From INSERM U127, Hôpital Lariboisière, 75475 Paris Cedex 10, France
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ABSTRACT |
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Increasing evidence suggests that mineralo- and glucocorticoids modulate cardiovascular homeostasis via the effects of circulating components generated within the adrenals but also through local synthesis. The aim of this study was to assess the existence of such a steroidogenic system in heart.
Using the quantitative reverse transcriptase-polymerase chain reaction,
the terminal enzymes of corticosterone and aldosterone synthesis
(11-hydroxylase and aldosterone synthase, respectively) were
detected in the rat heart. This pathway was shown to be physiologically active, since production of aldosterone, corticosterone, and their precursor, deoxycorticosterone, was detected in both the homogenate and
perfusate of isolated rat hearts using radioimmunoassay after Celite
column chromatography. Perfusion of angiotensin II or
adrenocorticotropin for 3 h increased aldosterone and
corticosterone production and decreased deoxycorticosterone, suggesting
that aldosterone and corticosterone are formed within the isolated
heart from a locally present substrate.
Chronic regulation of this intracardiac system was then examined. As in
adrenals cardiac 11-hydroxylase and aldosterone-synthase mRNAs
were independently regulated by 1 week's treatment with either low
sodium and high potassium diet (which increased aldosterone synthase
mRNA level only), angiotensin II (which raised level of both
mRNAs), or adrenocorticotropin (which stimulated the
11
-hydroxylase gene exclusively). Changes in cardiac steroid levels
during treatment were not directly related to their plasma levels
suggesting independent regulating mechanisms. This study, therefore,
provides the first evidence for the existence of an endocrine cardiac
steroidogenic system in rat heart and emphasizes its potential
physiological and pathological relevance.
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INTRODUCTION |
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Glucocorticoids (corticosterone in the rat and cortisol in humans)
and mineralocorticoids (mainly aldosterone in both species) are
synthesized from cholesterol, predominantly in the adrenal cortex. The
two forms of the cytochrome P-450 enzyme which catalyze the final step
of these synthetic pathways are encoded by two closely related genes
CYP11B1 and CYP11B2, respectively (1) but display
differences in their enzymatic activity, regulation, and tissular
distribution (2). P-450 11-hydroxylase
(11
-OHase)1 synthesizes
corticosterone from 11-deoxycorticosterone (DOC) in the zona
fasciculata reticularis and is mainly regulated by adrenocorticotropic
hormone (ACTH). P-450 aldosterone (Aldo)-synthase, which catalyzes
synthesis of aldosterone from DOC, is present only in the zona
glomerulosa. Its activity is principally controlled by angiotensin II
(Ang II) and potassium and more weakly by ACTH and sodium (3, 4). While
ACTH is a chronic inhibitor of aldosterone secretion, it is also a
potent stimulator of its synthesis in some acute conditions (5, 6). Two
other P-450c11 genes, CYP11B3 and CYP11B4 were
recently cloned from a rat genomic library (7). CYP11B3 was
97% identical to CYP11B1 and encoded an enzyme with
activities intermediate between those of 11
-OHase and Aldo-synthase (8), whereas CYP11B4 appeared to be a pseudogene (7).
Besides this classical adrenal biosynthetic pathway, extra-adrenal
sites of steroid hormone production have been identified (9), for
example in brain (10) and more recently in vessels. Indeed, aldosterone
and corticosterone production and 11-OHase and Aldo-synthase gene
expression have been demonstrated in mesenteric rat artery and in
endothelial and smooth muscle cells isolated from human pulmonary
artery (11-13). Moreover, Hatakeyama et al. (13) showed
that this vascular aldosterone potentiates Ang II-induced hypertrophy
of cultured vascular smooth muscle cells, suggesting a physiological
role for this locally generated steroid. To date, there is no
information regarding aldosterone or corticosterone synthesis within
the heart. However, Knox and Lockett (14) have previously demonstrated
that isolated hearts produce a substance whose physicochemical
properties are consistent with those of aldosterone. The detection of
3
-hydroxysteroid dehydrogenase activity (which produces progesterone
from pregnenolone) in rat heart also indicates the potential for
steroid metabolism in cardiac tissue (15).
