Sexual dimorphism in glomerular arginine transport affects nitric oxide generation in old male rats

2009 ◽  
Vol 297 (1) ◽  
pp. F80-F84 ◽  
Author(s):  
Idit F. Schwartz ◽  
Tamara Chernichovski ◽  
Natalia Krishtol ◽  
Avishai Grupper ◽  
Ido Laron ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Ex Vivo ◽  
Male Rats ◽  
Young Males ◽  
Young Females ◽  
Arginine Transport ◽  
Arginine Uptake ◽  
Oxide Synthase ◽  
Endothelial Nitric Oxide ◽  
Old Females

Animal models suggest that decreased renal endothelial nitric oxide synthase (eNOS) activity in old males promotes renal injury, whereas females are protected. We aimed to explore whether aging alters glomerular arginine uptake by CAT-1, the selective arginine supplier to eNOS in rats. Arginine uptake by glomeruli from young males (3 mo) was significantly higher than in young females. Old males (19 mo) exhibited a significant decrease in arginine transport compared with young males, whereas no differences were observed between old and young females. CAT-1 abundance remained unchanged in all experimental groups. The abundance of PKCα (CAT-1 inhibitor) was significantly augmented in young females vs. young males, old vs. young males, and in old females vs. old males. No differences in PKCα content were detected between old and young females. Phosphorylated PKCα was significantly increased in old rats from both genders. αTocopherol, a PKC inhibitor, produced a significant increase in arginine transport and restored NO generation in old males only. Ex vivo incubation of glomeruli from old males with PMA (PKC stimulant) significantly attenuated the effect of tocopherol on arginine uptake. In conclusion, attenuated glomerular arginine transport by CAT-1 contributes to the age-dependent, NO-deficient state in old male rats through upregulation of PKCα. In old females glomerular arginine transport is protected from the effects of PKCα by an unknown mechanism.

scholarly journals Pravastatin induces NO synthesis by enhancing microsomal arginine uptake in healthy and preeclamptic placentas

2019 ◽  
Vol 19 (1) ◽  
Author(s):  
Zita Pánczél ◽  
Zoltán Kukor ◽  
Dorina Supák ◽  
Bence Kovács ◽  
András Kecskeméti ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Hsp90 Inhibitor ◽  
Filtration Method ◽  
Enos Activity ◽  
Arginine Uptake ◽  
New Findings ◽  
Placental Blood ◽  
Oxide Synthase ◽  
Endothelial Nitric Oxide ◽  
Rapid Activation

Abstract Background Pravastatin, a known inducer of endothelial nitric-oxide synthase (eNOS) was demonstrated in human placenta, however the exact mechanism of it’s action is not fully understood. Since placental NO (nitric oxide) synthesis is of primary importance in the regulation of placental blood flow, we aimed to clarify the effects of pravastatin on healthy (n = 6) and preeclamptic (n = 6) placentas (Caucasian participants). Methods The eNOS activity of human placental microsomes was determined by the conversion rate of C14 L-arginine into C14 L-citrulline with or without pravastatin and Geldanamycin. Phosphorylation of eNOS (Ser1177) was investigated by Western blot. Microsomal arginine uptake was measured by a rapid filtration method. Results Pravastatin significantly increased total eNOS activity in healthy (28%, p<0.05) and preeclamptic placentas (32%, p<0.05) using 1 mM Ca2+ promoting the dissociation of a eNOS from it’s inhibitor caveolin. Pravastatin and Geldanamycin (Hsp90 inhibitor) cotreatment increased microsomal eNOS activity. Pravastatin treatment had no significant effects on Ser1177 phosphorylation of eNOS in either healthy or preeclamptic placentas. Pravastatin induced arginine uptake of placental microsomes in both healthy (38%, p < 0.05) and preeclamptic pregnancies (34%, p < 0.05). Conclusions This study provides a novel mechanism of pravastatin action on placental NO metabolism. Pravastatin induces the placental microsomal arginine uptake leading to the rapid activation of eNOS independently of Ser1177 phosphorylation. These new findings may contribute to better understanding of preeclampsia and may also have a clinical relevance.

scholarly journals Interaction of the endothelial nitric oxide synthase with the CAT-1 arginine transporter enhances NO release by a mechanism not involving arginine transport

