An Improved Chemiluminescence Assay Suggests NGN Nitric Oxide-Mediated Action of Lysophosphatidylcholine and Acetylcholine

Menon, N. K.; Wolf, Andreas; Zehetgruber, Manfred; Bing, Richard J.

doi:10.3181/00379727-217-rc2

Cited by 28 publications

(12 citation statements)

References 0 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Quantitative measurements of NO in medium were made with a Sievers Research Redox Chemiluminescence Detector, model NOA 270 (Sievers Research, Boulder, CO). The lower limit of detection by this method is 22 pmol (22,23).…”

Section: Methodsmentioning

confidence: 99%

Nitric Oxide-Induced Cytotoxicity: Involvement of Cellular Resistance to Oxidative Stress and the Role of Glutathione in Protection

et al. 1995

View full text Add to dashboard Cite

A series of experiments were designed to examine the potential cytotoxicity of nitric oxide (NO), or reactive species derived from NO, in HA1 fibroblasts and H202-resistant variants of this cell line, designated OC14 cells. A 1-h exposure at 37OC to a 1.7 mM bolus dose of NO, prepared in N2-gassed medium, significantly reduced clonogenic survival in the HA1 fibroblasts line to 60% of control cells treated with N2-gassed medium alone. The OC14 cells were found to be completely resistant (100% survival) to NO-mediated injury in comparable experiments. A second set of experiments was designed to determine the role of the intracellular antioxidant, glutathione, in protection against NO-mediated injury. Depletion of total glutathione resulted in a significant reduction in HA1 and OC14 clonogenic survival to 8% and 50% when compared with respective control cells. The effect of total glutathione depletion on NO-initiated toxicity in HA1 cells was dose-and cell-density dependent and was observed to occur within 5 min of exposure to NO. Further evidence of cytotoxicity was demonstrated by loss of trypan blue dye exclusion properties in glutathione-depleted HA1 cells after NO is a free radical molecule endogenously produced by a variety of cell types. NO has been demonstrated to participate in physiologic reactions involving regulation of vasomotor tone, neurotransmission, immune system function, and platelet aggregation (1-4). Recent clinical trials have demonstrated some efficacy for inhaled NO in treatment of neonatal pulmonary hypertension and adult respiratory distress syndrome (5-7). Paradoxically, elevated concentrations of atmospheric NO are known to be harmful in many animal models, and lethal exposures at high concentrations have been reported in both animals and humans (8-10). This same paradoxical beneficial and/or toxic effect of NO has been observed in vitro. Several Received August 10, 1993; accepted July 8, 1994. Correspondence: Dr. M. Whit Walker, Department of Neonatology, Greenville Hospital Systems, 701 Grove Road, Greenville, SC 29605.Supported by National Institutes of Health Grants HL42057, HL08841, HL51469, CA.51116, HD28810, GM08260, and DK38924.NO exposure. Other experiments demonstrated that nitrate and nitrite exposure produced no cytotoxicity in glutathione-depleted HA1 cells and that coincubation of NO-saturated medium with oxyhemoglobin inhibited NO-induced cytotoxicity in glutathione-depleted HA1 cells. These results demonstrate that 1 ) nitric oxide, or an NO-derived reactive nitrogen species other than nitrites or nitrates, is responsible for reduction in clonogenic survival and trypan blue dye exclusion capabilities in vitro; 2) biochemical pathways associated with cellular resistance to oxidative stress also confer resistance to NO-mediated injury in this cell model; and 3) total glutathione content determines a significant portion of cell sensitivity to NO-mediated cytotoxicity.

show abstract

Section: Methodsmentioning

confidence: 99%

Nitric Oxide-Induced Cytotoxicity: Involvement of Cellular Resistance to Oxidative Stress and the Role of Glutathione in Protection

et al. 1995

View full text Add to dashboard Cite

show abstract

“…17 This involved injecting 10µL of deproteinised plasma samples into vanadium chloride. 18 The nitrate and nitrite present in samples were reduced to NO, and quantified from a calibration curve based on prepared concentrations of sodium nitrate.…”

