Maintenance Of Cardiac Hypertrophy Research Articles

Integrating GRK2 and NFkappaB in the Pathophysiology of Cardiac Hypertrophy.

G protein coupled receptor kinase type 2 (GRK2) plays an important role in the development and maintenance of cardiac hypertrophy and heart failure even if its exact role is still unknown. In this study, we assessed the effect of GRK2 on the regulation of cardiac hypertrophy. In H9C2 cells, GRK2 overexpression increased atrial natriuretic factor (ANF) activity and enhanced phenylephrine-induced ANF response, and this is associated with an increase of NFκB transcriptional activity. The kinase dead mutant and a synthetic inhibitor of GRK2 activity exerted the opposite effect, suggesting that GRK2 regulates hypertrophy through upregulation of NFκB activity in a phosphorylation-dependent manner. In two different in vivo models of left ventricle hypertrophy (LVH), the selective inhibition of GRK2 activity prevented hypertrophy and reduced NFκB transcription activity. Our results suggest a previously undisclosed role for GRK2 in the regulation of hypertrophic responses and propose GRK2 as potential therapeutic target for limiting LVH.

Abstract 420: Delineating the Caspase-dependent Targets and Signal Pathways That Promote Pathological Cardiac Hypertrophy

Although cardiac hypertrophy is initially an adaptive response, chronic stress on the heart is a maladaptive process that inevitably leads to end-stage heart failure. Interestingly, this pathological process is also characterized by cell behaviors associated with apoptosis. We previously demonstrated the essential role of the intrinsic cell death pathway during cardiac hypertrophy; however, the caspase-dependent pathways and cleavage targets remain elusive. To this aim, we evaluated a myocyte enhancer factor 2 (MEF2) transcription factor inhibitor, histone deacetylase 3 (HDAC3), and gelsolin as potential caspase cleavage substrates involved in the induction and/or maintenance of cardiac hypertrophy. In vitro cleavage assays were completed with effector caspase and recombinant substrate protein which demonstrated caspase-dependent cleavage. HDAC3 cleavage was observed during early stages of hypertrophy and reduced in the presence of a caspase inhibitor. Luciferase assays demonstrated that the transcriptional activity of MEF2 is dependent on intact caspase function suggesting caspase-directed HDAC3 cleavage may serve as a novel regulatory mechanism to alleviate MEF2 suppression to engage the hypertrophy gene expression program. Unlike HDAC3, caspase mediated gelsolin cleavage occurs at latter stages and is coincident with the cytoskeletal alterations that occur during this process. As gelsolin is a potent actin capping/severing enzyme, we hypothesize that caspase-mediated gelsolin activation acts as a key regulatory step in the structural rearrangements that allow for hypertrophy to occur. We have generated adenoviral vectors containing caspase cleavage mutants and cleaved forms of HDAC3 and gelsolin and will discuss the impact of these modified substrates on the hypertrophy process in vitro and in vivo. Collectively, this work suggests that caspase signalling acts to engage both the transcriptional program and cytoskeletal accommodations that characterize cardiac hypertrophy. Importantly, these observations suggest that identification of inhibitors that suppress caspase activity and/or activity of its cognate substrates may offer novel therapeutic targets to limit the development of pathological hypertrophy.

Abstract P197: Reversible Acetylation of GSK3β Regulates the Development of Cardiac Hypertrophy

Glycogen synthase kinase 3β (GSK3β) is a critical regulator of diverse cellular functions involved in the maintenance of cellular structure, function and survival. In the heart, GSK3β has been shown to play a key role in antagonizing the development of cardiac hypertrophy. It has been shown that GSK3β activity is inhibited by phosphorylation at Ser9 during the development of cardiac hypertrophy. However, recent studies indicated that blocking the Ser9 phosphorylation alone is not sufficient to protect GSK3β activity, suggesting that alternative mechanisms might be involved in regulation of GSK3β activity. In the present study, we tested the hypothesis whether GSK3β is acetylated in established hypertrophy, which leads to inhibition of GSK3β activity, and thereby promoting progression and maintenance of cardiac hypertrophy. We examined acetylation, phosphorylation and catalytic activity of GSK3β in pressure overload model of cardiac hypertrophy at different stages of hypertrophy development. The results showed that GSK3β catalytic activity was significantly down-regulated throughout the course of hypertrophy development. However, the inhibitory phosphorylation of GSK3β was observed only at initial stages, but not at later stages of pressure overload (4 weeks). Interestingly, we observed a significantly increased acetylation of GSK3β, which corresponded to the reduced activity at this later stage of hypertrophy. We then determined the impact of GSK3β acetylation on its catalytic activity, by measuring the ability of GSK3β to phosphorylate the substrate glycogen synthase. The results showed that acetylation decreased the catalytic activity of the GSK3β by almost 3 fold. Next, we performed mass spectrometry to find out the target lysine residues of GSK3β. Proteomic analysis revealed that multiple lysine residues in different regions of GSK3β were acetylated. By using site directed mutagenesis, we found that acetylation inhibits the catalytic activity of GSK3β by suppressing its substrate binding ability, which is independent of inhibitory-Ser 9 phosphorylation. These studies for the first time show a phosphorylation independent mechanism regulating GSK3β activity during progression and maintenance of cardiac hypertrophy.

