Skip Navigation

Cardiovascular Research 2004 61(4):771-779; doi:10.1016/j.cardiores.2003.12.005
© 2004 by European Society of Cardiology
This Article
Right arrow Abstract Freely available
Right arrow FREE Full Text (PDF) Freely available
Right arrow E-letters: Submit a response
Right arrow Alert me when this article is cited
Right arrow Alert me when E-letters are posted
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to My Personal Archive
Right arrow Download to citation manager
Right arrowRequest Permissions
Right arrow Disclaimer
Google Scholar
Right arrow Articles by Mori, T.
Right arrow Articles by Perry, G. J
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Mori, T.
Right arrow Articles by Perry, G. J
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?

Copyright © 2004, European Society of Cardiology

Volume overload results in exaggerated cardiac hypertrophy in the atrial natriuretic peptide knockout mouse

Tatsuhiko Moria,b, Yiu-Fai Chena, Ji An Fenga, Tetsuya Hayashib, Suzanne Oparila and Gilbert J Perry*,a,c

aVascular Biology and Hypertension Program, Division of Cardiovascular Disease, Department of Medicine, University of Alabama at Birmingham, Birmingham, AL 35294, USA
bOsaka Medical College, Osaka, Japan
cCardiology Section, Birmingham VA Medical Center, Birmingham, AL 35233, USA

* Corresponding author. Cardiology Section, Birmingham Veteran's Administration Medical Center, 700 S 19th Street (111H), Birmingham, AL 35233, USA. Tel.: +1-205-934-1341; fax: +1-205-975-2566. perry{at}uab.edu

Received 13 June 2003; revised 4 December 2003; accepted 8 December 2003


   Abstract
Top
 Abstract
1. Introduction
 2. Methods
 3. Results
 4. Discussion
 References
 
Objective: Previous studies suggest that atrial natriuretic peptide (ANP) may act as an autocrine/paracrine factor to modulate cardiac hypertrophy in response to various stimuli. The effect of ANP deficiency on the response to volume overload has not previously been studied. We hypothesised that ANP deficient mice would develop excess cardiac hypertrophy in response to volume overload stress. Methods: Male homozygous ANP deficient (Nppa–/–) and wildtype (Nppa+/+) male mice maintained on either a normal salt (0.55% NaCl) or low salt (0.05% NaCl) diet from weaning were studied after 2 weeks of volume overload from an aorto-caval fistula (ACF). Unoperated littermates served as controls. Left ventricular (LV) structure and function was evaluated by echocardiography. Heart, LV, and lung weights were determined at sacrifice. Myocyte diameter was measured by morphometric analysis of fixed sections of the left ventricle. Results: BP, heart weight, and LV weight were increased in Nppa–/– vs. Nppa+/+ unoperated mice. Nppa–/– mice developed exaggerated heart and LV weight compared to Nppa+/+ mice following ACF. Increased myocyte diameter paralleled increased echo LV wall thickness following ACF in Nppa+/+ but not Nppa–/– mice fed with 0.55% NaCl, indicating that an alternate mechanism contributed to increased wall thickness in Nppa–/– mice. Mid-wall shortening was mildly depressed in the Nppa–/– vs. Nppa+/+ genotype following ACF with fed 0.55% NaCl. A 0.05% NaCl diet from weaning normalized BP, but did not prevent exaggerated cardiac enlargement and LV hypertrophy following ACF in Nppa–/– mice. Conclusions: ANP-deficient mice exhibited an exaggerated increase in heart and LV weight in response to volume overload, which was not prevented by normalization of blood pressure. The findings suggest that ANP is an important physiologic modulator of the cardiac hypertrophy induced by volume overload.

KEYWORDS Aorto-caval fistula; Cardiac enlargement; Echocardiography; Transgenic mice; Heart failure


   1. Introduction
Top
 Abstract
 1. Introduction
2. Methods
 3. Results
 4. Discussion
 References
 
Atrial natriuretic peptide (ANP) is a peptide hormone that has potent natriuretic, diuretic, vasodilator, sympatholytic, and renin- and aldosterone-suppressing activities and is involved in the regulation of volume and electrolyte balance and blood pressure [1,2]. Recent studies have demonstrated that ANP is also involved in the direct regulation of cardiac growth [3–9]. Natriuretic peptide receptors have been demonstrated to be present in the heart and in cardiac cells in vitro [3,4]. Further, several recent in vivo studies have suggested that ANP expression is inversely related to cardiac growth. Transgenic mice overexpressing ANP have lower heart weight, and blood pressure than wildtype mice [5,6]. Conversely, transgenic mice with homozygous disruption of the pro-ANP gene (Nppa) or the natriuretic peptide receptor A (NPR-A) gene exhibit significant cardiac enlargement, which is out of proportion to the modest elevations of blood pressure observed in these models [7–10]. Masciotra et al. [11] found an inverse correlation between left ventricular (LV) ANP and LV mass, independent of blood pressure, in recombinant inbred rat strains derived from the Wistar-Kyoto spontaneously hypertensive rat. ANP attenuates the growth response of neonatal cardiomyocytes to angiotensin II, endothelin I, and adrenergic stimuli, while ANP receptor blockade results in hypertrophy of neonatal cardiomyocytes [12–14]. The inverse relationship of ANP levels with cardiac growth and heart weight in these various models suggests that ANP may act as an autocrine/paracrine factor to modulate cardiac hypertrophy in response to various physiologic and pathophysiologic stimuli.

