From Cardiovascular Research, Institute of Physiology, University of
Zürich (B.S.O., M.R.T.); Cardiology, University Hospital Zürich
(T.F.L.); and Neurosurgery, University Hospital Bern (N.G.), Switzerland; and
the Department of Chemistry, Institute of Biotechnology, Oakland University,
Rochester, Minn (V.B., T.M.).
Correspondence to Thomas F. Lüscher, MD, Professor and Head of Cardiology, University Hospital Zürich, CH-8091 Zürich, Switzerland. E-mail karluh{at}usz.unizh.ch
Methods and Results—With a porphyrinic microsensor, NO release
was measured in atherosclerotic human carotid arteries and normal
mammary arteries obtained during surgery. eNOS protein expression was
analyzed by immunohistochemistry. In normal arteries, the
initial rate of NO release after stimulation with calcium ionophore
A23187 (10 µmol/L) was 0.42±0.05 (µmol/L)/s (n=10). In
contrast, the initial rate of NO release was markedly reduced in
atherosclerotic segments, to 0.08±0.04 (µmol/L)/s (n=10,
P<0.0001). NO peak concentration in normal arteries was
0.9±0.09 µmol/L (n=10) and in atherosclerotic segments,
0.1±0.03 µmol/L (n=10, P<0.0001). Reduced NO
release in atherosclerotic segments was accompanied by marked reduction
of immunoreactive eNOS in luminal endothelial cells,
although specific endothelial cell markers (CD31) were
present (n=13). Endothelial cells of vasa vasorum
of atherosclerotic segments, however, remained positive for eNOS, as
was the endothelium of normal arteries.
Conclusions—In clinically relevant human
atherosclerosis, eNOS protein expression and NO release
are markedly reduced. This may be involved in the progression of
atherosclerosis.
In endothelial cells, NO is formed from
L-arginine8 9 by
eNOS.10 eNOS is constitutively expressed and
activated by cell surface receptors or mechanical forces such
as shear stress and stretch.1 11 NO relaxes
vascular smooth muscle, inhibits platelet activation, and modulates
migration and growth of vascular smooth muscle. Furthermore, NO
regulates genes that lead to the expression of adhesion molecules for
monocytes.12 13 14
Indirect evidence suggests that alterations in the NO pathway might be
involved in endothelial dysfunction and
atherosclerosis. In
hypercholesterolemia and
atherosclerosis, endothelium-dependent
relaxation is reduced.3 4 5 Chronic administration
of L-arginine to hypercholesterolemic
animals improves endothelium-dependent
relaxation.6 In patients with coronary
artery disease, basal NO release seems to be impaired, as suggested by
a blunted response to L-NMMA.7 However, in human
atherosclerosis, no direct measurement of NO release or
NOS protein has been reported.
This study presents the first direct evidence that NO release and
eNOS expression are markedly reduced in clinically manifest human
atherosclerosis.
Porphyrinic NO Microsensor
Experimental Protocols
Immunohistochemistry
Calculations and Statistical Analysis
Expression of eNOS
Expression of Endothelial Cell Marker
Endothelial NO plays an important role in vascular
homeostasis, adhesion of white blood cells, and platelet
function.10 There are at least 3 major mechanisms
in the NO pathway that may lead to endothelial
dysfunction and consequently atherogenesis: (1) functional
abnormalities of NOS due to substrate or cofactor
deficiency10 18 ; (2) increased breakdown of
NO3 5 ; or (3) reduced expression of eNOS. Until
now, even with invasive in vivo studies in humans, no clear distinction
between these possibilities could be made.7
Moreover, studies of the NO pathway in rabbit models of
atherosclerosis gave unexpected results, ie, both eNOS
protein and mRNA are increased in the atherosclerotic aorta of these
animals, despite impaired endothelium-dependent
vascular relaxation (most likely due to inactivation of NO by
superoxide4 ). In contrast, our results provide
the first direct evidence that eNOS protein expression is markedly
diminished in advanced human atherosclerosis and NO
release is reduced. These findings are supported by studies in
explanted vein grafts in which eNOS expression is reduced only in
diseased segments.19 The differences in NO
release observed in normal and atherosclerotic segments were mirrored
by marked differences in NOS expression and are in line with previous
work showing potent endothelium-dependent relaxations
in normal mammary arteries20 and reduced
responses in atherosclerotic human coronary
arteries.21
We confirmed the presence of endothelial cells in
both normal and atherosclerotic segments by light microscopy and by
endothelial cell marker (CD31). A
heterogeneity in vascular beds is an unlikely
explanation for the observed differences, because both mammary and
carotid arteries are branches of the subclavian artery and
brachiocephalic trunk. Differences in NO production due to
anatomic heterogeneity in receptor expression could be
excluded by use of the receptor-independent agonist calcium ionophore.
