From the Department of Pharmacology, New York Medical College, Valhalla,
NY.
Correspondence to Alberto Nasjletti, MD, Department of Pharmacology, New York Medical College, Valhalla, NY 10595.
Several studies indicate that Ang II promotes
lipoxygenase-catalyzed production of
eicosanoids in vascular tissue. For example, Ang II was reported to
stimulate release of 12-HETE from segments of human umbilical
artery10 and to increase
12-lipoxygenase mRNA and protein along with 12-HETE
release in cultured porcine smooth muscle cells.1
In addition, the production of
lipoxygenase-derived HETEs by segments of thoracic
aorta was shown to increase in rats with Ang II–dependent
hypertension.6 11
Reports that lipoxygenase inhibitors
attenuate the vascular actions of Ang II9 10 12
and lower blood pressure in hypertensive
rats11 13 14 incriminate
lipoxygenase-derived eicosanoids in the mechanisms of
arterial hypertension. Because 12-HPETE and other
hydroperoxides arising from polyunsaturated fatty acids via
metabolism by lipoxygenase(s) inhibit
PGI2,5 15 excessive
expression of lipoxygenase or
lipoxygenases may promote elevation of blood pressure
by weakening the activity of antihypertensive mechanisms mediated by
PGI2.16 If so,
lipoxygenase inhibitors may lower blood
pressure in such settings by fostering production of
PGI2. Therefore, the present study was
designed to test the hypothesis that PGI2
participates in the implementation of the antihypertensive effect of
the lipoxygenase inhibitor baicalein in
rats with Ang II–induced hypertension.
Experiments were conducted on untreated normotensive rats,
sham-infused normotensive rats, and rats with Ang II–induced
hypertension. Rats with Ang II–induced hypertension were prepared as
previously described.17 Briefly, an Alzet osmotic
minipump (model 2002, Alza Corporation) filled with Ang II
([Ile5]Ang II; Sigma Chemical) was placed
through a 1-cm midline incision in the abdominal cavity of rats
anesthetized with methoxyflurane (Pitman-Moore); the nominal
infusion rate of Ang II was 200 ng/min. Sham-infused rats were prepared
by placing in the abdominal cavity an osmotic minipump filled with 0.01
mol/L acetic acid, the vehicle of Ang II.
Animals in protocols 1, 2, and 3 were instrumented with a chronic
arterial catheter. One day before minipump placement, rats
were anesthetized with pentobarbital sodium (60 mg/kg IP) and a
polyethylene cannula (PE-50) filled with saline solution (0.15 mol/L
NaCl) containing heparin (100 units/mL) was introduced through the left
femoral artery and advanced into the lower abdominal aorta. Animals in
protocols 2 and 3 also were fitted with a venous cannula (PE-50)
introduced through the left femoral vein and advanced into the lower
inferior vena cava. Both cannulas were tunneled
subcutaneously to an exit point at the nape of the neck and plugged
with steel pins until use. All rats received ampicillin (30 mg ·
kg-1 · 12 h-1 SC)
for 3 days after surgery.
Experiments were conducted 12 to 14 days after minipump placement. In
protocols 1, 2, and 3, the mean arterial pressure of awake
rats was measured via the femoral arterial cannula by means
of a pressure transducer (model P23XL; Statham Division, Gould Inc)
coupled to a polygraph (model 7D; Grass Instrument). In protocol 4,
systolic blood pressure was determined through tail
sphygmography with the use of a electrosphygmomanometer (Narco
Bio-System).
Experimental Protocols
Protocol 2 was designed to investigate the contribution of
vasodepressor prostanoids to the effects of baicalein and CDC on blood
pressure of rats with Ang II–induced hypertension. Hypertensive rats
were pretreated intravenously with
indomethacin (5 mg/kg bolus injection followed by
infusion at 5 mg/kg per hour) or drug/vehicle alone. Sixty minutes
later, baicalein was administered subcutaneously (60 mg/kg) to animals
pretreated with indomethacin (n=6) or vehicle alone
(n=8), and blood pressure was monitored over the next 120 minutes.