Several lines of evidence indicate that glucocorticoids and aldosterone
may influence cardiac function. (i) Glucocorticoid and
mineralocorticoid receptors have been identified in human and rodent
heart (16-18); (ii) 11-hydroxysteroid dehydrogenase, which converts
glucocorticoids to their inactive 11-keto metabolites and confers
mineralocorticoid specificity to aldosterone target tissues (19), has
also been detected in cardiac cells (20); (iii) aldosterone triggers
cardiac fibrosis (21, 22) and electrolyte imbalance (23, 24); and (iv)
glucocorticoids regulate the cardiac expression of a subset of
steroid-responsive genes (25, 26). We hypothesize that
mineralocorticoids and glucocorticoids modulate cardiac homeostasis not
only via the effects of circulating components generated within the
adrenal glands, but also through local synthesis. The aim of this study
was, therefore, to assess the existence of such a local system in the
rat heart.
To test this hypothesis, we investigated 11-OHase and Aldo-synthase
gene expression in rat heart by a quantitative polymerase chain
reaction after reverse transcription. We then measured basal aldosterone and corticosterone production in the isolated rat heart
using Celite column chromatography coupled with radioimmunoassay. Finally, we examined the regulation of this cardiac endocrine system
using 1 week of treatment with a low sodium/high potassium diet, Ang
II, or ACTH. This study provides direct evidence that local pathways of
aldosterone and corticosterone synthesis exist in rat heart.
Furthermore, this cardiac steroid production is regulated by the
classical stimuli of adrenal steroid biosynthesis.
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EXPERIMENTAL PROCEDURES |
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Animals
The study, which was conducted in accordance with both institutional guidelines and those formulated by the European community for the use of experimental animals (L358-86/609/EEC), was performed using 2-month-old male Wistar rats (Iffa Credo, Lyon, France). For chronic experiments (protocol 2 below), animals were randomly divided into four groups, each group receiving one of the following treatments: (i) a low sodium and high potassium group (0.01% Na+ and 2% K+ in chow); (ii) Ang II (100 ng/kg/min) infused via a subcutaneous osmotic minipump (Alzet 2002, Charles River, Paris, France); (iii) ACTH (5 ng/kg/min) infused by osmotic minipump; and (iv) a sham-operated group, with an osmotic minipump containing only physiological serum. Systolic blood pressure was measured by the tail-cuff method. After 1 week of treatment, rats were anesthetized by intraperitoneal injection of sodium pentobarbital (60 mg/kg), and blood was collected for measurements of aldosterone, corticosterone, and deoxycorticosterone concentration and renin activity.
Cardiac Perfusion
Hearts were excised and immediately dropped into ice-cold Krebs-Henseleit buffer. After cessation of beating, hearts were rapidly mounted on an aortic cannula, and retrograde perfusion was initiated as described previously (27). Briefly, hearts were perfused with modified Krebs-Henseleit solution (pH 7.4, 37 °C) containing (in mM) NaCl 120, KCl 5, NaHCO3 25, MgSO4 1.2, KH2PO4 1.2, CaCl2 1.2, glucose 11, sodium pyruvate 5, and bovine serum albumin 0.15% (fraction V, Pentex, Miles), oxygenated with a 95% O2, 5% CO2 gas mixture. Contractile parameters, oxygen consumption, and coronary flow were continuously recorded until cardiac perfusion was completed.
Experimental Protocol 1--
In this set of experiments, hearts
from control rats were perfused for 3 h under different
conditions. To wash out plasma components from the heart and allow
equilibration, hearts were perfused during the first 15 min at a
pressure of 60 mm Hg without recycling of the buffer. They were then
perfused for 3 h with 100 ml of recirculating buffer containing
either Ang II (108 M) or ACTH
(10
8 M) (Sigma, St. Quentin Fallavier,
France). Thus, time 0 of perfusion indicates the onset of Ang II or
ACTH perfusion. 10-ml samples of cardiac perfusate were collected at
times 0, 1, 2, and 3 h, and steroid concentrations were corrected
for volume variation due to sample removal.
