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Lacolley, P. et al. Prevention of aortic and cardiac fibrosis by spironolactone in old normotensive rats. J. Am. Coll. Cardiol. 37, 662-667

Department of Internal Medicine and INSERM U337, Broussais Hospital, Paris, France.
Journal of the American College of Cardiology (Impact Factor: 16.5). 03/2001; 37(2):662-7. DOI: 10.1016/S0735-1097(00)01129-3
Source: PubMed
ABSTRACT
Because the synthesis of aldosterone is mainly modulated by angiotensin II through type I receptor stimulation and because converting enzyme inhibition (CEI) does not modify aortic extracellular matrix in old normotensive rats, the aim of the present study was to determine whether inhibition of aldosterone formation was able to prevent aortic fibrosis in old Sprague-Dawley normotensive rats.
We have previously shown that long-term aldosterone antagonism prevents the age-related increase in aortic collagen accumulation in young spontaneously hypertensive rats, independent of blood pressure changes. In contrast, we reported that the positive effects of CEI in the prevention of aortic collagen accumulation were related to the inhibition of angiotensin II actions on angiotensin II type I receptors.
For this purpose, we studied the histomorphometric and stiffness (echo-tracking technique) changes of an eight-week treatment with the aldosterone antagonist spironolactone by comparison with placebo.
At the end of treatment, spironolactone in conscious animals did not change intra-arterial blood pressure, aortic and carotid wall thickness, and cardiac weight. Cardiac collagen density and, to a lesser extent, carotid collagen and elastin densities and contents were significantly decreased in association with an increase of carotid distensibility.
These results show that in old normotensive rats, spironolactone can markedly prevent cardiac and, to a lesser extent, arterial fibrosis and improve arterial stiffness, despite a lack of hypotensive effect.

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Prevention of Aortic and Cardiac Fibrosis
by Spironolactone in Old Normotensive Rats
Patrick Lacolley, MD, PHD, Michel E. Safar, MD, Bernadette Lucet, TECH,
Katia Ledudal, T
ECH, Carlos Labat, TECH, Athanase Benetos, MD, PHD
Paris, France
OBJECTIVES Because the synthesis of aldosterone is mainly modulated by angiotensin II through type I
receptor stimulation and because converting enzyme inhibition (CEI) does not modify aortic
extracellular matrix in old normotensive rats, the aim of the present study was to determine
whether inhibition of aldosterone formation was able to prevent aortic fibrosis in old
Sprague-Dawley normotensive rats.
BACKGROUND We have previously shown that long-term aldosterone antagonism prevents the age-related
increase in aortic collagen accumulation in young spontaneously hypertensive rats, indepen-
dent of blood pressure changes. In contrast, we reported that the positive effects of CEI in the
prevention of aortic collagen accumulation were related to the inhibition of angiotensin II
actions on angiotensin II type I receptors.
METHODS For this purpose, we studied the histomorphometric and stiffness (echo-tracking technique)
changes of an eight-week treatment with the aldosterone antagonist spironolactone by
comparison with placebo.
RESULTS At the end of treatment, spironolactone in conscious animals did not change intra-arterial
blood pressure, aortic and carotid wall thickness, and cardiac weight. Cardiac collagen density
and, to a lesser extent, carotid collagen and elastin densities and contents were significantly
decreased in association with an increase of carotid distensibility.
CONCLUSIONS These results show that in old normotensive rats, spironolactone can markedly prevent cardiac
and, to a lesser extent, arterial fibrosis and improve arterial stiffness, despite a lack of
hypotensive effect. (J Am Coll Cardiol 2001;37:662–7) © 2001 by the American College of
Cardiology
Aging is associated with an increased arterial stiffness (1).
Modifications of arterial structure and function are consid-
ered to be responsible for this alteration independently of
blood pressure changes. With age, aortic medial thickness
and cross-sectional area (CSA) of the aorta increase signif-
icantly, together with the development of extracellular
matrix, principally collagen (1). All these vascular changes,
which predominate on central arteries, can be theoretically
reversed by appropriate drug treatments. For instance,
aminoguanidine prevents the age-related increase of arterial
stiffness, without modifying wall thickness or the total
amount of extracellular matrix, an effect probably resulting
from changes in collagen cross-linking and advanced glyco-
sylation end products (2). In contrast, converting enzyme
inhibition (CEI), which prevents aortic collagen accumula-
tion in young spontaneously hypertensive rats (SHRs) (3),
has no comparable effect in old normotensive rats. In these
animals, CEI reduces significantly blood pressure and aortic
wall thickness but does not modify the amount of extracel-
lular matrix (4).
