From the Department of Medicine III (K.H., I.K., I.S., D.H., S.K., T.M.,
Y.Y.), University of Tokyo School of Medicine; Second Department of Internal
Medicine (K.K., H.M.), Kansai Medical University, Osaka; Lead Generation
Research Laboratories (T.S.), Tanabe Seiyaku Co, Ltd, Osaka; and Institute of
Applied Biochemistry (K.M.), University of Tsukuba, Ibaraki, Japan.
Correspondence to Issei Komuro, MD, Department of Medicine III, University of Tokyo School of Medicine, 7–3-1 Hongo, Bunkyo-ku, Tokyo 113, Japan. E-mail komuro-tky{at}umin.u-tokyo.ac.jp
Methods and Results—To determine whether AT1-mediated
signaling is indispensable for the development of pressure
overload–induced cardiac hypertrophy, pressure overload
was produced by constricting the abdominal aorta of AT1A
knockout (KO) mice. Quantitative reverse transcriptase–polymerase
chain reaction revealed that the cardiac AT1 (probably
AT1B) mRNA levels in AT1A KO mice were <10%
of those of wild-type (WT) mice and were not affected by pressure
overload. Chronic treatment with subpressor doses of Ang II increased
left ventricular mass in WT mice but not in KO mice.
Pressure overload, however, fully induced cardiac
hypertrophy in KO as well as WT mice. There were no
significant differences between WT and KO mice in expression levels of
fetal-type cardiac genes, in the left ventricular wall
thickness and systolic function as revealed by the
transthoracic echocardiogram, or in the
histological changes such as myocyte
hypertrophy and fibrosis.
Conclusions—AT1-mediated Ang II signaling is not
essential for the development of pressure overload–induced cardiac
hypertrophy.
The intracellular signals are evoked by Ang II through seven
transmembrane Ang II receptors.27 28 At
present, Ang II receptors are divided into two major subtypes,
AT1 and AT2 receptors, and
AT1 receptors are further subdivided into
AT1A and AT1B receptors,
which are the products of different genes.28
It is generally accepted from many studies using subtype-specific
receptor antagonists that most of the well-known Ang II
functions in the cardiovascular system are mediated by
AT1.27 28 Studies of gene
targeting have clearly shown that AT1A-mediated
Ang II signaling is essential for the maintenance of systemic
blood pressure.29 30 In contrast, the
physiological roles of AT2 in
the cardiovascular system were largely unknown. Recent
studies of AT2 KO mice, however, have revealed
that AT2 is also involved in the regulation of
systemic blood pressure.31 32 These results
suggest that genetically engineered mice are powerful tools to clarify
the pathophysiological roles of the RAS in
cardiovascular systems.
In the present study, to examine whether Ang II is essential for
the development of pressure overload–induced cardiac
hypertrophy, chronic pressure overload was produced in the
heart of AT1A KO mice. Chronic pressure overload
induced cardiac hypertrophy in KO mice as well as in WT
mice, although the administration of subpressor doses of Ang II does
not induce cardiac hypertrophy in KO mice. Pressure
overload fully induced all phenotypes observed in cardiac
hypertrophy, such as cardiomyocyte
hypertrophy, reprogramming of gene expression, and
perivascular fibrosis in both kinds of mice. These results suggest that
pressure overload can induce cardiac hypertrophy without
the AT1-mediated Ang II signaling pathways.