Likewise, rats pretreated with indomethacin (n=7) or
drug/vehicle alone (n=6) were injected subcutaneously with CDC (8
mg/kg), and blood pressure was recorded.
Protocol 3 was designed to investigate the contribution of
PGI2 to the effect of baicalein on blood pressure
of rats with Ang II–induced hypertension. Hypertensive rats were
injected subcutaneously with baicalein (60 mg/kg), followed 60 minutes
later by an intravenous injection of nonimmune rabbit serum
(0.3 mL; n=5) or rabbit serum containing antibodies directed against
5,6-dihydro-PGI2 (0.3 mL; n=5). Blood pressure
was monitored throughout the experiment. The serum containing
5,6-dihydro-PGI2 antibodies was donated by Dr
Lawrence Levine (Brandeis University). The method of immunization
against 5,6-dihydro-PGI2 and the characteristics
of the resulting antibodies were described
previously.19 20
5,6-Dihydro-PGI2 antibodies were reported to bind
PGI2 and neutralize the vasodepressor action of
PGI2.20 21
We conducted complementary studies to validate the effectiveness
and specificity of 5,6-dihydro-PGI2 antiserum to
block the vasodepressor effect of PGI2 in rats.
Experiments were conducted in awake Sprague-Dawley rats previously
instrumented with femoral arterial and venous cannulas to
measure blood pressure and administer drugs, respectively. The effect
of intravenous bolus injections of
PGI2 (2.0 µg/kg) on blood pressure was examined
before and at intervals after intravenous administration of
5,6-dihydro-PGI2 antiserum (0.3 mL; n=5). In
separate rats, the effect of intravenous bolus injections
of PGE2 (2.0 µg/kg) on blood pressure was
examined before and at intervals after intravenous
administration of 5,6-dihydro-PGI2 antiserum (0.3
mL; n=5).
Protocol 4 was designed to investigate the effect of in vivo treatment
with baicalein on release of 6-keto-PGF1
In additional experiments, rats with Ang II–induced hypertension of 12
days' duration were housed in metabolic cages and
subsequently injected with baicalein (n=8) or vehicle only (n=8) as
described in protocol 1. Thereafter, urine was collected for 3 hours to
measure renal excretion of prostanoids and 12-HETE. Similar experiments
were conducted in sham-infused normotensive rats injected with
baicalein (n=6) or vehicle (n=7).
Analytical Procedures
Measurement of Conversion of PGH2 to PGI2
by Rings of Thoracic Aorta
Measurement of PGE2 and 6-Keto-PGF1
Measurements of Eicosanoids in Urine
Statistical Analysis
The effects of baicalein on mean arterial pressure in
rats with Ang II–induced hypertension pretreated and not pretreated
with indomethacin are shown in Fig 2
Fig 3
The effects of intravenous bolus injections of
PGI2 (2.0 µg/kg) or PGE2
(2.0 µg/kg) on the mean arterial pressure of normotensive
rats before and after the intravenous administration of
5,6-dihydro PGI2 antiserum are shown in Fig 4
Fig 5
Fig 6
The Table
The acute antihypertensive effect of baicalein and CDC in rats
with Ang II–induced hypertension may be a functional consequence of
diminished production of lipoxygenase-derived
eicosanoids that mediate or facilitate vasoconstrictor
mechanisms.6 7 9 It also may be linked to
activation of a vasodilatory mechanism mediated by
PGI2 and/or to deactivation of a pressor
mechanism mediated by PGH2, both of which are
events caused by elimination of the inhibitory influence of
12-HPETE and other lipoxygenase products on
prostacyclin synthase.5 6 15 16 This study
demonstrates that baicalein and CDC do not reduce the blood pressure of
hypertensive rats pretreated with indomethacin. Because
indomethacin inhibits
cyclooxygenase without affecting the vascular
production of lipoxygenase-derived
HETEs,6 our results suggest that the acute
antihypertensive effect of these agents in rats with Ang II–induced
hypertension relies for its implementation on a prostanoid-mediated
mechanism. This conclusion derives additional support from experiments
demonstrating that the vasodepressor effect of baicalein in
hypertensive rats is reversed partially through treatment with
5,6-dihydro-PGI2 antiserum. The administration of
5,6-dihydro-PGI2 antiserum also attenuates
vasodepressor responsiveness to PGI2 but not to
PGE2, which is in agreement with a report that
antibodies directed to 5,6-dihydro-PGI2 bind and
neutralize the biological activities of
PGI2.20 21 The observation
that 5,6-dihydro-PGI2 antiserum causes partial
reversal of the antihypertensive effect of baicalein implicates
PGI2 in the implementation of such an effect.
