelial cells (HUVECs) proliferation, VEGF secretion, and nitric oxide (NO) production. These promoted effects of TSD were partly inhibited by treatment with PI3K inhibitor (LY294002) or eNOS inhibitor (L-NAME), respectively, and completely inhibited by treatment with LY294002 and L-NAME simultaneously. Western blot analysis findings further indicated that TSD medicated serum upregulated p-Akt and p-eNOS expressions, which were significantly inhibited by LY294002 or L-NAME and completely inhibited by both LY294002 and L-NAME; these results indicated that TSD medicated serum induced HUVECs VEGF expression via PI3K/Akt-eNOS signaling. TSD medicated serum contains hydroxysafflor yellow A, ferulic acid, and ligustilide detected by UPLC with standards, so these effect of TSD medicated serum may be associated with these three active compounds absorbed in serum.

Role of cytoskeleton in shear stress-induced endothelial nitric oxide production

1997 ◽  
Vol 273 (1) ◽  
pp. H347-H355 ◽  
Author(s):  
H. L. Knudsen ◽  
J. A. Frangos
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Shear Stress ◽  
Plasma Membrane ◽  
Cytochalasin D ◽  
Induced Response ◽  
No Production ◽  
Dependent Manner ◽  
Mechanochemical Transduction

To study the role of the cytoskeleton in mechanochemical transduction, human umbilical vein endothelial cells were exposed to cytoskeleton-disrupting or -stabilizing agents, and the flow-induced production of nitric oxide (NO) as monitored by intracellular levels of guanosine 3',5'-cyclic monophosphate (cGMP) was examined. A shear stress of 20 dyn/cm2 elevated cGMP levels approximately twofold relative to basal (stationary) levels (1.9 +/- 0.1 pmol cGMP in stationary controls; P < 0.01). Treatment with 1 microM phalloidin and 0.5 microM cytochalasin D did not significantly affect the flow-induced response (1.77 +/- 0.23 and 2.89 +/- 0.18 pmol cGMP in stationary controls, respectively), whereas disruption of microtubules with 0.5 microM colchicine significantly elevated the response (3.64 +/- 0.18 pmol cGMP in stationary controls; P < 0.01). The NO synthase inhibitor NG-amino-L-arginine abrogated all flow-induced elevations of cGMP, indicating that increased cGMP levels were mediated by NO. Cytoskeletal disruption with 0.2 microM cytochalasin D or 0.5 microM colchicine did not alter cGMP levels in response to 10 nM bradykinin. The role of the plasma membrane in mechanochemical transduction was examined by treatment with cholesteryl hemisuccinate, which attenuated the flow-induced response in a dose-dependent manner. In conclusion, the pathways of flow- and bradykinin-mediated NO production in endothelial cells did not require actin filament turnover or intact actin or microtubule cytoskeletons, and cholesterol, possibly by stiffening the plasma membrane, attenuated the flow response.

Podokinesis in endothelial cell migration: role of nitric oxide

1998 ◽  
Vol 274 (1) ◽  
pp. C236-C244 ◽  
Author(s):  
Eisei Noiri ◽  
Eugene Lee ◽  
Jacqueline Testa ◽  
James Quigley ◽  
David Colflesh ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Cell Migration ◽  
Endothelial Cell ◽  
No Synthase ◽  
Endothelial Cell Migration ◽  
Guidance Cues ◽  
Cell Matrix ◽  
Cell Matrix Adhesion

Previously, we demonstrated the role of nitric oxide (NO) in transforming epithelial cells from a stationary to locomoting phenotype [E. Noiri, T. Peresleni, N. Srivastava, P. Weber, W. F. Bahou, N. Peunova, and M. S. Goligorsky. Am. J. Physiol. 270 ( Cell Physiol. 39): C794–C802, 1996] and its permissive function in endothelin-1-stimulated endothelial cell migration (E. Noiri, Y. Hu, W. F. Bahou, C. Keese, I. Giaever, and M. S. Goligorsky. J. Biol. Chem. 272: 1747–1753, 1997). In the present study, the role of functional NO synthase in executing the vascular endothelial growth factor (VEGF)-guided program of endothelial cell migration and angiogenesis was studied in two independent experimental settings. First, VEGF, shown to stimulate NO release from simian virus 40-immortalized microvascular endothelial cells, induced endothelial cell transwell migration, whereas N G-nitro-l-arginine methyl ester (l-NAME) or antisense oligonucleotides to endothelial NO synthase suppressed this effect of VEGF. Second, in a series of experiments on endothelial cell wound healing, the rate of VEGF-stimulated cell migration was significantly blunted by the inhibition of NO synthesis. To gain insight into the possible mode of NO action, we next addressed the possibility that NO modulates cell matrix adhesion by performing impedance analysis of endothelial cell monolayers subjected to NO. The data showed the presence of spontaneous fluctuations of the resistance in ostensibly stationary endothelial cells. Spontaneous oscillations were induced by NO, which also inhibited cell matrix adhesion. This process we propose to term “podokinesis” to emphasize a scalar form of micromotion that, in the presence of guidance cues, e.g., VEGF, is transformed to a vectorial movement. In conclusion, execution of the program for directional endothelial cell migration requires two coexisting messages: NO-induced podokinesis (scalar motion) and guidance cues, e.g., VEGF, which imparts a vectorial component to the movement. Such a requirement for the dual signaling may explain a mismatch in the demand and supply with newly formed vessels in different pathological states accompanied by the inhibition of NO synthase.