Development of the heart in ANP−/− mice during early postnatal period

Disruption of the proANP gene results in marked cardiac hypertrophy, chronic and salt‐sensitive hypertension. It is unclear when cardiac hypertrophy in ANP−/− mice appears during development. The present study was designed to examine cardiac hypertrophy in ANP−/− mice during early postnatal development and correlate it with expression of the renin‐angiotensin (RAS) and the extracellular matrix (ECM) systems. Hearts were excised from 1–5 week old neonatal and 12 week adult male ANP+/+ and ANP−/− mice. Total mRNA was extracted from whole neonatal hearts and adult left ventricles, and evaluated using Northern blot analysis and quantitative RT‐PCR. Cardiac hypertrophy as indicated by an increase in heart/body weight ratio was evident in ANP−/− mice at all age groups. The most significant molecular changes were an increase in the expression of cardiac BNP mRNA level and a decrease in metalloproteinase 2 (MMP‐2) mRNA of ANP−/− as compared to the ANP+/+ mice. These data support the idea that the renin‐angiotensin system plays a minor role in the development of cardiac hypertrophy in the proANP gene disrupted mice and suggest the essential role of ANP and ECM in the development and maintenance of cardiac hypertrophy in the ANP−/− mice. Supported by the Heart and Stroke Foundation of Ontario.

Modulation of atrial contraction by PKA and PKC during the compensated phase of eccentric cardiac hypertrophy.

Calcium homeostasis is intimately regulated by protein kinase phosphorylation cascades that are also involved in the induction and maintenance of cardiac hypertrophy. In addition, the development of cardiac hypertrophy has been associated with alteration in the activation of the adrenergic system. Therefore, we investigated the specific role of protein kinase A (PKA) and C (PKC) on cardiac muscle contractile activity in the presence and absence of adrenergic stimulation. Isolated left atrial preparations from sham- and volume overload-induced cardiac hypertrophied rats were superfused with Tyrode and electrically stimulated at 0.75 Hz. Contraction was assessed in the basal and pre-stimulated (norepinephrine, 10(-9)M) states. Specific inhibitors, KT 5720 for PKA and Ro-32-0432 for PKC, were used. Peak tension development in left atria from sham-operated rats was more sensitive to PKC- than PKA-inhibition, whereas this differential sensitivity was abolished in the hypertrophied hearts. This difference was mainly due to an increase in the role of PKA in the contractile response. Developed peak tension by left atria from shunt rats was higher than that from sham rats, but when expressed to relative tissue mass, hypertrophied muscle showed weaker contraction than that from the sham group. In addition, the left atrial velocity of contraction in the sham is PKA-sensitive, while that of the shunt is PKC-sensitive. Furthermore, the velocity of relaxation shows dependency on both protein kinases, with PKC having a greater effect than PKA in the hypertrophied group. NE increased the PTD and the velocity of contraction (+dT/dt) through PKA and PKC dependent mechanisms, without affecting the velocity of relaxation (-dT/dt) in atrial muscle from sham rats. In contrast, during eccentric hypertrophy NE effectively reduced PTD as well as the -dT/dt through a PKC-dependent mechanism. The present study demonstrates that during early development of moderate eccentric cardiac hypertrophy there is: (1) a reduced specific peak tension developed due to an imbalance in the PKA and PKC activation; (2) a change in the protein kinase dependence of the velocity of contraction and relaxation from PKA to PKC with atrial hypertrophy; and (3) a negative inotropic response to adrenergic receptor stimulation. These functional responses may play a critical role in the cardiac performance during the progression of eccentric cardiac hypertrophy into the decompensated phase and heart failure.

