scholarly journals Germline GATA2 variant disrupting endothelial eNOS cell function and angiogenesis can be restored by c-Jun/AP-1 upregulation

Haematologica ◽  
2021 ◽  
Author(s):  
Giulio Purgatorio ◽  
Elisa Piselli ◽  
Giuseppe Guglielmini ◽  
Emanuela Falcinelli ◽  
Loredana Bury ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Transcription Factor ◽  
Ad Hoc ◽  
Cell Function ◽  
Clinical Manifestations ◽  
Enos Gene ◽  
No Production ◽  
Enos Expression ◽  
No Donor ◽  
Gata2 Deficiency

GATA2 is a transcription factor with key roles in hematopoiesis. Germline GATA2 gene variants have been associated with several inherited and acquired hematologic disorders, including myelodysplastic syndromes. Among the spectrum of GATA2 deficiency-associated manifestations thrombosis has been reported in 25% of patients, but the mechanisms are unknown. GATA2 was shown to be involved in endothelial nitric oxide synthase (eNOS) regulation and vascular development. We assessed eNOS expression and angiogenesis in patients with GATA2-deficiency. Platelets and blood outgrowth endothelial cells (BOEC) from GATA2-variant carriers showed impaired NO-production and reduction of eNOS mRNA and protein expression and of eNOS activity. GATA2 binding to the eNOS gene was impaired in BOEC from GATA2-deficiency patients, differently from control BOEC. GATA2-deficiency BOEC showed also defective angiogenesis, which was completely restored by treatment with the NO-donor S-nitroso-N-acetylpenicillamine (SNAP). Atorvastatin, but not resveratrol, largely restored eNOS expression, NO biosynthesis and neoangiogenesis in GATA2-deficient BOEC by a mechanism involving increased expression of the eNOS transcription factor AP-1/c-JUN, replacing GATA2 when the latter is inactive. Our results unravel a possible thrombogenic mechanism of GATA2 mutations, definitely establish the regulation of eNOS by GATA2 in endothelial cells and show that endothelial angiogenesis is strictly dependent on the eNOS/NO axis. Given the ability of atorvastatin to restore NO production and angiogenesis by GATA2-deficient endothelial cells, the preventive effect of atorvastatin on thrombotic events and possibly on other clinical manifestations of the syndrome related to deranged angiogenesis should be explored in patients with GATA2-deficiency in an ad hoc designed clinical trial.

Prunella vulgaris L. Upregulates eNOS Expression in Human Endothelial Cells

2010 ◽  
Vol 38 (03) ◽  
pp. 599-611 ◽  
Author(s):  
Ning Xia ◽  
Larissa Bollinger ◽  
Katja Steinkamp-Fenske ◽  
Ulrich Förstermann ◽  
Huige Li
Keyword(s):  
Gene Expression ◽  
Endothelial Cells ◽  
Therapeutic Potential ◽  
Chinese Herbs ◽  
Enos Gene ◽  
No Production ◽  
Prunella Vulgaris ◽  
Enos Expression ◽  
Salviae Miltiorrhizae ◽  
Salviae Miltiorrhizae Radix

The purported effects of "circulation-improving" herbs used in traditional Chinese medicine (TCM) show striking similarities with the vascular actions of nitric oxide (NO) produced by the endothelial NO synthase (eNOS). We have previously reported that Salviae miltiorrhizae radix and Zizyphi spinosae semen upregulate eNOS expression. In the present study, we studied the effect on eNOS gene expression of 15 Chinese herbs with potential effects on the vasculature, and identified Prunella vulgaris L. (PVL) (flowering spike) as a potent eNOS-upregulating agent. In EA.hy 926 cells, a cell line derived from human umbilical vein endothelial cells (HUVEC), an aqueous extract of PVL increased eNOS promoter activity, eNOS mRNA and protein expressions, as well as NO production in concentration- and time-dependent manners. We have previously shown that ursolic acid (a constituent of Salviae miltiorrhizae radix), betulinic acid (a compound present in Zizyphi spinosae semen), luteolin and cynaroside (ingredients of artichoke, Cynara scolymus L.) are capable of enhancing eNOS gene expression. These compounds are also present in significant quantities in PVL. Thus, PVL contains active principles that stimulate human eNOS gene expression, and such compounds may have therapeutic potential against cardiovascular diseases.