Experimental Protocol 2-- Hearts excised from rats treated for 1 week with a low Na+/high K+ diet, Ang II, or ACTH were perfused without recycling the buffer. After the initial wash out and equilibration (15 min), 10 ml of perfusate were collected, perfusion was stopped, and hearts were homogenized as described above.
Hormone Assay
Steroids were extracted from the collected perfusates using chloroform. Extracted perfusate and cardiac homogenate were dried under vacuum and redissolved in a 0.25 M phosphate buffer (pH 7) containing sodium azide (2 g/liter) with 3000 cpm of each tritiated steroid to calculate recovery. Steroids were separated in a column composed of Celite and silica gel (Celite 545, Silica Gel 60, Fluka, Paris, France) using a polarity gradient of isoacetane and ethyl acetate. Recovery ranged from 40 to 60%, and results were corrected for this. Levels of aldosterone, corticosterone, and deoxycorticosterone were determined in duplicate by radioimmunoassay using rabbit polyclonal antibodies. The cross-reactivity of each specified antibodies with different biological steroids has been previously described (29). Radioimmunoassay was performed in a 0.25 M phosphate buffer (pH 7), containing sodium azide (2 g/liter) and gelatin (1 g/liter). The accuracy of each series of assays was determined by the addition of various known amounts of unlabeled steroid. To verify that the hormones released into perfusate were not degraded in Krebs buffer, exogenous steroids were added during perfusion.
Incubation of Cardiac Homogenate with 11-[3H]Deoxycorticosterone
To confirm that immunoassays measured bona fide steroids and did
not cross-react with other steroidal compounds, cardiac homogenate was
incubated with [3H]DOC. Briefly, hearts from control rats
were perfused with or without Ang II (108 M)
as described in experimental protocol 1. After 3 h, perfusion was
stopped, and the heart was minced with scissors and washed twice in
physiological serum. The heart was then homogenized using a chilled
glass homogenizer in ice-cold buffer (pH 7.4) containing 0.25 M sucrose, 15 mM malate, 5 mM
MgCl2, 10 mM EGTA, 10 mM Tris-HCl, 10 mM KH2PO4, 0.075% bovine serum
albumin (fraction V, Pentex, Miles), and protease inhibitors (1 mM phenylmethysulfonyl fluoride, 1 µM
pepstatin, 1 µM leupeptin, and 0.1 µM
aprotinin). Insoluble matter was removed by centrifugation. The
supernatant was then incubated aerobically for 1 h at 37 °C in
2 ml of buffer (composition as above) containing 90 pmol of
deoxy-[1
,2
-3H]corticosterone (specific activity, 47 Ci/mmol; Amersham Corp., Les Ulis, France), as described previously
(30). Steroids were then extracted by chloroform and separated as
described under "Hormone Assay." Unlabeled steroids were added with
chloroform to assess steroid loss. The radioactivity of the
corresponding fraction was counted using a scintillation counter.
Aliquots of each fraction were used for radioimmunoassay to assay
unlabeled steroids and to calculate recovery. The protein concentration was determined according to the method of Bradford (28), with bovine
serum albumin used as a standard.
Total RNA Extraction
Each heart was separated into its four constituent chambers. Total RNA was extracted from these cardiac tissues and the adrenal glands according to Trizol reagent protocol (Life Technologies, Inc., Cergy Pontoise, France). The yields of total RNA extracted were similar in all four cardiac chambers, in control and treated hearts, and in nonperfused and perfused hearts. The quality of RNA was confirmed by ethidium bromide staining in 1% agarose gel.
Quantitative Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR)
Primers--
Oligonucleotide primers (Bioprobe Systems,
Montreuil, France) were chosen in homologous parts of the coding region
of 11-OHase and Aldo-synthase genes (Fig.