The effect of selective ANP deficiency on the cardiac response to volume overload has not previously been studied. The aorto-caval fistula (ACF) model in the mouse produces biventricular volume overload characterised by progressive increases in filling pressures and hypertrophic remodelling of both ventricles over time [15]. We hypothesised that Nppa–/– mice would develop excess cardiac enlargement and ventricular hypertrophy in response to volume overload stress independently of the effects of blood pressure. We found that Nppa–/– mice fed a basal salt diet had higher mean arterial pressure (MAP) and much larger, heavier hearts than Nppa+/+ mice at baseline, and an exaggerated hypertrophic response to the stress of ACF. A low salt diet eliminated the MAP difference between genotypes, but did not ameliorate the excess hypertrophy at baseline, or the exaggerated hypertrophic response to ACF. These findings support the concept that ANP plays an important counteregulatory role in modulating the LV hypertrophic response to hemodynamic stress.


   2. Methods
Top
 Abstract
 1. Introduction
 2. Methods
3. Results
 4. Discussion
 References
 
2.1 Animal preparation
Mice with homologous deletion of the pro-ANP gene originally generated in the laboratory of John et al., [7] and control Nppa+/+ mice of the 129xB6 strain were used in these studies. The animals were raised in our resident colony, which was founded with pathogen-free breeding pairs. Genotypes were identified with polymerase chain reaction assay of genomic DNA from tail snips after weaning as previously described [16]. All protocols were approved by the Institutional Animal Care and Use Committee at the University of Alabama at Birmingham and conformed with the Guide for the Care and Use of Laboratory Animals published by the US National Institutes of Health (NIH Publication No. 85-23, revised 1996).

In the initial set of experiments, male Nppa+/+ (n = 10) and Nppa–/– (n = 12) mice fed a standard mouse pellet diet (0.55% NaCl and 0.77% Ca2+, Harlan-Teklad, Madison, WI) from weaning underwent ACF surgery at 8–10 weeks of age. Male Nppa+/+ (n = 10) and Nppa–/– (n = 7) littermates served as unoperated controls. To create an ACF, a 2.5–3-mm infra-renal side-to-side anastomosis between the inferior vena cava and abdominal aorta was made using microsurgical techniques [15]. Shunt patency was verified visually by swelling of the inferior vena cava and by mixing of arterial and venous blood. All procedures were performed by a single surgeon. We have previously demonstrated that this surgical technique produces large reproducibly sized shunts [15]. Two weeks after surgery, ACF and unoperated controls were lightly anesthetized with tribromoethanol anesthesia (375 mg/kg, i.p.) and a carotid cannula inserted into the left carotid artery. Fifteen-micrometer colored microspheres were injected into the aorta via the carotid cannula for assessment of the severity of aorto-caval shunting, as previously described [15], after which the animals were euthanized by cervical dislocation and organs collected for organ weight and histology. These microspheres are too large to cross the capillary circulation, but can reach the venous circulation and lodge in the lung capillaries via the ACF. The percentage of injected microspheres recovered from the lungs thus reflects the degree of shunting across the ACF. We confirmed similar degrees of shunting in Nppa+/+ and Nppa–/– mice using this methodology (% of injected microspheres recovered in lungs 0.84±0.06, vs. 0.74±0.05, p = 0.21, numbers represent mean±S.E.M.). After our initial experiments demonstrated significant differences in heart and LV weight between genotypes following ACF, additional Nppa+/+ (n = 6) and Nppa–/– mice (n = 6) underwent creation of an ACF for echocardiographic assessment, and were compared to additional Nppa+/+ (n = 6) and Nppa–/– (n = 5) unoperated controls.

In order to minimize BP differences between groups, the above experiments were repeated in animals fed a low salt diet (0.05% NaCl, and 0.77% Ca2+, Harlan-Teklad) from weaning (n = 6–7 mice in each group). In this group of animals, microsphere injection was not performed. Echocardiography was performed on all animals in this group, with the exception of one Nppa–/– unoperated control, and two Nppa–/– ACF mice, who died during sedation for insertion of the carotid cannula.