At the early phase of development in SHRs, spironolac-
tone prevents the accumulation of aortic collagen with
minimal changes of blood pressure (5). This action may be
explained on the basis of aldosterone-mediated mecha-
nisms. First, aldosterone might act physiologically on spe-
cific mineralocorticoid receptors of large vessels (5). A
similar effect has been previously described in detail for the
myocardium, in which spironolactone reduces subendocar-
dial collagen without changing cardiac weight (68). Sec-
ond, aldosterone release is known to be modulated by the
renin-angiotensine-bradykinin systems (5–8). Studies of
old normotensive rats, in which minimal effects of CEI have
been observed on vascular extracellular matrix (4), may be
relevant to evaluate better the mechanisms of action of
spironolactone on large arterial vessels.
The purpose of the present study was to determine in old
Sprague-Dawley normotensive rats the preventive effect of
spironolactone on cardiac, aortic and carotid collagen accu-
mulation as judged by histomorphometry. Changes in
carotid arterial stiffness were studied in parallel using high-
resolution echo-tracking techniques.
METHODS
Twenty-six Sprague-Dawley rats (Iffa Credo, L’Abresle,
France) were housed in our animal room (five to seven per
cage), which was maintained at a temperature of 20° C to
22° C, a humidity of 55% to 65%, and a 12-h light/dark
cycle. The rats were fed a standard diet (0.13 mEq/g Na
and 0.205 mEq/g K
) and had free access to tap water. Rats
were randomly allocated to two groups (n 13 per group)
From the Department of Internal Medicine and INSERM U337, Broussais
Hospital, Paris, France. The study was performed with a grant from Assistance
Publique de Paris, the Institut National de la Sante´ et de la Recherche Me´dicale
(INSERM U337), the Association Claude Bernard, GPH-CV and Searle Pharma-
ceutical Company.
Manuscript received May 8, 2000; revised manuscript received August 18, 2000,
accepted October 2, 2000.
Journal of the American College of Cardiology Vol. 37, No. 2, 2001
© 2001 by the American College of Cardiology ISSN 0735-1097/01/$20.00
Published by Elsevier Science Inc. PII S0735-1097(00)01129-3
Page 1
and treated for eight weeks (seven days per week) starting at
the age of 22 months. One group received placebo, and the
other group received spironolactone at 200 mg/kg body
weight, a dosage previously tested in our laboratory (5). This
dose has also been used in our previous study in SHRs (5).
The different treatments were administered once daily by
gavage. The study was performed according to the interna-
tional guidelines recommended for animal experiments.
Arterial pressure and heart rate evaluation in conscious
rats. Animals were anesthetized with pentobarbital
(50 mg/kg body weight intraperitoneally). A catheter
(PE-50 fused to PE-10) was placed in the lower abdominal
aorta via the femoral artery. The catheter was filled with
heparinized saline (50 U/ml), tunneled under the skin of the
back, and excised between the scapulas. The animals were
then allowed to recover from anesthesia for 24 h in
individual cages. Then arterial pressure measurements were
performed in conscious, freely moving rats in their own cage
after at least a 30-min rest. Arterial pressure and heart rate
were evaluated 24 h after the last drug administration
between 8 and 10 AM. Mean blood pressure and heart rate
were recorded by means of a Statham P23 ID pressure
transducer (Gould) connected to a Gould Brush recorder
(model G 4133) according to previously described method
and standard ethical rules on animal experiments.
Carotid hemodynamic study. At the end of the eight-
week treatment the rats were anesthetized with 50 mg/kg
pentobarbital intraperitoneally. The femoral catheter
(0.9 mm inner diameter), filled with saline and coupled to a
Statham P2S1D pressure transducer (Gould Statham, Ox-
nar, California) was advanced toward the right common
carotid artery and placed in the middle of the lumen.
Changes in left carotid arterial diameter were determined
noninvasively using a high-precision A-mode ultrasonic
device described previously in man and rats (9,10).
Briefly, this device measures internal diameter and its
systolic-diastolic variations with a precision close to 50 and
1
m, respectively. This degree of resolution is made
possible by oversampling (5,000 arterial diameter measure-
ments/s) and averaging 16 consecutive values. Because this
frequency is established as asynchronous with the instru-
ment clock, the resolution of the measurement increases
with the square root of the number of independent time
intervals acquired. A 10-MHz focalized transducer is ster-
eotactically positioned over the common carotid artery, 1 cm
below carotid bifurcation, using gel as transmitting medium.
The artery is simply exposed and not dissected. The
Doppler technique is used to place the probe perpendicu-
larly to the arterial axis, in its largest cross-sectional dimen-
sion. After the transducer is switched to radio-frequency
mode, the backscattered echoes from both anterior and
posterior walls are visualized on an oscilloscopic screen. The
radio-frequency signals of both were exhibiting a high
signal-to-noise ratio and were then easily tagged by an
electronic tracker so that their movement could be derived.
The blood pressure is measured as described above on the
right common carotid artery, simultaneously to the deter-
mination of the left carotid arterial diameter, using a
Statham transducer (P23 Db) and a Gould processor
(Cleveland, Ohio).
