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Propofol Protects Against High GlucoseInduced

Endothelial Dysfunction in Human Umbilical Vein


Endothelial Cells
Minmin Zhu, MD, Jiawei Chen, MD, PhD, Zhiming Tan, MD, PhD, and Jing Wang, MD
BACKGROUND: Hyperglycemia, via peroxynitrite-mediated endothelial nitric oxide synthase
(eNOS) enzymatic uncoupling, induced endothelial dysfunction. Propofol has been reported to
improve high glucoseinduced endothelial dysfunction. However, its mechanisms of action
remain unclear. We hypothesized that propofol could improve hyperglycemia-induced endothelial
dysfunction by decreasing the peroxynitrite level and thus restoring eNOS coupling.
METHODS: At the end of 3 days of incubation in medium with 30 mM glucose, human
umbilical vein endothelial cells were treated with different concentrations (0.2, 1, 5, and 25
M) of propofol for different times (0.5, 1, 2, and 4 hours). In parallel experiments, cells
were cultured in 5 mM glucose for 3 days as a control. Nitric oxide (NO) production was
measured with a nitrate reductase assay. Superoxide anion (O
2

) accumulation was
measured with the reduction of ferricytochrome c and dihydroethidine uorescence assay.
The treatment that had maximal effect on 30 mM glucoseinduced NO production and O
2

accumulation was applied in the following studies to examine the underlying signaling
pathways. eNOS total protein, eNOS dimer and monomer expression, eNOS phosphorylation
at Ser
1177
, inducible NO synthase total protein, inducible NO synthase dimer and monomer
expression, peroxynitrite, and guanosine triphosphate cyclohydrolase I expression were
measured by Western blot. Tetrahydrobiopterin (BH
4
) level was measured with liquid
chromatographymass spectrometry.
RESULTS: Compared with 5 mM glucose treatment, 30 mM glucose signicantly decreased NO
production by 60% (P 0.001) and increased O
2

accumulation by 175% (P 0.0026), which


were both attenuated by propofol in a concentration- and time-dependent manner. Compared
with 5 mM glucose treatment, total eNOS protein expression was increased by 30 mM glucose
(P 0.001), whereas the ratio of eNOS dimer/monomer (P 0.0001) and eNOS phosphory-
lation (P 0.001) were decreased by 30 mM glucose. Propofol did not affect 30 mM
glucoseinduced total eNOS protein expression, but restored the ratio of eNOS dimer/monomer
(P 0.0005) and increased eNOS phosphorylation (P 0.001). 30 mM glucoseinduced O
2

accumulation was inhibited by the eNOS inhibitor hydrochloride. Furthermore, compared with 5
mM glucose treatment, 30 mM glucose decreased the BH
4
level (P 0.0001) and guanosine
triphosphate cyclohydrolase I expression (P 0.001), whereas it increased peroxynitrite level
(P 0.0003), which could all be reversed by propofol (P 0.0045, P 0.001, P 0.0001 vs
30 mM glucose treatment, respectively).
CONCLUSIONS: Propofol has benecial effects on 30 mM glucoseinduced NO reduction and
O
2

accumulation in human umbilical vein endothelial cells. This may be mediated through
inhibiting peroxynitrite-mediated BH
4
reduction, and restoring eNOS coupling. (Anesth Analg
2012;114:3039)
P
erioperative hyperglycemia, which may be caused
by a physiological stress response and excessive
glucose transfusion, is a common clinical metabolic
disorder in nondiabetic
1
as well as diabetic patients.
2
Within a normal glucose range, endothelial nitric oxide
synthase (eNOS) produces nitric oxide (NO), whereas in
hyperglycemia, eNOS produces superoxide anion (O
2