Moreover, the amount of NO released from normal arteries was comparable
to that of other normal vessels.17 Finally, it
should be noted that IMA samples were obtained from age-matched
patients with documented atherosclerosis in the
coronary circulation and angiographically normal mammary
artery.
The discrepancies between our findings in human
atherosclerosis and animal models might be explained by
the duration of the atherosclerotic process, ie, months in Watanabe
rabbits versus decades in patients. Furthermore, the severity of
clinically relevant lesions and species differences may play a role.
Indeed, vascular lesions of hypercholesterolemic
rabbits mimic human plaques only in part. Nevertheless, it is
conceivable that eNOS protein expression and NO release may be normal
or increased in early human atherosclerosis, whereas at
later stages NOS expression and NO release are reduced. Hence, abnormal
endothelium-dependent responses during early
atherosclerosis might be due to reduced substrate
and/or cofactor concentrations or increased superoxide formation. This
could explain why the administration of
L-arginine22 or the cofactor
tetrahydrobiopterin18 improves
endothelial function only in early stages of
atherosclerosis and in the microcirculation, where no
plaques develop, but not in epicardial coronary arteries of
patients with clinically relevant coronary
disease.22 Thus, at later stages of the
atherosclerotic process, when eNOS expression and NO production
are decreased, these therapies are no longer effective.
Received March 3, 1998;
revision received April 21, 1998;
accepted April 28, 1998.
2.
Ross R. The pathogenesis of
atherosclerosis: a perspective for the 1990s.
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4.
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Suematsu M, Hayashi Y, Itoh H, Yokoyama M. Endothelial
constitutive nitric oxide synthase protein and mRNA increased in rabbit
atherosclerotic aorta despite impaired
endothelium-dependent vascular relaxation.
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Billingham ME. Antiatherogenic effects of L-arginine in the
hypercholesterolemic rabbit. J Clin
Invest. 1992;90:1168–1172.
7.
Quyyumi AA, Dakak N, Mulcahy D, Andrews NP, Husain S,
Panza JA, Cannon RO. Nitric oxide activity in the atherosclerotic human
coronary circulation. J Am Coll Cardiol. 1997;29:308–317.[Abstract]
8.
Palmer RM, Ferrige AG, Moncada S. Nitric oxide release
accounts for the biological activity of
endothelium-derived relaxing factor. Nature. 1987;327:524–526.[Medline]
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Palmer RM, Ashton DS, Moncada S. Vascular
endothelial cells synthesize nitric oxide from
L-arginine. Nature. 1988;333:664–666.[Medline]
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10.
Cooke JP, Dzau VJ. Nitric oxide synthase: role in the
genesis of vascular disease. Annu Rev Med. 1997;48:489–509.[Medline]
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11.
Topper JN, Cai J, Falb D, Gimbrone MA Jr.
Identification of vascular endothelial genes
differentially responsive to fluid mechanical stimuli:
cyclooxygenase-2, manganese superoxide dismutase,
and endothelial cell nitric oxide synthase are
selectively up-regulated by steady laminar shear stress. Proc
Natl Acad Sci U S A. 1996;93:10417–10422.
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Khan BV, Harrison DG, Olbrych MT, Alexander RW, Medford
RM. Nitric oxide regulates vascular cell adhesion molecule 1 gene
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© 1998 American Heart Association, Inc.
Brief Rapid Communications
Reduced Endothelial Nitric Oxide Synthase Expression and Production in Human Atherosclerosis
Abstract
Top
Abstract
Introduction
Methods
Results
Discussion
References
Background—NO regulates vascular
tone and structure, platelets, and monocytes. NO is synthesized by
endothelial NO synthase (eNOS).
Endothelial dysfunction occurs in
atherosclerosis.
