| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
1 Merck Sharp & Dohme-Cardiovascular Research Center, 2 Institute for Surgical Research, and 3 Department of Medicine B, Rikshospitalet, University of Oslo, N-0027 Oslo, Norway
 |
ABSTRACT |
|---|
Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
|
|---|
Both myocardial
and plasma endothelin-1 (ET-1) are elevated in congestive heart failure
(CHF). However, the role played by endogenous ET-1 in the progression
of CHF remains unknown. The aim of the present study was to investigate
and correlate myocardial gene expression programs and left ventricular
(LV) remodeling during chronic ET-receptor antagonism in CHF rats.
After ligation of the left coronary artery, rats were randomized to
oral treatment with a nonselective ET-receptor antagonist (bosentan,
100 mg · kg
1 · day1,
n = 11) or vehicle (saline,
n = 13) for 15 days, starting 24 h
after induction of myocardial infarction. Bosentan substantially attenuated LV dilatation during postinfarction failure as evaluated by
echocardiography. Furthermore, bosentan decreased LV systolic and
end-diastolic pressures and increased fractional shortening. Myocardial
expression of preproET-1 mRNA and a fetal gene program characteristic
of myocardial hypertrophy were increased in the CHF rats and were not
affected by bosentan. Consistently, right ventricular-to-body weight
ratios, diameters of cardiomyocytes, and echocardiographic analysis
demonstrated a sustained hypertrophic response and a normalized
relative wall thickness after intervention with bosentan. Thus the
modest reduction of preload and afterload provided by bosentan
substantially attenuates LV dilatation, causing improved
pressure-volume relationships. However, the compensatory hypertrophic
response was not altered by ET-receptor antagonism. Therefore, ET-1
does not appear to play a crucial role in the mechanisms of myocardial
hypertrophy during the early phase of postinfarction failure.
endothelin; hypertrophy; ischemic heart failure
| |
INTRODUCTION |
|---|
|
Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
|
|---|
ENDOTHELIN-1 (ET-1) is a 21-amino acid peptide with diverse cardiovascular effects including potent vasoconstrictor properties on resistance and capacitance vessels (for review, see Refs. 14 and 25) as well as positive inotropic effects (11, 15). Furthermore, ET-1 has been shown to act as a potent growth factor in several cells in vitro, including fibroblasts (30) and cardiomyocytes (29). ET-1 has also been shown to induce several phenotypic markers of hypertrophy in isolated cardiomyocytes (29). However, the physiological and pathophysiological roles played by ET-1 in vivo are still poorly understood.
Plasma ET-1 is usually present at levels lower than those reported to exert significant biological effects and is regarded as spillover into the plasma from the tissues (5). According to the prevailing opinion, ET-1 acts as an autocrine/paracrine factor rather than as a hormone (25). However, increased levels of plasma ET-1 during congestive heart failure (CHF) have been demonstrated in both humans (24) and experimental animal models (2). Furthermore, plasma ET-1 has been shown to correlate closely to the functional classes of heart failure (4, 24) and has also been shown to be a strong and independent predictor of mortality in both acute (18) and chronic heart failure (19).
We and others have recently shown that myocardial preproendothelin-1 (ppET-1) mRNA is substantially upregulated during CHF (17a, 27) and that cardiomyocytes appear to synthesize ET-1 (1, 32). Recent evidence also suggests that such endogenous elevations of ET-1 may contribute to maintenance of cardiac function during CHF in rats (27). However, to what extent endogenous ET-1 plays a significant role in the pathophysiology of CHF still remains to be elucidated. Currently, a number of ET-receptor antagonists with different receptor subtype selectivities have become available. Such antagonists now offer possibilities to investigate both the biological and pathophysiological actions of ET-1. Indeed, in animal models and in patients with CHF, acute administration of ET-receptor antagonists have been shown to exert favorable hemodynamic responses (12, 31). Thus ET-receptor antagonists may represent a novel pharmacotherapeutic modality in the treatment of CHF. Two recent studies have demonstrated a substantial reduction of mortality after chronic administration of ET-receptor antagonists during ischemic heart failure in rats (16, 26). In a recent study, long-term administration of the nonselective ET-receptor antagonist bosentan attenuated left ventricular (LV) dilatation in postinfarction failure in rats (7). However, the mechanisms of these beneficial effects of ET-receptor antagonism remain to be determined.
Thus the aim of the present study was to investigate to what extent the beneficial effects of ET-receptor antagonism on LV remodeling could be attributed to a decrease in myocardial wall stress secondary to reduced preload and afterload, and to what extent ET-mediated autocrine/paracrine growth regulatory mechanisms are involved in the remodeling process. To address these mechanisms, a quantitative assessment of the alterations in LV dimensions and hemodynamics was correlated to myocardial gene expression of phenotypic markers of hypertrophy and to ppET-1 and angiotensinogen mRNA levels during the early phase of severe ischemic heart failure in rats after chronic treatment with bosentan, a nonpeptidic ETA-ETB-receptor antagonist.
| |
MATERIALS AND METHODS |
|---|
|
Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
|
|---|
Animal preparation. We used the left coronary artery-ligated rat model of CHF according to the method of Selye et al. (28) and the modifications of Pfeffer and colleagues (21). Male Wistar rats (300-350 g) were subjected to ligation of the left coronary artery, resulting in infarction of the LV free wall. Briefly, the rats were anesthetized with halothane and ventilated with the use of a rodent ventilator with a mixture of 30% O2-70% N2O and 1% halothane. A left thoracotomy was performed, the heart was exteriorized, and the proximal portion of the left coronary artery was rapidly ligated by an intramural 6-0 silk suture. The heart was subsequently returned to its normal position, and the thoracotomy was closed. Except for ligation of the coronary artery, sham-operated rats underwent an identical procedure. Surgical mortality was ~30% in the rats with myocardial infarction. The animal experiments, procedures, and housing were approved by the Hospital Board for Animal Research in accordance with the Norwegian Council for Animal Research.
Study protocol.
After ligation of the left coronary artery, rats were randomized to
treatment with bosentan (Ro 47-0203, Hoffmann-La Roche, Basel,
Switzerland) or saline, starting 24 h after induction of myocardial
infarction. Bosentan (100 mg · kg1 · day1)
was prepared fresh every day in saline and administered by gavage once
daily for 15 days. It has previously been shown that this oral dose of
bosentan blocks the pressor actions of ET-1 for more than 24 h (3, 7,
13). The treatment protocol was started 24 h after induction of
myocardial infarction to minimize the possibility of a direct effect of
bosentan on infarct size. Infarct size was determined by assessing the
segmental length of the scar tissue relative to the circumference of
the LV immediately after excision of the heart and by weighing the scar
tissue. All rats had infarct size >40% of the LV circumference. A
group of sham-operated rats received no treatment.