Development of Pressure Overload–Induced Cardiac
Hypertrophy
Quantitative RT-PCR Analysis
Chronic Administration of Subpressor Dose of Ang II
Northern Blot Analysis
Echocardiographic Analysis
Histological Analysis
Statistical Analyses
Effects of Subpressor Doses of Angiotensin II on
Development of Cardiac Hypertrophy
Cardiac Hypertrophy by Abdominal Aortic
Banding
Effects of Pressure Overload on Expression of Fetal-Type
Cardiac Genes
Morphological and Functional Changes in Hypertrophied
Hearts
Histological Changes in Hypertrophied
Hearts
We have developed an in vitro system of stretching cardiac myocytes
cultured on deformable silicon dishes17 18 and
have demonstrated that mechanical stretch induces activation of the
protein kinase cascade of phosphorylation, transient
expression of immediate early response genes, and an increase in
protein synthesis, resulting in cardiomyocyte
hypertrophy.17 18 19 20 21 These hypertrophic
responses to mechanical stretch were inhibited by pretreatment with
AT1
antagonists,22 23 suggesting that the
endogenous Ang II secreted in response to mechanical
stretch mediates the signaling pathway, leading to the development of
cardiomyocyte hypertrophy in vitro. However,
because treatment with AT1
antagonists only partially inhibited the hypertrophic
responses to mechanical stretch in cardiac myocytes, signaling pathways
other than those provoked by Ang II seem to be involved in the
formation of cardiac hypertrophy.22
In addition, it has been reported that there are both common and
divergent pathways between Ang II–and mechanical stretch–induced
signaling pathways.22 25 26 These results,
together with the findings that several humoral factors other than Ang
II can induce cardiac
hypertrophy,7 24 49 suggest that
AT1A-mediated Ang II signaling is not the only
essential factor for the development of mechanical stress–induced
cardiac hypertrophy.
Recent progress in mouse genetics has brought about great advances in
the understanding of regulatory mechanisms of the systemic circulatory
system. Targeted gene disruptions of
angiotensinogen,50
ACE,51
AT1A,29 30 and
AT231 32 have indicated that the
RAS plays a critical role in maintaining the homeostasis of systemic
circulation. In mice lacking the AT1A gene, blood
pressure was low and was not elevated with the administration of Ang II
(Fig 2A
This study suggests that there are two distinct pathways,
AT1-dependent and
AT1-independent pathways, in pressure
overload–induced cardiac hypertrophy and that the
AT1-independent pathway can fully substitute for
the AT1-dependent pathway. We recently observed
that acute pressure overload produced by transverse aortic banding
could induce acute hypertrophic responses in KO mice as well as WT
mice.53 It remains to be determined what factors
substitute for AT1A-mediated signaling in the
development of cardiac hypertrophy in
AT1A KO mice. We have demonstrated that ET-1 is
involved in stretch-induced cardiomyocyte
hypertrophy in vitro.24 Cultured
cardiac myocytes secrete ET-1 in response to passive mechanical
stretch, and ETA receptor activation mediates the
stretch-induced hypertrophic responses. In our present experimental
model, endogenous ET-1 may substitute for Ang II in the
development of cardiac hypertrophy. Moreover, we have
observed that passive stretch induced hypertrophic responses in
cultured cardiac myocytes prepared from KO mice as well as WT mice and
that tyrosine kinase inhibitors potently inhibited the
stretch-induced responses in KO cardiac myocytes but not in WT
cardiomyocytes.54 These results
suggest that some humoral factors that activate tyrosine
kinases are secreted in response to mechanical stress or that tyrosine
kinase pathways, which are usually inhibited by
AT1-mediated signals,55 may
be activated in the absence of AT1A.
In summary, by using AT1A KO mice, we
demonstrated that AT1A-mediated Ang II signaling
is not indispensable for the development of pressure overload–induced
cardiac hypertrophy and that signaling pathways other than
those provoked by Ang II can fully induce hypertrophic responses during
hemodynamic overload in the absence of
AT1. The identification of the
AT1-independent signaling pathways that are
involved in pressure overload–induced cardiac hypertrophy
will provide new insights into the development of novel therapeutic
strategies for cardiac hypertrophy.
Received October 31, 1997;
accepted December 2, 1997.
© 1998 American Heart Association, Inc.