1, panel A). The sense primer
was 5'-ACTCCGTGGCCTGAGACG-3' (position 363-381 bp, exon II) and the antisense primer 5'-CTGTGTGGTGGACTTGAA-3' (position 709-691 bp, exon
IV) according to the sequence published by Nomura et al. (1). We obtained a PCR product of 346 bp for each transcript after
RT-PCR amplification (Fig. 2A,
left panel).
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Internal Standard Preparation--
To quantify transcripts of
11-OHase and Aldo-synthase enzymes by RT-PCR, target mRNA was
coamplified with a defined concentration of specific internal standard
(cRNA). The PCR product was subcloned into a pSP(64) poly(A) vector
(Promega, Charbonnières, France). Sequence analysis of the
subcloned cDNA (Euro Séquences Gènes Services, Montigny
le Bretonneux, France) confirmed 100% identity with the published
sequence of 11
-OHase (1). Since the cardiac 11
-OHase mRNA
level was 7-fold higher than that of Aldo-synthase, we subcloned
11
-OHase cDNA. The 11
-OHase PCR product was then linearized
with NcoI and ligated with a 100-bp insert
(PvuI/ScaI fragment of pBluescript II SK
phagemid). The internal standard was obtained using SP6 RNA polymerase
after template linearization with PstI, and the
transcription reaction was performed using labeled UTP as a precursor.
Transcript concentration was determined after measurement of the
radioactivity incorporated into the RNA product. After RT-PCR, we
obtained a PCR product of 446 bp (Fig. 2A, left
panel).
Quantitative RT-PCR Protocol--
Total RNA was incubated with a
fixed amount of internal standard and 200 units of Moloney murine
leukemia virus RT (Life Technologies, Cergy pontoise, France) in a
20-µl reaction volume containing 20 mM Tris-HCl (pH 8.4),
50 mM KCl, 2.5 mM MgCl2, 10 mM dithiothreitol, 0.1 µg/ml bovine serum albumin, 1 mM dNTP, 0.2 µM oligo-p(dT)15 primer (Boehringer Mannheim, Meylan, France), and 50 units of RNase
inhibitor (Promega, Charbonnières, France). This reaction mixture
was incubated for 10 min at 25 °C, then for 75 min at 40 °C, and
the reaction was stopped by heating for 3 min at 94 °C. The
resultant single strand cDNA was amplified using 0.5 µM of each sense and antisense primer and 2.5 units of
Taq DNA polymerase (Boehringer Mannheim, Meylan, France) in
50 µl of 10 mM Tris-HCl (pH 8.4), 50 mM KCl,
3.75 mM MgCl2, 1 mM dNTP, and
0.01% gelatin. 32 and 30 amplification cycles were undertaken for
heart and adrenals, respectively, as follows: denaturation at 94 °C
for 1 min, annealing of primers at 61 °C for 1 min, and primer
extension at 72 °C for 1 min. A trace of [-P32]dCTP
was included in the PCR reaction for quantification of the different
products. The number of C residues present in each fragment (11
-OHase = 69, Aldo-synthase = 74, and internal
standard = 95) was taken into account for this quantification.
CYP11B3 mRNA amplification was carried out using the
same protocole but annealing of the CYP11B3 primer was
performed at 59 °C for 32 cycles.
Differentiation by Enzymatic Digestion of 11-OHase and
Aldo-Synthase PCR Products--
After RT-PCR amplification, we
obtained a PCR product of 346 bp for each transcript as expected (Fig.
2A, left panel). The two PCR products were then
size-differentiated using XhoI, which hydrolyzed the
Aldo-synthase PCR product into two fragments of 177 and 169 bp.
XhoI did not affect the PCR products of 11
-OHase or the
internal standard (Figs. 1, panel A, and 2A,
right panel). Digestion efficiency was verified in
preliminary experiments using other restriction enzymes
(TaqI, MaeII) and the hydrolyzed 11
-OHase PCR
product (PstI, PvuII).