From the two continuous signals of pulsatile changes in
arterial diameter (systolic diameter, diastolic diameter) and
blood pressure, the computerized acquisition system fits the
diameter-pressure curve within the diastolic-systolic range
of PP, and then calculates the arterial lumen CSA-pressure
curve, using an arc tangent function as previously described
(9,10). Mean diameter is integrated from the diameter-time
curve and mean CSA is deduced from this parameter.
Because the arterial and the PP signals were not determined
on the same side, we checked previously (9,10) that there
was no time delay between the diameter and the pressure
signals due to the electronic processing. In addition, we
verified in groups of anesthetized rats that the pressure-
CSA curve did not differ whether the diameter signal was
recorded on the right side and the pressure signal on the left
side of the common carotid artery, or the opposite design
was used.
The reproducibility of the method was studied in nine
Sprague-Dawley rats using the coefficient of variation (SD
expressed as a percentage of the mean of 10 successive
measurements). The reproducibility of carotid diameter
measurements and their systolic-diastolic variation was as-
sessed over five measurements, each performed by two
observers over a 30-min period. Under these conditions the
mean intraobserver coefficients of variation were 3 1 and
6 2%.
The distension of an artery (change in volume) during a
cardiac cycle depends on the elastic characteristics of the
vessel wall (and the surrounding tissue) and the local PP
(5,9,10). Local arterial cross-sectional distensibility, assum-
ing a constant length of a cylindrical vessel, is defined by the
percent systolic-diastolic change in luminal CSA for a given
change in intravascular pressure (P). In relation to the
nonlinearity of the mean blood pressure-CSA curve, arterial
distensibility decreases curvilinearly as mean blood pressure
increases. Thus, the distensibility-pressure curve over the
systolic-diastolic range was established by deriving the
equation of the mean blood pressure-CSA curve, allowing
the evaluation of distensibility at any given value of arbitrary
pressure. In this context, “operational” distensibility was
defined as the distensibility corresponding to the steady-
state mean arterial pressure of each animal. As for disten-
sibility, it was possible to define from the CSA- mean blood
pressure curve an operational value of compliance. Compli-
Abbreviations and Acronyms
CEI converting enzyme inhibition
CSA cross-sectional area
P change in pulse pressure
PP pulse pressure
SHRs spontaneously hypertensive rats
663
JACC Vol. 37, No. 2, 2001
Lacolley
et al.
February 2001:662–7
Aortic Fibrosis Prevention by Spironolactone in Old Normotensive Rats
Page 2
ance is calculated as CSA/P, where CSA represents the
absolute pulsatile change in carotid lumen CSA, and there-
fore the product of distensibility by CSA. The carotid
incremental elastic modulus is calculated as the ratio be-
tween the carotid lumen on wall thickness ratio and disten-
sibility, carotid medial thickness and CSA being measured
as detailed in the next section.
Histomorphometric study. Histomorphometric parame-
ters of the carotid artery and of the thoracic aorta were
measured according to the following procedure. At the end
of the hemodynamic study, and after median thoracotomy,
rats were exsanguinated via a catheter placed in the right
auricle while saline was injected into the femoral catheter.
When the liquid from the auricle ran clear, the circulatory
tract was rinsed with a 4% formaldehyde solution. The
animals died very shortly after the formaldehyde infusion
was started. After 1 or 2 min, a clamp was positioned on the
auricle and the fixation liquid infused for3hatapressure
equal to the mean blood pressure of each animal (3,5,11–
13). At the end of perfusion, left ventricular weight was
measured and the thoracic aorta and the carotid artery
dissected and preserved in a 4% formaldehyde solution until
the histological study was performed.
The different structures of the aortic and carotid media
were studied in a vascular segment longitudinally embedded
in paraffin. Three successive sagittal sections 5
m thick
were treated with specific stains to obtain a monochromatic
color associated with each structure of the vessel media.
Sirius red was used for collagen staining, orcein for elastin,
and hematoxylin after periodic acid oxidation for nuclear
staining. Histomorphometric methods have been previously
published in detail (3,5–8,11–13).
For the evaluation of the left ventricular subendocardial
collagen, each heart was cut perpendicularly to the apex to
base axis into four parts of the same thickness. These
specimens were dehydrated through graded ethanol solution
and embedded in paraffin. Two representative sections,
3
m each, were studied from each block, mounted on glass
slide and treated with Sirius red, used for collagen staining
of the left ventricle (14). Then the slides stained with the
Sirius red were examined at a magnification of 250.
Morphometric analysis was performed as described for
aortic segments.
Statistical analysis. Results are expressed as mean 1 SD.
Data were analyzed by one-way analysis of variance
(ANOVA). When F was 0.05, a Bonferroni test was
performed for intergroup comparisons. A value of p 0.05
was considered significant. Univariate correlations were
performed in the overall population using standard tech-
niques.