Basic Science Reports
Pressure Overload Induces Cardiac Hypertrophy in Angiotensin II Type 1A Receptor Knockout Mice
Abstract
Top
Abstract
Introduction
Methods
Results
Discussion
References
Background—Many studies have
suggested that the renin-angiotensin system plays an
important role in the development of pressure overload–induced cardiac
hypertrophy. Moreover, it has been reported that pressure
overload–induced cardiac hypertrophy is completely
prevented by ACE inhibitors in vivo and that the stored
angiotensin II (Ang II) is released from cardiac myocytes
in response to mechanical stretch and induces cardiomyocyte
hypertrophy through the Ang II type 1 receptor
(AT1) in vitro. These results suggest that the
AT1-mediated signaling is critical for the development of
mechanical stress–induced cardiac hypertrophy.
Key Words: hypertrophy • pressure • angiotensin • genes
Introduction
Top
Abstract
Introduction
Methods
Results
Discussion
References
A growing body
of evidence suggests that Ang II plays an important role in the
development of cardiac
hypertrophy.1 2 ACE
inhibitors or Ang II receptor antagonists
induce the regression and prevent the development of cardiac
hypertrophy both in experimental animal
models3 4 5 6 7 and in hypertensive
patients.8 9 10 Many studies have demonstrated
that hemodynamic overload activates the tissue
RAS in the heart.11 12 13 14 15 16 mRNA and/or protein
levels of renin,11
ACE,12 13
angiotensinogen,5 11 14 and Ang II
receptors15 16 have been reported to be increased
in hypertrophied hearts. In addition to these in vivo studies,
mechanisms by which mechanical stress induces cardiomyocyte
hypertrophy have also been investigated in vitro with
cultured cardiac myocytes. We have developed an in vitro system of
stretching cultured cardiac myocytes seeded on deformable silicon
dishes.17 18 Using this system, we demonstrated
that mechanical stretch of cultured cardiac myocytes activates
the phosphorylation cascade of protein kinases, induces
the expression of immediate early genes and fetal-type genes, and
increases the protein synthesis rate.17 18 19 20 21
Furthermore, we reported that Ang II plays a critical role in the
mechanical stretch–induced cardiomyocyte
hypertrophy.22 It was recently
reported that Ang II is stored in the secretory granules of cardiac
myocytes and that secretion is induced by mechanical
stress.23 However, mechanical stretch–induced
cardiomyocyte hypertrophy was not completely
prevented by Ang II receptor
antagonists,22 suggesting that there
might be signaling pathways other than those provoked by Ang II.
Indeed, endothelin-1 (ET-1) also plays a pivotal role in the
development of mechanical stress–induced cardiomyocyte
hypertrophy.24 Recent studies have
also demonstrated that there are some differences between Ang II–and
mechanical stress–induced signaling pathways in cardiac
myocytes.22 25 26 Therefore, it is questionable
whether Ang II is indispensable for the development of mechanical
stress–induced cardiac hypertrophy.
Methods
Top
Abstract
Introduction
Methods
Results
Discussion
References
Animals
AT1A KO mice (n=10) and WT mice (n=10), 18
weeks old, from the same genetic background were used in the
present study.29 Mice were housed under
climate-controlled conditions with a 12-hour light/dark cycle and were
provided with standard food and water ad libitum. All protocols were
approved by local institutional guidelines.
Pressure overload was produced by constriction of the abdominal
aorta as described previously.33 Briefly, mice
were anesthetized by injection of sodium pentobarbital (30
mg/kg IP). The abdominal aorta was constricted at the suprarenal level
with 7–0 nylon strings by ligation of the aorta with a blunted
29-gauge needle, which was pulled out thereafter. To monitor the
hemodynamic effects of aortic constriction at 40 days
after aortic banding, the left carotid artery was cannulated with
stretched PE 50 tubing, which was tunneled under the skin and
exteriorized posteriorly at the base of the neck. After the mice had
completely recovered from anesthesia, blood pressure was
measured in conscious mice under unrestrained conditions and was
recorded continuously over a period of 60 minutes by a polygraph
system (Nihon Koden Co). On the basis of the arterial blood
pressure recordings during this period, a mean value for
systolic pressure was calculated. After blood pressure was
recorded, hearts were excised, weighed, and subjected to further
analysis.