Myocardial enzymatic activity of renin and cathepsin D before and after bilateral nephrectomy.

A local renin-angiotensin system is present within the myocardium and can play a role in the initiation and maintenance of cardiac hypertrophy. The source of myocardial renin maybe direct cardiac renin gene expression, or plasma renin of renal origin. A primary indication that myocardial renin is derived from plasma renin of renal origin was from work showing that cardiac renin activity was no longer detected 30 hours after bilateral nephrectomy (BNX). However, more recent studies have been able to detect myocardial renin after BNX. We measured normal rat cardiac renin before and after 48-hour BNX using a myocardial renin assay with improved sensitivity. The myocardial renin assay was also used to assess normal rat cardiac myocyte renin levels. Since cardiac tissue contains cathepsin D, a lysosomal enzyme capable of renin-like activity, a rat cathepsin D assay was also developed to assess cathepsin D contribution to renin-like activity. Several artifacts were shown to contribute to myocardial renin-like enzymatic activity levels after BNX, including initial plasma renin stimulation during BNX surgery, assay pH, and cardiac cathepsin D activity. Myocardial renin concentration after 48-BNX was found to be only approximately 1% of normal control levels, and renin concentration in normal cardiac myocytes was only 2-fold greater than assay blanks. Both results were probably overestimated due to cathepsin D contamination. In conclusion, no evidence was found for myocardial renin synthesis in the normal adult rat heart, and myocardial renin decays to near zero levels after 48-hour BNX.

Matters of Sex

Despite the popular concept that heart disease preferentially affects the male population, in every year for the past 16 years, cardiovascular disease has killed more women than men.1 Unfortunately, a disparity still exists between men and women in the diagnosis and aggressive treatment of heart disease. The perception that women are relatively protected is reflected by the fact that women are much more fearful of dying from some form of cancer than from cardiovascular disease.1 Understanding any risk factors that predispose or protect the female population from cardiovascular and coronary disease is therefore a compelling goal for the research community. See p 1419 Left ventricular hypertrophy is a compensatory response of the heart to a variety of stresses, and it is a strong predictor of morbidity and mortality in individuals whether or not they have been diagnosed with cardiovascular disease. Sex differences in the development of this disease over time have been suggested by clinical studies dealing with patients who suffer from aortic stenosis or hypertension.2,3 For example, older female patients with aortic stenosis tend to have increased hypertrophy accompanied by greater concentric remodeling and preserved left ventricular function when compared with male patients. Premenopausal women, in fact, exhibit a thinner posterior wall, smaller left ventricular mass, and better load-dependent and load-independent cardiac function than age-matched men under conditions of mild essential hypertension.4 Population studies have shown an age-dependent increase in left ventricular mass in healthy normotensive women that is not seen in men, as determined by echocardiography.5 These and many other studies in both humans and animals suggest that estrogens can affect the remodeling of the heart. Estrogens are potent vasodilators; they act in this capacity by increasing nitric oxide production. However, recent findings indicate that estrogens have direct actions on the myocardium as …

Relevance of Angiotensin II for Cardiac Hypertrophy and Failure Induced by Cardiac Volume Overload

Angiotensin II can contribute to the development and maintenance of cardiac hypertrophy indirectly via its hemodynamic effects and directly via its cardiac trophic effects. Cardiac volume overload by aortocaval shunt increases plasma angiotensin II as well as angiotensin II generated by cardiac tissue. Plasma angiotensin II contributes to the cardiac volume overload via its hemodynamic effects, since blockers of the renin–angiotensin system attenuate the rise in left ventricular end-diastolic pressure (LVEDP). Increased cardiac angiotensin II, generated by and dependent on cardiac ACE, however, appears to drive the hypertrophic response. For a similar prevention of the rise in circulating angiotensin II and in LVEDP, only an angiotensin-converting enzyme (ACE) inhibitor with high affinity for cardiac ACE (quinapril) also prevents the rise in cardiac angiotensin II and—similar to an AT1 receptor blocker—the development of cardiac hypertrophy. An ACE inhibitor with low affinity for cardiac ACE (enalapril) does not prevent the rise in cardiac angiotensin II by aortocaval shunt and the development of cardiac hypertrophy. In the maintenance phase of cardiac volume overload, cardiac angiotensin II returns to normal and hypertrophic growth and cardiac remodeling cease. In this phase, enalapril causes regression of cardiac hypertrophy in relation to its hemodynamic effects and in a manner similar to an AT1 receptor blocker.

