Vol.2, No.12, 1360-1368 (2010)
doi:10.4236/ns.2010.212166
Copyright © 2010 SciRes. Openly accessible at http://www.scirp.org/journal/NS/
Natural Science
The effect of nebivolol on the production of nitric
oxide induced by bacterial lipopolysaccharide and
peptidoglycan in mice
Fadi El-Rami1, Hampartsoum Barsoumian1, Joseph Simaan2, Alexander M. Abdelnoor1*
1Department of Microbiology and Immunology, Faculty of Medicine, American University of Beirut, Beirut, Lebanon;
*Corresponding Author:
2Department of Pharmacology and Therapeutics, Faculty of Medicine, American University of Beirut, Beirut, Lebanon
Received 25 June 2010; revised 18 July 2010; accepted 2 August 2010.
ABSTRACT
Nitric oxide (NO) plays a pivotal role in main-
taining balance of physiological events in many
systems including the autonomic, cardiovas-
cular, hematological, and pulmonary systems.
Lipopolysaccharide (LPS) and peptidoglycan
(PGN), components of the outer cell membranes
of Gram-negative bacteria and cell walls of
Gram-positive bacteria respectively, are in-
criminated in NO-induced septic shock. Ne-
bivolol is a third generation β1-adrenoceptor
blocker with a vasodilatory property attributed
to enhanced availability of nitric oxide and re-
duction of cellular oxidative stress through an
unknown mechanism. The current study ex-
plored the hypothesis that if nebivolol enhances
the availability of NO, pretreatment with ne-
bivolol may enhance production of NO in re-
sponse to subsequent treatment with LPS and
PGN, an observation that may have relevance in
clinical septic shock. Groups of female BALB/c
mice each containing 12 mice (6-8 weeks old)
were injected intraperitoneally with LPS (30
µg/mouse), PGN (100 µg/mouse), nebivolol (0.25
µg/g, 0.35 µg/g, 0.7 µg/g), LPS and nebivolol
(0.25 µg/g), LPS and nebivolol (0.35 µg/g), LPS
and nebivolol (0.7 µg/g), PGN and nebivolol
(0.25 µg/g), PGN and nebivolol (0.35 µg/g). One
group of mice was injected with saline and an-
other served as control. Three mice from each
group were bled 1, 3, 6 and 9 hours post-injec-
tion, the blood was pooled and the nitrite serum
levels, reflecting NO concentration, were de-
termined using Greiss reagent. The following
results were obtained: 1) Treatment with saline
did not induce NO production; 2) LPS induced
NO production to a maximal limit of 545% at 9
hours as compared to treatment with saline; 3)
PGN did not induce NO production; 4) Nebivolol
at most doses and periods (7 out of 10 deter-
minations) increased NO production over a
range of 18-110% as compared to treatment with
saline; 5) Nebivolol enhanced LPS-induced
production of NO by 58% at a dose of 0.7 µg/gm
at 9 hours. It is concluded that nebivolol in-
duces NO production. At low doses nebivolol
initially appeared to have a suppressive or no
effect on NO production induced by LPS. In-
crease in the dose of nebivolol resulted in
augmentation of LPS-induced production of NO.
PGN, in the dose tested, did not have an effect
on NO production.
Keywords: Nebivolol; Peptidoglycan;
Lipopolysaccharide; Nitric Oxide;
β1-Adrenoceptor Blocker
1. INTRODUCTION
Since the early report by Furchgott and Zawadzki in
1980 that the endothelium produces a vasodilator sub-
stance, initially referred to as the endothelium-derived
relaxing factor, later demonstrated to be NO, extensive
research revealed that NO was a short-lived mediator of
numerous physiological as well as pathophysiological
phenomena [1]. NO is synthesized by vascular endothe-
lium, macrophages, neutrophils, Kupffer cells, brain
cells and other cell types through the enzymatic effect of
NO synthase on L-arginine [2]. NO, a very small lipo-
philic molecule with an ultra short half-life less than 5
seconds in biological tissues is a pleiotropic molecule
that is indispensable for various physiological functions
[3]. In the vascular bed, NO is a prominent vasodilator
that relaxes smooth muscle, a potent inhibitor of platelet

F. El-Rami et al. / Natural Science 2 (2010) 1360-1368
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aggregation, and a major mediator of proliferation of
vascular smooth muscle and endothelial cells via a cyclic
GMP-independent mechanism [4-6]. Furthermore, some
studies showed that NO reduces cardiomyocyte apop-
tosis and oxygen consumption despite reduced blood
flow in patients [7,8]. NO is a mediator of defense
mechanisms including its cytotoxic role against foreign
cells through functional disruption, and in tumor cells
through DNA damage and p53 accumulation. However,
elevated NO levels have deleterious effects. NO leads
to increased pain perception through stimulation of
pain-mediating sensory nerves, overproduction of per-
oxynitrite with negative effects on proteins and cell
function, S-nitrosylation of proteins such as transcription
factors, signaling kinases, ion channels and TGF- β,
which elicits conformational modifications leading to
loss of function [9-12].