2005 ◽  
Vol 386 (3) ◽  
pp. 567-574 ◽  
Author(s):  
Chunying LI ◽  
Wei HUANG ◽  
M. Brennan HARRIS ◽  
Jonathan M. GOOLSBY ◽  
Richard C. VENEMA
Keyword(s):  
Nitric Oxide ◽  
Nitric Oxide Synthase ◽  
Endothelial Nitric Oxide Synthase ◽  
Adaptor Protein ◽  
No Production ◽  
Arginine Transport ◽  
No Release ◽  
Oxide Synthase ◽  
Endothelial Nitric Oxide

eNOS (endothelial nitric oxide synthase) catalyses the conversion of L-arginine into L-citrulline and NO. Evidence has been presented previously that eNOS is associated with the CAT (cationic amino acid transporter)-1 arginine transporter in endothelial caveolae, and it has been proposed that eNOS–CAT-1 association facilitates the delivery of extracellular L-arginine to eNOS. Definitive proof of a protein–protein interaction between eNOS and CAT-1 is lacking, however, and it is also unknown whether the two proteins interact directly or via an adaptor protein. In the present study, we raised a polyclonal antibody against CAT-1, and show using reciprocal co-immunoprecipitation protocols that eNOS and CAT-1 do indeed form a complex in BAECs (bovine aortic endothelial cells). In vitro binding assays with GST (glutathione S-transferase)–CAT-1 fusion proteins and eNOS show that the two proteins interact directly and that no single CAT-1 intracellular domain is sufficient to mediate the interaction. Overexpression of CAT-1 in BAECs by adenoviral-mediated gene transfer results in significant increases in both L-arginine uptake and NO production by the cells. However, whereas increased L-arginine transport is reversed completely by the CAT-1 inhibitor, L-lysine, increased NO release is unaltered, suggesting that NO production in this in vitro model is independent of CAT-1-mediated transport. Furthermore, eNOS enzymic activity is increased in lysates of CAT-1-overexpressing cells accompanied by increased phosphorylation of eNOS at Ser-1179 and Ser-635, and decreased association of eNOS with caveolin-1. Taken together, these data suggest that direct interaction of eNOS with CAT-1 enhances NO release by a mechanism not involving arginine transport.

Differential regulation of glomerular arginine transporters (CAT-1 and CAT-2) in lipopolysaccharide-treated rats

2003 ◽  
Vol 284 (4) ◽  
pp. F788-F795 ◽  
Author(s):  
Doron Schwartz ◽  
Idit F. Schwartz ◽  
Ehud Gnessin ◽  
Yoram Wollman ◽  
Tamara Chernichovsky ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Mesangial Cells ◽  
Primary Cultures ◽  
Northern Blotting ◽  
Cationic Amino Acid ◽  
Arginine Transport ◽  
Arginine Uptake ◽  
Endothelial Nitric Oxide ◽  
In Vivo Administration

The decrease in glomerular filtration rate (GFR) that is characteristic of sepsis has been shown to result from inhibition of glomerular endothelial nitric oxide synthase (eNOS) by nitric oxide (NO) generated from the inducible isoform of NOS (iNOS). Although l-arginine is the sole precursor for NO biosynthesis, its intracellular availability in glomeruli from septic animals has never been investigated. Arginine uptake was measured in freshly harvested glomeruli from the following experimental groups: 1) untreated rats; 2) rats pretreated with LPS (4 mg/kg body wt, 4 h before experiments); 3) rats treated with LPS as above with eitherl- N 6-(1-iminoethyl)lysine hydrochloride (l-NIL), a selective iNOS antagonist, or 7-nitroindazole, a selective neuronal NOS antagonist; and 4) rats treated with l-NIL only. Both glomeular and mesangial arginine transport characteristics were found compatible with a y+ system. Arginine uptake was augmented in glomeruli from LPS-treated rats. Treatment with l-NIL completely abolished this effect whereas l-NIL alone had no effect. Similar results were obtained when primary cultures of rat mesangial cells were preincubated with LPS (10 μg/ml for 24 h) with or withoutl-NIL. Using RT-PCR, we found that in vivo administration of LPS resulted in a significant increase in glomerular cationic amino acid transporter-2 (CAT-2) mRNA expression whereas CAT-1 mRNA was undetected. Northern blotting further confirmed a significant increase in glomerular CAT-2 by LPS. In mesangial cells, the expression of both CAT-1 and CAT-2 mRNA was augmented after incubation with LPS. In conclusion, in vivo administration of LPS augments glomerular arginine transport through upregulation of steady-state CAT-2 mRNA while downregulating CAT-1 mRNA. These results may correspond to the changes in glomerular iNOS and eNOS activity in sepsis.

