Mechanistic Study of Hemoglobin Induced Platelet Activation

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 5262-5262
Author(s):  
Christine R. Carlisle ◽  
Mary Stahle ◽  
Ryan Vest ◽  
Roy Hantgan ◽  
Janet Lee ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Reactive Oxygen Species ◽  
Platelet Activation ◽  
Reactive Oxygen ◽  
Inhibitory Effect ◽  
Mechanistic Study ◽  
Blood Collection ◽  
Oxygen Species ◽  
Free Hemoglobin ◽  
No Donor

Abstract Abstract 5262 Nitric oxide (NO) has been previously shown to inhibit adenosine diphosphate (ADP)- and thrombin- mediated platelet activation. This inhibitory effect of NO is blocked by cell-free oxyhemoglobin (Villagra, Shiva et al. 2007). Cell-free hemoglobin (Hb) also appears to directly activate platelets even in the absence of ADP. However, the precise mechanisms for cell-free Hb-induced platelet activation have not been fully delineated. We hypothesized that oxyhemoglobin can promote platelet activation through one of three potential mechanisms: 1) direct apoprotein interaction with platelet receptors, 2) reactive oxygen species (ROS) production, and/or 3) scavenging of endogenous NO. Platelets were isolated from whole blood obtained from healthy volunteers (n=15) using sodium citrate blood collection tubes. Platelet activation was measured by examining immunofluorescent Pac-1 labeling by flow cytometry using activated glycoprotein IIb/IIIa as a measure of activation. In the presence of 10 uM concentration of NO donor mahmaNONOate, ADP-induced platelet activation was inhibited by 44%. Incubation of 100 uM cell-free oxyhemoglobin eliminated the inhibitory effect of NO. In addition, cell-free oxyhemoglobin (100 uM concentration) induced platelet activation by 17% above unstimulated controls even in the absence of ADP. Methemoglobin was used to test if direct Hb protein interaction with platelets was sufficient for platelet activation, independent of heme redox state. To produce methemoglobin, the ferrous iron in oxyhemoglobin was oxidized to its ferric form, thus methemoglobin is likely to preserve all direct protein interactions. However, unlike oxyhemoglobin, incubation of 100 uM methemoglobin did not induce platelet activation. Next, we tested whether reactive oxygen species production was responsible for hemoglobin-induced platelet activation. In the absence of hemoglobin, superoxide dismutase (SOD) (100 U/ml) and catalase (100 U/ml) reduced baseline platelet activation, verifying a ROS (Pignatelli, Pulcinelli et al. 1998). However, in the presence of oxyhemoglobin, SOD and catalase did not significantly abrogate the effect of Hb on platelet activation, suggesting that the ROS superoxide and hydrogen peroxide are not involved in hemoglobin-induced platelet activation. To determine if scavenging of endogenous NO was responsible for hemoglobin-induced platelet activation, platelets were incubated in the presence or absence of NO scavengers, carboxy-PTIO (5mM) and Fe-DTCS (3 mM), or nitric oxide synthase (NOS) inhibitor, LNMMA (100 uM). Results from these investigations suggest scavenging of endogenous NO was not responsible for hemoglobin-induced platelet activation. Overall our data suggest a complex mechanism for oxyhemoglobin-mediated platelet activation that is likely to involve more than a single contributing pathway. These data aid in elucidating the mechanism of platelet activation by cell-free hemoglobin and may provide novel insights to designing potential treatments to reduce the platelet activation underlying hypercoagulable states associated with a variety of pathological conditions including hemolytic anemias, stroke, and diabetes. This work was supported by NIH grants HL058091 and HL098032 Disclosures: Gladwin: Patents filed related to treating hemolysis.: Patents & Royalties. Kim-Shapiro:Patents filed related to treating hemolysis. Honororia from Sangart Inc.: Honoraria, Patents & Royalties.

scholarly journals Modulation of metabolic activity of phagocytes by antihistamines

