Free Radical Biology & Medicine 41 (2006) 295 – 301
www.elsevier.com/locate/freeradbiomed
Original Contribution
Plasma nitrite reserve and endothelial function in the
human forearm circulation
Tienush Rassaf a,1 , Christian Heiss b,1 , Ulrike Hendgen-Cotta a , Jan Balzer a , Simone Matern a ,
Petra Kleinbongard a , Andrew Lee b , Thomas Lauer a , Malte Kelm a,⁎
a
b
Department of Medicine, Division of Cardiology, University Hospital Aachen, 52074 Aachen, Germany
Department of Medicine, Division of Cardiology, University of California, San Francisco, CA 94143, USA
Received 19 November 2005; revised 20 March 2006; accepted 6 April 2006
Available online 26 April 2006
Abstract
Attenuation of endothelium-derived nitric oxide (NO) synthesis is a hallmark of endothelial dysfunction. Early detection of this disorder may
have therapeutic and prognostic implications. Plasma nitrite mirrors acute and chronic changes in endothelial NO-synthase activity. We
hypothesized that local plasma nitrite concentration increases during reactive hyperemia of the forearm, reflecting endothelial function. In healthy
subjects (n = 11) plasma nitrite and nitrate were determined at baseline and during reactive hyperemia of the forearm using reductive gas-phase
chemiluminescence and flow-injection analysis, respectively. Endothelium-dependent dilation of the brachial artery was measured as flowmediated dilation (FMD) using high-resolution ultrasound. Results were compared to patients with endothelial dysfunction as defined by reduced
FMD (n = 11). Reactive hyperemia of the forearm increased local plasma nitrite concentration from 68 ± 5 to 126 ± 13 nmol/L (p < 0.01), whereas
in endothelial dysfunction nitrite remained unaffected (116 ± 12 to 104 ± 10 nmol/L; n.s.), corresponding to nitrite reserves of 94 ± 21 and
−8 ± 4%. This was accompanied by a significantly greater increase in brachial artery diameter (FMD: 8.5 ± 0.4% vs 2.9 ± 0.5%, for healthy
subjects and endothelial dysfunction, respectively; p < 0.001). This observation suggests that nitrite changes reflect endothelial function.
Assessment of local plasma nitrite during reactive hyperemia may open new avenues in the diagnosis of vascular function.
© 2006 Elsevier Inc. All rights reserved.
Keywords: Nitric oxide; Nitrite; Endothelial function; Free radical
Introduction
Endothelial dysfunction is an early stage of arteriosclerosis
and has been attributed to impaired nitric oxide (NO)
bioactivity and enhanced formation of oxygen-derived free
radicals [1]. Taking into account that endothelial dysfunction is
at least in part reversible [2], an early diagnosis of this disorder
by assessing endothelial NO-synthase (eNOS) activity may
Abbreviations: BA, brachial artery; CAD, coronary artery disease; CVD,
cardiovascular disease; eNOS, endothelial isoform of nitric oxide synthase;
FMD, flow-mediated dilation of the BA; GTN, glycerol trinitrate induced
dilation of the BA; NO, nitric oxide; LDL, low-density lipoprotein; HDL, highdensity lipoprotein.
⁎ Corresponding author. Fax: +49 241 80 82 303.
E-mail address: [email protected] (M. Kelm).
1
Both authors contributed equally to this work.
0891-5849/$ - see front matter © 2006 Elsevier Inc. All rights reserved.
doi:10.1016/j.freeradbiomed.2006.04.006
have prognostic and therapeutic consequences. While direct
biochemical evidence for an impaired eNOS activity has been
obtained in experimental models, this approach is difficult in
humans so far. Once released into the vascular lumen,
endothelium-derived NO either undergoes oxidation or
participates in nitros(yl)ation reactions. The compounds
produced differ greatly in biological activity, concentration,
stability, and compartmentalization between plasma and blood
cells.
An increase in shear stress, i.e. the tangential force exerted
by the flow of blood over the surface of the endothelium, is the
strongest physiological stimulus of eNOS activity and leads to
increases in NO formation. Shear-stress-induced NO-dependent
dilation of the brachial artery (BA) can be measured
noninvasively as flow-mediated dilation (FMD) using highresolution ultrasound and is commonly used to characterize
endothelial function [3]. This ultrasonographic method
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T. Rassaf et al. / Free Radical Biology & Medicine 41 (2006) 295–301
quantifies the dilation of conduit arteries, e.g. brachial artery, in
response to physiologically relevant increases in laminar shear
stress induced by ischemic dilation of the downstream microvasculature. Increases in shear stress lead to a rapid activation of
endothelial NO-synthase with consecutive increases in NO
formation. Accordingly, FMD is largely abolished following
NOS inhibition [2] and therefore provides a valuable “read-out”
of local vascular NO availability. A biochemical approach to
determine eNOS activity is still missing.