Total RNA was prepared from the hearts of mice by use of RNA
STAT-60 (TEL-TEST "B," Inc) followed by digestion with DNase
(Takara Shuzo) to eliminate any contamination of genomic DNA. The
RT-PCR analysis for AT1 and
AT2 mRNA quantification was performed with the
deletion-mutated cRNA as described
previously.15 16 34 The amplification
efficiencies of target and competitor transcripts are equal under
optimal concentrations of competitor transcripts. The
oligonucleotide primers used for RT-PCR
analysis are as follows: for AT1,
5'-GAGTCCTGTTCCACCCGATCACCGATCAC-3' and
5'-GGATGACGCCCAGCTGAATCAGCACATCC-3'; for AT2,
5'-TTGCTGCCACCAGCAGAAAC-3' and 5'-GTGTGGGCCTCCAAACCATTGCTA-3'. The
sequence of these primers is identical to that of murine Ang II
receptor cDNA.35 36 To verify that equal amounts
of RNA were subjected to RT-PCR, GAPDH mRNA was also amplified with the
following primers: 5'-GGTCGGTGTGAACGGATTTG-3' and
5'-GACGGACACATTGGGGGTAG-3'. Denaturing (94°C for 45 seconds),
annealing (58°C for 1 minute), and extension (72°C for 1 minute)
reactions were performed for 30 cycles. Because the primers used for
the amplification of AT1 correspond to common
sequences between AT1A and
AT1B, both AT1A and
AT1B mRNAs were amplified. When PCR was performed
without the step of RT, no PCR product was amplified, indicating
that the product was not generated by the amplification of
contaminated genomic DNA. The range of concentrations of sample RNA and
internal control–deleted cRNA, as well as the number of amplification
cycles, was selected from within the exponential phase. To determine
the amount of mRNA, 5 µCi of [
-32P]dCTP
was included in the PCR reaction mixtures, and the incorporated
32P activity was measured with a scintillation
counter.
An osmotic minipump (model 2002, Alza Corp) was implanted
subcutaneously into mice. Subpressor doses of Ang II (100 ng ·
kg-1 ·
min-1)37 and 0.01 mol/L
acetic acid in saline or saline alone were administered for 14 days,
and arterial blood pressure was measured as described
above. After blood pressure was recorded under unrestrained
conditions, hearts were rapidly excised and weighed.
Ten micrograms of total RNA was separated on a 1.2%
agarose/formaldehyde gel and blotted onto Hybond-N membrane (Amersham
Co). cDNA of ANP, BNP, and SERCA38 and
oligonucleotide DNA of ß-MHC39
were used as probes. Hybridizing bands were quantified with a FUJIX
Bio-Imaging Analyzer BAS 2000 (Fuji Film Co).
Transthoracic echocardiography
was performed with HP Sonos 100 (Hewlett-Packard Co) with a 10-MHz
imaging transducer as described previously.40
Mice were anesthetized with ketamine (10 mg/kg IP) and
xylazine (15 mg/kg IP). After a good-quality two-dimensional image was
obtained, M-mode images of the left ventricle were recorded.
Intraventricular septum thickness,
end-diastolic left ventricular internal
diameter (EDD), end-systolic left ventricular
internal diameter (ESD), and left ventricular posterior
wall thickness were measured. All measurements were performed by use of
the leading edge–to–leading edge convention adopted by the American
Society of
Echocardiography.41 Percent
fractional shortening (%FS) was calculated as
%FS=[(EDD-ESD)/EDD]x100. End-diastolic volume and
end-systolic volume were automatically calculated on a cardiac
ultrasound machine. Left ventricular ejection fraction (EF)
was calculated by the cubed method as follows:
EF=[(EDD)3-
(ESD)3]/EDD3.