Peptidoglycan (PGN), a constituent of the bacterial
cell wall, is a polymer of N-acetylglucosamine and N-
acetylmuramic acid residues. Lipopolysaccharide (LPS)
is an amphipathic component of Gram-negative bacterial
outer membrane consisting of 3 parts: the O-antigen
(O-polysaccharide), the core polysaccharide and lipid A
[13]. Both LPS and PGN are believed to contribute to
hypotension and shock in bacteremias. PGN engages
Toll-like receptor 2 (TLR2) and activates the myeloid
differentiation factor 88 (MyD88)-independent pathway
and LPS engages TLR4 that results in activation of both
the MyD88-dependent and independent pathways. Both
pathways lead eventually to an inflammatory response,
mainly through the production of pro-inflammatory cy-
tokines and inducible nitric oxide synthase (iNOS)
[14-16].
Nebivolol is a novel third generation β1-adrenoceptor
blocker [17-19] advocated for the treatment of hyperten-
sion. It is unique among all β1-adrenoceptor blockers in
that it possesses a vasodilatory property attributed to
synthesis of NO [20,21] as well as to increasing NO
bioavailability by decreasing the oxidative stress [22-26].
In addition, Ladage et al. [27] reported that the endothe-
lium-dependent increase in NO induced by nebivolol
was due to stimulation of β3-adrenoceptors and estrogen
receptors. Clinically, nebivolol has been shown to protect
the heart from ischemia, arrhythmia and myocardial
infarction through NO, including limiting oxygen con-
sumption and maintaining cardiac contraction even with
a reduced blood flow, in addition to minimizing
cardiomyocyte apoptosis [28-30].
In bacteremias, bacterial cells release some of their
constituents such as LPS, lipoteichoic acid and PGN.
These constituents stimulate a number of cell types to
produce NO. The current study was undertaken to ex-
plore the effect of nebivolol on PGN- and LPS-induced
NO production, reflected by determination of serum ni-
trite concentration.
2. MATERIALS AND METHODS
2.1. Study Protocol
Experiments were performed on 132 female BALB/c
mice divided into 10 groups of 12 mice each and 2
groups of 6 mice each, treated with intraperitoneal injec-
tions of either saline, LPS, PGN, nebivolol or combina-
tions as summarized in Table 1. All the groups of mice,
except groups 7 and 10, were bled 1, 3, 6 and 9 hours
post-treatment. Prior to bleeding, mice were anesthetized
by an intraperitoneal injection of ketamine and xylazine.
The blood of 3 mice from each group at each time inter-
val was pooled, allowed clotting and the serum was
separated and stored at 4°C for nitrate/nitrite measure-
ment. Groups 7 and 10 were bled at 6 and 9 hours
post-treatment. Serum concentration of nitric oxide was
indirectly measured using the Greiss reagent which de-
termines the concentration of nitrite (NO2
−) and the ni-
trate (NO3
−), the final breakdown products of nitric ox-
ide. The Greiss reagent was used according to the
manufacturer’s instructions (Sigma Chemicals co., MO,
USA). NO has an ultra short half-life in blood that does
not exceed 5 seconds. It dissociates into two final end
products, namely nitrite (NO2
−) and nitrate (NO3−). Gre-
iss reagent changes nitrates to nitrites and measures the
nitrite concentration through a colorimetric reaction, the
concentration of nitrite reflecting the concentration of
NO. Spectrophotometric readings were done at a wave-
length 490nm using a microplate reader. Assays for the
determination of nitrate and nitrite were run twice as
each sample of blood was pooled from 3 mice that re-
ceived the same treatment at the same time interval. The
average of the two determinations was taken as a reflec-
tion of NO concentration and the standard deviation of
the two determinations was calculated indicated repro-
ducibility of the analysis. Comparisons between differ-
ent groups of mice and at different time intervals were
considered significant when there was no overlap be-
tween the means and standard deviations.