Section: Laboratory Studymentioning

confidence: 99%

Effect of aspirin, clopidogrel and dipyridamole on soluble markers of vascular function in normal volunteers and patients with prior ischaemic stroke

et al. 2006

View full text Add to dashboard Cite

A note on versions:The version presented here may differ from the published version or from the version of record. If you wish to cite this item you are advised to consult the publisher's version. Please see the repository url above for details on accessing the published version and note that access may require a subscription.For more information, please contact eprints@nottingham.ac.uk IL-6 also increases endothelial production of the fibrinolytic inhibitor, plasminogen activator inhibitor type 1 (PAI-1). We assessed the effects of aspirin, clopidogrel and dipyridamole on a panel of the above soluble markers of vascular function in normal volunteers and patients with a prior history of ischaemic stroke. We also assessed the effect of combining the three antiplatelet agents on the soluble markers since we have shown previously that three drugs may be superior to one or two in inhibiting platelet function in vitro. 15Page 4 of 15 METHODS DesignTwo randomised, outcome-blinded, multiway, crossover trials of antiplatelet therapy were performed in normal volunteers and patients with prior ischaemic stroke. The study design has been published previously 16 in a report of the effect of aspirin, clopidogrel and dipyridamole on the function of circulating platelets, phagocytes and their conjugates. SubjectsThe study protocol was approved by the local Research Ethics Committee. All subjects gave written informed consent and the trials were performed according to the that complete data were present for each person; laboratory data from withdrawing subjects are not included in the statistical analyses. Laboratory studySerum and plasma collected into 3.8% sodium citrate (1:9 volume) or 4.0 mmol/L EDTA were centrifuged (1,500g for 15min) within 1 hour of venepuncture. Aliquots were frozen at -80 o C and then assayed in batches using commercially available ELISA kits. CRP (high sensitivity kit, Kalon Biological, Hants, UK) and PDGF (R&D System, Abington, UK) were measured in serum; cGMP (R&D System), vWf (Corgenix, Cambridgeshire, UK), MCP-1 (R&D System) and PAI-1 (Technoclone TC, Surrey, UK)were analysed in citrated plasma. Statistical analysisThe primary analysis compared the presence and absence of each drug on soluble markers, i.e. aspirin versus no aspirin, clopidogrel versus no clopidogrel, and dipyridamole versus no dipyridamole. Secondary analyses compared triple therapy (ACD) versus all other drugs, as before. 16 Analyses were performed using the SAS statistical package.Page 6 of 15 RESULTSDemographic and clinical information for the subjects are given in table 1. The stroke patients are representative of our clinical stroke service and were significantly older and had more risk factors (more hypertension and smokers, higher systolic blood pressure, and elevated total cholesterol) than the volunteers (all p<0.05).When considering the individual effects of the three antiplatelet agents on soluble markers of vascular function, three observations were made. First, dipyridamole significantly reduced plasma vWf levels in ...

show abstract

“…The chemiluminescence assay for NO 2 -is typically 10-fold less sensitive than the assay for NO [44,60], perhaps reflecting incomplete reduction of NO 2 -to NO. Acids other than HCl (e.g.…”

Section: + 2hmentioning

confidence: 99%

“…Most investigators report the detection threshold of the chemiluminescence assay for NO is 20-50 pmol, in liquids [6,56,57] and 1 ppb in gas. Some authors report that the chemiluminescence/NO curve is linear over the range 300-3000 pmol [60,61], while others have found a degree of nonlinearity with increasing levels of NO [43]. …”

mentioning

confidence: 99%

“…Since virtually all water contains NO 2 -, the signal from background NO 2 -levels in the solvents should always be accounted for by background subtraction from the experimental value. This problem can be avoided by using special relatively NO 2 --free water.The chemiluminescence assay for NO 2 -is typically 10-fold less sensitive than the assay for NO [44,60], perhaps reflecting incomplete reduction of NO 2 -to NO. Acids other than HCl (e.g.…”

mentioning

confidence: 99%

See 1 more Smart Citation

Measurement of nitric oxide, nitrite and nitrate using a chemiluminescence assay:an update for the year 2000