2.2 Echo assessment of LV size and function
Two weeks after surgery, echocardiography (10–12.5-MHz vascular probe, Agilent Sonos 5500, Agilent Technologies, Andover, MA) with simultaneous monitoring of arterial pressure under tribromoethanol anesthesia (375 mg/kg, i.p.) was performed as previously described [15]. Echo LV mass, circumferential wall stress (CWS), and velocity of circumferential fiber shortening (VCFr) were calculated using standard techniques as previously reported [15,17,18]. Wall thickness was calculated as the average of septal and posterior wall thickness. VCFdif, the difference between observed VCFr and predicted VCFr at a given CWS, was used to normalize VCFr for CWS [15]. In order to correct for differences in wall thickness between genotypes, LV systolic myocardial function was assessed from LV midwall shortening (MWS) [19]. MWSdif, the difference between observed MWS and predicted MWS at a given CWS, was used to assess LV myocardial function normalized to wall stress, in a manner analogous to that described for VCFdif [15]. Predicted MWS was calculated utilizing a regression curve of MWS vs. CWS in 30 normal mice (predicted MWS=0.22–0.000273 x CWS) (unpublished data). Following the echo studies, animals were sacrificed by cervical dislocation, and the heart and lungs were quickly removed and weighed.

2.3 Histological analysis
LV cardiomyocyte diameter was measured as previously described [20]. Hearts were excised and were immediately immersed in 4% paraformaldehyde containing 0.25% glutaraldehyde and 4.5% sucrose (1). The samples were placed into 10% formaldehyde, and then dehydrated in graded concentrations of ethanol, immersed in xylene, and finally embedded in paraffin. From the paraffin blocks 5-µm-thick sections were cut, stained with hematoxylin–eosin (HE) and Malloy-azan, and examined by light microscopy. Morphometric analysis of each heart section was performed with a computer-based morphometric system. Five to six mice from each experimental group were included in the histological analysis. At least five cross-sections of each heart were examined, and the measurements were averaged for statistical analysis. All morphometric analyses were carried out by a single examiner, who was blinded with respect to the experimental group to which each sample belonged. To evaluate the mean diameter of LV cardiomyocytes, the shortest diameter of each cardiomyocyte was measured only in nucleated transverse sections stained with hematoxylin and eosin. One hundred fifty cardiomyocytes in each LV were measured using an ocular micrometer disc with a linear scale at a magnification of 400 x, and the average cardiomyocyte diameter of each specimen was calculated.

2.4 Statistical analysis
Results are expressed as means±S.E.M. Analyses were carried out with Sigma Stat software (Jandel Scientific, San Rafael, CA). Our primary statistical test was analysis of variance (ANOVA) [21]. Differences in mean values due to main effects, and interactions between these main effects were tested with a p<0.05 considered statistically significant. Post hoc comparisons were performed by pairwise multiple comparison using the Student–Newman–Keuls test. Higher body weight was noted in the Nppa–/– compared to Nppa+/+ mice on a normal salt diet. The effects of genotype and ACF on relative tissue weights, and echo measurements of LV size and wall thickness were adjusted for differences in body weight between groups by analysis of covariance (ANCOVA) with body weight as the covariate [22,23]. Descriptive statistics to be reported includes means, standard errors, linear regression and correlation coefficients.


   3. Results
Top
 Abstract
 1. Introduction
 2. Methods
 3. Results
4. Discussion
 References
 
3.1 Effect of ANP genotype on response to ACF, normal salt diet
The effects of ANP genotype and ACF operation on body weight, MAP and organ weights at sacrifice are shown in Table 1. MAP was higher in the Nppa–/– mice than in Nppa+/+ mice, and decreased in both genotypes following ACF. Body weight was significantly increased in the Nppa–/– compared to Nppa+/+ mice. Organ weights were therefore corrected for the differences in body weight using ANCOVA. Both uncorrected and corrected values are shown in Table 1. Heart weight and LV weight were greater in unoperated Nppa–/– than Nppa+/+ mice, and increased more in Nppa–/– compared to Nppa+/+ mice following ACF (ANCOVA corrected data, interaction of genotype x ACF, p<0.05) (Fig. 1). Lung weight did not differ significantly between genotypes in unoperated mice, but increased significantly more in Nppa–/– compared to Nppa+/+ mice following ACF (ANCOVA corrected data, interaction of genotype x ACF, p = 0.04).


View this table:
[in this window]
[in a new window]

  		Table 1 Effect of genotype and ACF on body weight (BW), mean arterial pressure (MAP), and organ weights

 

Figure 1
View larger version (23K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		Fig. 1 Effects of 2-week low (0.05% NaCl) and normal (0.55% NaCl) salt diet, ACF, and Nppa genotype on whole heart weight. The adjusted heart weight was determined by analysis of covariance (ANCOVA) with body weight as a covariate. There was an exaggerated increase in heart weight in the Nppa–/– compared to Nppa+/+ mice following ACF (p<0.001 genotype x ACF, two-way ANOVA). Results are means±S.E.M. *p<0.05 vs. respective Nppa+/+ groups, {dagger}p<0.05 vs. respective control unoperated groups.