RESULTS
Blood pressure and heart rate in conscious rats. Table 1
shows the mean values of intra-arterial mean and pulse
blood pressure measured in conscious animals at the end of
the eight-week treatment period, 24 h after the last gavage.
Spironolactone did not induce any significant change in
blood pressure and heart rate when compared to the placebo
group. Body weight was not affected by spironolactone.
None of the placebo- or spironolactone-treated animals
died throughout the treatment period nor during the exper-
imental study.
Carotid arterial mechanical and histomorphological
changes. For the same mean blood pressure, carotid artery
distensibility was significantly higher and carotid elastic
modulus significantly lower in spironolactone- than in
placebo-treated animals (Table 2), whereas there was no
significant change in mean and pulsatile diameter.
Table 3 shows that the carotid and aortic wall thickness
were unchanged by treatment, whereas carotid elastin and
collagen densities and contents were significantly decreased
in the spironolactone-treated rats. The elastin-to-collagen
ratio and the number and size of nuclei were unchanged in
the spironolactone group. Thoracic aorta was poorly mod-
ified, with the exception of a slight increase in elastin
density but not content.
Univariate correlations in the overall population showed
that carotid collagen density (%) was negatively correlated
with carotid distensibility (r
2
0.14; p 0.02) and
positively correlated with carotid artery elastic modulus
(r
2
0.20; p 0.032).
Cardiac structural parameters. Subendocardial collagen of
the left ventricular wall was markedly reduced (80%) by
spironolactone. Compared to placebo rats, the value was:
2.94 0.96 vs. 7.55 2.00 (p 0.0001). There was also
a significant decrease in left ventricular thickness (2.96
0.38 vs. 2.30 0.25 mm; p 0.0001), whereas total heart
Table 1. Body Weight, Intra-arterial Blood Pressure, and Heart
Rate (HR) Data in Conscious Animals
Placebo Spironolactone
Body weight (g) 689 61 638 93
Mean arterial pressure (mm Hg) 113 16 111 11
Pulse pressure (mm Hg) 48 7437
HR (beats/min) 388 37 381 35
Values are mean SD.
Table 2. Measurement of Carotid Hemodynamic Parameters
Placebo Spironolactone
Blood pressure (mm Hg)
Mean 111 17 107 10
Pulse 33 12 31 11
Heart rate (beats/min) 317 31 325 26
Carotid diameter (
m)
Mean 1.532 0.142 1.506 0.087
Pulse 0.071 0.033 0.080 0.009
Carotid compliance
(mm
2
,mmHg
1
10
3
)
4.05 1.79 5.31 1.05
Distensibility
(mm Hg
1
10
3
)
2.17 0.79 2.96 0.51*
Elastic modulus (kPa) 0.40 0.15 0.31 0.07*
Values are expressed as mean SD.
*p 0.05.
664 Lacolley
et al.
JACC Vol. 37, No. 2, 2001
Aortic Fibrosis Prevention by Spironolactone in Old Normotensive Rats
February 2001:662–7
Page 3
and left ventricular weight were not modified by spirono-
lactone (Table 4). A strong positive correlation between
carotid and left ventricular collagen (%) was observed in the
totality of the animals (r
2
0.23; p 0.007). Cardiac
collagen was also strongly correlated with left ventricular
thickness (r
2
0.40; p 0.0001). Both carotid and cardiac
collagen were not correlated with mean blood pressure.
DISCUSSION
In the present study we showed that in old Sprague-Dawley
rats, long-term spironolactone did not change blood pres-
sure, heart weight, or carotid and aortic medial thickness
significantly. In contrast, significant changes occurred in the
arterial extracellular matrix, involving a decrease in carotid
elastic and collagen densities and contents. Although these
vascular changes were less pronounced than the observed
decrease in subendocardial collagen density, they were
associated with a significant pressure-independent increase
of carotid distensibility and decrease of carotid incremental
elastic modulus.
Histomorphometric changes and blood pressure. Al-
though the role of mechanical factors as a determinant of
arterial hypertrophy has been well documented in hyperten-
sive animals and humans (14–16), there are still discrepan-
cies on the components of the arterial wall (vascular smooth
muscle, elastin, collagen) that are the most sensitive to
pressure load. There is little doubt that the degree of
hypertrophy of the arterial wall is strongly influenced by the
level of mean blood pressure according to the Laplace law:
1) several studies in the past have indicated a strong positive
relation between arteriolar smooth muscle mass and mean
blood pressure levels in untreated rats (14), and 2) after drug
treatment of hypertension, there is a parallelism between
blood pressure reduction and the decrease in size of arterial
smooth muscle (15,16). These observations agree with the
changes in medial thickness that we observed previously in
studies of SHRs treated by AT
1
blockade and CEI (3).