)
rather than NO,
3,4
resulting in endothelial dysfunction.
This phenomenon is known as eNOS uncoupling,
5
dur-
ing which eNOS is turned from a protective enzyme to a
producer of oxidative stress.
6
Tetrahydrobiopterin (BH
4
), a cofactor for NOS, is essen-
tial for the dimerization of NOS.
7,8
Peroxynitrite produced
by high glucose reduces BH
4
levels by oxidative degrada-
tion,
9
or by degrading guanosine triphosphate cyclohydro-
lase I (GTPCH I),
10
a rate-limiting enzyme of BH
4
synthesis.
As a result, eNOS becomes uncoupled and produces O
2

.
Excessive O
2

further reacts with NOto formmore peroxyni-


trite, thus forming a vicious circle. The IV anesthetic propofol
(2,6-diisopropylphenol) has been reported to improve high
glucoseinduced endothelial dysfunction.
11
However, its
mechanisms of action remain unclear. In the present study,
we hypothesized that propofol could improve high glucose
induced endothelial dysfunction by decreasing peroxynitrite
levels and thus restoring eNOS coupling.
From the Department of Anaesthesiology, Fudan University Shanghai
Cancer Centre, Shanghai; and Department of Oncology, Shanghai Medical
College, Fudan University, Shanghai, Peoples Republic of China.
Accepted for publication October 10, 2011.
The authors declare no conflicts of interest.
Reprints will not be available from the authors.
Address correspondence to Zhiming Tan, MD, PhD, Department of Anaes-
thesiology, Fudan University Shanghai Cancer Centre and Department of
Oncology, Shanghai Medical College, Fudan University, No. 270 DongAn
Rd., Shanghai, 200032, P.R. China. Address e-mail to tanzm@shca.org.cn.
Copyright 2012 International Anesthesia Research Society
DOI: 10.1213/ANE.0b013e31823f0c42
February 2012 Volume 114 Number 2 www.anesthesia-analgesia.org 303
METHODS
The investigation was approved by the Fudan University
Institutional Academic Committee. Human umbilical vein
endothelial cells (HUVECs) (Clonetics; Lonza, Basel, Switzer-
land) were cultured in Dulbeccos modified Eagle medium
with 5 mM glucose containing gentamicin, amphotericin-B,
and 10%fetal bovine serum. Cells were grown in an incubator
supplemented with 95% air and 5% CO
2
at 37C and subcul-
tured on reaching 90% confluence. The fifth passage of cells
was used in the present study. Five and 30 mM glucose were
widely used in in vitro studies to represent normal and high
glucose
12
; therefore, we used these 2 concentrations to treat
cells in this study. For the 5 mMor 30 mMglucose group, cells
were cultured in Dulbeccos modified Eagle medium with 5
mM or 30 mM glucose for 3 days. Propofol (Sigma, St. Louis,
MO) was dissolved in dimethyl sulfoxide and added in the
culture medium to achieve the appropriate concentration.
During general anesthesia, plasma concentrations of propofol
may range from 5 to 50 M
13
; therefore, in the present study,
HUVECs cultured in 30 mM glucose were treated with
different concentrations (0.2, 1, 5, and 25 M) of propofol for
different times (0.5, 1, 2, and 4 hours). In parallel experiments,
cells were cultured in 5 mM glucose for 3 days as control. The
number of cells used in each experiment was approximately
1 10
6
.
NO production was measured with a nitrate reductase
assay kit (Nanjing Jiancheng Bioengineering Institute, Nan-
jing City, P. R. China) according to the manufacturers
instructions. The results are shown as a percentage of the
control group.
O
2