Hemodynamic measurements.
Sixteen days after the induction of myocardial infarction or sham
operation, and 24 h after the last dose of bosentan or saline, the rats
were anesthetized and ventilated with the use of a rodent ventilator as
described above. A 2-F micromanometer-tipped catheter (model SPR-407,
Millar Instruments, TX) was inserted through the right carotid artery
and advanced into the LV. LV end-diastolic pressure (LVEDP), LV
systolic pressure (LVSP), and peak positive first derivative of
the LV pressure
(+dP/dtmax)
were subsequently recorded (CardioMed Flowmeter CM 4008, Medi-Stim,
Oslo, Norway). The time constant of isovolumic relaxation (
) was
calculated according to the logarithmic method (34).
Echocardiographic examination.
Immediately after the hemodynamic measurements, echocardiography was
performed using the fully digital Vingmed System FiVe (Vingmed Sound,
Horten, Norway), with a 7.5-MHz linear array transducer. The system
software was modified to allow for high-frame-rate recordings with an
area of interest <20 mm from the surface of the transducer. Images
were obtained by placing the transducer directly on the chest with the
rat in the supine position. All recordings were digitally transferred
to an on-line computer and stored on magnetic optical disks for
subsequent analysis using the EchoPac software (Vingmed Sound).
Two-dimensional long-axis and short-axis views of the LV were recorded
with a typical frame rate of 196 s1. The short-axis
dimensions were recorded at the level of the papillary muscles. After
gain settings were optimized, M-mode tracings were recorded at the same
level, and septal and posterior wall thickness as well as LV internal
dimensions were measured in both the M-mode and the two-dimensional
tracings. LV internal dimensions were recorded as the largest
anteroposterior diameter. In all cases this diameter was recorded
outside the infarcted area. The tracings were analyzed by one observer
(R. Bjønerheim), who had no knowledge of the study group. Examples
of the two-dimensional and M-mode recordings are shown in Fig.
1.
|
Tissue sampling and RNA extraction.
The rats were killed by excision of the heart. The atria were removed,
and the right and left ventricles were dissected, separated, and
weighed. The infarcted area (scar tissue) was excised from the
noninfarcted LV, carefully avoiding contamination of viable myocardial
tissue with necrotic tissue. The scar tissue was weighed to estimate
infarct size. The tissues were snap frozen in liquid N2 and stored at 
70°C.
Total RNA from the myocardial tissues was prepared by acid-phenol
extraction in the presence of chaotropic salts (TRIzol, GIBCO-BRL,
Gaithersburg, MD) and subsequent isopropanol-ethanol precipitation.
Histochemistry and morphometric analysis. Frozen myocardial tissue from nonischemic LV free wall was cut into 5-µm sections in a cryostat microtome. The sections were fixed in acetone for 10 min and subsequently stained with hematoxylin and eosin. Individual myocytes were subjected to morphometric analysis using a micrometric system (Olympus BX50, Olympus Optical, Tokyo, Japan) at a magnification of ×400. The cell diameter was measured at the site of the nucleus, and the recordings were restricted to myocytes containing nuclei in the center of the fiber in longitudinal sections. A total of 30 myocytes per animal were analyzed.
cDNA cloning and plasmid vector constructs.
A 390-bp Sac
I/Pvu II restriction fragment of the
rat ppET-1 cDNA was subcloned into the pBluescript SK+ vector
(Stratagene, La Jolla, CA) between the restriction sites
Sac I and
Sma I. With the use of DNA sequence
information published for the rat angiotensinogen DNA (GenBank
accession nos. L00093 and L00094), a 294-bp fragment of the
angiotensinogen cDNA [nucleotides (nt) 51-157, exon 4, and
nt 61-249, exon 5] was amplified by RT-PCR from mRNA isolated from the liver, subcloned into pBluescript SK+, and sequenced by the dideoxy chain-termination technique. Similarly, the rat glyceraldehyde-3-phosphate dehydrogenase (GAPDH), cardiac 
-actin, atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), myosin light chain-2 (MLC-2), and troponin I cDNA were amplified by
RT-PCR from rat myocardial tissues, and the skeletal -actin cDNA was
amplified from rat skeletal muscle. The cDNA fragments were
characterized by DNA sequence analysis using the dideoxy chain-termination method and subcloned into pBluescript SK+
[GAPDH, nt 458-994 (GenBank accession no. M17701); cardiac
-actin, nt 453-986 (GenBank accession no. X80130); ANP, nt
1-635 (GenBank accession no. X00665); BNP, nt 180-365
(GenBank accession no. M25297); MLC-2, nt 81-525 (GenBank
accession no. M11851); troponin I, nt 214-559 (GenBank accession
no. M92074); and skeletal -actin, nt 2581-3037 (GenBank
accession no. J00692)].
Synthesis of radiolabeled RNA probes. The pBluescript SK+/ppET-1 vector was linearized at the Sac I site and blunt ended with T4 DNA polymerase to allow the synthesis of an antisense ppET-1 RNA probe from the T7 promoter site (protected fragment 390 bp). For angiotensinogen riboprobe synthesis, pBluescript SK+/angiotensinogen was linearized at a unique Bst XI site (protected fragment 275 bp). GAPDH riboprobe was synthesized by linearizing pBluescript SK+/GAPDH at a unique Eco0109 I site (protected fragment 128 bp).
Continuously radiolabeled antisense RNA probes for RNase protection assay of ppET-1, angiotensinogen, and GAPDH mRNAs were generated by in vitro transcription using T3 or T7 RNA polymerase and [-32P]CTP (3,000 Ci/mmol, NEN, Boston, MA). Generally, linearized template DNA was
transcribed in the presence of 40 mmol/l Tris · HCl,
pH 7.5, 6 mmol/l MgCl2, 2 mmol/l
spermidine, 10 mmol/l NaCl, 10 mmol/l dithiothreitol, 500 µmol/l of
each of the unlabeled ribonucleotides (ATP, GTP, and UTP), 3 µmol/l
of [-32P]CTP
(specific activity ~800 Ci/mmol), 20 units of the RNase inhibitor
RNasin (Promega, Madison, WI), and 20 units of either T3 or T7 RNA
polymerase. After the transcription reaction was completed, the
template DNA was removed by digestion with RQ1 DNase I. Under the
conditions described, the RNA probes were labeled with high specific
activity [~0.5 × 109 counts/min (cpm)/µg].