Key Words: atherosclerosis • arteries • endothelium • nitric oxide • stroke
Introduction
Top
Abstract
Introduction
Methods
Results
Discussion
References
Atherosclerosis
accounts for half of the morbidity and mortality in Western countries.
Its pathogenesis, however, is not clear. Risk factors of
atherosclerosis, ie, LDL levels, diabetes mellitus,
hypertension, and smoking, are associated with vascular dysfunction,
including monocyte adhesion and invasion, smooth muscle proliferation
and migration, platelet activation, and extracellular matrix
formation.1 2 In animal models, the
endothelial NO pathway appears to be involved in
atherosclerosis.3 4 5 6 However,
little is known about this in human
atherosclerosis.7
Methods
Top
Abstract
Introduction
Methods
Results
Discussion
References
Blood Vessels
Thirty-five arteries of patients undergoing carotid atherectomy
or coronary bypass surgery were collected. For NO measurement,
carotid arteries from 10 patients (mean age, 66 years; range, 53 to 78
years) and IMAs (normal segments) from 10 age-matched control subjects
(mean age, 66 years; range, 54 to 75 years; P=NS) were used.
The presence or absence of atherosclerosis was
confirmed visually, microscopically, and by histology. The absence of
atherosclerosis in IMAs is in line with previous
studies.15 Immunohistochemistry for eNOS and CD31
was performed in 13 carotid arteries and 2 IMAs.
Measurements of NO were carried out with a porphyrinic
microsensor as described16 17 immediately after
surgical removal of segments (<30 minutes). The amperometric signal
was recorded with a chart recorder (Recom Electronic AG), and
NO concentration was determined from the measured current by means of a
calibration curve with NO standards. Standard NO solutions (1.8
mmol/L) were prepared from aqueous solution saturated with pure gaseous
NO (Garbagas).
Isolated vascular segments were cut longitudinally and pinned on
the bottom of organ chambers filled with fresh HBSS buffer. The active
tip of the L-shaped NO microsensor was placed on the
endothelial surface of the IMA or carotid strips. A
precision stereo zoom microscope (PZM) and a micromanipulator (M3301)
were used for positioning. Ten microliters of a 10 µmol/L
calcium ionophore A23187 solution was injected on the luminal side of
the vascular strips with a pneumatic picoinjector (PV820) positioned
with a micromanipulator (PZM, PV820, and M3301 are all from World
Precision Instruments). In each tissue sample, NO release was measured
at 3 different anatomic sites.
Serial paraformaldehyde-fixed paraffin sections
were mounted onto silane-coated slides, dewaxed and rehydrated, washed
in PBS, and incubated in 1 µg/mL anti-eNOS monoclonal antibodies
(Transduction Laboratories; 2 hours at room temperature or overnight at
4°C). Adjacent sections were incubated in a 1:100 dilution of primary
monoclonal antibodies for CD-31 (JC/70A; Dako) or preimmune control
serum for 2 hours at room temperature, washed in PBS for 10 minutes,
incubated in biotinylated secondary antibody for 30 minutes at room
temperature, and visualized by use of the avidin-biotin-peroxidase
labeling system (ABC-elite kit; Vector Laboratories). Sections were
counterstained with hematoxylin and mounted with Kaiser's
solution (Merck).
For statistical analysis, the initial rate of NO release
[slope of NO peak, (µmol/L)/s] and the maximal concentration of NO
produced (NO peak; µmol/L) were measured. In each tissue sample,
NO release was measured at 3 different anatomic sites, and the mean
value was calculated for n=1. In each set of experiments, n is the
number of blood vessels studied. Data are given as mean±SEM.
Statistical analysis was performed with unpaired Student's
t test. Values of P<0.05 were considered
statistically significant.
Results
Top
Abstract
Introduction
Methods
Results
Discussion
References
NO Release
Representative amperometric curves of NO release
are shown in Figure 1A
. In normal
segments, a rapid release of NO was observed after injection of 10
µmol/L calcium ionophore A23187 solution (Figure 1A, top). In
contrast, the initial rate of NO release and NO peak concentration were
significantly reduced in atherosclerotic carotid arteries (Figure 1A, bottom). The initial rate of NO release after calcium ionophore
administration was 0.42±0.05 (µmol/L)/s in normal arteries but only
0.08±0.04 (µmol/L)/s in atherosclerotic carotid arteries (Figure 1B, left; n=10, P<0.0001). NO peak concentration was
0.9±0.09 µmol/L in normal arteries but was reduced to
0.1±0.03 µmol/L in atherosclerotic arteries (Figure 1B, right;
n=10, P<0.0001).