McMurtry¹,

Kim²,

Dinh-Xuan³

et al. 2000

Analusis

View full text Add to dashboard Cite

This review deals with the measurement of nitric oxide (NO) using the chemiluminescence assay. A preliminary discussion is offered to explain the chemical basis for this assay and emphasize the importance of measuring NO. After reviewing some practical aspects and caveats of the chemiluminescence assay, we review its application in a variety of research and clinical settings, such as measurement of breath NO and serum nitrates. The importance of avoiding confounding effects of dietary nitrates when assessing the NO system in humans is discussed and new data are provided on the confounding effects of atmospheric NO on measurement of breath NO. The utility and ease of the chemiluminescence assay for these applications is contrasted with the challenge of using chemiluminescence for correlation of vascular tone and NO production, studies in which real-time measurements are required. Although the chemiluminescence assay is one of the most reliable, rapid and reproducible assays available, it is optimal for gas phase measurements and may be suboptimal for measurement under certain circumstances. This chapter complements several more general reviews of the methodology for measuring NO.Article available at http://analusis.edpsciences.org or http://dx.doi.org/10.1051/analusis:2000280455 Need for measuring NOIt is clear that NO is essential in a significant number of normal physiological processes. However, the scope of its importance also spans pathophysiology of disease, such as in cardiogenic shock [14,15]. Inhaled NO is used for diagnostic purposes, and has therapeutic value, in diseases such as pulmonary hypertension [16][17][18], asthma and chronic obstructive lung diseases [12,[19][20][21]. NO may even promote regression of diseases, such as atherosclerosis [22,23].In view of the broad biological importance of this molecule, it is not surprising that basic and clinical research pertaining to NO has been so prolific. However, a computerised search of the past five years' literature found NO measurements reported in only 1089 of 26123 articles dealing with NO. Since many scientists don't measure NO, perhaps we should make the case for measurement explicit.There are four major reasons why one should consider measuring NO: Non specificity of NO inhibitorsPharmacological experiments are useful in studying NO and its effects. Nevertheless, the non-specificity of NO inhibitors [24][25][26] impairs one's ability to draw sound mechanistic conclusions. For instance, N G -monomethyl-L-arginine (L-NMMA), a NOS antagonist, displays hemodynamic effects that are not solely mediated by its ability to inhibit NO synthesis [27]. L-NMMA can also be metabolized to yield NO. Furthermore, even high doses of NOS inhibitors do not completely prevent NO synthesis [28]. Without measuring NO one cannot be assured that the observed effects resulting from administration of a NOS inhibitor reflect the complete loss of NO, nor can one exclude non-specific effects of these inhibitors. The existence of multiple endothelium-derived vasodilatorsIt i...

show abstract

An Improved Chemiluminescence Assay Suggests NGN Nitric Oxide-Mediated Action of Lysophosphatidylcholine and Acetylcholine

Cited by 28 publications

References 0 publications

Nitric Oxide-Induced Cytotoxicity: Involvement of Cellular Resistance to Oxidative Stress and the Role of Glutathione in Protection

Nitric Oxide-Induced Cytotoxicity: Involvement of Cellular Resistance to Oxidative Stress and the Role of Glutathione in Protection

Effect of aspirin, clopidogrel and dipyridamole on soluble markers of vascular function in normal volunteers and patients with prior ischaemic stroke

Measurement of nitric oxide, nitrite and nitrate using a chemiluminescence assay:an update for the year 2000

Contact Info

Product

Resources

About