Synthesis of radiolabeled cDNA probes.
Continuously labeled cDNA probes for Northern blot analysis of ANP,
BNP, skeletal -actin, cardiac -actin, MLC-2, troponin I, and
GAPDH mRNA expression were radiolabeled with the random priming method
in the presence of
[-32P]dCTP
(specific activity ~6,000 Ci/mmol).
RNase protection assay. Total RNA (20 µg per assay) from the nonischemic myocardial tissue of the LV and from the right ventricle were mixed with the antisense ppET-1, angiotensinogen, and GAPDH RNA probes, coprecipitated with ethanol, and dissolved in hybridization buffer (80% deionized formamide, 100 mmol/l sodium citrate, 300 mmol/l sodium acetate, and 1 mmol/l EDTA, pH 6.4). The hybridization reaction was performed overnight at 43°C. The samples were subsequently treated with RNase (RNase A-RNase T) at 37°C for 30 min. This reaction was terminated with a commercial RNase inactivation-precipitation mixture (RPA II kit, Ambion, Austin, TX). The precipitated RNA was dissolved in 80% formamide, 0.1% xylene cyanol, 0.1% bromphenol blue, and 2 mmol/l EDTA and subjected to electrophoresis on a 5% denaturing polyacrylamide gel. Autoradiography of the gel was performed with the use of storage phosphor plates and a scanning PhosphorImager (445 SI, Molecular Dynamics, Sunnyvale, CA). Densitometric analysis of the bands was performed with the Image-Master software package (Pharmacia Biotech, Piscataway, NJ). The data were subsequently corrected for variations in RNA loading by normalizing the ppET-1 mRNA and angiotensinogen mRNA expression to the GAPDH mRNA levels.
Northern blot analysis.
Total RNA (20 µg/lane) from the noninfarcted part of the LV and from
the right ventricle of CHF-bosentan rats
(n = 11), CHF-saline rats
(n = 12), and sham-operated rats
(n = 6) were denatured in 50%
formamide and 6.5% formaldehyde and size fractionated with formaldehyde-agarose gel electrophoresis, transferred onto nylon filter
membranes by capillary blotting, and successively hybridized with
radiolabeled cDNA probes for ANP, BNP, skeletal -actin, cardiac
-actin, MLC-2, and troponin I. The nylon membranes were hybridized
at 42°C for 18 h in a hybridization buffer containing 5× SSC
(saline-sodium citrate), 5× Denhardt's solution, 50% formamide, 50 mmol/l sodium phosphate, pH 6.5, 125 µg/ml sonicated salmon sperm
DNA, and 2-3 × 106
cpm/ml cDNA probe (specific activity 1-2 × 109 cpm/µg). After the
hybridization, the filters were washed in 2× SSC-0.1% SDS at
room temperature and, finally, washed in 0.1× SSC-0.1% SDS at
60°C. Autoradiography and densitometric analysis of the bands were
performed as described in RNase protection
assay. To normalize for variations of the amount of
total RNA loaded on the gel and for variations of transfer
efficiencies, the same membranes were rehybridized with the GAPDH cDNA
as internal control.
RT-PCR of - and 
-myosin heavy chain
cDNA.
The relative amounts of cardiac - and -myosin heavy chain (MHC)
mRNAs were determined by simultaneous amplification of the two mRNA
species by "hot" (radioactively labeled) RT-PCR using assay
conditions recently described (9). One common pair of oligonucleotide
primers was used to amplify both the - and -MHC cDNA fragments.
With the assumption that this set of primers will anneal to identical
sequences in the - and -MHC transcripts with equal efficiencies
and that the length of elongation is identical, the amplified fragments
should be representative of the endogenous levels of mRNA for these two
transcripts. Determination of the relative proportions of the amplified
DNA fragments was done by differential restriction endonuclease
digestion with an enzyme for which a restriction site is found only in
the -MHC cDNA fragment. cDNA was synthesized from 1 µg of total
RNA isolated from the noninfarcted part of the left and right
ventricles. cDNA synthesis conditions and PCR conditions were the same
as previously described (9). The PCR amplifications were performed with
the following oligonucleotides identical to sequences of both - and
-MHC: forward primer, 5'-GCA GAC CAT CAA GGA CCT (nt
5,401-5,418 for -MHC, GenBank accession no. X15938; and nt
5,371-5,388 for -MHC, GenBank accession no. X15939), and
reverse primer, 5'-TGC TGC ACC TTG CGG AAC TTG (nt
5,742-5,722 for -MHC and nt 5,712-5,692 for -MHC,
complementary strand). Distinction between the - and -MHC
iso-mRNAs was achieved by digestion of the PCR reaction mixture with 10 units of Tru 91 (10 U/ml, Promega) in
a standard reaction buffer at 65°C for 90 min. This digestion
yields fragments of 309 bp for -MHC and 259 + 50 bp for -MHC.
After the addition of 80% formamide loading buffer, the fragments were
separated by electrophoresis on an 8% denaturing polyacrylamide gel.
Autoradiography of the gel was performed on storage phosphor plates and
a scanning PhosphorImager. Densitometric analysis of the bands were
performed with the Image-Master software (Pharmacia Biotech). The
results are expressed as ratios of -MHC mRNA to -MHC mRNA.