accumulation was determined by the reduction of


ferricytochrome c and ferridihydroethidine fluorescence
assay as described previously.
12
To measure ferricyto-
chrome c reduction, HUVECs were washed and incubated
at 37C for 30 minutes with Krebs-HEPES buffer containing
20 M ferricytochrome c (Sigma) with or without superox-
ide dismutase (Sigma). The absorbance was read at 550 nm
spectrophotometrically. Reduction of ferricytochrome c
with superoxide dismutase was subtracted from the values
without superoxide dismutase. To measure dihydroethi-
dine fluorescence, cells were washed, cultured with Krebs-
HEPES buffer containing 4 M dihydroethidine at 37C for
10 minutes, then washed and imaged with a confocal
microscope.
To measure protein expression, cells were lysed by cell
lysis buffer (Cell Signaling Technology, Danvers, MA)
containing 1 mM phenylmethanesulfonyl fluoride and pro-
tease inhibitor (Calbiochem). Equal amounts of protein
were separated by 8% or 12% sodium dodecyl sulfate
polyacrylamide gel electrophoresis (SDS-PAGE) and trans-
ferred to polyvinylidene difluoride membranes. To study
eNOS homodimer expression, low-temperature SDS-PAGE
was used as described previously.
12
After incubation in 5%
skim milk, the membranes were incubated with an appro-
priate dilution of primary antibody at 4C overnight. The
primary antibodies used were polyclonal antibody against
eNOS (Santa Cruz Biotechnology, Santa Cruz, CA), phos-
phorylated eNOS (ser
1177
) (Cell Signaling), inducible NOS
(iNOS) (BD Transduction Laboratories), nitro-tyrosine (Cell
Signaling), GTPCH I (Santa Cruz Biotechnology), and
-actin (Santa Cruz Biotechnology). After being incubated
with secondary antibody, the membranes were washed and
detected with the ECL system, and relative densities of the
Figure 1. Effects of propofol on 30 mM glucose
induced nitric oxide (NO) production and superoxide
anion (O
2

) accumulation. Cells were cultured in


medium with 30 mM glucose or 5 mM glucose for 3
days. At the end of 3 days of incubation, cells
cultured in 30 mM glucose were treated with differ-
ent concentrations (0.2, 1, 5, and 25 M) of
propofol for different times (0.5, 1, 2, and 4 hours).
Cells were cultured in 5 mM glucose as a control.
Compared with 5 mM glucose treatment, 30 mM
glucose could decrease NO production (A) and
increased O
2

accumulation (B), which could be


reversed by propofol in a concentration- and time-
dependent manner. Propofol solvent dimethyl
sulfoxide (DMSO) did not affect 30 mM glucose
induced NO production and O
2

accumulation (*P
0.05 vs 5 mMglucose; #P 0.05 vs 30 mMglucose,
n 5). Data are shown as mean SD. a.u.
arbitrary units.
Propofol Inhibits Endothelial Dysfunction in High Glucose
304 www.anesthesia-analgesia.org ANESTHESIA & ANALGESIA
protein bands were analyzed by Scan-gel-it software (UN-
SCAN-IT gel 6.0, Silk Scientific Inc., Orem, UT). Peroxyni-
trite could react with protein tyrosine residues to form
nitro-tyrosine; therefore, it was measured as a marker of
peroxynitrite production.
Cellular biopterin levels were measured as described
previously,
12
but with modifications. Briefly, cells were
pelleted by centrifugation, and lysed in cold extract buffer
(50 mM Tris-HCl, pH 7.4, 1 mM EDTA, 1 mM DTT). After
measuring protein concentration, protein was removed by
a 1:1 mixture of 1.5 M HClO
4
and 2 M H
3
PO
4
. Total
biopterin (BH
4
, dihydrobiopterin, biopterin) was deter-
mined after acid oxidation, whereas dihydrobiopterin and
biopterin were determined after alkali oxidation. Biopterin
was measured by liquid chromatographymass spectrom-
etry using the Agilent G6520A accurate-mass Q-TOF liquid
chromatographymass spectrometry system with a
ZORBAX Eclipse plus C18 column (2.1 50 mm). The BH
4
concentration was calculated by subtracting dihydrobiop-
terin and biopterin from total biopterin.
Data were obtained from at least 3 separately performed
experiments, and are presented as mean SD. Statistical
significance was determined in multiple comparisons
among independent groups of data in which analysis of
variance indicated the presence of significant difference. A
value of P 0.05 was considered significant.
RESULTS
In HUVECs, compared with 5 mM glucose treatment, 30
mM glucose caused a marked reduction of NO production
by 60% (P 0.001), and a significant increase of O
2