Validity of the RT-PCR Method--
(Data not shown.) Cardiac
mRNA concentrations were determined after 32 cycles,
i.e. within the exponential amplification phase of both
cardiac mRNA and standard RNA sequences. Indeed, a linear increase
in the number of log amplified target molecules was observed between 28 and 33 PCR cycles. The amplification efficiency was the same (93%) for
11-OHase, Aldo-synthase, and their specific internal standard as
demonstrated by their parallel amplification curves. The absence of
competition between endogenous and synthetic RNA was indicated by the
linear relationship between the amount of radioactivity incorporated
into the PCR products and the initial amount of RNA. In contrast, the
radioactivity incorporated into the internal standard of each tube
remained unchanged. These PCR tests were also performed for adrenal
tissue RNA.
Statistical Analysis
Statistical significance was estimated between two groups using one-way analysis of variance and group-to-group comparison using Student's t test. Tests were considered significant when p < 0.05. All values presented are the mean ± S.E.
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RESULTS |
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11-OHase and Aldo-Synthase mRNA Levels and Steroid
Production in Normal Hearts
Cardiac Gene Expression--
11-OHase and Aldo-synthase
mRNA levels were higher in atria than in ventricles (1.5- and
1.3-fold for the 11
-OHase mRNA level and 1.3- and 1.4-fold for
the Aldo-synthase mRNA level in right and left atria
versus right and left ventricles, respectively). 11
-OHase
mRNA levels were 7-fold higher than those of Aldo-synthase in each
cardiac cavity (Fig. 2, A and B).
CYP11B3 gene expression was undetectable in the heart of
2-month-old rats (Fig. 3). In contrast, a
low level of CYP11B3 mRNA was observed in the heart of
21-day-old rats. We also observed a similar developmental regulation in
the adrenal glands, consistent with previous reports (8).
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Cardiac Steroid Production-- Hearts of control rats were perfused for 15 min with Krebs buffer to wash out plasma steroids before measurement. They were then perfused for 3 h with Krebs buffer, in the presence or absence of Ang II or ACTH, according to protocol 1 as described under "Experimental Procedures."
Homogenate-- Aldosterone, corticosterone, and DOC were detected in the homogenate of isolated rat heart, until completion of perfusion, under baseline conditions (Fig. 4). Basal cardiac levels of these steroids were modified by Ang II and ACTH. Indeed, Ang II and ACTH enhanced aldosterone production by 3.5- and 3.4-fold, respectively and corticosterone production by 2- and 3-fold, respectively (Fig. 4, A and B). In contrast, DOC levels (Fig. 4C) fell 1.7- and 1.6-fold in response to 3 h of Ang II and ACTH perfusion, respectively.
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Perfusate-- Aldosterone, corticosterone, and DOC were all detected in the perfusate of isolated rat heart, until completion of perfusion, under baseline conditions (Fig. 5). Again, Ang II infusion produced a rise in levels of both aldosterone (8-fold) and corticosterone (1.4-fold). ACTH induced a rapid rise in aldosterone levels (4.9-fold, after 1 h of perfusion) (Fig. 5A) and a slower rise in corticosterone levels (1.5-fold, after 2 h of perfusion) (Fig. 5B). In contrast, DOC levels fell rapidly, 0.7-fold at 1 h, and were undetectable after 2 h in response to either Ang II or ACTH perfusion (Fig. 5C).
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Dose-response Curves--
The effects of increasing concentrations
of Ang II and ACTH on cardiac steroid production are showed in Fig.
6. Levels of both aldosterone and
corticosterone in cardiac homogenate rose from 1.8- to 4.6-fold and
1.6- to 3.9-fold, respectively, in response to increasing Ang II
concentrations from 109 to 10
7
M during 3 h of perfusion. In contrast, DOC levels
fell from 1.4-fold (10
9 M) to 5-fold
(10
7 M) in a dose-dependent
fashion. ACTH also raised aldosterone and corticosterone levels at a
concentration of 10
9 M (2.3- and 1.6-fold,
respectively) and 10
8 M (3.7- and 3-fold,
respectively). In contrast, ACTH led to a dose-dependent
1.4-fold (10
10 M) to 2.7-fold
(10
8 M) decrease in DOC levels. ACTH effects
on cardiac steroid production seemed maximal at 10
8
M. An ACTH and Ang II dose-related effect was also obtained
in cardiac perfusate (data not shown).