Rosiglitazone improves aortic arginine transport, through inhibition of PKCα, in uremic rats

2008 ◽  
Vol 295 (2) ◽  
pp. F471-F477 ◽  
Author(s):  
Merav Ingbir ◽  
Idit F. Schwartz ◽  
Alexander Shtabsky ◽  
Irina Filip ◽  
Ran Reshef ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Peroxisome Proliferator ◽  
Protein Nitration ◽  
Peroxisome Proliferator Activated Receptor ◽  
Beneficial Effects ◽  
Arginine Transport ◽  
Arginine Uptake ◽  
Ppar Agonists ◽  
Carbamyl Choline ◽  
Endothelial Nitric Oxide

Peroxisome proliferator-activated receptor (PPAR) agonists were shown to inhibit atherosclerosis through augmentation of endothelial nitric oxide synthase (eNOS) activity. In addition, rosiglitazone exerts a beneficial effect in chronic renal failure (CRF). Since l-arginine transport by CAT-1 (the specific arginine transporter for eNOS) is inhibited in uremia, we aimed to explore the effect of rosiglitazone on arginine transport in CRF. Arginine uptake by aortic rings was studied in control animals, rats, 6 wk following 5/6 nephrectomy (CRF) and rats with CRF treated with rosiglitazone. The decrease of arginine transport in CRF was prevented by rosiglitazone. Immunobloting revealed that CAT-1 protein was decreased in CRF but remained unchanged following rosiglitazone administration. Protein content of the membrane fraction of PKCα and phosphorylated CAT-1 increased significantly in CRF, effects that were prevented by rosiglitazone. PKCα phosphorylation was unchanged but significantly attenuated by rosiglitazone in CRF. Ex vivo administration of phorbol-12-myristate-13-acetate to rosiglitazone-treated CRF rats significantly attenuated the effect of rosiglitazone on arginine uptake. The decrease in cGMP response to carbamyl-choline (eNOS agonist) was significantly attenuated by rosiglitazone in CRF. Western blotting and immunohistochemistry analysis revealed that protein nitration was intensified in the endothelium of CRF rats and this was attenuated by rosiglitazone. In conclusion, rosiglitazone prevents the decrease in arginine uptake in CRF through both depletion and inactivation of PKCα. These findings are associated with restoration of eNO generation and attenuation of protein nitration and therefore may serve as a novel mechanism to explain the beneficial effects of rosiglitazone on endothelial function in uremia.

scholarly journals Sustained Maternal Smoking Triggers Endothelial-Mediated Oxidative Stress in the Umbilical Cord Vessels, Resulting in Vascular Dysfunction

Antioxidants ◽  
2021 ◽  
Vol 10 (4) ◽  
pp. 583
Author(s):  
Szabolcs Zahorán ◽  
Péter R. Szántó ◽  
Nikolett Bódi ◽  
Mária Bagyánszki ◽  
József Maléth ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Umbilical Cord ◽  
Maternal Smoking ◽  
Ex Vivo ◽  
Vascular Dysfunction ◽  
Redox Balance ◽  
Heavy Smoking ◽  
Long Term Effect ◽  
Oxide Synthase ◽  
Endothelial Nitric Oxide

Nitric oxide (NO) bioavailability is fundamental in the regulation of redox balance and functionality of the endothelium, especially in the case of the umbilical cord (UC), which has no innervation. The analysis of UC vessel-related complications could serve as a useful tool in the understanding of the pathophysiological mechanisms leading to neonatal cardiovascular disorders. Therefore, the aim of this study was to characterize the mechanisms that rule the severity of prenatal endothelial dysfunction, induced by the long-term effect of maternal smoking. Our analysis describes the initiation and the consequences of endothelial nitric oxide synthase (NOS3) deactivation, along with the up-regulation of possible compensatory pathways, using structural, molecular and biochemical approaches. This study was carried out on both the UC arteries and veins originated from neonates born to non-smoking and heavy-smoking mothers. The alterations stimulated by maternal smoking are vessel-specific and proportional to the level of exposure to harmful materials passed through the placenta. Typically, in the primarily exposed veins, an increased formation of reactive oxygen species and an up-regulation of the highly-efficient NOS2-NO producing pathway were detected. Despite all the extensive structural and functional damages, the ex vivo heat and cadmium ion-treated UC vein pieces still support the potential for stress response.

scholarly journals Increased l-arginine uptake and inducible nitric oxide synthase activity in aortas of rats with heart failure