accumulation by 175% (P 0.0026). We also found that


propofol could attenuate 30 mM glucoseinduced NO
reduction and O
2

accumulation in a concentration- and


time-dependent manner. Incubation of cells with 1 M
propofol for 1 hour caused a significant reversal of NO
reduction and O
2

accumulation (P 0.0015 for NO


production and P 0.0012 for O
2

accumulation vs 30 mM
glucose group; Fig. 1). This treatment was used in the
following experiments to study the signaling pathways
responsible for the protective effects of propofol.
Because NO production and O
2

accumulation may be
regulated by eNOS and iNOS, we determined the eNOS
and iNOS protein expression and dimerization. Compared
with the 5 mM glucose group, 30 mM glucose increased
total eNOS protein expression by 1.5-fold (P 0.001) (Fig.
2, A and C). We also found that the ratio of eNOS
dimer/monomer was significantly lower in the 30 mM
Figure 2. Effects of propofol on 30 mM glucose
induced endothelial nitric oxide synthase (eNOS)
and inducible nitric oxide synthase (iNOS) total
protein expression and dimerization. A and B, Cells
were cultured in either 5 mM glucose or 30 mM
glucose for 3 days with corresponding treatment.
Equal amounts of proteins (boiled or nonboiled) were
separated by sodium dodecyl sulfatepolyacrylamide
gel electrophoresis (SDS-PAGE) or low-temperature
SDS-PAGE, respectively, and immunoblotted with
polyclonal antibody to eNOS and iNOS. C and D,
Quantication of eNOS and iNOS band density (*P
0.05 vs 5 mMglucose; #P 0.05 vs 30 mMglucose,
n 5). E and F, Quantication of eNOS and iNOS
band density: ratio of dimer/monomer (*P 0.05 vs
5 mMglucose; #P 0.05 vs 30 mMglucose, n 5).
Data are shown as mean SD.
February 2012 Volume 114 Number 2 www.anesthesia-analgesia.org 305
glucose group than in the 5 mM glucose group (P 0.0001;
Fig. 2, A and E). Propofol 1 M did not affect total eNOS
protein expression, but restored the ratio of eNOS
dimer/monomer (P 0.0005 vs 30 mM glucose group; Fig.
2, A, C, and E). Compared with the 5 mM glucose group, 30
mM glucose increased total iNOS protein expression by
2.3-fold (P 0.001; Fig. 2, B and D). However, the ratio of
iNOS dimer/monomer was not affected by 30 mM glucose
(Fig. 2, B and F). Propofol 1 M had no effect on 30 mM
glucosemediated total iNOS protein expression or iNOS
dimerization (Fig. 2, B, D, and F).
The effect of 30 mM glucose and propofol on O
2

accumulation was confirmed by dihydroethidine fluores-


cence assay. We further observed that 30 mM glucose
induced O
2

accumulation was inhibited by hydrochloride,


an inhibitor of eNOS. When hydrochloride and propofol were
used simultaneously, there was no further reduction of O
2

accumulation (Fig. 3).


The activity of eNOS is also regulated by eNOS phos-
phorylation, and the most widely studied site is Ser
1177
.
Compared with the 5 mM glucose group, 30 mM glucose
treatment increased total eNOS expression, whereas it
decreased eNOS phosphorylation at Ser
1177
(P 0.001).
Although propofol had no effect on 30 mM glucose
induced total eNOS expression, it increased eNOS
phosphorylation in a concentration-dependent manner.
Compared with the 30 mM glucose group, incubation of
cells with 5 M propofol for 1 hour caused a significant
reversal of eNOS phosphorylation (P 0.001) and the ratio
of eNOS phosphorylation/total eNOS expression (P
0.001). However, the ratio of eNOS phosphorylation/total
eNOS expression of the 5 M and 25 M propofol groups
was still significantly lower than that of the 5 mM glucose
group (P 0.001; Fig. 4).
Furthermore, we examined the total biopterin and BH
4
levels in response to 30 mM glucose and propofol. Com-
pared with the 5 mM glucose group, 30 mM glucose
decreased total biopterin (P 0.0003), BH
4
levels (P
0.0001), and the ratio of BH
4
/total biopterin (P 0.0011),
which could all be reversed by 1 M propofol (P 0.0032,
P 0.0045, P 0.0076 vs 30 mM glucose group, respec-
tively; Fig. 5).
Compared with the 5 mM glucose group, 30 mM
glucose treatment increased nitro-tyrosine expression
(P 0.0003), whereas it decreased GTPCH I expression
(P 0.001), which could both be reversed by 1 M
propofol (P 0.001; P 0.0001 vs 30 mM glucose group,
respectively; Fig. 6).
DISCUSSION
The main finding of the present study is that propofol
could protect HUVECs against 30 mM glucoseinduced cell
dysfunction by increasing NO production and decreasing
O
2

