Doubts concerning functional endothelial nitric oxide synthase in human erythrocytes

Böhmer, Anke; Beckmann, Bibiana; Sandmann, Jörg; Tsikas, Dimitrios

doi:10.1182/blood-2011-11-393124

Cited by 37 publications

(25 citation statements)

References 10 publications

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“…20,22 It was important to confirm, therefore, that the protein identified as eNOS in our study was catalytically active, L-arginine:citrulline conversion was dependent on the presence of Ca 2ϩ /calmodulin, and activity was inhibited by L-NAME. Although we cannot exclude the additional presence of minor amounts of other NOS isoforms in RBCs it appears that the majority of NO produced in these cells was generated by an active eNOS.…”

Section: Isolation Isoform Identification and Activity Of Red Cell mentioning

confidence: 99%

“…21 Another recent study failed to detect increases in 15 N-labeled nitrite/nitrate after the addition of 15 N-L-Arg to intact RBCs. 22 RBCs have been shown to express a protein containing epitopes of an eNOS. 6,15,16,20,[23][24][25] However, positive staining with anti-iNOS antibodies has also been reported, 20,25 and others suggested that RBCs might express a novel NOS isoform.…”

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confidence: 99%

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Human red blood cells at work: identification and visualization of erythrocytic eNOS activity in health and disease

Cortese‐Krott

Rodriguez-Mateos

Sansone

et al. 2012

Blood

162

120

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IntroductionThe key event in the pathogenesis of arteriosclerosis is believed to be a dysfunction of the endothelium with disruption of vascular homeostasis, predisposing blood vessels to vasoconstriction, inflammation, leukocyte adhesion, thrombosis, and proliferation of vascular smooth muscle cells. Red blood cells (RBCs) are typically considered as shuttles of respiratory gases and nutrients for tissues, less so compartments important to vascular integrity. Patients with coronary artery disease (CAD) and concomitant anemia have a poorer prognosis after myocardial infarction, percutaneous coronary intervention, and coronary artery bypass grafting, and are more prone to developing heart failure with fatal outcomes. [1][2][3] Surprisingly, erythropoietin treatment fails to improve diagnosis, indicating that a compromised gas exchange/nutrient transport capacity of blood is insufficient to explain this outcome.Nitric oxide (NO) is an essential short-lived signaling/ regulatory product of a healthy endothelium that is critically important for vascular health. Decreased production and/or bioactivity of NO are a hallmark of endothelial dysfunction and have been shown to contribute to accelerated atherogenesis. In the cardiovascular system, NO is continuously produced in endothelial cells (ECs) by the type III isoform of NO synthase (eNOS, NOS3; EC 1.14.13.39). 4 In addition to endothelial cells, some circulating blood cells also contain eNOS.It is an accepted dogma that RBCs take up and inactivate endothelium-derived NO via rapid reaction with oxyhemoglobin to form methemoglobin and nitrate, thereby limiting NO available for vasodilatation. Yet it has also been shown that RBCs not only act as "NO sinks" but synthesize, store, and transport NO metabolic products. Under hypoxic conditions in particular, it has been demonstrated that RBCs induce NO-dependent vasorelaxation. 5,6 Mechanisms of release and potential sources of NO in RBCs are still a matter of debate, but candidates include iron-nitrosylhemoglobin, 7 S-nitrosohemoglobin, [8][9][10] and nitrite. The latter may form NO either via deoxyhemoglobin 5,11 or xanthine oxidoreductase (XOR)-mediated reduction, 6,12 or via spontaneous 12 and carbonic anhydrase-facilitated disproportionation. 13 Most of these processes show a clear oxygen-dependence, and several are favored by low oxygen tensions. The relative contribution of either mechanism to NO formation varies with oxygen partial pressure along the vascular tree. In addition, RBCs release ATP when subjected to hypoxia, providing an alternative vasodilatory pathway. 14 The publication costs of this article were defrayed in part by page charge payment. Therefore, and solely to indicate this fact, this article is hereby marked ''advertisement'' in accordance with 18 USC section 1734. 16,19 and citrulline 15,18 in the supernatant. However, Kang et al failed to measure citrulline production in RBC lysates, 20 maybe because of loss of cellular structures or cofactors important for activity. 21 Another recent study fai...

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Section: Isolation Isoform Identification and Activity Of Red Cell mentioning

confidence: 99%

mentioning

confidence: 99%

Human red blood cells at work: identification and visualization of erythrocytic eNOS activity in health and disease

Cortese‐Krott

Rodriguez-Mateos

Sansone

et al. 2012

Blood

162

120

View full text Add to dashboard Cite

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“…RBCs play an important role in the balance between generation and scavenging of nitric oxide (NO), which is a key regulator of vascular homeostasis [4]. Although findings on a functional RBC-NOS in humans are quite contradictory [5], recently it was hypothesized that human RBCs carry active NOS, producing NO even under normoxic conditions [6,7]. Of note, NO generation in RBC has been demonstrated to be influenced by various factors ex vivo, including insulin [7] and acetylcholine [8].…”