View larger version (18K):
[in a new window]
Figure 1. A, Amperograms of NO release from normal IMA (top)
and atherosclerotic carotid artery (bottom) to calcium ionophore A23187
(10 µmol/L) measured on endothelial surface with
a porphyrinic microsensor. B, Initial rate of NO release (left) and
maximal concentration of NO (right) to calcium ionophore A23187
(10 µmol/L) of isolated IMAs (solid bars) and carotid arteries
(open bars). Values are mean±SEM (P<0.0001,
n=10).
Labeling of normal arterial segments with monoclonal
antibody for eNOS revealed high-level expression of eNOS in luminal
endothelial cells (Figure 2A). In contrast, immunoreactive eNOS was
undetectable in luminal endothelial cells of carotid
segments with advanced atherosclerosis (Figure 3A). However, endothelial
cells of the vasa vasorum inside the atherosclerotic plaques were
strongly positive for eNOS staining (Figure 4A and 4C), indicating that the negative
staining in luminal endothelial cells was not due to
technical failure but rather reflects downregulation of the enzyme.
Smooth muscle cells of normal segments and of atherosclerotic carotid
arteries as well as foam cells and macrophages in the plaque
were negative for eNOS.
View larger version (98K):
[in a new window]
Figure 2. Immunohistology of normal IMA. Serial sections
were labeled with anti-eNOS antibody (A), control nonimmune serum (B),
or monoclonal anti-CD31 antibody (C). Positive staining is indicated by
dark purple color of luminal endothelial cells.
Positive staining for eNOS (A) demonstrated normal expression of eNOS
protein in CD31-positive (C) endothelial cells. Arrows
indicate position of internal elastic lamina. Sections were
counterstained with hematoxylin. Magnification x400. LU indicates
lumen.
View larger version (88K):
[in a new window]
Figure 3. Immunohistology of atherosclerotic carotid artery.
Serial sections were labeled with anti-eNOS antibody (A), control
nonimmune serum (B), or monoclonal anti-CD31 antibody (C). Positive
staining is indicated by dark purple color of luminal
endothelial cells. Negative staining for eNOS (A)
demonstrated diminished expression of eNOS protein in CD31-positive (C)
endothelial cells. Arrows indicate intact
endothelial cell layer in these areas. Sections were
counterstained with hematoxylin. Magnification x400. LU indicates
lumen.
View larger version (136K):
[in a new window]
Figure 4. Immunohistology of atherosclerotic carotid artery.
Serial sections were labeled with anti-eNOS antibody (A and C), control
nonimmune serum (B), or monoclonal anti-CD31 antibody (D). Positive
staining is indicated by dark purple color. Endothelial
cells of vasa vasorum inside plaque are positive for eNOS. Positive
labeling for CD31 confirms endothelial cells. Sections
were counterstained with hematoxylin. Magnification x400.
Anti-CD31 staining, a specific marker for
endothelial cells, was positive in all instances,
indicating the presence of endothelial cells (Figures 2C
, 3C, and 4D) and demonstrating that both eNOS-expressing cells in
normal arterial segments and eNOS-negative cells in
atherosclerotic carotid arteries were indeed
endothelial cells. The negative staining with control
serum demonstrated the specificity of both anti-eNOS and anti-CD31
antibodies (Figures 2B, 3B, and 4B).
Discussion
Top
Abstract
Introduction
Methods
Results
Discussion
References
This study provides the first direct evidence that NOS protein
expression and NO release are diminished in advanced human
atherosclerosis. Initial and peak NO release as
determined by direct in situ measurement with a porphyrinic sensor were
markedly reduced in atherosclerotic carotid arteries. The impaired NO
release was accompanied by a substantial reduction of immunoreactive
eNOS protein in luminal endothelial cells of
atherosclerotic plaques but not in vasa vasorum.