2011 ◽  
Vol 4 (1) ◽  
pp. 15-19 ◽  
Author(s):  
Antonin Lojek ◽  
Milan Číž ◽  
Michaela Pekarová ◽  
Gabriela Ambrožová ◽  
Ondřej Vašíček ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Reactive Oxygen Species ◽  
Metabolic Activity ◽  
Reactive Oxygen Species Generation ◽  
Reactive Oxygen ◽  
Inhibitory Effect ◽  
Oxygen Species ◽  
Antioxidative Properties ◽  
Enhanced Chemiluminescence ◽  
Inos Protein Expression

Modulation of metabolic activity of phagocytes by antihistaminesThe purpose of the study was to investigate the effects of H1-antihistamines of the 1stgeneration (antazoline, bromadryl, brompheniramine, dithiaden, cyclizine, chlorcyclizine, chlorpheniramine, clemastine) and the 2ndgeneration (acrivastine, ketotifen, and loratadine) on the respiratory burst of phagocytes. Reactive oxygen species generation in neutrophils isolated from rat blood was measured using luminol-enhanced chemiluminescence. Changes in nitrite formation and iNOS protein expression by RAW 264.7 macrophages were analysed using Griess reaction and Western blotting. The antioxidative properties of drugs in cell-free systems were detected spectrophotometrically, luminometrically, fluorimetrically, and amperometrically. The majority of the H1-antihistamines tested (bromadryl, brompheniramine, chlorcyclizine, chlorpheniramine, clemastine, dithiaden, and ketotifen) exhibited a significant inhibitory effect on the chemiluminescence activity of phagocytes. H1-antihistamines did not show significant scavenging properties against superoxide anion and hydroxyl radical, thus this could not contribute to the inhibition of chemiluminescence. H1-antihistamines had a different ability to modulate nitric oxide production by LPS-stimulated macrophages. Bromadryl, clemastine, and dithiaden were the most effective since they inhibited iNOS expression, which was followed by a significant reduction in nitrite levels. H1-antihistamines had no scavenging activity against nitric oxide. It can be concluded that the effects observed in the H1-antihistamines tested are not mediated exclusively via H1-receptor pathway or by direct antioxidative properties. Based on our results, antihistamines not interfering with the microbicidal mechanisms of leukocytes (antazoline, acrivastine and cyclizine) could be used preferentially in infections. Other antihistamines should be used, under pathological conditions accompanied by the overproduction of reactive oxygen species.

Nitric oxide and reactive oxygen species in the nucleus revisitedThis review is one of a selection of papers published in a Special Issue on Oxidative Stress in Health and Disease.

2010 ◽  
Vol 88 (3) ◽  
pp. 296-304 ◽  
Author(s):  
Chantale Provost ◽  
Faten Choufani ◽  
Levon Avedanian ◽  
Ghassan Bkaily ◽  
Fernand Gobeil ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Nitric Oxide ◽  
Reactive Oxygen Species ◽  
Reactive Oxygen ◽  
Prostaglandin E ◽  
Oxygen Species ◽  
Nuclear Level ◽  
No Donor ◽  
Envelope Membranes ◽  
Endothelial Nitric Oxide

Recent work from our group showed that the nuclear envelope membranes contain several G protein-coupled receptors, including prostaglandin E2 (EP3R) and endothelin-1 (ET-1) receptors. Activation of EP3R increased endothelial nitric oxide synthase (eNOS) RNA expression in nuclei. eNOS and inducible NOS (iNOS) are reported to also be present at the nuclear level. Furthermore, reactive oxygen species (ROS) were also localized at the nuclear level. In this review, we show that stimulation with NO donor sodium nitroprusside results in an increase of intranuclear calcium that was dependent on guanylate cyclase activation, but independent of MAPK. This increase in nuclear calcium correlated with an increase in nuclear transcription of iNOS. H2O2 and ET-1 increase both cytosolic and nuclear ROS in human endocardial endothelial cells and in human aortic vascular smooth muscle cells. This increase in ROS levels by H2O2 and ET-1 was reversed by the antioxidant glutathione. In addition, our results strongly suggest that cytosolic signalization is not only transmitted to the nucleus but is also generated by the nucleus. Furthermore, we demonstrate that oxidative stress can be sensed by the nucleus. These results highly suggest that ROS formation is also generated directly by the nucleus and that free radicals may contribute to ET-1 regulation of nuclear Ca2+ homeostasis.