We conducted a “proof of concept” study and attempted to
establish an index to assess eNOS capacity biochemically in
humans. Plasma nitrite, the main oxidation product of NO,
sensitively reflects acute [4] and chronic [5] changes in eNOS
activity in healthy subjects. We sought to determine the
functional reserve of eNOS activity and measured local plasma
nitrite concentrations in the antecubital vein at baseline and
during reactive hyperemia following 5 min of forearm ischemia
in young healthy subjects. Notably, we observed that plasma
nitrite increased by a factor of 2 and that the time course of
nitrite mirrored the increase in BA diameter. In a second series
of experiments, we show that individuals with endothelial
dysfunction, as defined by reduced flow-mediated dilation, do
not exhibit an increase in nitrite. The correlation between the
local nitrite increase and the degree of vasodilation suggest that
nitrite changes may reflect endothelial function.
Materials and methods
Study population
We studied 11 healthy volunteers and 11 patients with
endothelial dysfunction. The patients were recruited from the
Department of Medicine. A three-vessel CAD was diagnosed in
all patients by coronary angiography. Left ventricular function
was normal in all subjects. Hypertension was defined by
systolic blood pressure >140 mm Hg, diastolic blood pressure
>90 mm Hg [6], or current antihypertensive medication.
According to ADA guidelines [7], diabetes mellitus was
defined by glucose levels in plasma >126 mg/dL (fasting) or
current antidiabetic medication. Hypercholesterolemia was
defined by a total cholesterol level >240 mg/dL, LDLcholesterol levels >160 mg/dL, or use of cholesterol-lowering
medication [8]. The clinical characteristics are summarized in
Table 1. Participants fasted and refrained from smoking at least
12 h prior to and until completion of the investigation.
Medication with organic nitrates was discontinued prior to
investigations. Subjects were studied in a supine position in a
quiet, air-conditioned room (21 °C). The study protocol was
approved by the ethics committee of our university, and all
subjects gave written informed consent before participating in
the study.
Study protocol
We performed two sets of experiments. In the first set, we
determined the time course of brachial artery diameter and
blood flow velocity during reactive hyperemia and measured
Table 1
Characteristics of study groups
n (m/f)
Age (years)
BMI
Hypertension (n)
Dyslipidemia (n)
Diabetes mellitus (n)
Smoking (n)
CAD (n)
ACE inhibitors (n)
Statins (n)
β-Blockers (n)
Organic nitrates (n)
MAP [mm Hg]
Pack years
Cholesterol [mg/dL]
LDL [mg/dL]
HDL [mg/dL]
Glucose [mg/dL]
CRP [mg/dL]
BA diameter [mm] baseline
FMD [%]
GTN [%]
Average ± SE
Control
Endothelial
dysfunction
p
11 (6/5)
25 ± 1
22.0 ± 0.7
0
0
0
0
0
0
0
0
0
95 ± 4
0
130 ± 13
73 ± 3
79 ± 2
87 ± 2
0.3 ± 0
4.0 ± 0.2
8.5 ± 0.4
14.7 ± 0.7
11 (5/6)
62 ± 2
29.6 ± 1.2
11
11
6
7
11
11
11
11
11
91 ± 1
17.8 ± 4.7
146 ± 5
130 ± 13
55 ± 6
132 ± 11
0.3 ± 0.09
5.1 ± 0.2
2.9 ± 0.5
8.3 ± 1.0
n.s.
<0.001
n.s.
n.s.
<0.001
<0.001
<0.001
<0.05
<0.001
n.s.
<0.01
<0.001
<0.001
the kinetics of plasma nitrite during hyperemia of the forearm in
the draining antecubital vein (Fig. 1). In the second set, we
compared the increases in nitrite and brachial artery diameter of
healthy volunteers to those of patients with endothelial
dysfunction (Fig. 2).