Selected Abbreviations and Acronyms
eNOS
=
endothelial NO synthase
IMA
=
internal mammary artery
L-NMMA
=
NG-monomethyl-L-arginine
NOS
=
NO synthase
Acknowledgments
This work was supported by Swiss National Science Foundation
grants 32–51069.97, 32–45878.95, and 31–047119.96/1; the Swiss
Cardiology Foundation; and in part by grant HL-55397
from the US Public Health Service. The help of Drs D.S.
Hartman, K. Hischikawa, and M. Pech is appreciated.
References
Top
Abstract
Introduction
Methods
Results
Discussion
References
1.
Lüscher TF, Noll G. The
endothelium in coronary vascular control.
Heart Disease. 1995;Update 3:1–10.
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 V. STANGL, M. LORENZ, S. MEINERS, A. LUDWIG, C. BARTSCH, M. MOOBED, A. VIETZKE, H.-T. KINKEL, G. BAUMANN, and K. STANGL Long-term up-regulation of eNOS and improvement of endothelial function by inhibition of the ubiquitin-proteasome pathway FASEB J, February 1, 2004; 18(2): 272 - 279. [Abstract] [Full Text] [PDF]
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 J U Kim, H K Chang, S S Lee, J W Kim, K T Kim, S W Lee, and W T Chung Endothelial nitric oxide synthase gene polymorphisms in Behcet's disease and rheumatic diseases with vasculitis Ann Rheum Dis, November 1, 2003; 62(11): 1083 - 1087. [Abstract] [Full Text] [PDF]
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 B. B Chesebro, E. Blessing, C.-C. Kuo, M. E Rosenfeld, M. Puolakkainen, and L. A. Campbell Nitric oxide synthase plays a role in Chlamydia pneumoniae-induced atherosclerosis Cardiovasc Res, October 15, 2003; 60(1): 170 - 174. [Abstract] [Full Text] [PDF]
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 S. R. Archacki, G. Angheloiu, X.-L. Tian, F. L. Tan, N. DiPaola, G.-Q. Shen, C. Moravec, S. Ellis, E. J. Topol, and Q. Wang Identification of new genes differentially expressed in coronary artery disease by expression profiling Physiol Genomics, September 29, 2003; 15(1): 65 - 74. [Abstract] [Full Text] [PDF]
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 A.-L. Yang, C. J. Jen, and H.-i. Chen Effects of high-cholesterol diet and parallel exercise training on the vascular function of rabbit aortas: a time course study J Appl Physiol, September 1, 2003; 95(3): 1194 - 1200. [Abstract] [Full Text] [PDF]
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 K. A. Griffiths, M. A. Sader, M. R. Skilton, J. A. Harmer, and D. S. Celermajer Effects of raloxifene on endothelium-dependent dilation, lipoproteins, and markers of vascular function in postmenopausal women with coronary artery disease J. Am. Coll. Cardiol., August 20, 2003; 42(4): 698 - 704. [Abstract] [Full Text] [PDF]
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A. Zulli, R. E. Widdop, D. L. Hare, B. F. Buxton, and M. J. Black High Methionine and Cholesterol Diet Abolishes Endothelial Relaxation Arterioscler Thromb Vasc Biol, August 1, 2003; 23(8): 1358 - 1363. [Abstract] [Full Text] [PDF]
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S. Ulker, D. McMaster, P. P. McKeown, and U. Bayraktutan Impaired activities of antioxidant enzymes elicit endothelial dysfunction in spontaneous hypertensive rats despite enhanced vascular nitric oxide generation Cardiovasc Res, August 1, 2003; 59(2): 488 - 500. [Abstract] [Full Text] [PDF]
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O. Gorchakova, W. Koch, N. von Beckerath, J. Mehilli, A. Schomig, and A. Kastrati Association of a genetic variant of endothelial nitric oxide synthase with the 1 year clinical outcome after coronary stent placement Eur. Heart J., May 1, 2003; 24(9): 820 - 827. [Abstract] [Full Text] [PDF]
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 T. Ueda, S. Taniguchi, T. Kawata, K. Mizuguchi, M. Nakajima, and A. Yoshioka Does skeletonization compromise the integrity of internal thoracic artery grafts? Ann. Thorac. Surg., May 1, 2003; 75(5): 1429 - 1433. [Abstract] [Full Text] [PDF]