Section: Introductionmentioning

confidence: 99%

Nitric oxide synthetic pathway and cGMP levels are altered in red blood cells from end-stage renal disease patients

et al. 2016

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Red blood cells (RBCs) enzymatically produce nitric oxide (NO) by a functional RBC-nitric oxide synthase (RBC-NOS). NO is a vascular key regulatory molecule. In RBCs its generation is complex and influenced by several factors, including insulin, acetylcholine, and calcium. NO availability is reduced in end-stage renal disease (ESRD) and associated with endothelial dysfunction. We previously demonstrated that, through increased phosphatidylserine membrane exposure, ESRD-RBCs augmented their adhesion to human cultured endothelium, in which NO bioavailability decreased. Since RBC-NOS-dependent NO production in ESRD is unknown, this study aimed to investigate RBC-NOS levels/activation, NO production/bioavailability in RBCs from healthy control subjects (C, N = 18) and ESRD patients (N = 27). Although RBC-NOS expression was lower in ESRDRBCs, NO, cyclic guanosine monophosphate (cGMP), RBC-NOS Serine1177 phosphorylation level and eNOS/ Calmodulin (CaM)/Heat Shock Protein-90 (HSP90) interaction levels were higher in ESRD-RBCs, indicating increased enzyme activation. Conversely, following RBCs stimulation with insulin or ionomycin, NO and cGMP levels were significantly lower in ESRD-than in C-RBCs, suggesting that uremia might reduce the RBC-NOS response to further stimuli. Additionally, the activity of multidrug-resistance-associated protein-4 (MRP4; cGMPmembrane transporter) was significantly lower in ESRDRBCs, suggesting a possible compromised efflux of cGMP across the ESRD-RBCs membrane. This study for the first time showed highest basal RBC-NOS activation in ESRDRBCs, possibly to reduce the negative impact of decreased NOS expression. It is further conceivable that high NO production only partially affects cell function of ESRDRBCs maybe because in vivo they are unable to respond to physiologic stimuli, such as calcium and/or insulin.Keywords End-stage renal disease Á Red blood cells Á Nitric oxide Á cGMP Á Nitric oxide synthase Mario Bonomini, and Assunta Pandolfi have contributed equally to the study.Electronic supplementary material The online version of this article

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“…43 It has been recently suggested that erythrocytes contain functional eNOS, 44–46 although others have found no evidence of erythrocyte eNOS activity. 47 A variety of NOS inhibitors have also been shown to reduce red blood cell deformability, 44,48 although the mechanism by which NO may maintain deformability has yet to be determined. It is therefore possible that BH4 depletion could contribute to the poor erythrocyte deformability, impaired microvascular perfusion, insufficient oxygen delivery, and acidosis observed in patients with severe malaria.…”

Section: Discussionmentioning

confidence: 99%

Tetrahydrobiopterin Supplementation Improves Phenylalanine Metabolism in a Murine Model of Severe Malaria

Alkaitis

Ackerman

2016

ACS Infect. Dis.

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Tetrahydrobiopterin (BH4) is an essential cofactor for both phenylalanine hydroxylase and nitric oxide synthase. Patients with severe malaria have low urinary BH4, elevated plasma phenylalanine, and impaired endothelium-dependent vasodilation, suggesting that BH4 depletion may limit phenylalanine metabolism and nitric oxide synthesis. We infected C57BL/6 mice with Plasmodium berghei ANKA to characterize BH4 availability and to investigate the effects of BH4 supplementation. P. berghei ANKA infection lowered BH4 in plasma, erythrocytes, and brain tissue but raised it in aorta and liver tissue. The ratio of BH4 to 7,8-BH2 (the major product of BH4 oxidation) was decreased in plasma, erythrocytes, and brain tissue, suggesting that oxidation contributes to BH4 depletion. The continuous infusion of sepiapterin (a BH4 precursor) and citrulline (an arginine precursor) raised the concentrations of BH4 and arginine in both blood and tissue compartments. The restoration of systemic BH4 and arginine availability in infected mice produced only a minor improvement in whole blood nitrite concentrations, a biomarker of NO synthesis, and failed to prevent the onset of severe disease symptoms. However, sepiapterin and citrulline infusion reduced the ratio of phenylalanine to tyrosine in plasma, aortic tissue, and brain tissue. In summary, BH4 depletion in P. berghei infection may compromise both nitric oxide synthesis and phenylalanine metabolism; however, these findings require further investigation in human patients with severe malaria.

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Doubts concerning functional endothelial nitric oxide synthase in human erythrocytes

Cited by 37 publications

References 10 publications

Human red blood cells at work: identification and visualization of erythrocytic eNOS activity in health and disease

Human red blood cells at work: identification and visualization of erythrocytic eNOS activity in health and disease

Nitric oxide synthetic pathway and cGMP levels are altered in red blood cells from end-stage renal disease patients

Tetrahydrobiopterin Supplementation Improves Phenylalanine Metabolism in a Murine Model of Severe Malaria

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