We characterized the conduit arterial and resistance vessel
responses of the forearm during reactive hyperemia using highresolution ultrasound [9]. Reactive hyperemia was induced by
inflation (250 mm Hg, 5 min) of a blood pressure cuff placed
around the forearm of a subgroup of healthy young controls
(n = 5). Brachial artery diameter and blood flow velocity were
determined at baseline, immediately after cuff deflation (0 s),
and every 15 s for 90 s. In analogy to FMD, we sought to assess
the functional reserve of eNOS capacity biochemically and
determined plasma nitrite as an index of eNOS activity. The
nitrite concentration in the draining antecubital vein was
measured at rest, immediately after ischemia, and every 10 s
for 90 s. In the second set, we determined the response of the
brachial artery and plasma nitrite side by side in healthy
controls and patients with endothelial dysfunction. Plasma
nitrite, brachial artery diameter, and flow velocity were
determined at baseline, immediately after cuff deflation, and
at 50 s—the time point of maximum increase as determined in
the first set (Fig. 1).
Measurement of plasma nitrite and nitrate
Blood was drawn from a catheter placed in the antecubital
vein proximal to the blood pressure cuff on the forearm,
collected into a prechilled heparinized tube, and centrifuged
immediately for 10 min at 800g and 4 °C. Plasma levels of
nitrite were determined using a triiodide/ozone-based
T. Rassaf et al. / Free Radical Biology & Medicine 41 (2006) 295–301
297
ischemia of the forearm in relation to baseline values (NO2−Rest)
(NO2−[%] = NO2−RH-NO2−Rest/NO2−Rest).
Assessment of vascular diameter and flow velocity
The diameter of the brachial artery and flow velocity were
determined using high-resolution ultrasound with a 15-MHz
linear array transducer (Sonos 5500, Philips Medizin Systeme,
Hamburg, Germany). Briefly, measurements were taken
proximal to the antecubital fossa at rest and during reactive
hyperemia after 5 min of ischemia, essentially as described [2].
The protocol was in accordance with the published guidelines,
yielding a coefficient of variation of less than 1% [9]. Flow
velocity represents the mean angle-corrected Doppler flow
velocity at the center of the vessel. Flow-mediated dilation was
calculated as the peak relative diameter gain after cuff deflation
as compared to baseline values. Smooth muscle dilatory
Fig. 1. Kinetics of plasma nitrite and brachial artery diameter during reactive
hyperemia. (A) Brachial artery diameter and flow velocity during forearm
reactive hyperemia and (B) plasma nitrite in antecubital vein were measured in
healthy control subjects (n = 5). Ischemic vasodilation of the forearm circulation
is reflected by increased flow velocity (filled circles) when reperfusion begins.
The increased shear at the artery wall leads to brachial artery dilation. The
brachial artery dilates with a peak at 60 s. The time course of brachial artery
diameter is paralleled by the plasma nitrite concentration (open circles) with a
peak at 50 s. Data are reported as percentage change from baseline value before
induction of reactive hyperemia. X axis represents the time after deflation of a
blood pressure cuff placed around the forearm distal to the site of measurement.
Symbols depict mean values ± SE. * p < 0.05 vs baseline value at rest before
hyperemia.
chemiluminescence assay, essentially as described [10,11]. In
brief, plasma was immediately divided into two aliquots. One
aliquot was directly injected into the reaction mixture consisting
of 45 mmol/L potassium iodide and 10 mmol/L iodine in glacial
acetic acid at 60 °C actively purged with a helium stream in line
with an NO chemiluminescence analyzer (88 CLD 77am sp and
88 AM, Eco Physics, Duernten, Switzerland). The other aliquot
was treated with 1/10 volume of 5% sulfanilamide in 1 M HCl
to scavenge nitrite for 15 min and then injected. The difference
in the two peaks sensitively reflected the concentration of nitrite
in the plasma sample.
Nitrate was quantified after enzymatic reduction to nitrite by
nitrate reductase using a flow-injection analysis which is a
colorimetrical assay based on the Griess reaction as described in
detail elsewhere [12,13].
We calculated the nitrite reserve as relative nitrite increase
during reactive hyperemia at 50 s (NO2− RH) after a 5 min
Fig. 2. Nitrite and brachial artery diameter increase in controls (A; open
symbols) but not in patients with endothelial dysfunction (B; filled symbols)
during reactive hyperemia. Plasma nitrite and diameter were measured at
baseline and 50 and 60 s after 5 min of lower arm ischemia, respectively. Circles
depict individual values; squares depict mean values ± SE. *p < 0.05 vs baseline
value before hyperemia. # p < 0.05 vs respective value in endothelial
dysfunction group.