Previous reports show that vascular and renal production of
PGI2 are increased in rats with Ang II–dependent
hypertension.17 In concordance with such studies,
we found that rats made hypertensive through infusion of Ang II feature
increased circulating levels of 6-keto-PGF1
A major finding in this study is that the antihypertensive effect of
baicalein in rats with Ang II–induced hypertension is accompanied by
an increase in the rate of conversion of exogenous
PGH2 to PGI2 by aortic
rings, bringing it up to the rate of conversion found in aortic rings
from normotensive rats. Previously, it was reported that rings of
thoracic aorta taken from rats with aortic coarctation-induced
hypertension are impaired in their ability to convert exogenous
PGH2 to PGI2 and that this
impairment is corrected through exposure of the rings to baicalein or
CDC.6 The notion that baicalein promotes
metabolism of PGH2 to
PGI2 in models of Ang II–dependent hypertension
fits well with our findings that baicalein treatment of hypertensive
rats increases the release of 6-keto-PGF1
It is unlikely that baicalein stimulates PGI2
synthase directly because its administration to normotensive rats did
not increase the conversion of PGH2 to
PGI2 by aortic rings or increase the release of
6-keto-PGF1
Recent studies revealed that superoxide anion production is
increased in arterial vessels of rats with Ang II–induced
hypertension.24 25 It also is known that
prostacyclin synthase is susceptible to inhibition or inactivation by
reactive oxygen species26 27 28 and that
flavonoids, such as baicalein, and hydroxycinnamic acid derivatives,
such as CDC, possess antioxidant properties.29 30
Accordingly, these lipoxygenase inhibitors
also may promote vascular conversion of PGH2 to
PGI2 in rats with Ang II–induced hypertension by
scavenging oxygen radicals that are damaging to prostacyclin
synthase.
In summary, we found that baicalein and CDC lower blood pressure in
rats made hypertensive by long-term Ang II infusion but not in
normotensive rats. The antihypertensive effect of these
lipoxygenase inhibitors was prevented
through pretreatment with indomethacin. The
antihypertensive effect of baicalein was partially reversed by the
administration of 5,6-dihydro-PGI2 antiserum,
which binds PGI2 and blocks its vasodepressor
action. Treatment of the hypertensive rats with baicalein also caused
selective increases in the rate of conversion of exogenous
PGH2 to PGI2 by aortic
rings, release of 6-keto-PGF1
Received August 25, 1997;
first decision September 22, 1997;
accepted October 31, 1997.
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© 1998 American Heart Association, Inc.
Scientific Contributions
Prostaglandin I2 Contributes to the Vasodepressor Effect of Baicalein in Hypertensive Rats
Abstract
Top
Abstract
Introduction
Methods
Results
Discussion
References
Abstract—Lipoxygenase
inhibitors reduce blood pressure in hypertensive rats. The
vasodepressor effect of lipoxygenase
inhibitors may be related to increased production
of prostaglandin (PG) I2 since
lipoxygenase-derived fatty acid hydroperoxides inhibit
PGI2 synthase. This hypothesis was examined in rats made
hypertensive by infusion of angiotensin II (200 ng/min IP)
for 12 to 14 days. In hypertensive but not in normotensive rats, the
lipoxygenase inhibitor baicalein (60 mg/kg
SC) increased (P<.05) the conversion of exogenous
PGH2 to PGI2 by aortic segments, the release of
6-keto-PGF1
by aortic rings, the concentration of
6-keto-PGF1 in blood, and the renal excretion of
6-keto-PGF1. Treatment with baicalein did not affect the
blood pressure of normotensive rats but decreased the blood pressure of
hypertensive rats from 177±8 to 133±9 mm Hg after 120 minutes
(P<.05). Also, the lipoxygenase
inhibitor cinnamyl-3,4-dihydroxy--cyanocinnamate (8
mg/kg SC) was without effect on the blood pressure of normotensive rats
but decreased the blood pressure of hypertensive rats from 182±4 to
139±8 mm Hg (P<.05). However, the blood pressure
of hypertensive rats pretreated with indomethacin (5
mg/kg IV) was affected by neither baicalein nor
cinnamyl-3,4-dihydroxy--cyanocinnamate. Moreover, in hypertensive
rats in which baicalein had decreased blood pressure to 148±6
mm Hg, the administration of rabbit serum containing antibodies
against 5,6-dihydro-PGI2 (0.3 mL IV) partially reversed the
response to baicalein, increasing blood pressure to 179±7 mm Hg
within 20 minutes (P<.05). The antibodies also were
shown to block the vasodepressor effect of PGI2 but not of
PGE2. Collectively, these data suggest contribution of
PGI2 to the acute antihypertensive effect of baicalein in
rats with angiotensin II–induced hypertension.