2.2. Drugs and Reagents Used
LPS from Salmonella enterica serovar Minnesota
(Sigma Chemicals co., MO, USA, prepared as suspend-
sion of 30 µg/0.5 ml); PGN from Bacillus subtilis
(Sigma Chemicals co., MO, USA, prepared as a suspen-
sion of 100 µg/0.5 ml); nebivolol (Cipla, India, prepared
as dilutions of 0.25 µg/g-0.7 µg/g); ketamine (Pan-
pharma, France, prepared as dilution of 6 mg/0.5 ml);
xylazine (Interchemie, Holland, prepared as dilution of

F. El-Rami et al. / Natural Science 2 (2010) 1360-1368
Copyright © 2010 SciRes. http://www.scirp.org/journal/NS/Openly accessible at
1362
Table 1. Dose of preparations injected to different groups of mice.
Group Number Intraperitoneal Injection of Agent Number of mice Injection volume
Group 1 Negative control 12 0.5 ml
Group 2 Saline (vehicle used to solutions) 12 0.5 ml
Group 3 LPS (30 µg/ mouse) 12 0.5 ml
Group 4 PGN (100 µg/ mouse) 12 0.5 ml
Group 5 n ( 0.25 µg/g) 12 0.5 ml
Group 6 n (0.35 µg/g) 12 0.5 ml
Group 7 n ( 0.7 µg/g) 6 0.5 ml
Group 8 LPS (30 µg/mouse) + n (0.25 µg/g) 12 0.5 ml
Group 9 LPS (30 µg/mouse) + n (0.35 µg/g) 12 0.5 ml
Group 10 LPS (30 µg/mouse) + n ( 0.7 µg/g) 6 0.5 ml
Group 11 PGN (100 µg/mouse) + n (0.25 µg/g) 12 0.5 ml
Group 12 PGN (100 µg/mouse) + n (0.35 µg/g) 12 0.5 ml
n: nebivolol; LPS: lipopolysaccharide; PGN: peptidoglycan.
0.6 mg/0.5 ml); Greiss reagent (Sigma Chemicals co.,
MO, USA). Chemicals were dissolved in pyrogen free
saline, except for the anesthetics which were dissolved
in pyrogen free water. All solvents were confirmed py-
rogen free by the Limulus Amebocyte Lysate (LAL) test.
3. RESULTS
The changes in the serum concentration of nitrites
(µmole/l) in response to various treatments and time
intervals are summarized in Table 2.
3.1. Untreated and Saline-Treated Groups of
Mice
NO levels in mice that were untreated or injected with
saline were approximately the same (Table 2).
3.2. Group of Mice Treated with Nebivolol
Out of 10 assays in different doses at different time
intervals, nebivolol increased the nitrate/nitrite produc-
tion in 7 assays over a range of 18-170% as compared to
saline treated group (Tables 2,3, Figure 1).
3.3. Group of Mice Treated with LPS Alone
As compared to the saline-treated group, there was an
increase with nitrate/nitrite concentration of 93%, 71%,
356% and 545% at time intervals 1, 3, 6 and 9 hours
respectively (Tables 2,4, Figure 2).
3.4. Group of Mice Treated with PGN Alone
PGN had an inconsistent effect on nitrate/nitrite pro-
duction ranging from −27% to +29% at various time
intervals, as compared to saline treated group. (Tables 2,
4, Figure 2).
3.5. Group of Mice Treated with
LPS + Nebivolol
Out of 10 assays in different doses at different time
intervals, there was a decrease or no change in nitrate/
nitrite production in 9 assays, ranging from −4% to −65%
as compared to respective values in response to treat-
ment with LPS alone. However, at a dose of nebivolol of
0.7 µg/gm and an interval of 9 hours, there was an in-
crease in nitrate/nitrite production of 58% as compared
to the respective value after treatment with LPS alone,
implying that LPS-induced NO production is inhibited
by nebivolol in lowest doses at short intervals of expo-
sure and is potentiated at high doses and long interval of
exposure (Tables 2,5, Figure 3).