scholarly journals Hydrogen Sulfide Increases Nitric Oxide Production and Subsequent S-Nitrosylation in Endothelial Cells

2014 ◽  
Vol 2014 ◽  
pp. 1-8 ◽  
Author(s):  
Ping-Ho Chen ◽  
Yaw-Syan Fu ◽  
Yun-Ming Wang ◽  
Kun-Han Yang ◽  
Danny Ling Wang ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Hydrogen Sulfide ◽  
Protein Kinase ◽  
Fluorescent Probe ◽  
Acid Methyl Ester ◽  
High Specificity ◽  
Protein S ◽  
No Production ◽  
No Donor

Hydrogen sulfide (H2S) and nitric oxide (NO), two endogenous gaseous molecules in endothelial cells, got increased attention with respect to their protective roles in the cardiovascular system. However, the details of the signaling pathways between H2S and NO in endothelia cells remain unclear. In this study, a treatment with NaHS profoundly increased the expression and the activity of endothelial nitric oxide synthase. Elevated gaseous NO levels were observed by a novel and specific fluorescent probe, 5-amino-2-(6-hydroxy-3-oxo-3H-xanthen-9-yl)benzoic acid methyl ester (FA-OMe), and quantified by flow cytometry. Further study indicated an increase of upstream regulator for eNOS activation, AMP-activated protein kinase (AMPK), and protein kinase B (Akt). By using a biotin switch, the level of NO-mediated protein S-nitrosylation was also enhanced. However, with the addition of the NO donor, NOC-18, the expressions of cystathionine-γ-lyase, cystathionine-β-synthase, and 3-mercaptopyruvate sulfurtransferase were not changed. The level of H2S was also monitored by a new designed fluorescent probe, 4-nitro-7-thiocyanatobenz-2-oxa-1,3-diazole (NBD-SCN) with high specificity. Therefore, NO did not reciprocally increase the expression of H2S-generating enzymes and the H2S level. The present study provides an integrated insight of cellular responses to H2S and NO from protein expression to gaseous molecule generation, which indicates the upstream role of H2S in modulating NO production and protein S-nitrosylation.

scholarly journals Reduction of electrical coupling between microvascular endothelial cells by NO depends on connexin37

2009 ◽  
Vol 297 (1) ◽  
pp. H93-H101 ◽  
Author(s):  
Rebecca L. McKinnon ◽  
Michael L. Bolon ◽  
Hong-Xing Wang ◽  
Scott Swarbreck ◽  
Gerald M. Kidder ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Electrical Coupling ◽  
Electrical Current ◽  
Microvascular Endothelial Cells ◽  
No Production ◽  
No Donor ◽  
Junction Protein ◽  
Arteriolar Wall ◽  
Microvascular Endothelial ◽  
In Cells

We have previously shown that increased nitric oxide (NO) production in sepsis impairs arteriolar-conducted vasoconstriction cGMP independently and that the gap junction protein connexin (Cx) 37 is required for this conducted response. In the present study, we hypothesized that NO impairs interendothelial electrical coupling in sepsis by targeting Cx37. We examined the effect of exogenous NO on coupling in monolayers of cultured microvascular endothelial cells derived from the hindlimb skeletal muscle of wild-type (WT), Cx37 null, Cx40 null, and Cx43G60S (nonfunctional mutant) mice. To assess coupling, we measured the spread of electrical current injected in the monolayer and calculated the monolayer intercellular resistance (inverse measure of coupling). The NO donor 2,2′-(hydroxynitrosohydrazino) bis-ethanamine (DETA) rapidly and reversibly reduced coupling in cells from WT mice, cGMP independently. NO scavenger HbO2 did not affect baseline coupling, but it eliminated DETA-induced reduction in coupling. Reduced coupling in response to DETA was also seen in cells from Cx40 null and Cx43G60S mice, but not in cells from Cx37 null mice. DETA did not alter the expression of Cx37, Cx40, and Cx43 in WT cells analyzed by immunoblotting and immunofluorescence. Furthermore, neither the peroxynitrite scavenger 5,10,15,20-tetrakis(4-sulfonatophenyl)porphyrinato iron (III), superoxide scavenger Mn(III)tetrakis(4-benzoic acid)porphyrin chloride, nor preloading of WT cells with the antioxidant ascorbate affected this reduction. We conclude that NO-induced reduction of electrical coupling between microvascular endothelial cells depends on Cx37 and propose that NO in sepsis impairs arteriolar-conducted vasoconstriction by targeting Cx37 within the arteriolar wall.