scholarly journals Interaction between reactive oxygen species and nitric oxide in the microvascular response to systemic hypoxia

2002 ◽  
Vol 93 (4) ◽  
pp. 1411-1418 ◽  
Author(s):  
Dawn R. S. Steiner ◽  
Norberto C. Gonzalez ◽  
John G. Wood
Keyword(s):  
Nitric Oxide ◽  
Reactive Oxygen Species ◽  
Reactive Oxygen ◽  
Oxygen Species ◽  
Leukocyte Adherence ◽  
No Donor ◽  
Substrate Limitation ◽  
Systemic Hypoxia ◽  
Microvascular Response ◽  
Spermine Nonoate

Systemic hypoxia results in oxidative stress due to a change in the reactive oxygen species (ROS)-nitric oxide (NO) balance. These experiments explored two mechanisms for the altered ROS-NO balance: 1) decreased NO synthesis by NO synthase due to limited O2 substrate availability and 2) increased superoxide generation. ROS levels and leukocyte adherence in mesenteric venules of rats during hypoxia were studied in the absence and presence of an NO donor [spermine NONOate (SNO)] and of the NO precursorl-arginine. We hypothesized that if the lower NO levels during hypoxia were due to O2 substrate limitation,l-arginine would not prevent hypoxia-induced microvascular responses. Graded hypoxia (produced by breathing 15, 10, and 7.5% O2) increased both ROS (123 ± 6, 148 ± 11, and 167 ± 3% of control) and leukocyte adherence. ROS levels during breathing of 10 and 7.5% O2 were significantly attenuated by SNO (105 ± 6 and 108 ± 3%, respectively) andl-arginine (117 ± 5 and 115 ± 2%, respectively). Both interventions reduced leukocyte adherence by similar degrees. The fact that the effects of l-arginine were similar to those of SNO does not support the idea that NO generation is impaired in hypoxia and suggests that tissue NO levels are depleted by the increased ROS during hypoxia.

Programming blood pressure in adult SHR by shifting perinatal balance of NO and reactive oxygen species toward NO: the inverted Barker phenomenon

2005 ◽  
Vol 288 (4) ◽  
pp. F626-F636 ◽  
Author(s):  
Simona Racasan ◽  
Branko Braam ◽  
Hein A. Koomans ◽  
Jaap A. Joles
Keyword(s):  
Nitric Oxide ◽  
Blood Pressure ◽  
Reactive Oxygen Species ◽  
Adult Life ◽  
Reactive Oxygen ◽  
Superoxide Production ◽  
Oxygen Species ◽  
No Donor ◽  
Wistar Kyoto ◽  
Inducible Nos