3.6. Groups of Mice Treated with
PGN + Nebivolol
Treatment with nebivolol in different doses and at
different intervals produced an inconsistent effect of
PGN-induced effect on nitrate/nitrite production varying
from no change in two assays, a decrease of −13% to
−18% in two assays and an increase in four assays of

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Table 2. Total concentration of nitrite in serum, reflecting serum concentration of nitric oxide, under control condi-
tions and in response to various treatments at different time intervals post-challenge.
Concentration of nitrite in serum (µmol/L)
Series Hours post-treatment 1 3 6 9
1 Control 41.5 ± 2.4 43.9 ± 2.4 51.1 ± 0 46.3 ± 2.4
2 Saline 31.2 ± 4.1 48.0 ± 0 44.9 ± 2.4 32.9 ± 2.4
3 LPS: 30 µg /0.5 ml 60.4 ± 11.5 82.3 ± 16.5 204.9 ± 17.4 212.9 ± 9.3
4 PGN: 100 µg /0.5 ml 39.8 ± 4.5 34.7 ± 2.2 29.9 ± 5.4 40.9 ± 10.1
5 n : 0.25 µg/g 37.5 ± 2.4 39.2 ± 1.0 43.8 ± 2.2 42.1 ± 2.9
6 n : 0.35 µg/g 44.7 ± 2.7 58.8 ± 4.7 74.9 ± 10.7 88.8 ± 7.3
7 n : 0.7 µg/g ND ND 32.6 ± 1.0 39.3 ± 3.3
8 LPS+ n: 0.25 µg/g 29.1 ± 1.7 28.7 ± 1.2 113.6 ± 5.7 211.7 ± 8.0
9 LPS+ n: 0.35 µg/g 57.4 ± 10.0 63.3 ± 1.3 175.3 ± 2.2 202.0 ± 11.9
10 LPS+ n: 0.7 µg/g ND ND 197.0 ± 7.7 335.7 ± 7.8
11 PGN+ n: 0.25 µg/g 39.2 ± 3.9 40.2 ± 5.7 34.7 ± 1.2 34.0 ± 3.6
12 PGN+ n: 0.35 µg/g 33.3 ± 4.7 39.9 ± 3.6 26.1 ± 1.3 40.15 ± 1.2
Serum nitrite concentrations were analyzed in duplicate and the average of the two determinations was taken as a reflection of NO concentra-
tion and the standard deviation of the two determinations indicated reproducibility of the analysis. LPS: lipopolysaccharide; PGN: pepti-
doglycan; n: nebivolol; ND: not determined.
Figure 1. Nitrite levels, reflecting NO levels, at 1, 3, 6 and 9 hours after injection with either saline or nebivolol (n).
Columns represent mean of duplicate analysis and the bars standard deviation of the duplicate analysis. Differences
between concentrations of nitrite were considered significant when the means and the standard deviations did not
overlap. n: nebivolol.

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Table 3. Percentage change in nitrite concentration in response to treatment with nebivolol as compared to treatment
with saline at different time intervals post-challenge.
Dose of nebivolol (µg/g) Percent change at hour 1Percent change at hour 3Percent change at hour 6 Percent change at hour 9
0.25 +19 −18 −2.3 +27
0.35 +45 +23 +67 +170
0.7 ND ND −27 +18
ND = not determined
Table 4. Percentage change in nitrite concentration in response to treatment with LPS and PGN as compared to
treatment with saline at different time intervals post-challenge.
Dose (µg/mouse)
LPS PGN
Percent change at hour 1Percent change at hour 3Percent change at hour 6 Percent change at hour 9
30 0 +93 +71 +356 +545
0 100 +29 −27 −33 +24
LPS: lipopolysaccharide, PGN: peptidoglycan.
Table 5. Percentage change in nitrite concentration in response to treatment with LPS and nebivolol in different doses,
as compared to treatment with LPS alone at different time intervals post-challenge.
Dose
LPS
(µg/mouse) N (µg/g)
Percent change at hour 1Percent change at hour 3Percent change at hour 6 Percent change at hour 9
30 0.25 −51 −65 −44 0
30 0.35 −5 −23 −16 −5
30 0.7 ND ND −4 +58
ND = not determined; N = nebivolol; LPS: lipopolysaccharide.
14% to 17% in different concentrations and time inter-
vals as compared to respective values of treatment with
PGN alone (Tables 2,6, Figure 4).