Statistical analysis. All data are presented as means ± SE. Statistical analysis was assessed by two-tailed, unpaired Student's t-test. The data groups were tested for equality of variances using Levene's test. P values <0.05 were considered to be statistically significant.
| |
RESULTS |
|---|
|
Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
|
|---|
Effects of bosentan on infarct size and myocardial hypertrophy. As shown in Table 1, the ratios of ventricular weight to body weight were significantly increased in both CHF groups compared with the sham-operated group (P < 0.05), demonstrating compensatory ventricular hypertrophy after myocardial infarction in rats. Sixteen days after myocardial infarction, the LV weight-to-body weight ratio was increased by 13 and 24% in the CHF-saline group and the CHF-bosentan group, respectively, compared with that in the sham-operated controls (P < 0.05). The right ventricular weight-to-body weight ratio was increased by 65% in both the CHF-saline group and the CHF-bosentan group compared with that in the sham-operated group (P < 0.05). Thus no significant differences in right ventricular weight-to-body weight ratios between the two treatment groups were observed, indicating sustained hypertrophic response during intervention with the ET-receptor antagonist bosentan. Furthermore, the diameter of the cardiomyocytes in sections of LV tissue was similar in the CHF-bosentan and CHF-saline groups (13.23 ± 0.1 and 13.30 ± 0.1 µm, respectively), demonstrating maintained cellular hypertrophy in the CHF-bosentan group.
|
Effects of bosentan on ventricular pressures and dimensions during CHF. The hemodynamic characteristics of the groups are described in Table 1. The hemodynamic abnormalities were characteristic of those previously reported in CHF rats with large myocardial infarctions and substantial LV impairment. Thus LVEDP was significantly increased and LVSP was significantly decreased in the CHF rats compared with the sham-operated group (P < 0.05). Treatment with bosentan, however, caused significant reductions of both LVEDP (from 23.2 ± 0.7 to 19.8 ± 1.6 mmHg) and LVSP (from 107 ± 3 to 99 ± 2 mmHg) compared with saline in the CHF rats (P < 0.05).
LV geometry in the different groups was analyzed by echocardiography, and the characteristics are described in Table 2. The echocardiographic analyses showed evidence of a substantial increase in LV intracavitary diameters in rats with myocardial infarction compared with diameters in sham-operated rats. LV end-diastolic diameter (LVEDD) and LV end-systolic diameter (LVESD) were both markedly increased in the CHF-saline group (M-mode short-axis tracings: 8.7 ± 0.20 and 7.7 ± 0.21 mm, respectively) compared with the sham-operated group (5.7 ± 0.13 and 3.9 ± 0.14 mm, P < 0.05). Treatment with bosentan significantly attenuated LV dilatation (LVEDD, 7.6 ± 0.29 mm; LVESD, 6.3 ± 0.32 mm; P < 0.05). Thus LV end-diastolic and end-systolic dilatation were 35 and 36% less in the CHF-bosentan group than in the CHF-saline group. As shown in Table 2, consistent dimensions were recorded in both the M-mode tracings and the two-dimensional long- and short-axis views, demonstrating similar reductions of LV dilatation in the CHF-bosentan group compared with the CHF-saline group. Interventricular septum end-diastolic thickness (IVSEDT) was significantly decreased in the CHF-saline group compared with the sham-operated group (1.4 ± 0.06 vs. 2.1 ± 0.06 mm, P < 0.05). However, IVSEDT in the bosentan-treated CHF group (2.2 ± 0.08 mm) was not statistically different from that in the sham-operated control group. Less prominent reductions of LV posterior wall end-diastolic thickness (PWEDT) were observed in the CHF-saline group compared with the sham-operated controls (1.9 ± 0.06 vs. 2.1 ± 0.05 mm). PWEDT in the CHF-bosentan group (2.1 ± 0.05 mm) was not statistically different from that in the sham-operated group. With regard to the substantial attenuation of LV dilatation in the bosentan-treated CHF rats, relative wall thickness [(IVSEDT + PWEDT)/LVEDD] was almost normalized in the CHF-bosentan group compared with the CHF-saline group (Table 3).
|
|
Effects of bosentan on LV systolic function during CHF.
Peak +dP/dt was decreased in both
untreated and bosentan-treated CHF rats compared with sham-operated
rats (P < 0.05, Table 1). However,
+dP/dtmax in CHF
rats treated with bosentan was not statistically different from that in
CHF rats treated with saline. As demonstrated in Fig.
2A,
coronary artery ligation caused a rightward and downward displacement
of the systolic pressure-diameter relationship. However, bosentan
shifted the systolic pressure-diameter relationship leftward,
indicating sustained or even improved LV contractility after treatment
with bosentan. Furthermore, LV fractional shortening [(LVEDD LVESD)/LVEDD] was markedly decreased in the CHF groups.
However, treatment with bosentan improved systolic function, causing a
50% increase in fractional shortening compared with untreated CHF rats
(18 ± 1% in the CHF-bosentan group vs. 12 ± 1% in the
CHF-saline group, P < 0.05).
|
Effects of bosentan on LV diastolic function during CHF.
was significantly increased after ligation of the left coronary
artery (Table 1). However, no significant effect on was observed
after intervention with bosentan. As shown in Fig.
2B, induction of severe CHF was
associated with a substantial rightward and upward shift of the
end-diastolic pressure-diameter relationship. Chronic treatment with
bosentan during CHF attenuated the rightward and upward displacement of
the end-diastolic pressure-diameter relationship, demonstrating
improved diastolic function.
Effects of bosentan on myocardial expression of ppET-1 mRNA and angiotensinogen mRNA. RNase protection assays were performed to assess regulation of myocardial ppET-1 and angiotensinogen mRNA levels in the nonischemic regions of the left and right ventricles at the end of the treatment protocol 16 days after induction of myocardial infarction. In concordance with previous reports (17a, 27), significant elevations of myocardial ppET-1 mRNA levels were found in the left (Fig. 3) and right ventricles (Fig. 6) of the CHF-saline group compared with the sham-operated group (4-fold and 3-fold elevations, respectively, P < 0.05). However, the myocardial ppET-1 mRNA levels of the left and the right ventricles of the bosentan-treated rats were not statistically different from those of the CHF-saline group, indicating that ET-receptor blockade does not affect induction of ppET-1 mRNA. As shown in Fig. 3, angiotensinogen mRNA levels in the left ventricle were found to be expressed at very low levels in sham-operated rats. Myocardial angiotensinogen mRNA levels did not appear to be regulated in the CHF rats 16 days after induction of myocardial infarction, irrespective of treatment protocol.
|
Effects of bosentan on induction of a hypertrophic gene program
during CHF.
Myocardial ANP, BNP, -MHC, and skeletal -actin mRNAs are
expressed at very low levels in the normal adult rat ventricle. However, these genes are reactivated in the process of hypertrophy of
the myocardium associated with CHF. Thus these mRNAs are frequently used as molecular markers of the hypertrophic phenotype. Northern blot
analysis and hot RT-PCR were performed to investigate the expression of
these mRNA markers. Characteristically, myocardial ANP, BNP, and
skeletal -actin mRNA expression were substantially induced in both
the left (Fig. 4) and the right ventricles
(Fig. 6) in untreated CHF rats. In the LV, myocardial expression of ANP, BNP, and skeletal -actin mRNAs were increased 16-, 5-, and 2-fold, respectively, in the untreated CHF group compared with the
sham-operated controls (P < 0.05).