 
Echo LV mass data correlated well with LV mass at sacrifice (r = 0.895, p<0.0001). Echo measurements of chamber dimension and wall thickness were corrected for body weight by ANCOVA. Both raw and corrected data are shown in Table 2. The excess LV mass in unoperated Nppa–/– vs. Nppa+/+ mice was entirely accounted for by increased wall thickness, as LV end diastolic dimension did not differ significantly between genotypes (Fig. 2). Echocardiography demonstrated a similar pattern of hypertrophy between genotypes following ACF, with increases in both LV end diastolic dimension and wall thickness contributing to LV hypertrophy in both genotypes (Table 2, Fig. 2). Morphometric analysis revealed increased LV myocyte diameter in Nppa–/– vs. Nppa+/+ mice, but in contrast to the echo wall thickness data, the percent increase in myocyte thickness was higher in the Nppa+/+ relative to Nppa–/– mice following ACF (p<0.001 for genotype, p<0.001 for control vs. ACF operation, p<0.001 genotype x ACF, Table 3 and Figs. 3 and 4Go). This suggests that factors other than myocyte thickening contributed to the increase in wall thickness in Nppa–/– mice following ACF.


View this table:
[in this window]
[in a new window]

  		Table 2 Effect of genotype and ACF on echo parameters of LV size and function

 

Figure 2
View larger version (22K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		Fig. 2 Contribution of increased wall thickness and increased LV size to ventricular hypertrophy in Nppa+/+ and Nppa–/– mice under normal salt and low salt conditions. Excess hypertrophy in unoperated Nppa–/– mice is entirely accounted for by increased AWT on both normal and low salt diets. Following ACF, AWT increased to a similar extent in both genotypes fed either a normal or low salt diet. There was an exaggerated increase in LVEDD in Nppa–/– vs. Nppa+/+ mice on a low salt diet (p = 0.02 ACF x genotype). LVEDD trended higher in Nppa–/– vs. Nppa+/+ mice on a normal salt diet (p = 0.18 ACF x genotype). Results are means±S.E.M. Values are corrected for difference in BW by ANCOVA and normalized to Nppa+/+ unoperated control mice. AWT—Average wall thickness. LVEDD—Left ventricular end-diastolic dimension.

 

View this table:
[in this window]
[in a new window]

  		Table 3 Effect of genotype and ACF on LV myocyte diameter (µm)

 

Figure 3
View larger version (93K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		Fig. 3 Representative sections from Nppa+/+ and Nppa–/– mice, demonstrating effect of dietary salt intake and ACF on LV histology. Values are means±S.E.M. of cardiomyocyte diameter measured at the shortest diameter of each cardiomyocyte in nucleated transverse sections. Numbers in parenthesis represent numbers of mice per group. Con=Control.

 

Figure 4
View larger version (24K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		Fig. 4 Relative changes in echo wall thickness and myocyte diameter, normalized to Nppa+/+ control mice, under normal salt and low salt conditions. The percentage change in wall thickness following ACF roughly tracked the change in myocyte diameter under low salt, but not normal salt conditions. In the ANP-deficient mice on a normal salt diet, wall thickness increased more than could be accounted for by the increase in LV myocyte diameter, whereas in the ANP replete mice, myocyte diameter increased relatively more than wall thickness. These findings indicate that the mechanism of increased wall thickness in response to volume overload differs between genotypes under normal salt conditions. Under low salt conditions, changes in wall thickness roughly parallel changes in myocyte diameter in response to volume loading by ACF. AWT—Average wall thickness (echo). Myo diam—myocyte diameter, histology.

 
There was no effect of either genotype or ACF on VCFr, a measure of LV chamber function. VCFdif, which normalises VCFr for CWS, was higher in the Nppa–/– than Nppa+/+ mice, and trended lower (p = 0.056) following ACF. The apparent increased VCFdif in the Nppa–/– mice likely reflects the increased wall thickness in those animals. MWS is a better method of comparing contractility between hearts that differ in wall thickness. MWS was lower in Nppa–/– mice than Nppa+/+ following ACF (interaction term p = 0.03; p<0.05, Nppa–/– ACF vs. Nppa+/+ ACF, Student–Newman–Keuls Test for multiple comparisons). MWSdif was calculated by comparing observed MWS to MWS predicted for observed wall stress, in order to correct for differences in wall stress between groups. MWSdif was lower in Nppa–/– mice than Nppa+/+ following ACF (interaction term p = 0.01; p<0.05, Nppa–/– ACF vs. Nppa+/+ ACF, Student–Newman–Keuls test for multiple comparisons), indicating that differences in wall stress do not account for the observed differences in MWS between groups.