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T. Rassaf et al. / Free Radical Biology & Medicine 41 (2006) 295–301
function was determined 4 min after sublingual application of
400 μg of glycerol trinitrate (Nitrolingual mite, Pohl, Germany).
Statistical analysis
Results are expressed as means ± standard error (SE).
Repeated-measurements analysis of variance was used to
estimate intraindividual and between-group effects. Pairwise
comparisons were corrected by the Bonferroni confident
interval. Normal distribution was estimated using the Kolmogorov–Smirnov test. Statistical significance was assumed if a
null hypothesis could be rejected at p = 0.05. All analyses were
performed with SPSS 11.0.1 (SPSS Inc., Chicago, IL).
Results
Characteristics of study groups
The study group consisted of 11 young (25 ± 1 years)
healthy volunteers without cardiovascular risk factors (hypertension, hypercholesterolemia, smoking, diabetes; Table 1).
The blood pressure and clinical lab parameters were within
normal limits. In contrast, the patients (n = 11) were
significantly older (62 ± 2 years), had more than three risk
factors, three-vessel coronary artery disease, and endothelial
dysfunction as defined by FMD. Pack years (a measure of
cigarette smoking over someone's lifetime; calculated as the
number of packs per day times the number of years that a
person has smoked), cholesterol, LDL, and fasting plasma
glucose were significantly greater and HDL was significantly
lower than those for controls. Vasodilation following GTN
was significantly attenuated in patients with endothelial
dysfunction as compared to that in healthy subjects. Whereas
controls were not taking regular medication, all patients were
on oral therapy with ACE inhibitors, statins, β-blockers, and
organic nitrates (Table 1). Participants fasted and refrained
from smoking at least 12 h prior to and until completion of
the investigation. Medication was discontinued the day before
investigation.
In our laboratory, the normal value for FMD as determined in
subjects without major cardiovascular risk factors (hypertension, smoking, hypercholesterolemia, diabetes) is 6.5 ± 0.4%
(95% confidence interval for the mean: 5.8–7.3%). Therefore,
we used the lower boundary limit of 5.8% as cutoff to diagnose
endothelial dysfunction, [2,14–16].
a relaxation of the peripheral resistance vessels as induced by
forearm ischemia and generates shear stress on the endothelium
of the brachial artery. Consistent with shear-stress-induced
vasodilation secondary to activation of eNOS, the diameter of
the brachial artery increased progressively during reactive
hyperemia with maximal values at 45–60 s (open symbols in
Fig. 1A).
The time course of plasma nitrite was qualitatively similar to
the time course of brachial artery diameter changes during
hyperemia (Fig. 1B). Nitrite was instantaneously increasing
after completion of ischemia and reached maximal values at
50 s. We calculated the nitrite reserve as percentage concentration increase at 50 s in relation to baseline values. The local
nitrite concentration increased from 49 ± 9 to 164 ± 21 nmol/L.
The absolute increase in nitrite of 115 nmol/L is equivalent to a
nitrite reserve of 235%. No significant changes in plasma nitrate
were seen (23 ± 7 to 21 ± 8 μmol/L; n.s.).
Nitrite fails to increase during reactive hyperemia in
endothelial dysfunction
In another set of experiments, we compared the increases in
nitrite and brachial artery diameter in controls and in patients
with endothelial dysfunction (n = 11 each, Figs. 2 and 3).
Surprisingly, baseline nitrite levels were significantly lower in
controls (68 ± 5 nmol/L) than in patients (116 ± 12 nmol/L),
whereas no difference was seen in baseline nitrate levels
(26.8 ± 2.4 μmol/L vs 26.1 ± 1.8 μmol/L; control vs endothelial
dysfunction; n.s.). At 50 s after release of lower arm occlusion,
Time course of brachial artery flow velocity, brachial diameter,
and plasma nitrite
We compared the time courses of brachial artery diameter
and blood flow velocity during reactive hyperemia in a
subgroup of controls (n = 5). Baseline diameter and flow
velocity of the brachial artery were 4.0 ± 0.3 mm and 20 ± 4 cm/
s, respectively. Immediately after deflation of the blood pressure
cuff, the flow velocity was significantly increased (57 ± 12 cm/
s), decreased gradually, and reached baseline values at 60 s
(filled symbols in Fig. 1A). The increased flow velocity reflects
Fig. 3. Percentage increase in plasma nitrite during reactive hyperemia of the
forearm in individuals with and without endothelial dysfunction as defined by
flow-mediated dilation of the brachial artery (FMD) of <5.8%.