Key Words: angiotensin II • prostaglandin I2 • prostacyclin synthase • 12-lipoxygenase • lipoxygenase inhibitors
Introduction
Top
Abstract
Introduction
Methods
Results
Discussion
References
Vascular tissues
contain a lipoxygenase that catalyzes the
oxygenation of C12 of arachidonic
acid.1 2 3 The product of this reaction is
12-HPETE, which undergoes spontaneous or peroxidase-catalyzed reduction
to 12-HETE.4 Both 12-HPETE and 12-HETE are
capable of influencing vascular functions. 12-HPETE inhibits vascular
PGI2 synthase activity5 and
was reported to increase the expression of arachidonic
acid–induced, PGH2-mediated constrictor
responses in rings of rat aorta.6 12-HETE was
shown to depolarize renal arterial smooth muscle
cells,7 increase the protein content of cultured
porcine aortic smooth muscle cells,8 and
facilitate the stimulatory actions of Ang II and vasopressin on calcium
transients in cultured smooth muscle.9
Methods
Top
Abstract
Introduction
Methods
Results
Discussion
References
Animals
Male Sprague-Dawley rats (Charles River Labs) weighing 275 to
300 g were used in all the experiments. The animals were housed in
group cages or in individual cages as appropriate and were fed a
standard chow (Ralston Purina). All protocols were approved by the
Institutional Animal Care and Use Committee.
Protocol 1 was designed to examine the effect on blood pressure
of treatment with lipoxygenase inhibitors
baicalein or CDC6 12 14 18 (BIOMOL Research
Laboratories). On the day of the experiment, 12 to 14 days after
minipump placement, rats with Ang II–induced hypertension were
injected subcutaneously with baicalein (60 mg/kg; n=8), CDC (8 mg/kg;
n=9), or vehicle only (sesame oil, 2.0 mL/kg of body wt; n=11).
Sham-infused normotensive rats also were injected with baicalein (60
mg/kg SC; n=6), CDC (8 mg/kg SC; n=4), or sesame oil vehicle (2.0 mL/kg
of body wt; n=4). Mean arterial pressure was monitored
before and after treatment. from
rings of descending thoracic aorta, conversion of exogenous
PGH2 to PGI2 by rings of
descending thoracic aorta, concentration of prostanoids in venous
blood, and renal excretion of 12-HETE and prostanoids. On the day of
the experiment, sham-infused rats and rats with Ang II–induced
hypertension were injected with baicalein (n=6) or vehicle only (n=6)
as described in protocol 1. One hour later, the rats were
anesthetized with pentobarbital sodium (60 mg/kg IP), the
abdominal cavity was exposed through a midline incision, the
inferior vena cava was punctured with an 18-gauge needle to
sample blood (1 ml) for measurement of prostanoids, and the descending
thoracic aorta was excised and cut into ring segments (3 mm in
length). The aortic rings were used immediately to assess release of
6-keto-PGF1 and ability to convert exogenous
PGH2 to PGI2.