accumulation, which might be achieved by regulating


eNOS coupling.
eNOS activity is regulated by a complicated process. At
the cellular level, it could be modulated by the transforma-
tion between eNOS dimer and monomer,
14
and by eNOS
expression
15
as well as phosphorylation.
16
Previous studies
indicated that high glucose could reduce eNOS dimeriza-
tion
12
and eNOS phosphorylation
17
at Ser
1177
in endothelial
cells, thus decreasing NO production and increasing O
2

accumulation. This was consistent with our findings. We


detected that 30 mM glucose decreased the ratio of eNOS
dimer/monomer and eNOS phosphorylation, leading to
NO reduction and O
2

accumulation. Interestingly, we
found that 30 mM glucose increased total eNOS expression.
We also found that 30 mM glucoseinduced eNOS protein
Figure 3. Effects of propofol on 30 mM
glucoseinduced superoxide anion (O
2

)
accumulation is similar to hydrochloride,
an endothelial nitric oxide synthase inhib-
itor. Cells were cultured in either 5 mM or
30 mM glucose for 3 days. At the end of
3 days of incubation, cells cultured in 30
mM glucose were treated with propofol (1
M) or hydrochloride (100 M), or propo-
fol (1 M) plus hydrochloride (100 M)
for 1 hour. After treatment, cells were
incubated with 4 M dihydroethidine for
10 minutes. Compared with 5 mM glu-
cose treatment, the intensity of uores-
cence (red) of human umbilical vein
endothelial cells in 30 mM glucose was
enhanced, and was attenuated by propo-
fol, or hydrochloride, or propofol plus
hydrochloride. Each experiment was re-
peated 3 times.
Propofol Inhibits Endothelial Dysfunction in High Glucose
306 www.anesthesia-analgesia.org ANESTHESIA & ANALGESIA
mainly existed in monomeric form. The inconsistency be-
tween total eNOS expression and NO production may be
explained by the imbalance between eNOS dimer and
monomer, favoring the transformation from dimer to
monomer. eNOS dimerization is regulated by intracellular
BH
4
levels,
7,8
which depend on a balance among de novo
synthesis, BH
4
oxidation, and recycling of dihydrobiopterin
to BH
4
.
18
De novo synthesis of BH
4
is affected by the level
of its rate-limiting enzyme GTPCH I.
19
It was reported
10
that the inhibition of BH
4
synthesis by high glucose
induced peroxynitrite is mainly mediated through proteasome-
dependent degradation of GTPCH I. Laursen et al.
9
also
reported that peroxynitrite could reduce BH
4
levels by
oxidation. Our data were consistent with theirs, indicating
that high glucose could decrease GTPCH I and BH
4
levels,
thus resulting in eNOS uncoupling.
In the present study, we found that 30 mM glucose
induced O
2

accumulation was attenuated by hydrochlo-


ride (eNOS inhibitor), suggesting that uncoupled eNOS
was an important source of O
2

accumulation in HUVECs
in response to 30 mM glucose. Although uncoupled iNOS
was also indicated to be a main resource of O
2

accumu-
lation,
20
in the present study, the ratio of iNOS dimer/
monomer was not altered by 30 mM glucose. We therefore
believe that iNOS was minimally involved in the 30 mM
glucoseinduced O
2