4. DISCUSSION
Nebivolol is a novel third generation β1-adrenoceptor
blocker [17-20] advocated for the treatment of hyperten-
sion. It is unique among all β1-adrenoceptor blockers in
that it possesses a vasodilatory property attributed to
synthesis of NO [21,22] as well as to increasing NO
bioavailability by decreasing the oxidative stress [23-27].
The observation that nebivolol increases the concentra-
tion of nitrite in 7 out of 10 as- says using different doses
and at different time intervals, reflecting an increase in
the NO levels, is confirmatory to observations reported
by others [31-33].
LPS and PGN have been associated with increased
production of NO during sepsis, where NO has been a
major contributor to vascular collapse, a major cause of
mortality in septic shock cases [34,35]. LPS but not
PGN induced NO synthesis. This can be attributed partly
to the different signaling pathways induced by each.
There are at least two pathways that LPS activates, both
of which lead to the production of NO. In the first, LPS
engages TLR4 expressed by macrophages and neutron-
phils. As a result 2 intracellular signaling pathways, the
MyD88-dependent and independent pathways are acti-
vated and lead to the production of pro-inflammatory
cytokines and NO [36-40]. In the second pathway, LPS
induces the production of gamma interferon which in
turn interacts with its receptor expressed on several cell
types [41]. The intracellular events that follow lead to
the activation of the transcription factor, IRF-1, and
production of NO. In support of these results, previous
studies have shown that LPS, rather than PGN, induced
effectively the cytokine expression machinery (TNF-α,
IL-1 α, IL-12, IL-23, IFN-γ, CCL-2, CCL-5) 3 hours
after LPS treatment shifting to a Th1 response as evi-
denced by the high IFN-γ/IL-4 ratio and by the immense

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delayed type hypersensitive response, while PGN fa-
vored the Th2 type [42]. Moreover, Fahmi et al. [43]
have shown that lipoteichoic acid rather than PGN is the
potent inducer of NO upon severe infection with Gram-
positive bacteria and that PGN augmented the lipo-
teichoic acid-NO inducing activity. It is worth noting
that Ida et al. [44] reported that propanolol, a beta
adrenergic receptor antagonist, did not influence cyto-
kine release by lipoteichoic acid.
Nebivolol at doses 0.25 and 0.35 µg/g given in com-
bination with LPS had no, or a suppressive effect on NO
production induced by LPS. Arai et al. [45] reported that
beta adrenergic receptor antagonists (ICI-188551, be-
taxolol, timolol and metipranolol) did not influence NO
production induced by LPS. However, nebivolol at a
dose 0.7 µg/g potentiated NO production induced by
LPS at 9 hours post-injection, implying that the effect of
nebivolol on LPS-induced NO production is dose and
time-dependent. These results could in part be explained
by the study of Broeder et al. [18] who reported that ne-
bivolol does not induce the production of NO. Rather, its
metabolites do so. It appears that a time factor is in-
volved, taking about 9 hours for the production of me-
tabolites which contribute to the delayed effect of ne-
bivolol. It can be hypothesized that intact nebivolol has a
suppressive effect on NO production induced by LPS,
and its metabolites that take about 9 hours to be formed
enhance NO production induced by LPS.
Table 6. Percentage change in nitrite concentration in response to treatment with PGN and nebivolol in different
doses, as compared to treatment with PGN alone at different time intervals post-challenge.
Dose
PGN
(µg/mouse) N (µg/g)
Percent change at hour 1Percent change at hour 3Percent change at hour 6Percent change at hour 9
100 0.25 0 +14 +17 +17
100 0.35 −18 +14 −13 0
100 0.7 ND ND ND ND
ND = not determined; N = nebivolol; LPS: lipopolysaccharide.
Figure 2. Nitrite levels, reflecting NO levels, at 1, 3, 6 and 9 hours after injection of saline, LPS and PGN. Columns
represent mean of duplicate analysis and the bars standard deviation of the duplicate analysis. Differences between
concentrations of nitrite were considered significant when the means and the standard deviations did not overlap. LPS:
lipopolysaccharide (30 µg/mouse); PGN: peptidoglycan (100 µg/mouse).

F. El-Rami et al. / Natural Science 2 (2010) 1360-1368
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1366
Figure 3. Nitrite levels, reflecting NO levels, at 1, 3, 6 and 9 hours after injection of LPS or LPS with nebivolol (n).