Measurement of 6-Keto-PGF1 Release by Rings of
Thoracic Aorta
Aortic rings were incubated in Krebs' bicarbonate buffer (2.0
mL) containing arachidonic acid (10 µmol/L) for
15 minutes at 37°C in an atmosphere of 95%
O2/5% CO2. The amount of
6-keto-PGF1 in the medium, an estimate of
PGI2 release, was analyzed as previously
described through enzyme immunoassay of unextracted samples with
reagents purchased from Cayman Chemical.22 The
results are expressed as picomoles of
6-keto-PGF1 released during the 15-minute
incubation period per milligram of dry tissue.
6-Keto-PGF1 could not be detected in samples
generated through incubation of aortic rings denatured by heating at
100°C for 5 minutes.
Aortic rings were preincubated for 20 minutes at 37°C in
Krebs' bicarbonate buffer gassed with 95%
O2/5% CO2 and containing
indomethacin (10 µmol/L) to inhibit
cyclooxygenase.6 The rings
were then transferred to 20-mL vials containing fresh buffer (2.0 mL)
for incubation at 37°C for 3 minutes in the presence and absence of
exogenous PGH2 (1 µmol/L). The
concentration of 6-keto-PGF1 in the incubation
media was measured through enzyme immunoassay of unextracted
samples.22 The concentration of
6-keto-PGF1 in medium derived from incubation
of aortic rings in indomethacin-containing buffer
without exogenous PGH2 was <3% of the
concentration in medium derived from incubations carried out in the
presence of exogenous PGH2. Hence, 
97% of the
6-keto-PGF1 in the incubation medium with
PGH2 arises from exogenous
PGH2. Results of the conversion of
PGH2 to PGI2 are expressed
as picomoles of 6-keto-PGF1 formed during a
3-minute incubation per milligram of dry tissue. No conversion of
exogenous PGH2 to PGI2
could be detected in control incubations using aortic rings denatured
by heating at 100°C for 5 minutes. Hence, when
cyclooxygenase is inhibited by
indomethacin, the conversion of exogenous
PGH2 to PGI2 by aortic
rings reflects the tissue activity of PGI2
synthase.6
in Blood
Blood (1 mL) was drawn from the inferior vena cava
into a syringe containing 4 mL of ice-cold ethanol and
indomethacin (10 µg/mL). The mixture was stored at
-20°C for 24 hours, followed by centrifugation at
1500g for 10 minutes, evaporation of the supernatant under a
stream of nitrogen, and reconstitution of the residue in 2 mL of 0.1
mol/L formic acid. PGE2 and
6-keto-PGF1 in the formic acid solution were
further purified with passage through a column of octadecylsilyl silica
(Sep-Pak C-18 cartridges; Waters Associates) according to a published
procedure,23 followed by quantification through
enzyme immunoassay.22 The results are expressed
as picomoles of PGE2 or
6-keto-PGF1 per milliliter of blood.
The contents of PGE2,
6-keto-PGF1, and 12-HETE in 3-hour urine
samples were determined after purification with passage of the samples
through a column of octadecylsilyl silica.23
PGE2 and 6-keto-PGF1
were measured with enzyme immunoassay.22 12-HETE
was measured through radioimmunoassay using reagents and a protocol
provided by PerSeptive Diagnostics. Data on urinary
excretion of eicosanoids are expressed as picomoles of eicosanoid per 3
hours.
Results are expressed as mean±SEM. ANOVA followed by the
Newman-Keuls a posteriori test was applied to the analysis of
data on the effect of drugs on blood pressure and for comparisons among
rats with Ang II–induced hypertension and sham-infused controls. Data
on the effect of baicalein on 6-keto-PGF1
release from aortic tissue, blood level of prostaglandins,
urinary eicosanoid excretion, and vascular conversion of
PGH2 to PGI2 were
analyzed with unpaired Student's t test. The null
hypothesis was rejected at a level of P<.05.
Results
Top
Abstract
Introduction
Methods
Results
Discussion
References
Fig 1
displays data on mean
arterial pressure before and after the administration of
baicalein or sesame oil vehicle only to sham-infused normotensive rats
and rats with Ang II–induced hypertension of 12 to 14 days' duration.