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Conversion of [3H]DOC to [3H]Corticosterone and [3H]Aldosterone in Cardiac Homogenate-- In control heart, 18 and 5% of [3H]DOC was converted to [3H]corticosterone and [3H]aldosterone, respectively (58.93 ± 4.54 and 15.36 ± 1.14 pmol/mg of protein/h for corticosterone and aldosterone, respectively). Ang II increased the conversion of [3H]DOC to [3H]corticosterone and [3H]aldosterone by 1.9-fold (111.88 ± 27.82 and 29.55 ± 3.10 pmol/mg of protein/h for corticosterone and aldosterone, respectively; both p < 0.01 versus control values).
Effects of Chronic Treatment with Low Na+/High K+ Diet, Ang II, or ACTH
Rats were treated for 1 week with a low Na+/high K+ diet, Ang II, or ACTH. At the time of sacrifice, hearts were excised and perfused according to protocol 2.
Anatomical and Physiological Data-- Systolic blood pressure increased after 1 week of Ang II (58%) and ACTH (17%) treatment (Table I). Ang II-treated rats developed moderate left ventricular hypertrophy (23% increase in left ventricular weight/right ventricular weight ratio).
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Plasma Hormones in Control and Treated Rats-- Plasma aldosterone levels (Table II) were increased by a low Na+/high K+ diet (3.5-fold) and treatment with Ang II (4.2-fold), but reduced by ACTH (0.8-fold). Plasma corticosterone concentrations were increased by all three treatments (1.9-, 3.6-, and 2.3-fold for low Na+/high K+, Ang II, and ACTH, respectively). Treatments with Ang II and ACTH raised plasma levels of DOC (4.2- and 2.3-fold, respectively). Plasma renin activity was affected only by a low Na+/high K+ diet, which caused a significant increase (37%).
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11-OHase and Aldo-Synthase mRNA Levels in Treated
Hearts--
Figs. 7 and
8 (A and B)
illustrate the changes in left ventricle 11
-OHase and Aldo-synthase
mRNA levels in response to a low Na+/high
K+ diet, Ang II, or ACTH. A low Na+/high
K+ diet increased Aldo-synthase mRNA levels by 2-fold,
whereas 11
-OHase mRNA levels remained unchanged. Ang II
increased the concentrations of 11
-OHase and Aldo-synthase mRNA
4- and 3.4-fold, respectively. ACTH raised the 11
-OHase mRNA
level 3.2-fold but had no effect on Aldo-synthase mRNA. Similar
regulations were found in the right ventricle (Fig. 8, C and
D).
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Cardiac Steroid Production in Treated Hearts-- Hearts from rats treated for 1 week with a low Na+/high K+ diet, Ang II, or ACTH were perfused for 15 min to wash out plasma steroids. Concentrations of aldosterone, corticosterone, and DOC were then measured in the homogenate and perfusate (Table III). A low Na+/high K+ diet raised levels of aldosterone by 3.8- and 11.3-fold and corticosterone by 3.7- and 1.7-fold in both homogenate and perfusate, respectively. Ang II treatment increased concentrations of aldosterone (4.4- and 16.7-fold), corticosterone (2.9- and 2.5-fold), and DOC (3.6- and 4.9-fold) in both homogenate and perfusate, respectively. Interestingly, ACTH enhanced aldosterone production 4.5-fold in the homogenate and 15-fold in the perfusate, but decreased plasma aldosterone levels (0.8-fold) (see Table II).