In contrast, aortic collagen accumulation is poorly sensi-
tive to the changes of mean blood pressure. Studies in SHRs
in vivo (3) have shown that i) aortic collagen is reduced with
CEI but not with dihydralazine for the same decrease in
medial thickness and the same mean blood pressure reduc-
tion, and ii) aortic collagen accumulation is diminished even
with nonantihypertensive doses of the CEI. It seems likely
that in SHRs, collagen accumulation, which reflects the
presence of a stiff wall material, is rather related to the level
of pulsatile pressure than to the level of mean arterial
pressure. Studies in verapamil-treated SHRs have shown
that aortic structural changes were associated with a sub-
stantial decrease of PP but not of mean arterial pressure (17).
In the present study, we showed that the decrease in
blood pressure produced by spironolactone in old Sprague-
Dawley rats was not significant. It might be argued that, in
this investigation, intra-arterial mean blood pressure was
measured 24 h after the last drug administration, and that,
with spironolactone, mean blood pressure at the drug’s peak
effect should be lower. However, in a previous pilot exper-
iment, we showed that the mean blood pressure reduction
3 h after drug administration did not exceed 15 3mmHg
in SHRs (5). Thus, in agreement with several previous
studies (8,18,19), it is safe to conclude that only a minimal
or a lack of mean blood pressure reduction was obtained in
our normotensive animals under spironolactone. This find-
ing contrasts with the reduction in cardiac and carotid
collagen observed during the present investigation and
concords with several results previously reported for cardiac
fibrosis (68,18,19). Two specific arguments suggest that
the mechanism of reduction of cardiovascular fibrosis under
spironolactone is poorly influenced by mechanical factors.
First, a triterpene acid derived from Centilla asiatica,a
licorice root derivative that is chemically similar to aldoste-
rone, has been found to enhance collagen synthesis in
human skin fibroblasts (20). Second, another mineralocor-
ticoid hormone, deoxycorticosterone, has been shown to
increase collagen synthesis in minced rat heart tissue (21).
Histomorphometric changes and aldosterone antago-
nism. In the literature, several in vitro investigations indi-
cate that aldosterone acts directly on large arterial vessels.
First, immunohistochemical methods have shown that the
intensity of staining of mineralocoticoid receptors within
the vascular wall predominates in the aorta and decreases
markedly with the size of the arteries (22). Second, endog-
enous vascular synthesis of aldosterone occurs in the rat
mesenteric artery, even after adrenalectomy (23–26). Inter-
estingly, the vascular endothelium should be particularly
involved in this synthesis (25). Finally, a direct and rapid
Table 3. Histomorphometric Data of the Thoracic Aorta and of
the Carotid Artery
Placebo Spironolactone
Thoracic aorta
Medial thickness (
m) 139.4 12.2 132.0 9.5
Elastin density (%) 23.98 1.56 25.81 3.00*
Collagen density (%) 16.78 1.51 16.17 1.04
Elastin content (
m
2
/mm) 33,299 1,983 33,758 2,275
Collagen content (
m
2
/mm) 23,486 3,739 21,379 2,392
Carotid artery
Medial thickness (
m) 55.3 5.9 57.7 8.1
Elastin density (%) 22.25 1.88 20.21 2.02**
Collagen density (%) 18.41 2.22 16.40 1.31**
Elastin content (
m
2
/mm) 12,235 888 11,533 820*
Collagen content (
m
2
/mm) 10,239 1,938 9,502 1,874*
Values are mean SD.
*p 0.05. **p 0.001.
Table 4. Spironolactone: Cardiac Structural Data
Placebo Spironolactone
Subendocardial collagen density (%) 7.55 2.00 2.94 0.96**
Heart weight/body weight ratio 2.5 0.4 2.4 0.4
Left ventricular weight/body
weight ratio
2.0 0.3 2.0 0.3
1 SD.
**p 0.001.
665
JACC Vol. 37, No. 2, 2001
Lacolley
et al.
February 2001:662–7
Aortic Fibrosis Prevention by Spironolactone in Old Normotensive Rats
Page 4
effect of aldosterone on sodium transport has also been
described in vascular smooth muscle cells (27–30), involving
in particular the Na
/H
antiport and the Na
,K
-
ATPase pump (31).
Numerous studies have shown that myocardial fibrosis in
response to chronic mineralocorticoid excess and salt load-
ing is independent of the degree of hypokalemia, hyperten-
sion, and cardiac hypertrophy (8,18,19). Furthermore, low-
dose spironolactone administration has been shown to offset
the effects of aldosterone on cardiac fibrosis with minimal
changes in blood pressure and cardiac mass (5,8). In the
present study, spironolactone caused little change in blood
pressure and cardiac hypertrophy but a striking reduction in
cardiac collagen accumulation was observed. This reduction
was much more pronounced than that observed on the
carotid artery. One possibility is that collagen content (but
not density) is simply higher in the heart than in the arteries,
resulting in a more substantial lowering under spironolac-
tone, according to the low of initial value. Another possi-
bility is that the number and/or sensitivity of mineralocor-
ticoid receptors differ markedly in the heart and in the
vessels. An indirect argument in favor of this possibility is
that these receptors are in higher number in larger than in
smaller arteries (22), resulting in a significant increase in
distensibility under spironolactone without significant
change in mean arterial pressure and hence arteriolar resis-
tance.