Before treatment, blood pressure was 103±6 and 177±8 mm Hg
(P<.05) in sham-infused rats and Ang II–infused rats,
respectively. Treatment with baicalein caused blood pressure to fall
progressively in rats with Ang II–induced hypertension, reaching a
level of 133±9 mm Hg (P<.05) after 120 minutes. In
contrast, treatment with baicalein was without effect on blood pressure
in sham-infused normotensive rats. Likewise, treatment with CDC did not
affect the blood pressure of sham-infused normotensive rats (103±6 and
100±10 mm Hg before and 120 minutes after CDC, respectively) but
decreased (P<.05) the blood pressure of rats with Ang
II–induced hypertension from 182±4 to 142±10 and 139±8 mm Hg
after 60 and 120 minutes, respectively. The administration of
drug/vehicle only did not affect the blood pressure of sham-infused
normotensive rats or of rats with Ang II–induced hypertension.
View larger version (29K):
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Figure 1. Mean arterial pressure (MAP)
before and after treatment with baicalein ( 
; 60 mg/kg SC) or sesame
oil vehicle (
; 2.0 mL/kg SC) in sham-infused normotensive rats (A)
and rats with Ang II–induced hypertension of 12 to 14 days' duration
(B). Results are mean±SEM; n indicates number of rats.
*P<.05 relative to corresponding data in rats treated
with sesame oil vehicle only.. Before baicalein administration, blood
pressure was comparable in hypertensive rats pretreated (178±6
mm Hg) and not pretreated (176±8 mm Hg) with
indomethacin. Treatment with baicalein decreased
(P<.05) blood pressure to 132±11 mm Hg in
hypertensive rats without indomethacin pretreatment. In
contrast, treatment with baicalein had little or no effect on the blood
pressure of hypertensive rats pretreated with
indomethacin. Like baicalein, the administration of CDC
did not affect the blood pressure of hypertensive rats pretreated with
indomethacin (176±5 and 177±6 mm Hg before and
120 minutes after CDC, respectively) but decreased (P<.05)
the blood pressure of hypertensive rats without
indomethacin pretreatment from 184±5 to 138±13 and
144±8 mm Hg after 60 and 120 minutes, respectively.
View larger version (28K):
[in a new window]
Figure 2. Mean arterial pressure (MAP) before
and after treatment with baicalein (60 mg/kg SC) in rats with Ang
II–induced hypertension not pretreated (A) or pretreated (B) with
indomethacin (5 mg/kg IV bolus plus infusion at 5 mg/kg
per hour). Results are mean±SEM; n indicates number of rats.
*P<.05 relative to control values before
treatment. shows a comparison of the
effects of nonimmune serum and 5,6-dihydro-PGI2
antiserum on the blood pressure of rats with Ang II–induced
hypertension that were pretreated with baicalein. The injection of
nonimmune serum did not modify the acute antihypertensive response to
baicalein. In contrast, the injection of
5,6-dihydro-PGI2 antiserum caused blood pressure
to increase (P<.05), partially reversing the acute
antihypertensive effect of baicalein in the hypertensive rats.
View larger version (22K):
[in a new window]
Figure 3. Effect of 5,6-dihydro-PGI2 antiserum
( , 0.3 mL IV, n=5) or nonimmune serum (, 0.3 mL IV, n=5) on mean
arterial pressure of rats with Ang II–induced hypertension
pretreated with baicalein (60 mg/kg SC). Results are mean±SEM; n
indicates number of rats. *P<.05 relative to data
obtained just before the injection of antiserum or nonimmune
serum.. Before the injection of the antiserum,
PGI2 and PGE2 caused blood
pressure to fall promptly by 36±1 and 22±2 mm Hg, respectively,
followed by a return to preinjection levels within the next 3 to 4
minutes. After injection of the antiserum, vasodepressor responsiveness
to PGI2 but not to PGE2 was
attenuated (P<.05) for up to 90 minutes.
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Figure 4. Reductions in mean arterial pressure
(MAP) induced by bolus injections of PGI2 (2.0 µg/kg IV,
n=5; top) or PGE2 (2.0 µg/kg IV, n=5; bottom) in
normotensive rats before (open bars) and after (filled bars) the
administration of 5,6-dihydro-PGI2 antiserum (0.3 mL IV).