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T. Cyrus, Y. Yao, J. Rokach, L. X. Tang, and D. Pratico Vitamin E Reduces Progression of Atherosclerosis in Low-Density Lipoprotein Receptor-Deficient Mice With Established Vascular Lesions Circulation, February 4, 2003; 107(4): 521 - 523. [Abstract] [Full Text] [PDF]
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M. Aikawa, S. Sugiyama, C. C. Hill, S. J. Voglic, E. Rabkin, Y. Fukumoto, F. J. Schoen, J. L. Witztum, and P. Libby Lipid Lowering Reduces Oxidative Stress and Endothelial Cell Activation in Rabbit Atheroma Circulation, September 10, 2002; 106(11): 1390 - 1396. [Abstract] [Full Text] [PDF]
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M. G. Modena, L. Bonetti, F. Coppi, F. Bursi, and R. Rossi Prognostic role of reversible endothelial dysfunction in hypertensive postmenopausal women J. Am. Coll. Cardiol., August 7, 2002; 40(3): 505 - 510. [Abstract] [Full Text] [PDF]
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 G. Ghilardi, M. L. Biondi, M. DeMonti, M. Bernini, O. Turri, F. Massaro, E. Guagnellini, and R. Scorza Independent Risk Factor for Moderate to Severe Internal Carotid Artery Stenosis: T786C Mutation of the Endothelial Nitric Oxide Synthase Gene Clin. Chem., July 1, 2002; 48(7): 989 - 993. [Abstract] [Full Text] [PDF]
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C. J. Jen, H.-P. Chan, and H.-i. Chen Chronic Exercise Improves Endothelial Calcium Signaling and Vasodilatation in Hypercholesterolemic Rabbit Femoral Artery Arterioscler Thromb Vasc Biol, July 1, 2002; 22(7): 1219 - 1224. [Abstract] [Full Text] [PDF]
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K. G. Lamping Enhanced Contractile Mechanisms in Vasospasm: Is Endothelial Dysfunction the Whole Story? Circulation, April 2, 2002; 105(13): 1520 - 1522. [Full Text] [PDF]
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 D. Hurlimann, F. Ruschitzka, and T.F. Luscher The relationship between the endothelium and the vessel wall Eur. Heart J. Suppl., February 1, 2002; 4(suppl_A): A1 - A7. [Abstract] [PDF]
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M.C. Verhaar, E. Stroes, and T.J. Rabelink Folates and Cardiovascular Disease Arterioscler Thromb Vasc Biol, January 1, 2002; 22(1): 6 - 13. [Abstract] [Full Text] [PDF]
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L. V. d'Uscio, L. A. Smith, and Z. S. Katusic Hypercholesterolemia Impairs Endothelium-Dependent Relaxations in Common Carotid Arteries of Apolipoprotein E-Deficient Mice Stroke, November 1, 2001; 32(11): 2658 - 2664. [Abstract] [Full Text] [PDF]
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H. E. von der Leyen and V. J. Dzau Therapeutic Potential of Nitric Oxide Synthase Gene Manipulation Circulation, June 5, 2001; 103(22): 2760 - 2765. [Full Text] [PDF]
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L. V. d'Uscio, T. A. Baker, C. B. Mantilla, L. Smith, D. Weiler, G. C. Sieck, and Z. S. Katusic Mechanism of Endothelial Dysfunction in Apolipoprotein E-Deficient Mice Arterioscler Thromb Vasc Biol, June 1, 2001; 21(6): 1017 - 1022. [Abstract] [Full Text] [PDF]
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G. Li, S. Tokuno, P. Tahepold, J. Vaage, C. Lowbeer, and G. Valen Preconditioning protects the severely atherosclerotic mouse heart Ann. Thorac. Surg., April 1, 2001; 71(4): 1296 - 1303. [Abstract] [Full Text] [PDF]
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 H M Snow, F Markos, D O'Regan, and K Pollock Characteristics of arterial wall shear stress which cause endothelium-dependent vasodilatation in the anaesthetized dog J. Physiol., March 15, 2001; 531(3): 843 - 848. [Abstract] [Full Text] [PDF]
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C. P. Tiefenbacher, T. Bleeke, C. Vahl, K. Amann, A. Vogt, and W. Kubler Endothelial Dysfunction of Coronary Resistance Arteries Is Improved by Tetrahydrobiopterin in Atherosclerosis Circulation, October 31, 2000; 102(18): 2172 - 2179. [Abstract] [Full Text] [PDF]