scholarly journals Salidroside Stimulates Mitochondrial Biogenesis and Protects against H2O2-Induced Endothelial Dysfunction

2014 ◽  
Vol 2014 ◽  
pp. 1-13 ◽  
Author(s):  
Shasha Xing ◽  
Xiaoyan Yang ◽  
Wenjing Li ◽  
Fang Bian ◽  
Dan Wu ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Transcription Factor ◽  
Endothelial Dysfunction ◽  
Mitochondrial Biogenesis ◽  
Umbilical Vein ◽  
Adenosine Monophosphate ◽  
No Production ◽  
Peroxisome Proliferator ◽  
Atp Production

Salidroside (SAL) is an active component ofRhodiola roseawith documented antioxidative properties. The purpose of this study is to explore the mechanism of the protective effect of SAL on hydrogen peroxide- (H2O2-) induced endothelial dysfunction. Pretreatment of the human umbilical vein endothelial cells (HUVECs) with SAL significantly reduced the cytotoxicity brought by H2O2. Functional studies on the rat aortas found that SAL rescued the endothelium-dependent relaxation and reduced superoxide anion (O2∙-) production induced by H2O2. Meanwhile, SAL pretreatment inhibited H2O2-induced nitric oxide (NO) production. The underlying mechanisms involve the inhibition of H2O2-induced activation of endothelial nitric oxide synthase (eNOS), adenosine monophosphate-activated protein kinase (AMPK), and Akt, as well as the redox sensitive transcription factor, NF-kappa B (NF-κB). SAL also increased mitochondrial mass and upregulated the mitochondrial biogenesis factors, peroxisome proliferator-activated receptor gamma-coactivator-1alpha (PGC-1α), and mitochondrial transcription factor A (TFAM) in the endothelial cells. H2O2-induced mitochondrial dysfunction, as demonstrated by reduced mitochondrial membrane potential (Δψm) and ATP production, was rescued by SAL pretreatment. Taken together, these findings implicate that SAL could protect endothelium against H2O2-induced injury via promoting mitochondrial biogenesis and function, thus preventing the overactivation of oxidative stress-related downstream signaling pathways.

P1, P4-diadenosine 5′-tetraphosphate induced DNA synthesis in mechanically injured cultured endothelial cells

2003 ◽  
Vol 217 (1) ◽  
pp. 21-26 ◽  
Author(s):  
J E Woodell ◽  
M LaBerge ◽  
E M Langan ◽  
R H Hilderman
Keyword(s):  
Endothelial Cells ◽  
Dna Synthesis ◽  
Fold Increase ◽  
Companion Paper ◽  
No Production ◽  
No Donor ◽  
Pharmacological Agents ◽  
Aortic Endothelial Cells ◽  
Cell Dysfunction ◽  
Cell Migration And Proliferation

Rapid re-endothelialization following balloon angioplasty can reduce restenosis by inhibiting smooth muscle cell migration and proliferation. However, formation of a neointima layer following angioplasty can be inhibited due to endothelial cell dysfunction and denudation. In a companion paper, it has been illustrated that mechanical loading causes a decrease in DNA synthesis in bovine aortic endothelial cells (BAECs) thus rendering them dysfunctional. The purpose of this study was to overcome BAEC dysfunction by incubation with pharmacological agents to increase DNA synthesis. Previous studies demonstrated that the adenosine dinucleotides Ap4A and Ap2A induced nitric oxide (NO) production from BAEC while Ap3A, Ap5A and Ap6A did not. This paper demonstrates that Ap4 and Ap2 A induce a 1.46- and 1.16-fold increase in DNA synthesis in mechanically stressed BAECs respectively, while Ap3A, Ap5A and Ap6A do not. Additionally, NOC-18, a slow NO release NO donor, significantly increases DNA synthesis in mechanically stressed BAECs without affecting unloaded cells. These results are consistent with NO inducing DNA synthesis in mechanically stressed BAECs.