scholarly journals Protective Role of Nitric Oxide in a Model of Thrombotic Microangiopathy in Rats

2001 ◽  
Vol 12 (10) ◽  
pp. 2088-2097
Author(s):  
JING SHAO ◽  
TOSHIO MIYATA ◽  
KOEI YAMADA ◽  
NORIO HANAFUSA ◽  
TAKEHIKO WADA ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Thrombotic Microangiopathy ◽  
Periodic Acid ◽  
Minor Role ◽  
Protective Mechanism ◽  
No Production ◽  
Nos Inhibitor ◽  
Inducible Nos

Abstract. A new model of thrombotic microangiopathy (TMA) was previously developed, and it was demonstrated that endothelial nitric oxide (NO) synthase (NOS) is upregulated in glomeruli in this model. It was hypothesized that the synthesis of NO, a potent vasodilator and platelet inhibitory factor, is induced as a defense mechanism. The goal of this study was to clarify the role of NO in this model.Ex vivoexperiments using Western blotting and functional assays demonstrated upregulation of endothelial NOS in isolated glomeruli from TMA rats. Inin vivoexperiments, five groups of rats were studied, including rats with TMA treated with vehicle,NG-nitro-L-arginine methyl ester (L-NAME) (a NOS inhibitor), or L-N6-(1-iminoethyl)lysine (L-NIL) (a specific inducible NOS inhibitor) and normal control rats treated with vehicle or L-NAME. Blood urea nitrogen levels, BP, urinary nitrate/nitrite excretion, and proteinuria were measured. Histologic assessments using periodic acid-Schiff staining and immunohistologic studies with markers for endothelium, platelets, fibrin, cell proliferation, and apoptosis were also performed. L-NAME inhibition of NO synthesis in rats with TMA resulted in more severe glomerular and tubulointerstitial injury, which was accompanied by thrombus formation and a marked loss of endothelial cells, with more apoptotic cells. These changes were associated with severe renal function deterioration. In contrast, these features were less pronounced in the vehicle- or L-NIL-treated rats with TMA and were absent in the control animals. In conclusion, inhibition of NO production in this model of TMA markedly exacerbated renal injury. The absence of effects with L-NIL treatment suggests a minor role for inducible NOS in this model. These results suggest that production of NO, most likely by endothelial cells, is an important protective mechanism in TMA.

scholarly journals Bradykinin stimulation of nitric oxide production is not sufficient for gamma-globin induction

2014 ◽  
Vol 142 (3-4) ◽  
pp. 189-196 ◽  
Author(s):  
Vladan Cokic ◽  
Tijana Suboticki ◽  
Bojana Beleslin-Cokic ◽  
Milos Diklic ◽  
Pavle Milenkovic ◽  
...  
Keyword(s):  
Gene Expression ◽  
Nitric Oxide ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Globin Gene ◽  
No Production ◽  
Dependent Manner ◽  
Erythroid Progenitors ◽  
Globin Gene Expression ◽  
Stimulation Of

Introduction. Hydroxycarbamide, used in therapy of hemoglobinopathies, enhances nitric oxide (NO) production both in primary human umbilical vein endothelial cells (HUVECs) and human bone marrow endothelial cell line (TrHBMEC). Moreover, NO increases ?-globin and fetal hemoglobin levels in human erythroid progenitors. Objective. In order to find out whether simple physiologic stimulation of NO production by components of hematopoietic microenvironment can increase ?-globin gene expression, the effects of NO-inducer bradykinin were examined in endothelial cells. Methods. The study was performed in co-cultures of human erythroid progenitors, TrHBMEC and HUVECs by ozone-based chemiluminescent determination of NO and real-time quantitative RT-PCR. Results. In accordance with previous reports, the endogenous factor bradykinin increased endothelial cell production of NO in a dose- and time-dependent manner (0.1-0.6 ?M up to 30 minutes). This induction of NO in HUVECs and TrHBMEC by bradykinin was blocked by competitive inhibitors of NO synthase (NOS), demonstrating NOS-dependence. It has been shown that bradykinin significantly reduced endothelial NOS (eNOS) mRNA level and eNOS/?-actin ratio in HUVEC (by twofold). In addition, bradykinin failed to increase ?-globin mRNA expression in erythroid progenitors only, as well as in co-culture studies of erythroid progenitors with TrHBMEC and HUVEC after 24 hours of treatment. Furthermore, bradykinin did not induce ?/? globin ratio in erythroid progenitors in co-cultures with HUVEC. Conclusion. Bradykinin mediated eNOS activation leads to short time and low NO production in endothelial cells, insufficient to induce ?-globin gene expression. These results emphasized the significance of elevated and extended NO production in augmentation of ?-globin gene expression.

Sign in / Sign up

Export Citation Format

Share Document