2001 ◽  
Vol 280 (2) ◽  
pp. H859-H867 ◽  
Author(s):  
Peter B. Stathopulos ◽  
Xiangru Lu ◽  
Ji Shen ◽  
Jeremy A. Scott ◽  
James R. Hammond ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Heart Failure ◽  
Amino Acid ◽  
Nitric Oxide Synthase ◽  
No Production ◽  
Cationic Amino Acid ◽  
Arginine Transport ◽  
Arginine Uptake ◽  
Oxide Synthase

l-Arginine crosses the cell membrane primarily through the system y+ transporter. The aim of this study was to investigate the role of l-arginine transport in nitric oxide (NO) production in aortas of rats with heart failure induced by myocardial infarction. Tumor necrosis factor-α levels in aortas of rats with heart failure were six times higher than in sham rats ( P < 0.01). l-Arginine uptake was increased in aortas of rats with heart failure compared with sham rats ( P < 0.01). Cationic amino acid transporter-2B and inducible (i) nitric oxide synthase (NOS) expression were increased in aortas of rats with heart failure compared with sham rats ( P < 0.05). Aortic strips from rats with heart failure treated with l-arginine but not d-arginine increased NO production ( P < 0.05). The effect ofl-arginine on NO production was blocked byl-lysine, a basic amino acid that shares the same system y+ transporter withl-arginine, and by the NOS inhibitor N G-nitro-l-arginine methyl ester (l-NAME). Treatment with l-lysine andl-NAME in vivo decreased plasma nitrate and nitrite levels in rats with heart failure ( P < 0.05). Our data demonstrate that NO production is dependent on iNOS activity andl-arginine uptake and suggest that l-arginine transport plays an important role in enhanced NO production in heart failure.

scholarly journals Adventitial Expression of Recombinant Endothelial Nitric Oxide Synthase Gene Reverses Vasoconstrictor Effect of Endothelin-1

1999 ◽  
Vol 19 (9) ◽  
pp. 1029-1037 ◽  
Author(s):  
Hisashi Onoue ◽  
Masato Tsutsui ◽  
Leslie Smith ◽  
Timothy O'Brien ◽  
Zvonimir S. Katusic
Keyword(s):  
Nitric Oxide ◽  
Nitric Oxide Synthase ◽  
Receptor Antagonist ◽  
Endothelial Nitric Oxide Synthase ◽  
Ex Vivo ◽  
Enos Gene ◽  
Endothelin 1 ◽  
Basilar Arteries ◽  
Oxide Synthase ◽  
Endothelial Nitric Oxide

The present study was designed to determine the effect of recombinant endothelial nitric oxide synthase (eNOS) gene expression on reactivity of canine basilar arteries to endothelin-1 (ET-1). Experiments were performed ex vivo. The arteries were exposed (30 minutes at 37°C) to adenoviral vectors encoding eNOS gene (AdCMVeNOS) or β-galactosidase reporter gene (AdCMVβ-Gal). Twenty-four hours after transduction, transgene expression was evident mainly in the vascular adventitia. Rings of control (nontransduced), AdCMVβ-Gal- and AdCMVeNOS-transduced arteries with and without endothelium were suspended for isometric tension recording. Levels of guanosine 3′,5′ -cyclic monophosphate (cGMP) were measured by radioimmunoassay. During contractions to uridine 5′-triphosphate, ET-1 (10−10 to 3×10−9 mol/L) caused further increase in tension in control and AdCMVβ-Gal-transduced arteries. In contrast, ET-1 caused concentration-dependent relaxations of AdCMVeNOS-transduced arteries. The relaxations to ET-1 in AdCMVeNOS-transduced arteries were endothelium-independent They were abolished by NG-nitro-L-argininemethyl ester or by chemical treatment of adventitia with paraformaldehyde before gene transfer. ET-1 (10−9 mol/L) significantly increased intracellular cGMP levels in AdCMVeNOS-transduced arteries without endothelium. In arteries transduced with AdCMVeNOS, higher concentrations (10−9 to 3×10−8 mol/L) of ET-2 also caused relaxations, whereas ET-3 and sarafotoxin, a selective ETB receptor agonist, did not produce any relaxations. The relaxations to ET-1 in AdCMVeNOS-transduced arteries were strongly reduced by BQ-123 (10−7 mol/L), an ETA receptor antagonist, but were not affected by BQ-788 (3×10−7 mol/L), an ETB receptor antagonist These results suggest that genetically modified adventitia can produce nitric oxide and cause relaxations in response to ET-1 via activation of ETA receptors. Our findings support a novel concept that successful transfer and expression of recombinant eNOS gene can lead to a qualitative change in responsiveness to vasoconstrictor substances.

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