T. Rassaf et al. / Free Radical Biology & Medicine 41 (2006) 295–301
plasma nitrite significantly increased in controls to
126 ± 13 nmol/L (94 ± 21% increase) but remained unaffected
in patients with endothelial dysfunction at 104 ± 10 nmol/L
(−8 ± 4%). The baseline brachial artery diameter was
significantly greater in endothelial dysfunction patients
(5.1 ± 0.2 mm) than in controls (4.0 ± 0.2 mm). In both
groups, the diameter of the brachial artery increased significantly to 4.3 ± 0.2 and 5.3 ± 0.2 mm, respectively, at 60 s after
deflation of the blood pressure cuff around the forearm. The
increase in brachial artery diameter (FMD) was significantly
greater in controls than in endothelial dysfunction patients
(8.5 ± 0.4% vs 2.9 ± 0.5%, p < 0.001).
Discussion
The key findings of the present study are that (i) the plasma
nitrite in the antecubital vein of healthy subjects increases
during reactive hyperemia of the forearm, (ii) the changes in
plasma nitrite reflect changes in flow-mediated dilation, and (iii)
the increase in nitrite is abolished in patients with endothelial
dysfunction.
Origin of plasma nitrite
Plasma nitrite levels are conserved across various mammalian species, including humans, in the range of 150–600 nmol/L
[17]. Apart from plasma, nitrite is also transported within red
blood cells [18]. The net concentration of nitrite in plasma is a
result of its formation and consumption. Several routes of
formation of nitrite exist in mammals. Nitrite is an oxidation
product of endothelium-derived NO. We and others [5,17] have
recently shown that up to 70–90% of circulating plasma nitrite
is derived from eNOS activity in humans and other mammals.
Moreover, nitrite is present in food, especially in processed
meat, in which nitrite is used to prevent botulism [19].
Furthermore, plasma nitrite increases after ingestion of large
amounts of inorganic nitrate. This increase is entirely due to
enterosalivary circulation of nitrate (as much as 25% is actively
taken up by the salivary glands) and reduction to nitrite by
commensal bacteria [19]. This nitrite enters the circulation
when saliva is swallowed [20]. The formation of nitrite is
counterbalanced by several pathways of elimination. Nitrite can
be oxidized to nitrate by oxyhemoglobin [21] in a reaction that
is by far slower than the oxidation of NO to nitrite. In addition,
nitrite can be reduced to NO under acidic conditions [22–25].
However, this will occur only at a pH of less than 7, which is
seen in tissues during ischemia [26]. Moreover it has been
shown that xanthine oxidase may reduce nitrite to NO [27].
Nitrite reduction by xanthine oxidase is greatly enhanced at low
oxygen tensions and acidic conditions such as those seen during
ischemia. Furthermore, nitrite is recycled back into bioactive
NO via reduction by desoxyhemoglobin [22]. It is suggested
that this mechanism ensures an autoregulated NO generation in
regions of poor oxygenation where desoxyhemoglobin predominates [28].
The different routes of formation and metabolism result in a
biologically relevant steady state concentration of nitrite. Given
299
that the plasma nitrite pool is under regulative control, we here
show that plasma nitrite increases during reactive hyperemia
and that this increase correlates with NO-dependent flowmediated dilation of the upstream brachial artery.
Nitrite and endothelial function
FMD (8.5 ± 0.4%) was significantly higher in young
healthy volunteers than in patients with cardiovascular risk
factors (2.9 ± 0.5%), consistent with endothelial dysfunction
in these patients. The measurement of flow-mediated dilation
of the brachial artery as a noninvasive endothelial function
test in humans has been used by numerous groups to
monitor endothelial function. This ultrasound method quantifies the dilation of conduit arteries in response to
physiologically relevant increases in laminar shear stress
induced by ischemic dilation of the downstream microvasculature. Increases in shear stress lead to a rapid activation of
eNOS with consecutive increases in NO formation. Accordingly, FMD is largely abolished following NOS inhibition
and therefore provides a valuable “read-out” of local vascular
NO availability. In analogy to FMD, we sought to
biochemically determine the capacity of eNOS activity.