Altered l-arginine metabolism results in increased nitric oxide release from uraemic endothelial cells

2002 ◽  
Vol 103 (1) ◽  
pp. 31-41 ◽  
Author(s):  
Raj C. THURAISINGHAM ◽  
Norman B. ROBERTS ◽  
Mark WILKES ◽  
David I. NEW ◽  
A. Claudio MENDES-RIBEIRO ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
In Vivo Model ◽  
No Production ◽  
Arginine Metabolism ◽  
Enos Expression ◽  
Nitric Oxide Release ◽  
Excessive Consumption ◽  
And Control

Results regarding the nitric oxide (NO) system in uraemia are contradictory. l-Arginine, the precursor of NO, is also metabolized by arginase to form ornithine and urea. In the present study, endothelial NO production and arginine metabolism in uraemia were assessed. In addition an in vivo model was used to examine excess consumption of NO in uraemia. NO and amino acid measurements were made from basal and stimulated (by bradykinin) uraemic and control endothelial cells. Reverse-transcriptase PCR was used to assess endothelial NO synthase (eNOS) and inducible NOS (iNOS) expression. Finally, aortae of uraemic rats were stained for nitrotyrosine (a marker of peroxynitrite). Basal uraemic cells produced more NO than the control cells. l-Arginine levels were greater in uraemic (supernatants/cells), but ornithine levels were higher in control (supernatants/cells). Following stimulation, NO levels in supernatants were similar, but the rise in NO production was greater in control compared with uraemic cells; l-arginine levels still remained higher in uraemic supernatants/cells. Differences in ornithine concentration (supernatants/cells) disappeared following bradykinin stimulation, due to a rise in ornithine levels in the uraemic group. There was no difference in eNOS expression, nor was iNOS detected in either group. Only aortae from uraemic rats showed evidence for nitrotyrosine staining. These studies demonstrated increased basal NO release in uraemic endothelial cells, perhaps by inhibition of arginase and hence diversion of arginine to the NO pathway. The increased NO produced under basal conditions may be inactive due to excessive consumption, resulting in peroxynitrite formation. Interestingly, bradykinin appears to restore arginase activity in uraemia, resulting in normalization of NO production.

scholarly journals Activation of Endothelial Nitric Oxide Synthase by the Angiotensin II Type 1 Receptor

Endocrinology ◽  
2006 ◽  
Vol 147 (12) ◽  
pp. 5914-5920 ◽  
Author(s):  
Hiroyuki Suzuki ◽  
Kunie Eguchi ◽  
Haruhiko Ohtsu ◽  
Sadaharu Higuchi ◽  
Sudhir Dhobale ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Angiotensin Ii ◽  
Endothelial Dysfunction ◽  
Enos Gene ◽  
No Production ◽  
Hypertrophic Response ◽  
Cgmp Production ◽  
At1 Antagonist

Enhanced angiotensin II (AngII) action has been implicated in endothelial dysfunction that is characterized as decreased nitric oxide availability. Although endothelial cells have been reported to express AngII type 1 (AT1) receptors, the exact role of AT1 in regulating endothelial NO synthase (eNOS) activity remains unclear. We investigated the possible regulation of eNOS through AT1 in bovine aortic endothelial cells (BAECs) and its functional significance in rat aortic vascular smooth muscle cells (VSMCs). In BAECs infected with adenovirus encoding AT1 and in VSMCs infected with adenovirus encoding eNOS, AngII rapidly stimulated phosphorylation of eNOS at Ser1179. This was accompanied with increased cGMP production. These effects were blocked by an AT1 antagonist. The cGMP production was abolished by a NOS inhibitor as well. To explore the importance of eNOS phosphorylation, VSMCs were also infected with adenovirus encoding S1179A-eNOS. AngII did not stimulate cGMP production in VSMCs expressing S1179A. However, S1179A was able to enhance basal NO production as confirmed with cGMP production and enhanced vasodilator-stimulated phosphoprotein phosphorylation. Interestingly, S1179A prevented the hypertrophic response similar to wild type in VSMCs. From these data, we conclude that the AngII/AT1 system positively couples to eNOS via Ser1179 phosphorylation in ECs and VSMCs if eNOS and AT1 coexist. However, basal level NO production may be sufficient for prevention of AngII-induced hypertrophy by eNOS expression. These data demonstrate a novel molecular mechanism of eNOS regulation and function and thus provide useful information for eNOS gene therapy under endothelial dysfunction.