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11-OHase and Aldo-Synthase mRNA Levels in Control and
Treated Adrenal Glands--
As in the heart, basal adrenal
concentration of 11
-OHase mRNA was 7.4-fold higher than that of
Aldo-synthase mRNA (Fig. 8, E and F). Similar
regulatory changes were observed in the adrenals as in left and right
ventricles. A low Na+/high K+ diet increased
Aldo-synthase mRNA levels only (4.2-fold). Treatment with Ang II
raised the concentrations of both 11
-OHase and Aldo-synthase mRNA 2- and 8.5-fold, respectively, whereas treatment with ACTH stimulated 11
-OHase gene expression exclusively (3.9-fold), as described previously (4, 31, 32).
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DISCUSSION |
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The main results of this study are: (i) 11-OHase and
Aldo-synthase, but not CYP11B3, genes are expressed in the
heart of the 2-month-old rat; (ii) cardiac tissue produces the steroid hormones, aldosterone, corticosterone, and DOC; (iii) this myocardial system is regulated by the same stimuli as the adrenals (sodium, potassium, Ang II, and ACTH); and (iiii) cardiac and plasma steroid concentrations seem independently regulated.
Evidence for a Cardiac System of Steroid Synthesis--
In this
study, we demonstrate cardiac expression of genes encoding key enzymes
involved in the biosynthesis of adrenal gluco- and mineralocorticoids.
Cardiac concentration of 11-OHase and Aldo-synthase mRNAs was
approximately 1000-fold lower in the heart than in the adrenals.
However, the total amount of both mRNA molecules in the whole heart
was only 100-fold lower than in the adrenal glands. Such a ratio is
comparable with that of angiotensin-converting enzyme mRNA, whose
total quantity is about 150-fold lower in heart than in the lungs, one
of the main sources for this enzyme (33). We could not detect
CYP11B3 mRNA (the third P-450c11 gene) in the hearts of
2-month-old rats. However, CYP11B3 was expressed in cardiac
tissue 21 days after birth. Thus, cardiac CYP11B3 gene expression is developmentally regulated, as previously described in the
adrenal glands (8).
Regulation of Cardiac Steroid Synthesis-- To assess the putative physiological role of this cardiac steroidogenic system, we investigated its regulation in response to chronic changes in sodium and potassium diet, Ang II, or ACTH, all of which control steroid biosynthesis in the adrenal glands (2).
Seven days of low Na+/high K+ diet resulted in increased concentrations of aldosterone and corticosterone in the heart associated with a rise in the level of Aldo-synthase mRNA but not 11Possible Role for Cardiac Aldosterone and Corticosterone
Productions--
The detection of glucocorticoid and mineralocorticoid
receptors in the heart (16, 17), as well as 11-hydroxysteroid
dehydrogenase activity (20), strongly support the possibility of
specific actions of aldosterone and corticosterone in the rat heart.
The discovery of a local steroidogenic system that responds on both short and long term physiological stimuli suggests paracrine or autocrine actions for these cardiac-generated steroids. Furthermore, the higher aldosterone concentration in heart than in plasma supports also a putative physiological role.
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ACKNOWLEDGEMENTS |
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We thank Dr. L. Rappaport and Dr. A. Carayon for helpful discussions and Dr. B. Prendergast for kind help in preparing the manuscript. We also thank T. Dakhli for animal handling.
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FOOTNOTES |
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* This study was supported by grants from INSERM.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
Recipient of a fellowship from the Ministère de la Recherche
et de l'Enseignement Supérieur.
§ Present address: Service de Biochimie, CHU Pitié-Salpétrière, 75634, Paris Cedex 13, France.
¶ To whom correspondence should be addressed: U127-INSERM, Hôpital Lariboisière, 41 Bd de la Chapelle, 75475 Paris Cedex 10, France. Tel.: 33-0142858065; Fax: 33-0148742315; E-mail: claude.delcayre{at}inserm.lrb.ap-hop-paris.fr.
1 The abbreviations used are: OHase, hydroxylase; DOC, 11-deoxycorticosterone; ACTH, adrenocorticotropic hormone; Aldo, aldosterone; Ang, angiotensin; RT, reverse transcriptase; PCR, polymerase chain reaction; bp, base pair(s).
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REFERENCES |
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