Results are mean±SEM; n indicates number of rats.
*P<.05 relative to control data obtained before
antiserum administration. displays data on the conversion of
exogenous PGH2 to PGI2 by
rings of descending thoracic aorta taken from normotensive and
hypertensive rats with and without baicalein treatment. In animals
without baicalein treatment, the conversion of
PGH2 to PGI2 by aortic
rings of normotensive rats surpassed (P<.05) that by aortic
rings of hypertensive rats. Baicalein treatment of the hypertensive
rats increased (P<.05) the conversion of exogenous
PGH2 to PGI2 by aortic
rings, whereas baicalein treatment of normotensive rats did not.
View larger version (20K):
[in a new window]
Figure 5. Conversion of exogenous PGH2 to
PGI2 (measured as 6-keto-PGF1 ) by rings of
descending thoracic aorta during incubation for 3 minutes at 37°C in
indomethacin-containing Krebs' bicarbonate buffer. The
aortic rings were obtained from sham-infused normotensive rats and rats
with Ang II–induced hypertension treated with baicalein (60 mg/kg SC)
or sesame oil vehicle only. Results are mean±SEM; n indicates number
of rats. *P<.05 relative to data in vehicle-treated
rats. shows data on the release of
6-keto-PGF1 from the rings of descending
thoracic aorta taken from normotensive and hypertensive rats with and
without baicalein treatment. In animals without baicalein treatment,
the release of 6-keto-PGF1 from aortic rings
incubated in medium containing arachidonic acid was
higher (P<.05) in hypertensive than in normotensive rats.
Baicalein treatment of the hypertensive rats increased further
(P<.05) the release of 6-keto-PGF1
from aortic rings, whereas baicalein treatment of normotensive rats was
without effect.
View larger version (20K):
[in a new window]
Figure 6. Release of 6-keto-PGF1 from rings
of descending thoracic aorta incubated for 15 minutes at 37°C in
Krebs' bicarbonate buffer containing arachidonic acid.
Aortic rings were obtained from sham-infused normotensive rats and rats
with Ang II–induced hypertension treated with baicalein (60 mg/kg SC)
or sesame oil vehicle. Results are mean±SEM; n indicates number of
rats. *P<.05 relative to data in vehicle-treated
rats. displays data on blood prostaglandins and urinary
excretion of eicosanoids in normotensive and hypertensive rats with and
without baicalein treatment. In animals without baicalein treatment,
the blood concentration of 6-keto-PGF1 and the
urinary excretion rate of 6-keto-PGF1 and
12-HETE were higher (P<.05) in hypertensive than in
normotensive rats. Baicalein treatment of the hypertensive rats
increased (P<.05) further the blood concentration and
urinary excretion of 6-keto-PGF1 while
decreasing (P<.05) the blood concentration of
PGE2 and the urinary excretion of 12-HETE.
Baicalein treatment of normotensive rats did not modify the blood
concentration and urinary excretion of
6-keto-PGF1 and PGE2 but
tended to decrease 12-HETE urinary excretion.