In conclusion, the goal of the present study was not to
perform an age-dependent/dose-dependent set of exper-
iments with respect to arterial stiffness and spironolac-
tone treatment. It was only to assess that even in old
normotensive rats, spironolactone is able to prevent an
age-induced increase in extracellular matrix accumulation
together with an improvement of arterial elasticity and
that this finding was independent of blood pressure
changes. Because in old normotensive rats, a similar effect
has not been observed under chronic converting enzyme
inhibition, the action of spironolactone might be consid-
ered as specific. Thus, spironolactone treatment may be
proposed in situations involving a development of extra-
cellular matrix as during aging and hypertension. In the
latter situation, particularly in essential hypertension in
which a positive statistical association has been observed
between increased arterial stiffness and increased aldoste-
rone (32), the clinical relevance of this finding should be
investigated.
Acknowledgment
We thank Maryse Deboute´ for her excellent assistance.
Reprint requests and correspondence: Dr. Michel Safar, Me´-
decine Interne 1, Hoˆpital Broussais, 96 rue Didot, 75674 Paris
Cedex 14, France. E-mail: michel.safar@brs.ap-hop-paris.fr.
REFERENCES
1. Yin F. The aging vasculature and its effects on the heart. In: Weisfeldt
M, editor. The Aging Heart. New York: Raven Press, 1990:137–213.
2. Corman B, Duriez M, Poitevin P, et al. Aminoguanidine prevents
age-related arterial stiffening and cardiac hypertrophy. Proc Natl Acad
SciUSA1998;95:1301–6.
3. Albaladejo P, Bouaziz H, Duriez M, et al. Angiotensin converting
enzyme inhibition prevents the increase in aortic collagen in rats.
Hypertension 1994;23:7482.
4. Michel JB, Heudes D, Michel O, et al. Effect of chronic ANG
I-converting enzyme inhibition on aging processes. II. Large arteries.
Am J Physiol 1994;267:R124–35.
5. Benetos A, Lacolley P, Safar ME. Prevention of aortic fibrosis by
spironolactone in spontaneously hypertensive rats. Arterioscler
Thromb Vasc Biol 1997;17:1152–6.
6. Weber KT, Brilla CG. Pathological hypertrophy and cardiac intersti-
tium: fibrosis and renin-angiotensin-aldosterone system. Circulation
1991;83:184965.
7. Weber KT, Sun Y, Guarda E. Structural remodeling in hypertensive
heart disease and the role of hormones. Hypertension 1994;23(part
2):869–77.
8. Brilla CG, Zhou G, Matsubara L, Weber KT. Collagen metabolism in
cultured adult rat cardiac fibroblasts: response to angiotensin II and
aldosterone. J Mol Cell Cardiol 1994;26:809–20.
9. Lacolley P, Ghodsi N, Glazer E, et al. Influence of graded changes in
vasomotor tone on the carotid arterial mechanics in live spontaneously
hypertensive rats. Br J Pharmacol 1995;115:1235–44.
10. Tardy Y, Meister JJ, Perret F, Waeber B, Brunner HR. Assessment of
the elastic behaviour of peripheral arteries from non-invasive measure-
ment of their diameter-pressure curves. Clin Phys Physiol Meas
1991;12:39–54.
11. Kratky RG, Lo DK, Roach MR. Quantitative measurement of fixation
rate and dimension changes in the aldehyde/pressure fixed canine
carotid artery. Blood Vess 1991;28:386–95.
12. Lee RMKW. Preservation of in vivo morphology of blood vessels for
morphometric studies. Scanning Microsc 1987;1:1298–93.
13. Valmsley JG, Gore RW, Dacey RG Jr, Damon DN, Duling BR.
Quantitative morphology of arterioles from the hamster cheek pouch
related to mechanical analysis. Microvasc Res 1982;24:249–71.
14. Richer C, Mulder P, Fornes P, Domergue V, Heudes D, Giudicelli
J-F. Long-term treatment with trandolapril opposes cardiac remodel-
ing and prolongs survival after myocardial infraction in rats. J Cardio-
vasc Pharmacol 1992;20:147–56.
15. Owens GK. Influence of blood pressure on the development of aortic
medial smooth muscle hypertrophy in spontaneously hypertensive rats.
Hypertension 1987;9:17887.
16. Levy BI, Michel JB, Salzman JL, et al. Effects of chronic inhibition of
converting enzyme on mechanical and structural properties of arteries
in rat renovascular hypertension. Circ Res 1988;63:227–39.
17. Kof I, Safar ME, Labat C, Lacolley P, Benetos A, Mourad JJ.
Arterial structural changes with verapamil in spontaneously hyperten-
sive rats. Am J Hypertens 1999;12:732–8.