For histological analysis, hearts were
fixed with 10% formalin by perfusion fixation. Fixed hearts were
embedded in paraffin, sectioned at 4-µm thickness, and stained with
hematoxylin-eosin for overall morphology or by the van Gieson method
for collagen. Myocyte cross-sectional area was measured from sections
stained with hematoxylin-eosin, and suitable cross sections were
defined as having nearly circular capillary profiles and nuclei. To
determine the degree of collagen fiber accumulation, we selected five
fields randomly and calculated the ratio of van Gieson–stained
fibrosis area to total myocardium area as described
previously with the image analysis software NIH IMAGE (NIH,
Research Service Branch).40
All results are expressed as mean±SEM. Multiple comparisons
among three or more groups were carried out by two-way ANOVA and
Fisher's exact test for post hoc analyses. A value of
P<.05 was considered statistically significant.
Results
Top
Abstract
Introduction
Methods
Results
Discussion
References
Quantitative RT-PCR Analysis of AT1 and
AT2 mRNA in Murine Hearts
Because AT1A mRNA levels were too low to be
detected by Northern blot analysis or RNase protection assay,
we subjected mRNA levels of Ang II receptors to semiquantitative
analysis by competitive RT-PCR methods as described
previously.15 16 34 mRNA levels of cardiac
AT1 gene in KO mice were <10% of those in WT
mice (Fig 1A
). Because the PCR
products of AT1 in AT1A
KO mice were completely digested by HincII (data not shown),
this slight expression of AT1 observed in KO
hearts should represent AT1B gene
transcripts.42 In WT mice, cardiac
AT1 mRNA levels were significantly higher in
hypertrophied hearts than in control hearts (1.5-fold;
P<.05), whereas no significant difference was observed in
the amount of AT2 mRNA levels between control and
hypertrophied hearts (Fig 1B), in good agreement with the previous
study of murine hypertrophied hearts.43 In
contrast, cardiac mRNA levels of both AT1 and
AT2 were not affected by pressure overload in KO
mice (Fig 1A and 1B).
View larger version (59K):
[in a new window]
Figure 1. Cardiac AT1 and AT2 mRNA
levels before and 40 days after aortic banding. RT-PCR analysis
was performed to evaluate mRNA levels of AT1 (A) and
AT2 (B). Normalized values of AT1 and
AT2 in WT heart before aortic banding are arbitrarily
expressed as 100% (right). RT-PCR was performed with primers specific
for murine AT1 and AT2 in reaction mixture
containing [ -32P]dCTP. Amplified DNA was
electrophoresed in 1% agarose gel and stained with ethidium bromide
(left). AT1 and AT2 mRNA levels were normalized
by deletion-mutated cRNA amplified at same time. *P<.05
vs sham-operated WT mice. 
AT1 and AT2
indicate amplified DNA from deleted cRNA as internal controls.
Molecular weight marker (MWM) is also shown.
AT1A and AT1B are 96%
identical at amino acid levels and pharmacologically indistinguishable
from each other.28 Because RT-PCR
analysis has revealed that trace amounts of
AT1 (probably AT1B) exist
in the heart of KO mice, the possible involvement of
AT1B in the development of cardiac
hypertrophy was examined. For this purpose, we continuously
administered a subpressor dose (100 ng ·
kg-1 · min-1) of
Ang II into WT and KO mice for 2 weeks. Systolic blood pressure
(Fig 2A) and body weight (data not shown)
were not changed by the administration of Ang II in any mice. The Ang
II treatment, however, markedly increased the heart weight/body weight
ratio in WT mice but not in KO mice (Fig 2B). Microscopic
analysis revealed that cross-sectional areas of cardiac
myocytes were increased only in WT mice (Fig 2C). In addition, Ang II
treatment induced upregulation of the ANP gene only in WT mice (Fig 2D). These results strongly suggest that in KO hearts, Ang II does not
evoke enough signals through AT1 to lead to the
development of cardiac hypertrophy.