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J. P. Cooke Does ADMA Cause Endothelial Dysfunction? Arterioscler Thromb Vasc Biol, September 1, 2000; 20(9): 2032 - 2037. [Abstract] [Full Text] [PDF]
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K. M. Channon, H. Qian, and S. E. George Nitric Oxide Synthase in Atherosclerosis and Vascular Injury : Insights From Experimental Gene Therapy Arterioscler Thromb Vasc Biol, August 1, 2000; 20(8): 1873 - 1881. [Abstract] [Full Text] [PDF]
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 L. R. Queen, B. Xu, K. Horinouchi, I. Fisher, and A. Ferro {beta}2-Adrenoceptors Activate Nitric Oxide Synthase in Human Platelets Circ. Res., July 7, 2000; 87(1): 39 - 44. [Abstract] [Full Text] [PDF]
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G. Valen, G. K Hansson, A. Dumitrescu, and J. Vaage Unstable angina activates myocardial heat shock protein 72, endothelial nitric oxide synthase, and transcription factors NF{kappa}B and AP-1 Cardiovasc Res, July 1, 2000; 47(1): 49 - 56. [Abstract] [Full Text] [PDF]
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U. Landmesser, R. Merten, S. Spiekermann, K. Buttner, H. Drexler, and B. Hornig Vascular Extracellular Superoxide Dismutase Activity in Patients With Coronary Artery Disease : Relation to Endothelium-Dependent Vasodilation Circulation, May 16, 2000; 101(19): 2264 - 2270. [Abstract] [Full Text] [PDF]
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 H. Li and U. Förstermann Structure-Activity Relationship of Staurosporine Analogs in Regulating Expression of Endothelial Nitric-Oxide Synthase Gene Mol. Pharmacol., March 1, 2000; 57(3): 427 - 435. [Abstract] [Full Text]
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T. J. Anderson Assessment and treatment of endothelial dysfunction in humans J. Am. Coll. Cardiol., September 1, 1999; 34(3): 631 - 638. [Full Text] [PDF]
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W. Linz, P. Wohlfart, B. A Scholkens, T. Malinski, and G. Wiemer Interactions among ACE, kinins and NO Cardiovasc Res, August 15, 1999; 43(3): 549 - 561. [Full Text] [PDF]
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P. A. Davis, L. Calo, T. F. Luscher, and B. S. Oemar Is Atherosclerosis a No NO State? • Response Circulation, May 25, 1999; 99 (20): 2709 - 2712. [Full Text] [PDF]
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 J. E. Kal, I. Vergroesen, and H. B. van Wezel The Effect of Nitroglycerin on Pacing-Induced Changes in Myocardial Oxygen Consumption and Metabolic Coronary Vasodilation in Patients with Coronary Artery Disease Anesth. Analg., February 1, 1999; 88(2): 271 - 271. [Abstract] [Full Text] [PDF]
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M. Barton, C. C. Haudenschild, L. V. d'Uscio, S. Shaw, K. Munter, and T. F. Luscher Endothelin ETA receptor blockade restores NO-mediated endothelial function and inhibits atherosclerosis in apolipoprotein E-deficient mice PNAS, November 24, 1998; 95(24): 14367 - 14372. [Abstract] [Full Text] [PDF]
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V. Sauzeau, H. Le Jeune, C. Cario-Toumaniantz, A. Smolenski, S. M. Lohmann, J. Bertoglio, P. Chardin, P. Pacaud, and G. Loirand Cyclic GMP-dependent Protein Kinase Signaling Pathway Inhibits RhoA-induced Ca2+ Sensitization of Contraction in Vascular Smooth Muscle J. Biol. Chem., July 7, 2000; 275(28): 21722 - 21729. [Abstract] [Full Text] [PDF]
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M. Eto, C. Barandier, L. Rathgeb, T. Kozai, H. Joch, Z. Yang, and T. F. Luscher Thrombin Suppresses Endothelial Nitric Oxide Synthase and Upregulates Endothelin-Converting Enzyme-1 Expression by Distinct Pathways: Role of Rho/ROCK and Mitogen-Activated Protein Kinase Circ. Res., September 28, 2001; 89(7): 583 - 590. [Abstract] [Full Text] [PDF]
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