18. Robert V, Van Thiem N, Cheav SL, Mouas C, Swynghedauw B,
Delcayre C. Increased cardiac types I and III collagen mRNAs in
aldosterone-salt hypertension. Hypertension 1994;24:306.
19. Young M, Fullerton M, Dilley R, Funder J. Mineralcorticoids,
hypertension and cardiac fibrosis. J Clin Invest 1994;93:257883.
20. Marquat FX, Bellon G, Gillery P, Wegrowski Y, Borel JP. Stimulation
of collagen synthesis in fibroblast cultures by a triterpene extracted
from Centella asiatica. Connect Tissue Res 1990;24:107–20.
21. Ooshima A, Fuller GC, Cardinale GJ, Spector S, Udenfried S.
Increased collagen synthesis in blood vessels of hypertensive rats and
its reversal by antihypertensive agents. Proc Natl Acad SciUSA
1974;71:3019–23.
22. Lombes M, Oblin ME, Gasc JM, Baulieu EE, Farman N, Bonvalet
JP. Immunohistochemical and biochemical evidence of a cardiovascu-
lar mineralocorticoid receptor. Circ Res 1992;71:503–10.
23. Lockett MF. Hormonal actions of the heart and the lungs on the
isolated kidney. J Physiol 1967;193:661–9.
24. Kornel L, Kanamariapudi N, Travers T, et al. Studies on high affinity
binding of mineralo- and glucocorticoids in rabbit aorta cytosol. J
Steroid Biochem 1982;16:245–64.
666 Lacolley
et al.
JACC Vol. 37, No. 2, 2001
Aortic Fibrosis Prevention by Spironolactone in Old Normotensive Rats
February 2001:662–7
Page 5
25. Takeda Y, Miyamori I, Yoneda T, et al. Production of aldosterone in
isolated rat blood vessels. Hypertension 1995;25:170–3.
26. Bunder JW, Pearc PT, Smith R, Campvell J. Vascular type I
aldosterone binding sites are physiological mineralocorticoid receptors.
Endocrinology 1989;125:2224–36.
27. Worcel M, Moura AM. Arterial effects of aldosterone and antimin-
eralocorticoid compounds on the mechanism of action. J Steroid
Biochem 1987;27:865–9.
28. Llaurado JG, Madden JA, Smith GA. Some effects of aldosterone on
sodium kinetics and distribution in porcine arterial wall. Am J Physiol
1983;244:R553–7.
29. Frideman SM. Evidence for an enhanced transmembrane sodium
(Na
) gradient induced by aldosterone in the incubated rat tail artery.
Hypertension 1983;4:230–7.
30. Garwitz ET, Jones AW. Altered arterial ion transport and its reversal
in aldosterone hypertensive rat. Am J Physiol 1982;243:H927–33.
31. Ikeda U, Hyman R, Smith TW, Medford RM. Aldosterone-mediated
regulation of Na
,K
-ATPase gene expression in adults and neonatal
rat cardiocytes. J Biol Chem 1991;266:1205866.
32. Blacher J, Amah G, Girerd X, et al. Association between increased
plasma levels of aldosterone and decreased systemic arterial compliance
in subjects with essential hypertension. Am J Hypertens 1997;10:
1326–34.
667
JACC Vol. 37, No. 2, 2001
Lacolley
et al.
February 2001:662–7
Aortic Fibrosis Prevention by Spironolactone in Old Normotensive Rats
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  • Source
    • "A study by Park et al. demonstrated that serum aldosterone is significantly associated with central aortic PWV in 438 hypertensive patients, suggesting a possible role for aldosterone in developing central aortic stiffness and increased PWV in hypertensive patients [77] . Studies on animal models showed a positive action of MRA on arterial and myocardial wall stiffness by reducing fibrosis [58,75,76,78] . Similar results have been showed in human subjects: PWV decreased significantly after six months spironolactone treatment in hypertensive elderly subjects [79]. "
    [Show abstract] [Hide abstract] ABSTRACT: Aldosterone is involved in various deleterious effects on the cardiovascular system, including sodium and fluid retention, myocardial fibrosis, vascular stiffening, endothelial dysfunction, catecholamine release and stimulation of cardiac arrhythmias. Therefore, aldosterone receptor blockade may have several potential benefits in patients with cardiovascular disease. Mineralocorticoid receptor antagonists (MRAs) have been shown to prevent many of the maladaptive effects of aldosterone, in particular among patients with heart failure (HF). Randomized controlled trials have demonstrated efficacy of MRA in heart failure with reduced ejection fraction, both in patients with NYHA functional classes III and IV and in asymptomatic and mildly symptomatic patients (NYHA classes I and II). Recent data in patients with heart failure with preserved ejection fraction are encouraging. MRA could also have anti-arrhythmic effects on atrial and ventricular arrhythmias and may be helpful in patient ischemic heart disease through prevention of myocardial fibrosis and vascular damage. This article aims to discuss the pathophysiological effects of aldosterone in patients with cardiovascular disease and to review the current data that support the use of MRA in heart failure.