3.2 Effect of ANP genotype on response to ACF, low salt diet
There was no significant difference in body weight between Nppa–/– and. Nppa+/+ maintained on a low salt diet. However, a small increase in weight following ACF was noted in both genotypes. MAP did not differ significantly between genotypes or between ACF and control animals maintained on a low salt diet (Table 1). However, even in the absence of significant differences in MAP between genotypes, both heart weight and LV weight were greater in Nppa–/– than Nppa+/+ at baseline and following ACF, and increased significantly more in Nppa–/– vs. Nppa+/+ mice following ACF (genotype x ACF, p<0.01, ANCOVA corrected data) (Fig. 1 and Table 1). This was accompanied by a trend toward increased lung weight in the Nppa–/– vs. Nppa+/+ following ACF (genotype x ACF, p = 0.08; ANCOVA corrected data) (Table 1). LV end diastolic dimension did not differ between unoperated Nppa+/+ and Nppa–/– mice, but increased more in Nppa–/– compared to Nppa+/+ mice following ACF (genotype x ACF, p<0.001) (Table 2). Wall thickness by echo was increased in Nppa–/– vs. Nppa+/+ unoperated mice, and increased in both groups following ACF, but the relative increase in wall thickness following ACF did not differ between genotypes (genotype x ACF, p = 0.39). However, by LV histology, myocyte diameter was increased in Nppa–/– vs. Nppa+/+ mice, and in ACF vs. control mice, and demonstrated an exaggerated increase in the Nppa–/– mice following ACF (p<0.001 for genotype, p<0.001 for control vs. ACF operation, p<0.001 genotype x ACF). MWS was decreased in Nppa–/– vs. Nppa+/+ mice. However, unlike the normal salt ACF mouse model, MWS and MWSdif did not worsen further in the Nppa–/– genotype relative to the Nppa+/+ genotype following ACF (interaction term p = 0.56).


   4. Discussion
Top
 Abstract
 1. Introduction
 2. Methods
 3. Results
 4. Discussion
References
 
Consistent with prior reports, MAP and heart weight are increased in Nppa–/– mice compared to Nppa+/+ controls fed standard mouse chow [7,16,24]. The present study demonstrates exaggerated cardiac enlargement and ventricular hypertrophy in the Nppa–/– mice following ACF. The excess LV enlargement was accompanied by evidence of mild LV dysfunction in the Nppa–/– mice at baseline, which worsened and was accompanied by pulmonary congestion following ACF. A low salt diet resulted in similar MAP in the two genotypes, but did not prevent the cardiac enlargement or left ventricular hypertrophy in the Nppa–/– mice at baseline, or the exaggerated cardiac enlargement, left ventricular hypertrophy and pulmonary congestion in response to ACF.

The present study suggests that ANP may play an important role in modulating the hypertrophy in response to volume overload, with a more excessive hypertrophy occurring in its absence. Left atrial ANP mRNA is elevated as early as 2 weeks post-ACF in the rat, and rises further over the ensuing 4 weeks [25]. Similarly, left ventricular ANP mRNA is elevated seven-fold at 1, 2, 3 and 7 days following creation of an ACF in the rat [26]. We have previously demonstrated increased LV angiotensin II in response to ACF in the mouse, which would tend to favor cardiac hypertrophy [15]. Activation of the sympathetic nervous system and the cardiac and systemic renin–angiotensin system, and elevation of LA and LV endothelin-1 and its receptor, have also been demonstrated in the rat following ACF [27–29]. The findings in the present study suggest that activation of these trophic influences in the absence of ANP results in excessive hypertrophy, and is consistent with the observation that ANP opposes the growth promoting effects of angiotensin II, endothelin-1, and norepinephrine in isolated myocytes and fibroblasts [13,14].

Morphometric analysis in the present study demonstrated that cross-sectional myocyte diameter is increased in the Nppa–/– relative to the Nppa+/+ mouse consuming either normal salt or low salt diet. The change in myocyte diameter roughly paralleled the change in echocardiographic wall thickness in mice on a low salt diet. However, the mechanism of increased wall thickness differed between genotypes on a normal salt diet. Nppa+/+ mice demonstrated a greater increase in myocyte diameter relative to wall thickness following ACF, whereas Nppa–/– mice demonstrated a greater increase in wall thickness relative to myocyte diameter. We have previously demonstrated increased expression of the extracellular matrix molecules periostin, osteopontin, collagen I and III, and thrombospondin in Nppa–/– relative to the Nppa+/+ mice exposed to experimental pressure overload due to transverse aortic banding [30]. The time course and direction of extracellular matrix changes in the ACF model in the mouse is unknown. In the rat, some investigators have reported decreased collagen volume fraction from 6 h to 1 week post-ACF, with recovery to normal levels by 2 weeks [31], whereas others have found a persistent decrease in collagen volume fraction as late as 4–10 weeks post-AVF [32]. Our findings of a greater percentage increase in myocyte diameter relative to wall thickness following ACF in Nppa+/+ mice on a normal salt diet suggest a relative decrease in extracellular matrix 2 weeks post-ACF in this group. The explanation for the opposite finding in Nppa–/– mice on a normal salt diet, i.e., a greater increase in wall thickness relative to myocyte diameter, is unclear. Increased myocyte number, increased hyperplasia of non-myocyte cells, increased synthesis or decreased breakdown of extracellular matrix components, or increased tissue water in the Nppa–/– vs. the Nppa+/+ ACF mice could account for this discrepancy. Further study is needed to elucidate the mechanism underlying the disparity between change in myocyte thickness and change in wall thickness observed between genotypes following ACF.