Columns represent mean of duplicate analysis and the bars standard deviation of the duplicate analysis. Differences
between concentrations of nitrite were considered significant when the means and the standard deviations did not
overlap. LPS: lipopolysaccharide(30 µg/mouse); n: nebivolol.
Figure 4. Nitrite levels, reflecting NO levels, at 1, 3, 6 and 9 hours after injection of PGN or PGN with nebivolol (n).
Columns represent mean of duplicate analysis and the bars standard deviation of the duplicate analysis. Differences
between concentrations of nitrite were considered significant when the means and the standard deviations did not
overlap. PGN: peptidoglycan (100 µg/mouse); n: nebivolol.
Understanding the relationship between nebivolol and
PGN on NO production machinery is complex due to the
interrelationships among NO induction pathways. Some
studies have indicated that beta blockers do not interact
through the MyD88-dependent or the MyD88-independent
pathways and that the MyD88-dependent pathway that
leads to NO production after PGN engages TLR2, is
unaffected by the administration of beta blockers [46].
Other findings have shown that beta blockers affect PGN
through other PGN recognition receptors such as CD14,
nucleotide oligomerization domain (Nod)-containing
proteins, a family of PGN recognition proteins, and

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PGN-lytic enzymes [47,48].
In conclusion it appears that nebivolol induces the
production of NO. When given with LPS in low dose it
suppresses LPS-induced production of NO, whereas a
high dose at long interval of time enhances this effect.
PGN had no significant effect on NO production and
nebivolol was without additional effect.
REFERENCES
[1] Furchgott, R.F., Zawadzki, J.V. (1980) The obligatory
role of endothelial cells in the relaxation of arterial
smooth muscle by acetylcholine. Nature, 288, 373-376.
[2] Ignarro, L. (1990) Nitric Oxide: A novel transduction
mechanism for transcellular communication. Hyper- ten-
sion, 16, 477-483.
[3] Hobbs, A., Higgs, A. and Moncada, S. (1999) Inhibition
of nitric oxide synthase as a potential therapeutic target.
Annual Review Pharmacology Toxicology, 39, 191-220.
[4] Maffei, A. and Lembo, G. (2009) Nitric oxide mecha-
nisms of nebivolol. Therapeutic Advances in Cardiovas-
cular Disease, 10, 1-11.
[5] Falciani, M., Rinaldi, B. D., Agostino, B., Mazzeo, F.,
Rossi, S. and Nobili, B. (2001) Effects of nebivolol on
human platelet aggregation. Journal of Cardiovascular
Pharmacology, 38, 922-929.
[6] Ignarro, J., Sisodia, M., Trinh, K., Bedrood, S., Wu, G.,
Wei, L. and Buga, G. (2002) Nebivolol inhibits vascular
smooth muscle cell proliferation by mechanisms involv-
ing nitric oxide but not cyclic GMP. Nitric Oxide, 7,
83-90.
[7] Mercanoglu, G., Safran, N., Gungor, M., Pamukcu, B.,
Uzun, H. and Sezgin, C. (2008) The effects of nebivolol
on apoptosis in a rat infarct model. Circulation Journal,
72, 660-670.
[8] Togni, M., Vigorito, F., Windecker, S., Abrecht, L., We-
naweser, P. and Cook, S. (2007) Does the beta-blocker
nebivolol increase coronary flow reserve? Cardiovascu-
lar Drugs and Therapy, 21, 99-108.
[9] Hancock, C.M. and Walsh, D.A. (2008) Modulation of
pain in osteoarthritis: the role of nitric oxide. Clinical
Journal of Pain, 24, 353-365.
[10] Szabo, C. and Radi, R. (2007) Peroxynitrite: biochemis-
try, pathophysiology and development of therapeutics.
Nature Reviews Drug Discovery, 6, 662-680.
[11] Mackenzie, I., Rutherford, D. and MacDonald, T. (2008)
Nitric Oxide and cardiovascular effects: new insights in
the role of nitric oxide for the management of os-
teoarthritis. Arthritis Research & Therapy, 10, 2.
[12] Guix, F., Uribesalgo, I., Coma, M. and Munoz, F. (2005)
The physiology and pathophysiology of nitric oxide in
the brain. Progress in Neurobiology, 76, 126-152.
[13] Raetz, C.R.. (1990) Biochemistry of endotoxin. The An-
nual Review of Biochemistry, 59, 129-170.