Similar elevations of the mRNA levels of ANP, BNP, and skeletal
-actin were observed in the right ventricle of untreated CHF rats
(3-fold, 7-fold, and 2-fold, respectively) compared with the
sham-operated group (P < 0.05).
However, no significant differences of myocardial ANP, BNP, and
skeletal -actin mRNA levels were observed between the CHF-bosentan
group and the CHF-saline group. The ratios of -MHC to -MHC mRNA
(-MHC/-MHC) were dramatically elevated in the CHF rats compared
with the sham-operated controls (Figs. 5
and 6), indicating induction of -MHC
mRNA. In the LV, -MHC/-MHC was increased 22-fold in the
CHF-bosentan group and 11-fold in the CHF-saline group compared with
that in sham-operated rats (P < 0.05). The difference in -MHC/-MHC between bosentan-treated CHF
rats and untreated CHF rats was not statistically significant (P = 0.082). Similar increments in
myocardial -MHC/-MHC were observed in myocardial tissues from the
right ventricles of both bosentan-treated CHF rats and untreated CHF
rats. Overall, myocardial expression of the mRNA markers of a
hypertrophic ventricular phenotype was not attenuated by bosentan
during the early phase of ventricular remodeling after myocardial
infarction in rats. Thus the hypertrophic response during CHF in rats
clearly did not appear to be hampered by ET-receptor blockade.
|
|
|
Effects of bosentan on expression of constitutive myocardial genes.
Expression of the genes encoding the contractile proteins MLC-2,
troponin I, and cardiac -actin was investigated using Northern blot
analysis. High levels of these constitutive mRNAs were observed in both
the left (Fig. 4) and the right ventricles (Fig. 6) of sham-operated
rats. MLC-2, troponin I, and cardiac -actin mRNA levels did not
appear to be regulated during CHF and were not affected by treatment
with bosentan when analyzed at the end of the intervention 16 days
after induction of myocardial infarction, indicating that these genes
are not regulated by ET.
| |
DISCUSSION |
|---|
|
Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
|
|---|
Ligation of the left coronary artery in rats is associated with induction of myocardial hypertrophy in the noninfarcted regions of the heart. This process is considered a beneficial adaptation compensating at least partially for the loss of contractile tissue. As demonstrated in the present study, both gross myocardial hypertrophy of the viable myocardium as well as induction of several phenotypic markers of hypertrophy were not affected by the treatment with bosentan. Despite improved hemodynamic responses, the ventricular-to-body weight ratios, the cardiomyocyte diameters, and the echocardiographic analysis of LV wall thickness as well as the hypertrophic gene program all consistently demonstrate that the hypertrophic response during CHF is not affected by bosentan. These findings may seem at odds with the expected outcome. However, it is important to bear in mind that the CHF rats treated with bosentan still exhibited significantly impaired hemodynamics, indicating that the compensatory mechanisms leading to hypertrophy would remain activated. Furthermore, treatment with bosentan did not affect myocardial expression of ppET-1 mRNA. The ppET-1 mRNA levels remained elevated during the treatment, indicating increased tissue ET-1. However, such elevations of tissue ET-1 cannot play a crucial role in the mechanism of hypertrophy during ischemic heart failure in rats because the high dose of bosentan employed in the present study would effectively block all the actions of ET-1 (3, 7, 13). In contrast to our data, Sakai et al. (26) recently reported amelioration of LV hypertrophy in CHF rats after ET-receptor antagonism. Although their study did not provide a quantitative assessment of the effects of ET-receptor antagonism on LV hypertrophy, several reasons may account for the apparent differences observed. First of all, the different findings may be due to different study protocols. Our study was designed to investigate the mechanisms of ET-receptor antagonism during the early phase of ventricular remodeling after myocardial infarction. Thus the effects of a 15-day treatment protocol with a mixed ETA-ETB-receptor antagonist were recorded at day 16 after induction of myocardial infarction to enable correlation of structural changes to regulation of gene expression programs. On the other hand, Sakai and colleagues (26) performed a 12-wk treatment protocol with BQ-123, a selective ETA-receptor antagonist. Thus it could be argued that the amelioration of LV hypertrophy after ETA-receptor antagonism observed by Sakai and colleagues could be due to compensatory activation of myocardial ETB receptors. However, ETB receptors have been reported (17) to mediate hypertrophic signals in cardiomyocytes in vitro. Thus the role of the ETB receptor in the process of myocardial remodeling needs to be addressed in future studies. Furthermore, administration of BQ-123 was started at day 10 after induction of myocardial infarction. Thus it is conceivable that the different findings of the two studies are due to modulation of different adaptive processes operating at various stages of myocardial remodeling. Consequently, it is important to keep in mind that the present study addresses the effects of ET-receptor antagonism at a single point in time, i.e., the first 16 days after myocardial infarction.
Apoptotic cardiomyocyte death has been shown to take place after myocardial infarction (10). Therefore, it could be hypothesized that apoptosis was more prominent in the CHF-saline rats than in CHF-bosentan rats as a result of ET-induced apoptosis through the ET receptors. Thus enhanced apoptosis could have counterbalanced and masked an increase in myocardial hypertrophy in the CHF-saline group. However, the diameters of the cardiomyocytes were similar in the CHF groups, indicating no significant alterations of hypertrophic response after intervention with bosentan. Thus the small but significant increase in LV-to-body weight ratio in bosentan-treated CHF rats could be explained by lower levels of apoptosis in the LV after ET-receptor antagonism. Although apoptosis has been shown to occur in failing myocardial tissue, the significance of this process as to the loss of myocardial tissue still needs to be established.
LV cavity size has been shown to be a major predictor of mortality in patients with CHF (35). The substantial attenuation of LV dilatation observed in the present study after intervention with bosentan may therefore explain the beneficial effects of ET-receptor antagonism on mortality in rats. Mulder and colleagues (16) recently reported a minor decrease in LV cavity size in CHF rats treated with bosentan. However, in the latter study, intervention with bosentan was initiated 7 days after ligation of the left coronary artery. In this context, the first few weeks after induction of ischemic heart failure in rats are associated with critical and substantial alterations in LV geometry (20). Dilatation of the LV may be a result of slippage and elongation of the cardiomyocytes (33). This process of slippage and elongation has been shown (20) to have already started within 2 days after myocardial infarction and rapidly progresses during the first weeks. Our findings indicate that early intervention with an ET-receptor antagonist after myocardial infarction may prove to be particularly beneficial. As shown in the present study, LV cavitary volume-to-mass ratio was reduced after treatment with bosentan. Previous evidence from other treatment protocols indicates that intervention leading to attenuation in LV volume without a proportional reduction in cardiac mass is associated with a more favorable ventricular performance and prolongation in survival (22).