scholarly journals Phytoestrogen Genistein Up-Regulates Endothelial Nitric Oxide Synthase Expression Via Activation of cAMP Response Element-Binding Protein in Human Aortic Endothelial Cells

Endocrinology ◽  
2012 ◽  
Vol 153 (7) ◽  
pp. 3190-3198 ◽  
Author(s):  
Hongwei Si ◽  
Jie Yu ◽  
Hongling Jiang ◽  
Hazel Lum ◽  
Dongmin Liu
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Endothelial Nitric Oxide Synthase ◽  
No Production ◽  
Enos Expression ◽  
Camp Response Element ◽  
Aortic Endothelial Cells ◽  
Oxide Synthase ◽  
Endothelial Nitric Oxide ◽  
Human Aortic Endothelial Cells

We previously reported that genistein, a phytoestrogen, up-regulates endothelial nitric oxide synthase (eNOS) and prevents hypertension in rats that are independent of estrogen signaling machinery. However, how genistein regulates eNOS expression is unknown. In the present study, we show that genistein enhanced eNOS expression and NO synthesis in primary human aortic endothelial cells. Inhibition of extracellular signal regulated kinase, phosphoinositol-3 kinase, or protein kinase C did not affect genistein-enhanced eNOS expression and NO synthesis. However, chemical inhibition of protein kinase A (PKA) or adenoviral transfer of the specific endogenous PKA inhibitor gene completely abolished PKA activity and genistein-stimulated eNOS expression and NO production. Accordingly, genistein induced PKA activity and subsequent phosphorylation of cAMP response element (CRE)-binding protein (CREB) at Ser133. Suppression of CREB by small interfering RNA transfection abolished genistein-enhanced eNOS expression and NO production. Consistently, deletion of the CRE site within human eNOS promoter eliminated genistein-stimulated eNOS promoter activity. These findings provide the first evidence to our knowledge that genistein may play a beneficial role in vascular function through targeting the PKA/CREB/eNOS/NO signaling pathway.

scholarly journals Evidence for Production and Functional Activity of Nitric Oxide in Seminiferous Tubules and Blood Vessels of the Human Testis1

1997 ◽  
Vol 82 (12) ◽  
pp. 4154-4161 ◽  
Author(s):  
R. Middendorff ◽  
D. Müller ◽  
S. Wichers ◽  
A. F. Holstein ◽  
M. S. Davidoff
Keyword(s):  
Nitric Oxide ◽  
Blood Vessels ◽  
Sertoli Cells ◽  
Cell Function ◽  
Soluble Guanylyl Cyclase ◽  
Guanosine Monophosphate ◽  
Human Testis ◽  
Seminiferous Tubules ◽  
No Production ◽  
No Donor

Previous studies have demonstrated that nitric oxide (NO) influences Leydig cell function. Here we provide evidence for NO production and activity in seminiferous tubules and blood vessels of the human testis. By immunohistochemistry, the soluble guanylyl cyclase (sGC), the intracellular NO receptor, and the second messenger, cyclic guanosine monophosphate (cGMP), were detected in myofibroblasts of the peritubular lamina propria in Sertoli cells, as well as in endothelial and smooth muscle cells of testicular blood vessels. Performed with isolated tubules and blood vessels, the biological activity of sGC could be proved by cGMP generation in response to treatments with the NO donor, sodium nitroprusside. The endothelial and neuronal subtypes of NO synthase (NOS) were localized immunohistochemically to the same cell types that express sGC and cGMP. In isolated tubules and vessels, the presence of endothelial NOS and neuronal NOS was confirmed by immunoblotting, and NOS activity was demonstrated by decreased cGMP production upon incubation with the NOS inhibitor l-nitro arginine methylester. These findings show that peritubular cells, Sertoli cells, and testicular blood vessels may be sites of NO production and activity, possibly involved in relaxation of seminiferous tubules and blood vessels to modulate sperm transport and testicular blood flow, respectively.

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