The “programming hypothesis” proposes that an adverse perinatal milieu leads to adaptation that translates into cardiovascular disease in adulthood. The balance between nitric oxide (NO) and reactive oxygen species (ROS) is disturbed in cardiovascular diseases, including hypertension. Conceivably, this balance is also disturbed in pregnancy, altering the fetal environment; however, effects of perinatal manipulation of NO and ROS on adult blood pressure (BP) are unknown. In spontaneously hypertensive rats (SHR), NO availability is decreased and ROS are increased compared with normotensive Wistar-Kyoto rats, and, despite the genetic predisposition, the perinatal environment can modulate adult BP. Our hypothesis is that a disturbed NO-ROS balance in the SHR dam persistently affects BP in her offspring. Dietary supplements, which support NO formation and scavenge ROS, administered during pregnancy and lactation resulted in persistently lower BP for up to 48 wk in SHR offspring. The NO donor molsidomine and the superoxide dismutase mimic tempol-induced comparable effects. Specific inhibition of inducible nitric oxide synthase (NOS) reduces BP in adult SHR, suggesting that inducible NOS is predominantly a source of ROS in SHR. Indeed, inducible NOS inhibition in SHR dams persistently reduced BP in adult offspring. Persistent reductions in BP were accompanied by prevention of proteinuria in aged SHR. We propose that in SHR the known increase in ANG II type 1 receptor density during development leads to superoxide production, which enhances inducible NOS activity. The relative shortage of substrate and cofactors leads to uncoupling of inducible NOS, resulting in superoxide production, activating transcription factors that subsequently again increase inducible NOS expression. This vicious circle probably is perpetuated into adult life.

scholarly journals Nitric oxide synthase inhibition causes acute increases in glomerular permeability in vivo, dependent upon reactive oxygen species

2016 ◽  
Vol 311 (5) ◽  
pp. F984-F990 ◽  
Author(s):  
Julia Dolinina ◽  
Kristinn Sverrisson ◽  
Anna Rippe ◽  
Carl M. Öberg ◽  
Bengt Rippe
Keyword(s):  
Nitric Oxide ◽  
Reactive Oxygen Species ◽  
High Performance ◽  
Reactive Oxygen ◽  
Size Exclusion ◽  
Oxygen Species ◽  
Glomerular Permeability ◽  
No Donor ◽  
Filtration Barrier

There is increasing evidence that the permeability of the glomerular filtration barrier (GFB) is partly regulated by a balance between the bioavailability of nitric oxide (NO) and that of reactive oxygen species (ROS). It has been postulated that normal or moderately elevated NO levels protect the GFB from permeability increases, whereas ROS, through reducing the bioavailability of NO, have the opposite effect. We tested the tentative antagonism between NO and ROS on glomerular permeability in anaesthetized Wistar rats, in which the left ureter was cannulated for urine collection while simultaneously blood access was achieved. Rats were systemically infused with either l-NAME or l-NAME together with the superoxide scavenger Tempol, or together with l-arginine or the NO-donor DEA-NONOate, or the cGMP agonist 8-bromo-cGMP. To measure glomerular sieving coefficients (theta, θ) to Ficoll, rats were infused with FITC-Ficoll 70/400 (mol/radius 10–80 Å). Plasma and urine samples were analyzed by high-performance size-exclusion chromatography (HPSEC) for determination of θ for Ficoll repeatedly during up to 2 h. l-NAME increased θ for Ficoll70Å from 2.27 ± 1.30 × 10−5 to 8.46 ± 2.06 × 10−5 ( n = 6, P < 0.001) in 15 min. Tempol abrogated these increases in glomerular permeability and an inhibition was also observed with l-arginine and with 8-bromo-cGMP. In conclusion, acute NO synthase inhibition in vivo by l-NAME caused rapid increases in glomerular permeability, which could be reversed by either an ROS antagonist or by activating the guanylyl cyclase-cGMP pathway. The data strongly suggest a protective effect of NO in maintaining normal glomerular permeability in vivo.

scholarly journals Interaction between reactive oxygen species and hormones during the breaking of embryo dormancy in Sorbus pohuashanensis by exogenous nitric oxide

2021 ◽  
Author(s):  
Hao Wang ◽  
Shuoran Tang ◽  
Jianan Wang ◽  
Hailong Shen ◽  
Ling Yang
Keyword(s):  
Nitric Oxide ◽  
Reactive Oxygen Species ◽  
Reactive Oxygen ◽  
Ethylene Biosynthesis ◽  
Oxygen Species ◽  
No Donor ◽  
Hormone Synthesis ◽  
Ros Accumulation ◽  
Embryo Dormancy ◽  
Exogenous Nitric Oxide