View this table:
[in a new window]
Table 1. Effect of Baicalein on Blood Prostaglandins and
Urinary Excretion of Eicosanoids in Sham-Infused Normotensive Rats and
Rats With Ang II–Induced Hypertension
Discussion
Top
Abstract
Introduction
Methods
Results
Discussion
References
The results of the present study demonstrate that treatment
with baicalein or CDC lowers blood pressure in rats with Ang
II–induced hypertension but not in normotensive rats. The study also
demonstrates that the renal excretion of 12-HETE is greater in
hypertensive than in normotensive rats and that baicalein treatment of
the hypertensive rats reduces urinary 12-HETE excretion to levels not
different from those in normotensive rats, which is in agreement with
reports that the drug inhibits
lipoxygenase(s).6 12 18 These
findings are consistent with involvement of products of
lipoxygenase activity in the mechanisms underlying Ang
II–dependent hypertension in rats. This conclusion also receives
support from reports that Ang II promotes vascular expression of
12-lipoxygenase,1 tissue
production of lipoxygenase products is
increased in models of Ang II–dependent
hypertension,6 11 and
lipoxygenase-derived eicosanoids contribute directly or
indirectly to the vascular actions of Ang
II.7 9 10 12 ,
elevated urinary excretion of 6-keto-PGF1, and
enhanced release of 6-keto-PGF1 from rings of
thoracic aorta during incubation in medium containing
arachidonic acid. Paradoxically, the ability of aortic
rings to metabolize exogenous PGH2 to
PGI2 was reduced in rats with Ang II–induced
hypertension, implying that vascular PGI2
synthase activity is reduced in the hypertensive rats. In these
animals, the increased release of 6-keto-PGF1
from aortic rings may be linked to overproduction of
PGH2, with the resulting elevation in cellular
PGH2 concentration driving up
PGI2 production in the face of reduced
PGI2 synthase activity. from
aortic segments incubated in medium containing
arachidonic acid, the concentration of
6-keto-PGF1 in blood, and the renal excretion
of 6-keto-PGF1. These effects of baicalein are
not driven by mechanisms that promote formation of all prostanoids
because baicalein treatment of hypertensive rats elicited reduction of
blood PGE2 levels and did not affect the renal
excretion of PGE2. from aortic rings incubated with
arachidonic acid, the blood concentration of
6-keto-PGF1, or the renal excretion of
6-keto-PGF1. The effects of baicalein on the
status of PGI2 formation and levels in rats with
Ang II–induced hypertension may be the result of interference with the
production of endogenous factors capable of
disrupting the metabolism of PGH2 to
PGI2 by PGI2 synthase. In
this context, reports that PGI2 synthase is
inhibited by 12-HPETE and other hydroperoxides derived from
polyunsaturated fatty acids via metabolism by
lipoxygenase are particularly
significant.5 6 15 Because Ang II stimulates
expression of 12-lipoxygenase,1
the administration of lipoxygenase
inhibitors baicalein or CDC to rats with Ang II–induced
hypertension may promote vascular conversion of
PGH2 to PGI2 by minimizing
the formation of lipoxygenase-derived fatty acid
hydroperoxides.6 from aortic
rings, concentration of 6-keto-PGF1 in blood,
and renal excretion of 6-keto-PGF1. These data
suggest a contribution of PGI2 to the acute
antihypertensive effect of baicalein in rats with Ang II–induced
hypertension. A vasodepressor prostanoid also appears to contribute to
the antihypertensive effect of CDC in rats with Ang II–induced
hypertension. Baicalein and CDC may promote PGI2
formation by interfering with the production of one or more
inhibitors of PGI2 synthase,
including lipoxygenase-derived fatty acid
hydroperoxides and reactive oxygen species.
Selected Abbreviations and Acronyms
Ang II
=
angiotensin II
CDC
=
cinnamyl-3,4-dihydroxy-
-cyanocinnamate
12-HETE
=
12-hydroxyeicosatetraenoic acid
12-HPETE
=
12-hydroperoxyeicosatetraenoic acid
PGE2
=
prostaglandin E2
PGF1
=
prostaglandin F1
PGH2
=
prostaglandin H2
PGI2
=
prostaglandin I2, or prostacyclin
TXA2
=
thromboxane A2
TXB2
=
thromboxane B2
Acknowledgments
This work was supported by grants 5-PO1-HL-4300 and HL-18579
from the National Institutes of Health. We thank Dr Lawrence Levine
from Brandeis University for donating the
5,6-dihydro-PGI2 antiserum used in these studies.
We also thank Chiara Kimmel-Preuss and Jennifer Brown for
secretarial assistance.
References
Top
Abstract
Introduction
Methods
Results
Discussion
References
1.
Natarajan R, Gu JL, Rossi J, Gonzales N, Lanting
L, Xu L, Nadler J. Elevated glucose and angiotensin II
increase 12-lipoxygenase activity and expression in
porcine aortic smooth muscle cells. Proc Natl Acad Sci
U S A. 1993;90:4947–4951., 15-hydroxy-9-11
(epoxymethano)prosta-5,13-dienoic acid and
15-hydroxy-11,9(epoxymethano) prosta-5,13-dienoic acid.
Prostagland Med. 1979;2:177–189.
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