accumulation.
Propofol is a widely used IV anesthetic and its proper-
ties have been studied in the last decades. Propofol has
been shown to be a peroxynitrite scavenger,
21
and could
reduce peroxynitrite through 2 different mechanisms: di-
rectly by scavenging peroxynitrite
21
and indirectly by scav-
enging O
2
.
22
It was reported
23
that propofol may protect
endothelial cells by reacting with peroxynitrite and reduc-
ing the interaction between peroxynitrite and tyrosine
residues, and the formation of toxic substance. In the
present study, we found that the cell-protective concentra-
tion of propofol was 1 M. However, Mathy-Hartert et al.
23
suggested that the cell-protective effect was observed at 30
M propofol. In their study, 3-morpholinosydnonimine
hydrochloride, a compound that slowly releases O
2

and
NO, was used to form peroxynitrite, whereas we used 30
mM glucose to induce peroxynitrite. Because different
stimuli may cause different amounts of peroxynitrite pro-
duction, it is possible that a different concentration of
propofol was required to provide a protective effect. Our
data were consistent with those of Mathy-Hartert et al.,
Figure 4. Effects of propofol on 30 mM glucoseinduced endothelial
nitric oxide synthase (eNOS) phosphorylation (P-eNOS). A, Cells were
cultured in either 5 mM or 30 mM glucose for 3 days with
corresponding treatment. Equal amounts of proteins were separated
by sodium dodecyl sulfatepolyacrylamide gel electrophoresis and
immunoblotted with polyclonal antibody to eNOS phosphorylation at
Ser
1177
and eNOS. B, Quantication of eNOS phosphorylation and
total eNOS band density (*P 0.05 vs 5 mM glucose; #P 0.05 vs
30 mM glucose, n 5; open bars represent total eNOS expression,
and lled bars represent eNOS phosphorylation). C, Quantication of
eNOS phosphorylation/total eNOS expression (*P 0.05 vs 5 mM
glucose; #P 0.05 vs 30 mM glucose, n 5). Data are shown as
mean SD.
Figure 5. Effects of propofol on 30 mM glucoseinduced total
cellular biopterin and tetrahydrobiopterin (BH
4
) levels. A, 30 mM
glucose decreased total biopterin and BH
4
levels, which could be
reversed by propofol (*P 0.05 vs 5 mM glucose; #P 0.05 vs 30
mM glucose, n 3; lled bars represent total cellular biopterin, and
open bars represent BH
4
). B, 30 mM glucose decreased the ratio of
BH
4
/total cellular biopterin, which could also be reversed by propofol
(*P 0.05 vs 5 mM glucose; #P 0.05 vs 30 mM glucose, n 3).
Data are shown as mean SD.
February 2012 Volume 114 Number 2 www.anesthesia-analgesia.org 307
demonstrating that the protective effect of propofol was
mediated by scavenging peroxynitrite. Previous studies
indicated that peroxynitrite could reduce BH
4
levels,
9,10
and results in eNOS uncoupling, thus creating positive
feedback. In the present study, we found propofol, via
decreasing peroxynitrite production (Fig. 6), increased the
levels of BH
4
and GTPCH I (Figs. 5 and 6), restored 30 mM
glucoseinduced eNOS uncoupling (Fig. 2, A and E) and
NO reduction (Fig. 1A) as well as O
2

accumulation (Fig.
1B), thus inhibiting positive feedback. Moreover, treatment
of cells with hydrochloride or propofol showed a similar
reduction of 30 mM glucoseinduced O
2

accumulation
(Fig. 3). These findings strongly indicated that propofol
restored 30 mM glucoseinduced NO reduction and O
2