Because the molecular markers of myocardial hypertrophy were similar in the CHF-bosentan and the CHF-saline groups, it seems unlikely that the beneficial effects of bosentan on LV geometry are caused by a direct action of bosentan on the myocardial tissue. However, due to the capacity of bosentan to elicit favorable hemodynamic responses during CHF, it seems most likely that the attenuation of LV dilatation is due to reduced preload and afterload and a subsequent decrease in myocardial wall stress. As indicated by the modest decrease in LVSP and the marked decrease in LVESD and LVEDD, systolic and diastolic wall tensions were markedly reduced. In the bosentan-treated CHF group, wall thickness of the viable myocardium was preserved, whereas the untreated CHF group showed considerable LV wall thinning in the nonischemic region. Therefore, not only wall tension but also wall stress was markedly reduced in the bosentan-treated CHF rats. Although we have not calculated wall stress because of asymmetry of the LV after myocardial infarction, all parameters determining wall stress were significantly reduced in the CHF rats after ET-receptor antagonism, implying decreased wall stress. The reduction in wall stress implies that bosentan decreased the load on the individual myocardial fibers and therefore reduced the stimulus to further dilatation. However, myocardial wall stress in the bosentan-treated CHF rats was presumably still significantly increased compared with that in sham-operated rats. Thus the stimulus to myocardial hypertrophy would still be present in the bosentan-treated CHF rats. The decrease in myocardial wall stress in the bosentan-treated CHF rats may exert an additional beneficial effect. A decrease in wall stress subsequently leads to reduced myocardial oxygen demand, which is a favorable effect in the setting of ischemic heart failure. Furthermore, it is conceivable that the beneficial effects of ET-receptor antagonism on coronary blood flow may have increased oxygen supply of the noninfarcted myocardium and indirectly contributed to improve cardiac function.
Previous evidence indicates that two separate properties of ET-receptor
antagonists may mediate the capacity to reduce LVEDP and preload.
First, ET-receptor antagonists have been shown to cause venodilation in
addition to arterial vasodilation (6, 8). Second, it has recently been
shown (23) that the ETA-receptor antagonist BQ-123 significantly enhances early LV relaxation and lowers
LVEDP in normal guinea pigs without altering systolic performance. However, in the present study, treatment with bosentan did not affect
, indicating that ET-receptor antagonism does not improve active
ventricular relaxation during severe CHF. Thus the mechanisms of the
decrease of preload after intervention with bosentan appears to be due
to venodilation rather than a direct effect on myocardial relaxation
during diastole.
Endothelin-1 has been shown to elicit a powerful inotropic response in isolated myocardial fibers (11). Thus ET-receptor antagonism could, at least theoretically, adversely affect LV function in vivo. Consequently, the putative beneficial effects of ET-receptor antagonism during CHF would depend on the capacity of ET-receptor blockers to reduce pulmonary and systemic vascular tone. Despite the anticipation that bosentan might impair myocardial contractility, the present study clearly demonstrates that systolic function was maintained or even improved. First, +dP/dtmax was not significantly altered after intervention with bosentan. Second, fractional shortening was vastly improved in the CHF-bosentan group. Third, the plot of LVSP vs. LVESD is consistent with maintained or improved contractility. Thus, although we were not able to measure cardiac output in the present study, the structural and dynamic data strongly indicate that systolic function was maintained or even improved with bosentan.
In conclusion, in the present study, early intervention with the nonselective ET-receptor antagonist bosentan during postinfarction failure in rats markedly attenuated LV dilatation and improved LV pressure-volume relationships. In addition, fractional shortening was increased and myocardial contractility maintained. The mechanism of these beneficial effects of bosentan appeared to be a substantial decrease in myocardial wall stress, attributed to decreases in both preload and afterload and LV diameter as well as to the preservation of myocardial wall thickness. ET-receptor antagonism by bosentan did not reduce gene expression markers of myocardial hypertrophy. Therefore, the favorable effects of bosentan during ischemic heart failure were caused by a reduction in the hemodynamic load, leading to ventricular dilatation without altering compensatory myocardial hypertrophy. Thus the data do not support an autocrine/paracrine role of ET-1 in the mechanisms of myocardial hypertrophy in the early phase of postinfarction failure.
| |
ACKNOWLEDGEMENTS |
|---|
We thank Dr. Dag Sørensen for scientific advice and helpful discussions on the CHF rat model and Drs. Leif Erik Vinge and Stig Urheim for scientific discussions and advice.
| |
FOOTNOTES |
|---|
This study was supported by grants from the Norwegian Council for Cardiovascular Diseases, the National Research Council, Novo Nordic Foundation, Johan Egebergs Medical Fund, and Medinnova SF.
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. §1734 solely to indicate this fact.
Address for reprint requests: H. Attramadal, Institute for Surgical Research, Rikshospitalet, N-0027 Oslo, Norway.
Received 4 February 1998; accepted in final form 12 May 1998.
| |
REFERENCES |
|---|
|
Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
|
|---|
1.
Arai, M.,
A. Yoguchi,
T. Iso,
T. Takahashi,
S. Imai,
K. Murata,
and
T. Suzuki.
Endothelin-1 and its binding sites are upregulated in pressure overload cardiac hypertrophy.
Am. J. Physiol.
268 (Heart Circ. Physiol. 37):
H2084-H2091,
1995
2.
Cavero, P. G.,
W. L. Miller,
D. M. Heublein,
L. L. Aarhus,
and
J. C. Burnett, Jr.
Endothelin in experimental congestive heart failure in the anesthetized dog.
Am. J. Physiol.
259 (Renal Fluid Electrolyte Physiol. 28):
F312-F317,
1990
3.
Clozel, M.,
V. Breu,
G. A. Gray,
B. Kalina,
B. M. Loffler,
K. Burri,
J. M. Cassal,
G. Hirth,
M. Muller,
W. Neidhart,
and
H. Ramuz.
Pharmacological characterization of bosentan, a new potent orally active nonpeptide endothelin receptor antagonist.