[14] Akira, S. and Takeda, K. (2004) Toll-like receptor
signaling. Nature Reviews Immunology, 4, 499-511.
[15] Jiang, Z., Georgel, P., Du, X., Shamel, L., Sovath, S.,
Mudd, S., Huber, M., Kalis, C., Keck, S., Galanos, C.,
Freudenberg, M. and Beutler, B. (2005) CD14 is required
for MyD88-independent LPS signaling. Nature Immu-
nolog, 6, 565-570.
[16] Kitchens, R.L., Thompson, P.A., O’Keefe, G.E. and
Munford, R.S. (2000) Plasma constituents regulate LPS
binding to, and release from, the monocyte cell surface.
Journal of Endotoxin Research, 6, 477-482.
[17] Mangrella, M., Rossi, F., Fici, F. and Rpssi, F. (1998)
Pharmacology of nebivolol. Pharmacological Research,
38, 6.
[18] Broeders, M., Doevendans, P.A. and Van Der Zee, R.
(2000) Nebivolol: A third generation beta-blocker that
augments vascular no release. Circulation, 102, 677-684.
[19] Maffei, A., Pardo, A., Carangi, R., Lembo, G., Carullo, P.,
Poulet, R., Gentile, M. and Vecchione, G. (2007) Ne-
bivolol induces Nitric Oxide release in the heart through
inducible Nitric Oxide synthase activation. Hypertension,
50, 652- 656.
[20] Weiss, R. (2006) Nebivolol: A novel beta-blocker with
nitric Oxide-induced vasodilatation. Vascular Health and
Risk Management, 2, 303-308.
[21] Veverka, A. and Salinas, J. (2007) Nebivolol in treatment
of chronic heart failure. Vascular Health and Risk Man-
agement, 3, 647-654.
[22] Agabiti-Rosei, E., Porteri, E. and Rizzoni, D. (2009)
Arterial stiffness, hypertension, and rational use of ne-
bivolol. Vascular Health and Risk Management, 5,
353-360.
[23] Zanchetti, A. (2004) Clinical pharmacodynamics of ne-
bivolol. Blood Pressure Supplies, 1, 17-32.
[24] Gupta, S. and Wright, H.M. (2008) Nebivolol: A highly
selective beta1-adrenergic receptor blocker that causes
vasodilation by increasing Nitric Oxide. Cardiovascular
Therapeutics, 26, 189-202.
[25] Papademetriou, V. (2008) Beta blockers in management
of hypertension: Focus on nebivolol. Expert Review of
Cardiovascular Th erapy, 6, 471-479.
[26] Judy, W.M. Cheng, B.S. (2009) Nebivolol: A third-gen-
eration P-Blocker for hypertension. Clinical Therapeu-
tics, 31, 3.
[27] Ladage, D., Brixius, K., Hoyer, H., Steingen, C., Wessel-
ing, A. and Malan, D. (2006) Mechanisms underlying
nebivolol-induced endothelial Nitric Oxide synthase ac-
tivation in human umbilical vein endothelial cells. Clini-
cal and Experimental Pharmacology and Physiology, 33,
720-724
[28] Ignarro, J., Sisodia, M., Trinh, K., Bedrood, S., Wu, G.,
Wei, L. and Buga, G. (2002) Nebivolol inhibits vascular
smooth muscle cell proliferation by Mechanisms involv-
ing Nitric Oxide but not cyclic GMP. Nitric Oxide, 7,
83-90.
[29] Maffei, A. and Lembo, G. (2009) Nitric Oxide mecha-
nisms of nebivolol. Therapeutic Advances in Cardiovas-
cular Disease, 10, 1-11.
[30] Togni, M., Vigorito, F., Windecker, S., Abrecht, L., We-
naweser, P. and Cook, S. (2007) Does the beta-blocker
nebivolol increase coronary flow reserve? Cardiovascu-
lar Drugs and Therapy, 21, 99-108.
[31] Maffei, A., Vecchione, C., Aretini, A., Poulet, R., Bet-
tarini, U. and Gentile, M.T. (2006) Characterization of
nitric oxide release by nebivolol and its metabolism.
American Journal of Hypertension, 19, 579-586.
[32] Supajatura, V., Ushio, H., Nakao, A., Akira, S., Okumura,
K., Ra, C. and Ogawa, H. (2002) Differential responses
of mast cell Toll-like receptors 2 and 4 in allergy and in-

F. El-Rami et al. / Natural Science 2 (2010) 1360-1368
Copyright © 2010 SciRes. http://www.scirp.org/journal/NS/Openly accessible at
1368
nate immunity. The Journal of Clinical Investigation, 109,
1351-1359.