accumulation via inducing eNOS coupling. We therefore


believe that eNOS recoupling is a main mechanism respon-
sible for the protective effect of propofol against 30 mM
glucoseinduced cell dysfunction.
Propofol could also activate eNOS and increase NO
production by inducing eNOS phosphorylation at Ser
1177
in normal conditions.
24
We consistently observed that 30
mM glucose decreased eNOS phosphorylation at Ser
1177
,
which could be reversed by propofol in a concentration-
dependent manner (Fig. 4). Note that incubation of cells
with 5 M, rather than 1 M, propofol significantly in-
creased eNOS phosphorylation at Ser
1177
and the ratio of
eNOS phosphorylation/total eNOS expression. Consider-
ing the fact that the protective concentration of propofol
against 30 mM glucoseinduced cell dysfunction was 1 M
(Fig. 1) and this concentration had no effect on eNOS
phosphorylation (Fig. 4), we therefore believe that eNOS
phosphorylation is not the main mechanism responsible for
the protective effect of propofol against 30 mM glucose
induced cell dysfunction.
This study has some limitations. First, the study was
implemented in HUVECs, and there may be some differ-
ences from in vivo systems in studying drug effectiveness
and toxicity. Next, HUVECs were exposed in 30 mM
glucose, which is rarely encountered during surgery or in
the intensive care unit. Moreover, considering the fact that
we mainly focused on the protective effect of propofol
against severe hyperglycemia in the present study, we did
not examine whether similar abnormalities in eNOS uncou-
pling occur under more clinically relevant glucose concen-
trations (e.g., 1520 mM glucose) and whether propofol
causes salutary actions under such conditions. Further
studies in in vivo and ex vivo models are necessary to
verify the present results. Nevertheless, this knowledge
may provide a rationale to use propofol to attenuate or
avoid an adverse impact in patients with hyperglycemia,
hypertension,
25
and ischemia/reperfusion,
26
which are as-
sociated with increased peroxynitrite production.
In summary, the present study suggested that 30 mM
glucose, by inducing peroxynitrite and decreasing BH
4
levels, induced eNOS uncoupling, leading to oxidative
stress and cell dysfunction. More importantly, our study
demonstrated that propofol, via scavenging peroxynitrite,
attenuated 30 mM glucoseinduced eNOS uncoupling, and
restored endothelial function.
DISCLOSURES
Name: Minmin Zhu, MD.
Contribution: This author helped design the study, conduct
the study, analyze the data, and write the manuscript.
Attestation: Minmin Zhu has seen the original study data,
reviewed the analysis of the data, approved the final manu-
script, and is the author responsible for archiving the study
files.
Name: Jiawei Chen, MD, PhD.
Contribution: This author helped analyze the data and write
the manuscript.
Attestation: Jiawei Chen has seen the original study data,
reviewed the analysis of the data, and approved the final
manuscript.
Name: Zhiming Tan, MD, PhD.
Contribution: This author helped design the study, analyze
the data, and write the manuscript.
Figure 6. Effects of propofol on 30 mM glucoseinduced nitro-
tyrosine and guanosine triphosphate cyclohydrolase I (GTPCH I)
expression. A, Cells were cultured in either 5 mM glucose or 30 mM
glucose for 3 days with corresponding treatment. Equal amounts of
proteins were separated by sodium dodecyl sulfatepolyacrylamide
gel electrophoresis and immunoblotted with polyclonal antibody to
nitro-tyrosine, or immunoblotted with polyclonal antibody to GTPCH I.
B, Quantication of nitro-tyrosine band density (*P 0.05 vs 5 mM
glucose; #P 0.05 vs 30 mM glucose, n 5). C, Quantication of
GTPCH I band density (*P 0.05 vs 5 mM glucose; #P 0.05 vs 30
mM glucose, n 5). Data are shown as mean SD.
Propofol Inhibits Endothelial Dysfunction in High Glucose
308 www.anesthesia-analgesia.org ANESTHESIA & ANALGESIA
Attestation: Zhiming Tan has seen the original study data,
reviewed the analysis of the data, and approved the final
manuscript.
Name: Jing Wang, MD.
Contribution: This author helped write the manuscript.
Attestation: Jing Wang has seen the original study data,
reviewed the analysis of the data, and approved the final
manuscript.
This manuscript was handled by: Marcel E. Durieux,
MD, PhD.
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