View larger version (28K):
[in a new window]
Figure 2. Effects of chronic Ang II administration.
Subpressor dose of Ang II (100 ng · kg-1 ·
min-1 SC) was continuously infused into mice by an osmotic
minipump for 14 days. A, Arterial blood pressure.
*P<.005 vs WT mice. B, Heart weight (mg)/body weight
(g) ratio. *P<.05 vs WT sham-operated mice. C,
Cross-sectional areas of cardiac myocytes before and 14 days after Ang
II infusion. Normalized values in WT mice before pressure overload are
arbitrarily expressed as 1.0. *P<.001 vs sham-operated
mice of each group. D, Representative
autoradiograms of Northern blot analysis of ANP
gene. Similar results were obtained from three independent experiments.
Ethidium bromide staining of 18S ribosomal RNA is presented to
show that loaded mRNA is equal and intact. Sham indicates sham-operated
mice; Ang II, Ang II–infused mice.
Many laboratories have reported that cardiac
hypertrophy is induced by constriction of the abdominal
aorta through activation of the RAS.5 6 7 At 40
days after operation, blood pressure was still elevated in both WT and
KO mice. The increase in systolic blood pressure was not
significantly different between WT and KO mice (WT, 37±7 mm Hg;
KO, 43±7 mm Hg), although the baseline blood pressure of KO mice
was significantly lower than that of WT mice (WT, 127±5 mm Hg;
KO, 92±6 mm Hg; P<.001) (Fig 3A), consistent with a previous
report.29 Unexpectedly, the heart weight/body
weight ratio was markedly increased not only in WT mice but also in KO
mice, and the degree of increase in this ratio was almost identical
between WT and KO mice (WT, 29% increase; KO, 33% increase) (Fig 3B),
indicating that pressure overload can increase left
ventricular weight without the
AT1-mediated signaling.
View larger version (13K):
[in a new window]
Figure 3. Effects of chronic pressure overload produced by
abdominal aortic banding. A, Arterial blood pressure.
*P<.005 vs sham-operated mice of each group;
**P<.001 vs WT sham-operated mice. B, Heart weight
(mg)/body weight (g) ratio. *P<.001 vs sham-operated
mice of each group.
Upregulation of fetal-type genes and downregulation of the SERCA
gene are well-established genetic responses to pressure overload
produced by abdominal aortic banding.17 38 44 Ang
II has been reported to be involved in the reprogramming of gene
expressions induced by mechanical stress.2 5 6 We
therefore examined the expression of these genes in hearts of WT and KO
mice. Abdominal aortic banding induced upregulation of fetal-type
cardiac genes such as ANP, BNP, and ß-MHC genes and downregulation of
SERCA gene in hearts of both mice (Fig 4). There were no significant differences
in expression levels of these genes between WT and KO mice (Fig 4).
These results suggest that pressure overload induces the reprogramming
of gene expression in the heart irrespective of the presence of
AT1-mediated Ang II signaling.
View larger version (82K):
[in a new window]
Figure 4. Effects of chronic pressure overload on cardiac
gene expressions. Representative
autoradiograms of Northern blot analysis.
Similar results were obtained in three independent experiments.
Ethidium bromide staining of 18S ribosomal RNA is presented to
show that loaded mRNA is equal and intact.
To determine the changes in left ventricular size and
function, transthoracic
echocardiography was performed before and 40 days
after aortic banding. The left ventricular wall was thicker
and left ventricular internal dimension was significantly
larger in pressure overloaded hearts than in sham-operated hearts of
both WT mice and KO mice (Fig 5, Table). In addition, almost the same
degree of decrease in percent fractional shortening and ejection
fraction was observed in both animal groups (Table).