We measured MWS to assess differences in myocardial contractility between groups. MWS corrects for differences between endocardial and midwall stress, and is a more appropriate means than VCF to compare myocardial function between groups with differing levels of ventricular hypertrophy [19,33]. We found evidence of depressed LV MWS in the Nppa–/– vs. the Nppa+/+ genotype on a normal salt diet, with further deterioration in the Nppa–/– relative to the Nppa+/+ genotype following ACF. The findings suggest the development of early ventricular dysfunction in the hypertrophied hearts of the Nppa–/– mice. Similar depression of ventricular function as assessed by MWS has been described in patients with LVH due to essential hypertension [19,33]. A low salt diet attenuated the difference in MWS between genotypes (p = 0.07 after correction for wall stress), and prevented further deterioration in the Nppa–/– following ACF.

Interpretation of the differential effect of genotype on cardiac size and function in response to ACF in this study is complicated by baseline differences in cardiac size and blood pressure between genotypes. We repeated the experiments on a low salt diet in an attempt to eliminate these baseline differences. A small, statistically nonsignificant difference in anesthetized MAP (~9 mm Hg) persisted between genotypes on a low salt diet in the present study. We have previously observed a 6-mm Hg (nonsignificant) difference in MAP between Nppa+/+ and Nppa–/– genotypes in conscious animals on a low salt diet (Chen, unpublished data). Although the MAP difference was not statistically significant in either study, the power to detect a difference of this magnitude in the present study was small given the variability of MAP and the number of animals studied (power ~0.20 for alpha of 0.05). John et al. [7] reported no significant difference in MAP between Nppa–/– and Nppa+/+ mice on 0.5% NaCl (standard mouse chow) in their original description of this genotype, but in fact did observe a 9-mm Hg (nonsignificant) difference between genotypes. In a subsequent report, this group reported a 14-mm Hg difference between Nppa–/– and +/+ mice following a low salt diet (0.008%) for 1 week [24]. The data from these various studies suggest a small difference in MAP between Nppa–/– and Nppa+/+ cannot be excluded, even on a low salt diet. Nonetheless, it seems unlikely that the small MAP difference observed can account for the major differences in heart weight and wall thickness in Nppa–/– vs. Nppa+/+ mice on a low salt diet.

We observed significant increased body weight in the Nppa–/– vs. Nppa+/+ mice maintained on a normal salt diet. The mechanism underlying this difference is unknown. Preliminary observations in our laboratory indicate that %body fat is not increased in Nppa–/– vs. Nppa+/+ mice (Chen, unpublished data). We corrected for these differences by ANCOVA with body weight as the covariate. The difference in body weight between genotypes was eliminated by a low salt diet. The mechanism underlying this effect is uncertain. However, we do not believe that fluid retention entirely accounts for the observed weight differences on a normal salt diet, as the Nppa–/– mice appear grossly larger than their Nppa+/+ counterparts.

In summary, ANP-deficient mice are characterized by salt-sensitive hypertension and concentric LV hypertrophy. Following ACF, Nppa–/– mice exhibit LV dysfunction and exaggerated heart weight and LV weight relative to Nppa+/+ mice. Salt restriction decreases MAP in Nppa–/– to levels similar to those observed in wildtypes, but does not eliminate the exaggerated heart weight and LV weight in either control or ACF mice. The findings suggest that ANP is an important physiologic modulator of the cardiac enlargement induced by volume overload.


   Acknowledgements
 
This work was supported in part by National Heart, Lung, and Blood Institute grants HL-44195, HL-50147, HL-45990, HL-07457, and HL-56046.