J. Pharmacol. Exp. Ther.
270:
228-235,
1994
4.
Cody, R. J.,
G. J. Haas,
P. F. Binkley,
Q. Capers,
and
R. Kelley.
Plasma endothelin correlates with the extent of pulmonary hypertension in patients with chronic congestive heart failure.
Circulation
85:
504-509,
1992
5.
Creminon, C.,
Y. Frobert,
A. Habib,
J. Maclouf,
C. Patrono,
P. Pradelles,
and
J. Grassi.
Enzyme immunometric assay for endothelin using tandem monoclonal antibodies.
J. Immunol. Methods
162:
179-192,
1993[Medline].
6.
D'Orleans-Juste, P.,
A. Claing,
T. D. Warner,
M. Yano,
and
S. Telemaque.
Characterization of receptors for endothelins in the perfused arterial and venous mesenteric vasculatures of the rat.
Br. J. Pharmacol.
110:
687-692,
1993[Medline].
7.
Fraccarollo, D.,
K. Hu,
P. Galuppo,
P. Gaudron,
and
G. Ertl.
Chronic endothelin receptor blockade attenuates progressive ventricular dilation and improves cardiac function in rats with myocardial infarction: possible involvement of myocardial endothelin system in ventricular remodeling.
Circulation
96:
3963-3973,
1997
8.
Ihara, M.,
T. Fukuroda,
T. Saeki,
M. Nishikibe,
K. Kojiri,
H. Suda,
and
M. Yano.
An endothelin receptor (ETA) antagonist isolated from Streptomyces misakiensis.
Biochem. Biophys. Res. Commun.
178:
132-137,
1991[Medline].
9.
Kaddoura, S.,
J. D. Firth,
K. R. Boheler,
P. H. Sugden,
and
P. A. Poole-Wilson.
Endothelin-1 is involved in norepinephrine-induced ventricular hypertrophy in vivo. Acute effects of bosentan, an orally active, mixed endothelin ETA and ETB receptor antagonist.
Circulation
93:
2068-2079,
1996
10.
Kajstura, J.,
W. Cheng,
K. Reiss,
W. A. Clark,
E. H. Sonnenblick,
S. Krajewski,
J. C. Reed,
G. Olivetti,
and
P. Anversa.
Apoptotic and necrotic myocyte cell deaths are independent contributing variables of infarct size in rats.
Lab. Invest.
74:
86-107,
1996[Medline].
11.
Kasai, H.,
M. Takanashi,
C. Takasaki,
and
M. Endoh.
Pharmacological properties of endothelin receptor subtypes mediating positive inotropic effects in rabbit heart.
Am. J. Physiol.
266 (Heart Circ. Physiol. 35):
H2220-H2228,
1994
12.
Kiowski, W.,
G. Sutsch,
P. Hunziker,
P. Muller,
J. Kim,
E. Oechslin,
R. Schmitt,
R. Jones,
and
O. Bertel.
Evidence for endothelin-1-mediated vasoconstriction in severe chronic heart failure.
Lancet
346:
732-736,
1995[Medline].
13.
Li, J. S.,
and
E. L. Schiffrin.
Effect of chronic treatment of adult spontaneously hypertensive rats with an endothelin receptor antagonist.
Hypertension
25:
495-500,
1995
14.
Masaki, T.
Possible role of endothelin in endothelial regulation of vascular tone.
Annu. Rev. Pharmacol. Toxicol.
35:
235-255,
1995[Medline].
15.
Moravec, C. S.,
E. E. Reynolds,
R. W. Stewart,
and
M. Bond.
Endothelin is a positive inotropic agent in human and rat heart in vitro.
Biochem. Biophys. Res. Commun.
159:
14-18,
1989[Medline].
16.
Mulder, P.,
V. Richard,
G. Derumeaux,
M. Hogie,
J. P. Henry,
F. Lallemand,
P. Compagnon,
B. Macè,
E. Comoy,
B. Letac,
and
C. Thuillez.
Role of endogenous endothelin in chronic heart failure: effect of long-term treatment with an endothelin antagonist on survival, hemodynamics, and cardiac remodeling.
Circulation
96:
1976-1982,
1997
17.
Mullan, D. M.,
D. Bell,
E. J. Kelso,
and
B. J. McDermott.
Involvement of endothelin (ET)A and ETB receptors in the hypertrophic effects of ET-1 in rabbit ventricular cardiomyocytes.
J. Cardiovasc. Pharmacol.
29:
350-359,
1997[Medline].
17a.
Øie, E.,
L. E. Vinge,
T. Tønnessen,
H. K. Grøgaard,
H. Kjekshus,
G. Christensen,
O. A. Smiseth,
and
H. Attramadal.
Transient, isopeptide-specific induction of myocardial endothelin-1 mRNA during congestive heart failure in rats.
Am. J. Physiol.
273 (Heart Circ. Physiol. 42):
H1727-H1736,
1997
18.
Omland, T.,
R. T. Lie,
A. Aakvaag,
T. Aarsland,
and
K. Dickstein.
Plasma endothelin as a prognostic indicator of 1-year mortality after acute myocardial infarction.
Circulation
89:
1573-1579,
1994
19.
Pacher, R.,
B. Stanek,
M. Hulsmann,
J. Koller-Strametz,
R. Berger,
M. Schuller,
E. Hartter,
E. Ogris,
B. Frey,
G. Heinz,
and
G. Maurer.
Prognostic impact of big endothelin-1 plasma concentrations compared with invasive hemodynamic evaluation in severe heart failure.
J. Am. Coll. Cardiol.
27:
633-641,
1996[Abstract].
20.
Pfeffer, J. M.,
M. A. Pfeffer,
P. J. Fletcher,
and
E. Braunwald.
Progressive ventricular remodeling in rat with myocardial infarction.
Am. J. Physiol.
260 (Heart Circ. Physiol. 29):
H1406-H1414,
1991
21.
Pfeffer, M. A.,
J. M. Pfeffer,
M. C. Fishbein,
P. J. Fletcher,
J. Spadaro,
R. A. Kloner,
and
E. Braunwald.
Myocardial infarct size and ventricular function in rats.
Circ. Res.
44:
503-512,
1979
22.
Pfeffer, M. A.,
J. M. Pfeffer,
C. Steinberg,
and
P. Finn.
Survival after an experimental myocardial infarction: beneficial effects of long-term therapy with captopril.
Circulation
72:
406-412,
1985
23.