View larger version (98K):
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Figure 5. Representative charts of
transthoracic M-mode echocardiograms. A, WT mice at 40 days
after sham operation; B, KO mice at 40 days after sham operation; C, WT
mice at 40 days after aortic banding; D, KO mice at 40 days after
aortic banding. IVS indicates interventricular septum;
LVID, left ventricular internal dimension; and PW,
posterior wall.
View this table:
[in a new window]
Table 1. Parameters in WT and KO Mice
It has been reported that Ang II plays an important role in the
development of ventricular remodeling, including
cardiomyocyte hypertrophy and fibrosis, through
AT1.45 46 47 48 Microscopic
analysis revealed that cross-sectional areas of cardiac
myocytes were increased both in KO mice and in WT mice (Fig 6A), indicating that each cardiac myocyte
was increased in size by pressure overload without
AT1 signaling. Perivascular fibrosis was
prominent after pressure overload in KO hearts as well as in WT hearts
(Fig 6B). The fibrotic area in KO hearts was almost the same as that in
WT hearts after pressure overload (Fig 6C). In addition, the thickness
of arterial walls was also increased in hearts of both
kinds of mice (Fig 6B). These results suggest that pressure overload
induces not only cardiac hypertrophy but also myocardial
fibrosis, even when the heart lacks signaling through
AT1.
View larger version (28K):
[in a new window]
Figure 6. Light microscopic analysis. A,
Cross-sectional areas of cardiac myocytes before and 40 days after
aortic banding. Normalized values in WT mice before pressure overload
are arbitrarily expressed as 1.0. *P<.001 vs
sham-operated mice of each group. B, Histological
examination of left ventricular fibrosis (van Gieson
staining). Perivascular collagen accumulation (red) is prominent in
both hearts after pressure overload. Original magnification, x160. C,
Relative area of fibrosis before and 40 days after aortic banding.
Normalized values in sham-operated heart are arbitrarily expressed as
1. *P<.001 vs sham-operated mice of each group.
Discussion
Top
Abstract
Introduction
Methods
Results
Discussion
References
Many studies have demonstrated that treatment with ACE
inhibitors or AT1
antagonists efficiently prevents and/or reduces pressure
overload–induced cardiac hypertrophy in both experimental
and clinical studies.3 4 5 6 7 8 9 10 In addition, all
components of the RAS are upregulated by pressure overload in the
heart.5 11 12 13 14 15 16 All these results suggest that the
RAS plays a critical role in the development of mechanical
stress–induced cardiac hypertrophy. In the present
study, however, we have demonstrated that hemodynamic
overload fully induces cardiac hypertrophy in genetically
engineered mice that lack the AT1A gene,
suggesting that AT1-mediated Ang II signaling is
not indispensable for the development of pressure overload–induced
cardiac hypertrophy. 
), suggesting that AT1A plays a critical
role in the Ang II–mediated regulation of blood pressure. Although the
RAS is highly activated in AT1A KO mice,
any histological abnormalities are not detectable in
the heart.29 30 This study also demonstrates that
slightly expressed AT1B was not functional in the
development of Ang II–induced cardiac hypertrophy (Fig 2B
through 2D). Although our experiments do not prove the role of
AT1B, these findings suggest that
AT1 is not involved in the development of cardiac
hypertrophy in AT1A KO mice.
Therefore, we tried to dissect the roles of
AT1-dependent and -independent signaling pathways
in the development of mechanical stress–induced cardiac
hypertrophy by creating pressure overload in
AT1A KO mice. At 40 days after abdominal aortic
banding, pressure overload induced almost the same degree of cardiac
hypertrophy in both KO and WT mice, which is
consistent with a recent report.52
Cardiac hypertrophy in KO mice was associated with the
reexpression of fetal-type cardiac genes and ventricular
remodeling, such as perivascular fibrosis. Unexpectedly, in the
abdominal aortic banding model, all these changes were
indistinguishable between WT hearts and KO hearts. Although these
results do not rule out the possibility that Ang II plays a critical
role in forming cardiac hypertrophy in a
physiological context, it strongly suggests that
cardiac hypertrophy can be induced by pressure overload
without Ang II signaling through AT1A. Therefore,
we speculate that blockade of AT1 with
pharmacological agents may not have any effects on the development of
cardiac hypertrophy in KO mice.