   Notes
 
Time for primary review 31 days


   References
Top
 Abstract
 1. Introduction
 2. Methods
 3. Results
 4. Discussion
 References
 

  1. Espiner E.A. Contemporary Endocrinology: Natriuretic Peptides in Health and Disease. Samson W.K., Levin E.R., eds. (1997) Totowa, NJ: Humana Press. 123–146.
  2. Flynn T.G. Contemporary Endocrinology: Natriuretic Peptides in Health and Disease. Samson W.K., Levin E.R., eds. (1997) Totowa, NJ: Humana Press. 1–20.
  3. Cao L., Gardner D.G. Natriuretic peptides inhibit DNA synthesis in cardiac fibroblasts. Hypertension (1995) 25:227–234.[Abstract/Free Full Text]
  4. Lin X., Hanze J., Heese F., Sodmann R., Lang R.E. Gene expression of natriuretic peptide receptors in myocardial cells. Circ. Res. (1995) 77:750–758.[Abstract/Free Full Text]
  5. Klinger J.R., Petit R.D., Curtin L.A., et al. Cardiopulmonary responses to chronic hypoxia in transgenic mice that overexpress ANP. Appl. Phys. (1993) 75:198–205.
  6. Barbee R.W., Perry B.D., Re R.N., Murgo J.P., Field L.J. Hemodynamics in transgenic mice with overexpression of atrial natriuretic peptide. Circ. Res. (1994) 74:747–751.[Abstract/Free Full Text]
  7. John S.W.M., Krege J.H., Oliver P.M., et al. Genetic decreases in atrial natriuretic peptide and salt-sensitive hypertension. Science (1995) 267:679–681.[Abstract/Free Full Text]
  8. Oliver P.M., Fox J.E., Kim R., et al. Hypertension, cardiac hypertrophy, and sudden death in mice lacking natriuretic peptide receptor A. Proc. Natl. Acad. Sci. (1997) 94:14730–14735.[Abstract/Free Full Text]
  9. Kishimoto I., Rossi K., Garbers D.L. A genetic model provides evidence that the receptor for atrial natriuretic peptide (guanylyl cyclase-A) inhibits cardiac ventricular hypertrophy. Proc. Natl. Acad. Sci. (2001) 98:2703–2706.[Abstract/Free Full Text]
  10. Knowles J.W., Exposito G., Mao L., et al. Pressure-independent enhancement of cardiac hypertrophy in natriuretic peptide receptor A-deficient mice. J. Clin. Invest. (2001) 107:975–984.[Web of Science][Medline]
  11. Masciotra S., Picard S., Deschepper C.F. Cosegregation analysis in genetic crosses suggests a protective role for atrial natriuretic factor against ventricular hypertrophy. Circ. Res. (1999) 84:1453–1458.[Abstract/Free Full Text]
  12. Horio T., Nishikimi T., YToshihara F., Matsuo H., Takishita S., Kangawa K. Inhibitory regulation of hypertrophy by endogenous atrial natriuretic peptide in cultured cardiac myocytes. Hypertension (2000) 35:19–24.[Abstract/Free Full Text]
  13. Fujisaki H., Ito H., Hirata Y., et al. Natriuretic peptides inhibit angiotensin II-induced proliferation of rat cardiac fibroblasts by blocking endothelin-1 gene expression. J. Clin. Invest. (1995) 96:1059–1065.[Web of Science][Medline]
  14. Calderone A., Thaik C.M., Takahashi N., Chang D.L.F., Colucci W.S. Nitric oxide, atrial natriuretic peptide, and cyclic GMP inhibit the growth-promoting effects of norepinephrine in cardiac myocytes and fibroblasts. J. Clin. Invest. (1998) 101:812–818.[Web of Science][Medline]
  15. Perry G.J., Mori T., Wei C., et al. Genetic variation in angiotensin converting enzyme does not prevent the development of cardiac hypertrophy or upregulation of angiotensin II in response to aorto-caval fistula. Circulation (2001) 103:1012–1016.[Abstract/Free Full Text]
  16. Sun J.Z., Chen S.J., Li G., Chen Y.F. Hypoxia reduces atrial natriuretic peptide clearance receptor gene expression in ANP knockout mice. Am. J. Physiol. (2000) 279:L511–L519.[Web of Science]
  17. Manning W., Wei J., Katz S., Litwin S., Douglas P. In vivo assessment of LV mass in mice using high-frequency cardiac ultrasound: necropsy validation. Am. J. Physiol. (1994) 266:H1672–H1675. [Heart Circ Physiol 35].[Web of Science][Medline]
  18. Gaasch W.H., Battle W.E., Oboler A.A., Banas J.S., Levine H.J. Left ventricular stress and compliance in man with special reference to normalized ventricular function curves. Circulation (1972) 45:746–762.[Abstract/Free Full Text]
  19. De Simone G.D., Devereux R.B., Koren M.J., Mensah G.A., Casale P.N., Laragh J.H. Midwall left ventricular mechanics. An independent predictor of cardiovascular risk in arterial hypertension. Circulation (1996) 93:259–265.[Abstract/Free Full Text]
  20. Hayashi T., Ijiri Y., Toko H., et al. Increased digitalis-like immunoreactive substances in patients with hypertrophic cardiomyopathy. Eur. Heart J. (2000) 21:296–305.[Abstract/Free Full Text]
  21. Milliken G.A., Johnson D.E. Analysis of Messy Data. (1984) New York: Van Nostrand, Reihold.
  22. Packard G.C., Boardman T.J. New Directions in Ecological Physiology. Feder M.E., Bennett A.F., Burggren W.W., Huey R.B., eds. (1987) Cambridge: Cambridge Univ. Press. 216–239.
  23. Packard G.C., Boardman T.J. The misuse of ratios, indices, and percentages in ecophysiological research. Physiol. Zool. (1988) 61:1–9.
  24. John S.W., Veress A.T., Honrath U., Chong C.K., Peng L., Smithies O. Blood pressure and fluid-electrolyte balance in mice with reduced or absent ANP. Am. J. Physiol. (1996) 271:R109–R114.[Web of Science][Medline]
  25. Yamakawa H., Imamura T., Matsuo T., et al. Diastolic wall stress and ANG II in cardiac hypertrophy and gene expression induced by volume overload. Am. J. Physiol. (2000) 279:H2939–H2946.[Web of Science]
  26. Su X., Brower G., Janicki J.S., Chen Y.F., Oparil S.O., Dell'Italia L.J. Differential expression of natriuretic peptides and their receptors in volume overload cardiac hypertrophy in the rat. J. Mol. Cell. Cardiol. (1999) 31:1927–1936.[CrossRef][Web of Science][Medline]
  27. Willenbrock R., Stauss H., Scheuermann M., Osterziel K.J., Unger T., Dietz R. Effect of chronic volume overload on baroreflex control of heart rate and sympathetic nerve activity. Am. J. Physiol. (1997) 273:H2580–H2585.[Web of Science][Medline]
  28. Ruzicka M., Skarda V., Leenen F.H. Effects of ACE inhibitors on circulating versus cardiac angiotensin II in volume overload-induced cardiac hypertrophy in rats. Circulation (1995) 92:3568–3573.[Abstract/Free Full Text]
  29. Brown L.A., Nunez D.J., Brookes C.I.O., Wilkins M.R. Selective increase in enthothelin-1 and endothelin A receptor subtype in the hypertrophied myocardium of the aorto-venacaval fistula rat. Circ. Res. (1995) 29:768–774.
  30. Wang D., Oparil S., Feng J.A., et al. Effects of pressure overload on extracellular matrix expression in the heart of the atrial natriuretic peptide-null mouse. Hypertension (2003) 42:88–95.[Abstract/Free Full Text]
  31. Brower G.L., Chancey A.L., Thanigaraj S., Matsubara B.B., Janicki J.J. Cause and effect relationship between myocardial mast cell number and matrix metalloproteinase activity. Am. J. Physiol, Heart Circ. Physiol. (2002) 283:H518–H525.[Abstract/Free Full Text]
  32. Ruzicka M., Keeley F.W., Leenen F.H. The renin–angiotensin system and volume overload-induced changes in cardiac collagen and elastin. Circulation (1994) 90:1989–1996.[Abstract/Free Full Text]
  33. Ganau A., Devereux R.B., Roman M.J., et al. Patterns of left ventricular hypertrophy and geometric remodeling in essential hypertension. J. Am. Coll. Cardiol. (1992) 19:1550–1558.[Abstract]