Prendergast, B. D.,
P., B Anning,
M. J. Lewis,
and
A. M. Shah.
Regulation of left ventricular relaxation in the isolated guinea-pig heart by endogenous endothelin.
Cardiovasc. Res.
33:
131-138,
1997
24.
Rodeheffer, R. J.,
A. Lerman,
D. M. Heublein,
and
J. C. Burnett, Jr.
Increased plasma concentration of endothelin in congestive heart failure in humans.
Mayo Clin. Proc.
67:
719-724,
1992[Medline].
25.
Rubanyi, G. M.,
and
M. A. Polokoff.
Endothelins: molecular biology, biochemistry, pharmacology, physiology, and pathophysiology.
Pharmacol. Rev.
46:
325-415,
1994[Medline].
26.
Sakai, S.,
T. Miyauchi,
M. Kobayashi,
I. Yamaguchi,
K. Goto,
and
Y. Sugishita.
Inhibition of myocardial endothelin pathway improves long-term survival in heart failure.
Nature
384:
353-355,
1996[Medline].
27.
Sakai, S.,
T. Miyauchi,
T. Sakurai,
Y. Kasuya,
M. Ihara,
I. Yamaguchi,
K. Goto,
and
Y. Sugishita.
Endogenous endothelin-1 participates in the maintenance of cardiac function in rats with congestive heart failure: marked increase in endothelin-1 production in the failing heart.
Circulation
93:
1214-1222,
1996
28.
Selye, H.,
E. Bajusz,
S. Grasso,
and
P. Mendell.
Simple techniques for the surgical occlusion of coronary vessels in the rat.
Angiology
11:
398-407,
1960.
29.
Shubeita, H. E.,
P. M. McDonough,
A. N. Harris,
K. U. Knowlton,
C. C. Glemotski,
J. H. Brown,
and
K. R. Chien.
Endothelin induction of inositol phospholipid hydrolysis, sarcomere assembly, and cardiac gene expression in ventricular myocytes. A paracrine mechanism for myocardial cell hypertrophy.
J. Biol. Chem.
265:
20555-20562,
1990
30.
Takuwa, N.,
Y. Takuwa,
M. Yanagisawa,
K. Yamashita,
and
T. Masaki.
A novel vasoactive peptide endothelin stimulates mitogenesis through inositol lipid turnover in Swiss 3T3 fibroblasts.
J. Biol. Chem.
264:
7856-7861,
1989
31.
Teerlink, J. R.,
B. M. Loffler,
P. Hess,
J. P. Maire,
M. Clozel,
and
J. P. Clozel.
Role of endothelin in the maintenance of blood pressure in conscious rats with chronic heart failure. Acute effects of the endothelin receptor antagonist Ro 47-0203 (bosentan).
Circulation
90:
2510-2518,
1994
32.
Tønnessen, T.,
A. Giaid,
D. Saleh,
P. A. Naess,
M. Yanagisawa,
and
G. Christensen.
Increased in vivo expression and production of endothelin-1 by porcine cardiomyocytes subjected to ischemia.
Circ. Res.
76:
767-772,
1995
33.
Weisman, H. F.,
D. E. Bush,
J. A. Mannisi,
M. L. Weisfeldt,
and
B. Healy.
Cellular mechanisms of myocardial infarct expansion.
Circulation
78:
186-201,
1988
34.
Weiss, J. L.,
J. W. Frederiksen,
and
M. L. Weisfeldt.
Hemodynamic determinants of the time-course of fall in canine left ventricular pressure.
J. Clin. Invest.
58:
751-760,
1976.
35.
White, H. D.,
R. M. Norris,
M. A. Brown,
P. W. Brandt,
R. M. Whitlock,
and
C. J. Wild.
Left ventricular end-systolic volume as the major determinant of survival after recovery from myocardial infarction.
Circulation
76:
44-51,
1987
This article has been cited by other articles:
|
|
 |
|
  N. S. Dhalla, M. R. Dent, P. S. Tappia, R. Sethi, J. Barta, and R. K. Goyal Subcellular Remodeling as a Viable Target for the Treatment of Congestive Heart Failure Journal of Cardiovascular Pharmacology and Therapeutics, March 1, 2006; 11(1): 31 - 45. [Abstract] [PDF]
|
|
|
|
 |
|
 S. A. R. Paiva, L. S. Matsubara, B. B. Matsubara, M. F. Minicucci, P. S. Azevedo, A. O. Campana, and L. A. M. Zornoff Retinoic Acid Supplementation Attenuates Ventricular Remodeling after Myocardial Infarction in Rats J. Nutr., October 1, 2005; 135(10): 2326 - 2328. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. B. Murray, J. D. Gardner, G. L. Brower, and J. S. Janicki Endothelin-1 mediates cardiac mast cell degranulation, matrix metalloproteinase activation, and myocardial remodeling in rats Am J Physiol Heart Circ Physiol, November 1, 2004; 287(5): H2295 - H2299. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. E. Vinge, E. Oie, Y. Andersson, H. K. Grogaard, G. O. Andersen, and H. Attramadal Myocardial distribution and regulation of GRK and beta -arrestin isoforms in congestive heart failure in rats Am J Physiol Heart Circ Physiol, December 1, 2001; 281(6): H2490 - H2499. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
B. K. Podesser, D. A. Siwik, F. R. Eberli, F. Sam, S. Ngoy, J. Lambert, K. Ngo, C. S. Apstein, and W. S. Colucci ETA-receptor blockade prevents matrix metalloproteinase activation late postmyocardial infarction in the rat Am J Physiol Heart Circ Physiol, March 1, 2001; 280(3): H984 - H991. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 G. Torre-Amione, J. B. Young, J.-B. Durand, B. Bozkurt, D. L. Mann, I. Kobrin, and C. M. Pratt Hemodynamic Effects of Tezosentan, an Intravenous Dual Endothelin Receptor Antagonist, in Patients With Class III to IV Congestive Heart Failure Circulation, February 20, 2001; 103(7): 973 - 980. [Abstract] [Full Text] [PDF]
|
|
|
|
|
;
n = 10), CHF-saline rats (
;
n = 13), and CHF-bosentan rats (
;
n = 10). Both LV systolic and
end-diastolic pressures (LVSP and LVEDP) and dimensions (LVSD and
LVEDD) were significantly decreased in CHF rats after intervention with
bosentan (P < 0.05). Means (
) ± SE are shown for each group.