Plasma nitrite concentrations were determined at baseline
and during reactive hyperemia in the forearm. Completion of
ischemia almost instantaneously increased plasma nitrite in
the forearm with a peak 50 s after cuff release. The time
course of nitrite mirrored the change in brachial artery
diameter. Interestingly, the regional nitrite peak preceded the
maximum dilation of the brachial artery and decreased faster
(Fig. 1). This suggests that the increase in nitrite is related to
the temporal shear-stress-induced activation of eNOS in the
upstream brachial artery. Shear-stress-induced release of NO
dilates the brachial artery. Part of the NO may be converted
to nitrite and is therefore detectable downstream of the
antecubital vein. This is further corroborated by the fact that
patients, who showed impaired dilatory responses of the
brachial artery, also lacked nitrite increases.
Nevertheless, the present study cannot exclude the possibility that there may be other nitrite sources in the forearm and
that nitrite may be involved in the observed vascular responses.
We have recently shown that nitrite affects cyclic GMP
production [29] and that tissues contain huge amounts of nitrite
[30]. It can therefore not be excluded that part of the nitrite
detected is released from the forearm and that nitrite dilated the
brachial artery and is therefore involved in the flow-mediated
dilation. Moreover, nitrite may also have been transported from
the conduit artery along the vascular tree to the resistance
arteries. Low pO2 levels together with sufficient concentrations
of nitrite may allow deoxyhemoglobin to act as a nitritereductase and as a vasodilator [28,31,32]. Although the patients
showed higher plasma nitrite concentrations at baseline, they
failed to show further nitrite increases during hyperemia.
Potentially, elevated NO scavenging in patients with cardiovascular disease may impair NO generation not only from
eNOS but also from other sources including nitrite. Whether
the reason for the abolished nitrite increase in endothelial
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dysfunction is impairment in eNOS activity, a microvascular
disease with reduction of peripheral flow reserve in hypertensive patients [33], or accelerated consumption of nitrite is not
clear. Measurements of pO2, pCO2, lactate, and arterial-venous
gradients of nitrite are needed to identify the mechanism of
nitrite increase.
Plasma nitrite reserve and diagnostic relevance
We measured local increases in plasma nitrite following
reactive hyperemia of the forearm in healthy volunteers and
compared this to patients with endothelial dysfunction. Whereas
all control subjects responded to forearm ischemia with an
increase in regional plasma nitrite (Fig. 2), we saw no increase
in patients with endothelial dysfunction.
To conduct a proof of concept study examining the
significance of eNOS capacity for the assessment of endothelial function, we compared healthy young subjects to old
patients with documented three-vessel coronary artery disease
and more than three cardiovascular risk factors (i.e. hypertension, diabetes, dislipoproteinemia). Larger studies considering
gender, age, and the respective cardiovascular risk factor are
needed to establish a cutoff value for nitrite for use in clinical
routine.
Combined with new techniques to conserve nitrite in blood
samples [34] our approach opens new avenues for a simple
method to assess endothelial function. This method is
independent of the operator and does not need complex
ultrasound techniques. This would allow the routine assessment
of endothelial function in patients at increased cardiovascular
risk. Moreover, conserving nitrite in samples allows shipping
and transport over long distances and therefore opens new
possibilities for multi center randomized control trials. Formerly,
nitrate (or NOx = the sum of nitrite and nitrate) has been used in
an attempt to assess endothelial function [35]. The reliability of
this approach, however, requires critical reassessment. Plasma
nitrate levels are influenced by a variety of NOS-independent
factors [36]. Furthermore, the high background concentration of
nitrate and its relatively long half-life in comparison to that of
nitrite explain the low sensitivity and therefore limit the usability
of nitrate for the quantification of eNOS activity.
We here show that the local forearm plasma nitrite
concentration fails to increase in patients with endothelial
dysfunction during reactive hyperemia. Applying a biochemical
approach to determine eNOS capacity, we here open new
pathways in the diagnosis and assessment of endothelial
dysfunction.
Acknowledgments
TR and UHC are supported by a grant from the
Forschungskommission of the Heinrich-Heine-UniversityDuesseldorf, Germany. MK and PK are supported by the
Deutsche Forschungsgemeinschaft (SFB 612). This work was
supported by awards from the American Heart Association
(CH, AL). The authors thank Dr. Yerem Yeghiazarians for
discussions and support.
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