Add to CiteULike CiteULike   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us    What's this?


This article has been cited by other articles:


Home page
 HypertensionHome page
C. Matsumoto, T. Hayashi, K. Kitada, C. Yamashita, M. Miyamura, T. Mori, A. Ukimura, M. Ohkita, D. Jin, S. Takai, et al.
Chymase Plays an Important Role in Left Ventricular Remodeling Induced by Intermittent Hypoxia in Mice
Hypertension, July 1, 2009; 54(1): 164 - 171.
[Abstract] [Full Text] [PDF]

Home page
 Am. J. Physiol. Heart Circ. Physiol.Home page
T. Hayashi, C. Yamashita, C. Matsumoto, C.-J. Kwak, K. Fujii, T. Hirata, M. Miyamura, T. Mori, A. Ukimura, Y. Okada, et al.
Role of gp91phox-containing NADPH oxidase in left ventricular remodeling induced by intermittent hypoxic stress
Am J Physiol Heart Circ Physiol, May 1, 2008; 294(5): H2197 - H2203.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow FREE Full Text (PDF) Freely available
Right arrow E-letters: Submit a response
Right arrow Alert me when this article is cited
Right arrow Alert me when E-letters are posted
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to My Personal Archive
Right arrow Download to citation manager
Right arrowRequest Permissions
Right arrow Disclaimer
Google Scholar
Right arrow Articles by Mori, T.
Right arrow Articles by Perry, G. J
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Mori, T.
Right arrow Articles by Perry, G. J
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?