AbstractThe breaking of dormancy mediated by reactive nitrogen species (RNS) is related to the accumulation of reactive oxygen species (ROS) in germinating embryos but the underlying mechanism is unclear. The objectives of this study were: (1) to explore the relationship between RNS-mediated dormancy release and ROS accumulation in germinating embryos of Sorbus pohuashanensis; and, (2) to investigate the relationships among germination time, ROS metabolism, and endogenous hormone synthesis. We studied the effects of exogenous nitric oxide (NO) donor sodium nitroprusside (SNP), the NO scavenger (4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (PTIO), abscisic acid (ABA), the exogenous ethylene donor ethrel, and the ethylene receptor inhibitor 2,5-norbornadien (NBD) on embryo germination and seedling growth. Embryos were released from dormancy by pretreatment with NO or ethylene, which was related to increased ethylene biosynthesis and decreased ABA levels. Breaking of dormancy by SNP was related to increased levels of ethylene, hydrogen peroxide, and glutathione, increased activities of superoxide dismutase and glutathione peroxidase, and decreased levels of ABA, superoxide anions, and malondialdehyde. These effects of nitric oxide were especially significant in seedling hypocotyls and radicles. These results demonstrate that NO can break S. pohuashanensis embryo dormancy by inducing ethylene biosynthesis, and that this signalling pathway is closely related to ROS accumulation and the antioxidant defence response.

The Role of Reactive Oxygen Species, Kinases, Hydrogen Sulfide, and Nitric Oxide in the Regulation of Autophagy and Their Impact on Ischemia and Reperfusion Injury in the Heart

2020 ◽  
Vol 16 ◽  
Author(s):  
Andrey Krylatov ◽  
Leonid Maslov ◽  
Sergey Y. Tsibulnikov ◽  
Nikita Voronkov ◽  
Alla Boshchenko ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Reactive Oxygen Species ◽  
Hydrogen Sulfide ◽  
Ischemia Reperfusion ◽  
Reactive Oxygen ◽  
Ischemia And Reperfusion ◽  
Oxygen Species ◽  
Ischemia And Reperfusion Injury ◽  
Adaptive Process

: There is considerable evidence in the heart that autophagy in cardiomyocytes is activated by hypoxia/reoxygenation (H/R) or in hearts by ischemia/reperfusion (I/R). Depending upon the experimental model and duration of ischemia, increases in autophagy in this setting maybe beneficial (cardioprotective) or deleterious (exacerbate I/R injury). Aside from the conundrum as to whether or not autophagy is an adaptive process, it is clearly regulated by a number of diverse molecules including reactive oxygen species (ROS), various kinases, hydrogen sulfide (H2S) and nitric oxide (NO). The purpose this review is to address briefly the controversy regarding the role of autophagy in this setting and to examine a variety of disparate molecules that are involved in its regulation.

Reactive Oxygen Species and Platelet Activation in Reperfusion Injury

Circulation ◽  
1997 ◽  
Vol 95 (4) ◽  
pp. 787-789 ◽  
Author(s):  
Rosemarie C. Forde ◽  
Desmond J. Fitzgerald
Keyword(s):  
Reactive Oxygen Species ◽  
Reperfusion Injury ◽  
Platelet Activation ◽  
Reactive Oxygen ◽  
Oxygen Species

Plasmonic Enhancement of Nitric Oxide Generation

Nanoscale ◽  
2021 ◽  
Author(s):  
Rachael Knoblauch ◽  
Chris Geddes
Keyword(s):  
Nitric Oxide ◽  
Reactive Oxygen Species ◽  
Reactive Nitrogen Species ◽  
Reactive Oxygen ◽  
Reactive Nitrogen ◽  
Oxygen Species ◽  
Nitrogen Species ◽  
Plasmonic Enhancement ◽  
Cancer Treatments

While the utility of reactive oxygen species in photodynamic therapies for both cancer treatments and antimicrobial applications has received much attention, the inherent potential of reactive nitrogen species (RNS) including...

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