[33] Tzemos, N., Lim, P.O. and MacDonald, T.M. (2001)
Nebivolol reverses endothelial dysfunction in essential
hypertension. A randomized, double-blind, crossover
study. Circulation, 104, 511-14.
[34] MacMicking, J.D., Nathan, C., Hom, G., Chartrain, N.,
Fletcher, D.S., Trumbauer, M., Stevens, K., Xie, Q.W.,
Sokol, K. and Hutchinson, N. (1997) Altered responses
to bacterial infection and endotoxic shock in mice lack-
ing inducible nitric oxide synthase. Cell, 8, 641-650.
[35] Cobb, J.P. and Danner, R.L. (1996) Nitric oxide and sep-
tic shock. The Journal of the American Medical Associa-
tion, 275, 1192-1196.
[36] Akira, S. and Takeda, K. (2004) Toll-like receptor
signaling. Nature Reviews Immunology, 4, 499-511.
[37] Bagchi, A., Herrup, E., Warren, E., Trigilio, J., Shin, H.,
Valentine, C. and Hellman, J. (2007) MyD88-dependent
and MyD88-independent pathways in synergy, priming,
and tolerance between TLR agonists. The Journal of
Immunology, 178, 1164-1171.
[38] Kawai, T., Adachi, O., Ogawa, T., Takeda, K. and Akira.
S. (1999) Unresponsiveness of MyD88-deficient mice to
endotoxin. Immunity, 11, 115-122.
[39] Kreutz, M., Ackermann, U., Hauschildt, S., Krause, S.W.,
Riedel, D., Bessler, W. and Andreesen, R. (1997) A com-
parative analysis of cytokine production and tolerance
induction by bacterial lipopeptides, lipopolysaccharides
and Staphyloccous aureus in human monocytes. Immu-
nology, 92, 396-401.
[40] Poltorak, A., He, X., Smirnova, I., Liu, M.Y., Huffel,
C.V., Du, X., Birdwell, D., Alejos, E., Silva, M. and
Galanos, C. (1998) Defective LPS signaling in C3H/HeJ
and C57BL/10ScCr mice: mutations in Tlr4 gene. Sci-
ence, 282, 2085-2088.
[41] Guix, F., Uribesalgo, I., Coma, M. and Munoz, F. (2005)
The physiology and pathophysiology of nitric oxide in
the brain. Progress in Neurobiology ,76, 126-152
[42] Manni, M. and Maestroni, G. (2008) Sympathetic nerv-
ous modulation of the skin innate and adaptive immune
response to peptidoglycan but not lipopolysaccharide:
Involvement of β-adrenoceptors and relevance in in-
flamematory diseases. Brain, Behavior, and Immunity,
22, 80-88.
[43] Fahmi, H., Ancuta, P. and Chaby, R. (1996) Preexposure
of mouse peritoneal macrophages to lipopolysaccharide
and other stimuli enhances nitric oxide response to sec-
ondary stimuli. Inflam mation Research, 45, 347-356.
[44] Ida, A.J., Giebelen, Masja, L., Mark, C.D., Joost, C.M.,
Meijers, M.L., Christian, D., Sonja, von A. and Tom van
der P. (2008) Endogenous adrenergic receptors inhibit
LPS-induced pulmonary cytokine release and coagula-
tion. AJRCMB Articles in Press.
[45] Arai, K., Wood, J.P. and Osborne, N.N. (2003) Beta-
adrenergic receptor agonists and antagonists counteract
LPS-induced neuronal death in retinal cultures by differ-
ent mechanisms. Brain Research, 985, 176-86.
[46] Palsson-McDermott, E. and O’Neill, L. (2004) Signal
transduction by the lipopolysaccharide receptor, Toll-like
receptor-4. Immunology, 113, 153-162.
[47] Dziarski, R. and Gupta, D. (2005) Peptidoglycan recog-
nition in innate immunity. Journal of Endotoxin Research,
11, 304-310.
[48] Dziarski, R. and Gupta, D. (2005) Staphylococcus aureus
peptidoglycan is a toll-like receptor 2 activator: A re-
evaluation. Infection and Immunity, 73, 5212-5216.