Selected Abbreviations and Acronyms
Ang II
=
angiotensin II
ANP
=
atrial natriuretic peptide
AT1
=
angiotensin II type 1 receptor
AT1A
=
angiotensin II type 1A receptor
AT1B
=
angiotensin II type 1B receptor
AT2
=
angiotensin II type 2 receptor
BNP
=
brain natriuretic peptide
ET-1
=
endothelin-1
KO
=
knockout
MHC
=
myosin heavy chain
PCR
=
polymerase chain reaction
RAS
=
renin-angiotensin system
RT
=
reverse transcriptase
SERCA
=
sarcoplasmic reticulum Ca2+-ATPase
WT
=
wild-type
Acknowledgments
This work was supported by a grant-in-aid for scientific
research and developmental scientific research from the Ministry of
Education, Science, and Culture and a grant from the Study Group of
Molecular Cardiology Japan (Dr Komuro). We are very
grateful for the help of Sanae Ogawa and Dr Kenjiro Kimura for
histology.
References
Top
Abstract
Introduction
Methods
Results
Discussion
References
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T. Arimoto, Y. Takeishi, H. Takahashi, T. Shishido, T. Niizeki, Y. Koyama, R. Shiga, N. Nozaki, O. Nakajima, K. Nishimaru, et al. Cardiac-Specific Overexpression of Diacylglycerol Kinase {zeta} Prevents Gq Protein-Coupled Receptor Agonist-Induced Cardiac Hypertrophy in Transgenic Mice Circulation, January 3, 2006; 113(1): 60 - 66. [Abstract] [Full Text] [PDF]
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S. Kawano, T. Kubota, Y. Monden, N. Kawamura, H. Tsutsui, A. Takeshita, and K. Sunagawa Blockade of NF-{kappa}B ameliorates myocardial hypertrophy in response to chronic infusion of angiotensin II Cardiovasc Res, September 1, 2005; 67(4): 689 - 698. [Abstract] [Full Text] [PDF]
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P. Bridgman, M. A. Aronovitz, R. Kakkar, M. I. Oliverio, T. M. Coffman, W. M. Rand, M. A. Konstam, M. E. Mendelsohn, and R. D. Patten Gender-specific patterns of left ventricular and myocyte remodeling following myocardial infarction in mice deficient in the angiotensin II type 1a receptor Am J Physiol Heart Circ Physiol, August 1, 2005; 289(2): H586 - H592. [Abstract] [Full Text] [PDF]
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A. H. Maass, K. Ikeda, S. Oberdorf-Maass, S. K.G. Maier, and L. A. Leinwand Hypertrophy, Fibrosis, and Sudden Cardiac Death in Response to Pathological Stimuli in Mice With Mutations in Cardiac Troponin T Circulation, October 12, 2004; 110(15): 2102 - 2109. [Abstract] [Full Text] [PDF]
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Y. Li, T. Ha, X. Gao, J. Kelley, D. L. Williams, I. W. Browder, R. L. Kao, and C. Li NF-{kappa}B activation is required for the development of cardiac hypertrophy in vivo Am J Physiol Heart Circ Physiol, October 1, 2004; 287(4): H1712 - H1720. [Abstract] [Full Text] [PDF]
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P. Niu, T. Shindo, H. Iwata, S. Iimuro, N. Takeda, Y. Zhang, A. Ebihara, Y. Suematsu, K. Kangawa, Y. Hirata, et al. Protective Effects of Endogenous Adrenomedullin on Cardiac Hypertrophy, Fibrosis, and Renal Damage Circulation, April 13, 2004; 109(14): 1789 - 1794. [Abstract] |