    Full-text · Article · Mar 2014 · IJC Heart and Vessels
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    • "It has been suggested that, irrespective of BP control, blocking vascular mineralocorticoid receptors may prevent or attenuate negative effects of aldosterone on the structure and function of the vascular wall (Lacolley et al. 2001; Mahmud and Feely 2005; Benetos et al. 1997). Thus, spironolactone treatment has been purported to improve vascular function via BPindependent , antifibrotic mechanisms (Lacolley et al. 2001; Benetos et al. 1997). "
    [Show abstract] [Hide abstract] ABSTRACT: Spironolactone is thought to improve aortic stiffness via blood pressure (BP) independent (antifibrotic) effects, but the exact mechanism is unknown. We used metabolomics and hemodynamic measures to reveal the underlying actions of spironolactone in people with a hypertensive response to exercise (HRE). Baseline and follow-up serum samples from 115 participants randomized to 3 months spironolactone (25 mg/day) or placebo were analysed using liquid chromatography/mass spectrometry and nuclear magnetic resonance spectroscopy. Hemodynamic measures recorded at baseline and follow-up included aortic pulse wave velocity (stiffness) and 24 h ambulatory BP. Aortic stiffness was significantly reduced by spironolactone compared with placebo (−0.18 ± 0.17 vs 0.30 ± 0.16 m/s; p < 0.05), but this was no longer significant after adjustment for the change in daytime systolic BP (p = 0.132). Further, the change in aortic stiffness was correlated with the change in daytime and 24 h systolic BP (p < 0.05). Metabolomics detected 42 features that were candidate downstream metabolites of spironolactone (no endogenous metabolites), although none were correlated with changes in aortic stiffness (p > 0.05 for all). However, the spironolactone metabolite canrenoate was associated with the change in daytime systolic BP (r = −0.355, p = 0.017) and 24 h pulse pressure (r = −0.332, p = 0.026). This remained highly significant on multiple regression and was independent of age, body mass index and sex. Canrenoate appears to be an active metabolite with BP-dependent effects on the attenuation of aortic stiffness in people with HRE. This finding, together with the lack of change in endogenous metabolites, suggests that the antifibrotic effects of spironolactone could be BP-dependent.
    Full-text · Article · Feb 2014 · Metabolomics
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    • "It has been suggested that, irrespective of BP control, blocking vascular mineralocorticoid receptors may prevent or attenuate negative effects of aldosterone on the structure and function of the vascular wall (Lacolley et al. 2001; Mahmud and Feely 2005; Benetos et al. 1997). Thus, spironolactone treatment has been purported to improve vascular function via BPindependent , antifibrotic mechanisms (Lacolley et al. 2001; Benetos et al. 1997). We used metabolomics to study metabolites in an unbiased, untargeted manner and investigate whether spironolactone causes metabolic perturbations that might explain arterial wall effects of spironolactone in individuals with HRE. "
    [Show abstract] [Hide abstract] ABSTRACT: Background: Spironolactone is an aldosterone antagonist that is thought to have antifibrotic effects (e.g. reduces aortic stiffness), but little is known about the underlying mechanisms. This could occur through direct effects on the endogenous aldosterone pathway or via active metabolites derived from spironolactone. In this study we aimed to determine the physiological actions of downstream metabolites of spironolactone. Methods: Baseline and follow-up serum samples from 115 participants (54 +/- 9years) randomized to three months spironolactone (25 mg/d) or placebo were analysed by liquid chromatography/mass spectrometry (LC/MS) using a high resolution LTQ-Orbitrap. Physiological variables recorded at baseline and follow-up included aortic stiffness (pulse wave velocity; PWV) and 24 hour ambulatory BP. Actions of spironolactone metabolites were assessed by correlation with the change in physiological variables after three months spironolactone therapy. Results: Spironolactone significantly reduced aortic PWV and 24 hour systolic BP (P < 0.05 for both). LC/MS detected 43 individual features that were candidate downstream metabolites of spironolactone, although none were correlated with changes in aortic PWV (P > 0.05 for all). However, the spironolactone metabolite canrenoate was associated with the change in daytime systolic BP (r = -0.415, P = 0.007). This remained highly significant on multiple regression ([beta]=-0.408, P = 0.009) and was independent of age, body mass index and sex (adjusted R2 = 0.268, P < 0.021). Conclusions: Downstream drug metabolites of spironolactone do not appear to have significant antifibrotic effects, as measured by attenuation of aortic stiffness. However, canrenoate appears to be an active metabolite of spironolactone with regard to ambulatory BP lowering effects.
    Full-text · Article · Sep 2012 · Journal of Hypertension
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