Nitric oxide inhibits isoprenaline-induced positive inotropic effects in normal, but not in hypertrophied rat heart.

Evidence has accumulated that, in the rat heart, nitric oxide (NO) inhibits beta-adrenoceptor-mediated positive inotropic effects. The aim of this study was to investigate whether this effect of NO may be altered in cardiac hypertrophy. For this purpose we studied the effects of the NO-donor SNAP (S-nitroso-N-acetyl-D,L-penicillamine) on isoprenaline-induced positive inotropic effects in left ventricular strips from three models of cardiac hypertrophy: a) 12-16 weeks old male spontaneously hypertensive rats (SHR) vs. age-matched normotensive Wistar-Kyoto (WKY) rats, b) six weeks old male Wistar WKY-rats sub-totally nephrectomized (SNX) 7 weeks after SNX vs. sham-operated rats (SOP) and c) four weeks old male Wistar WKY-rats supra-renal aortic-banded (AOB, band diameter 1.0 mm) 8 weeks after AOB vs. SOP. In all three models of cardiac hypertrophy the heart weight/body weight ratio was significantly higher than in their respective controls. On isolated electrically driven ventricular strips isoprenaline (10(-10)-10(-5) M) caused concentration-dependent increases in force of contraction. Maximal increases (Emax) were similar in SHR vs. WKY-rats, but reduced in SNX- (2.9+/-0.29 vs. 5.1+/-0.34 mN, p<0.01) and AOB-rats (2.3+/-0.37 vs. 4.2+/-0.33 mN, p<0.01). In control rats (WKY and the respective SOP) the NO-donor SNAP (10(-5) M) caused a significant rightward-shift of the concentration-response curve for isoprenalinel; this rightward-shift could be inhibited by methylene blue (10(-5) M). In ventricular strips of SHR, SNX- and AOB-rats, however, 10(-5) M SNAP failed to significantly affect isoprenaline-induced positive inotropic effect. We conclude that in cardiac hypertrophy effects of NO are attenuated. Such an impairement of the NO-system could contribute to the development and/or maintenance of cardiac hypertrophy.

Alterations of Angiotensin II Receptor Contents in Hypertrophied Hearts

Tsukuba hypertensive mice, which carry the human genes for renin and angiotensinogen, show cardiac hypertrophy as well as hypertension due to activation of the renin-angiotensin system (RAS). Here, we compared the cardiac angiotensin II (Ang II) receptor contents in these and normotensive control mice by means of ligand binding studies and competitive reverse transcription-polymerase chain reaction analyses. The content of the Ang II receptor type 1 (AT1) was significantly higher at both the protein (2.5-fold; p < 0.01) and mRNA (1.4-fold; p < 0.05) levels in the hypertensive mice than that in control mice. Almost identical levels of the Ang II receptor type 2 (AT2) expression were identified at the mRNA levels in the two types of mice, although the levels were less than 20% of those of AT1 mRNA in control mice. These results suggest that AT1 in the heart is upregulated in response to Ang II-induced hypertrophic change and that, in particular, the upregulation of AT1 in particular contributes to the development and/or maintenance of cardiac hypertrophy in conjunction with the increase in Ang II production, because AT1 is responsible for cardiac hypertrophy related to the RAS.

Antihypertensive drug therapy and regression of left ventricular hypertrophy: a review with a focus on diuretics*

Regression of LVH is a reasonable adjunctive goal of blood pressure treatment, given the well described risks of hypertrophy and the possible benefits inherent in its reversal. Data suggest that any of the presently recommended agents for initial monotherapy, i.e. diuretics, beta adrenergic inhibitors, ACE inhibitors, or calcium blockers, are effective in achieving regression of hypertrophy if blood pressure lowering is achieved. While there may be other factors that play a role in the genesis and maintenance of cardiac hypertrophy in the hypertensive subject, it would appear that blood pressure elevation is probably the most important one. If reduction of blood pressure to normotensive levels can be achieved and maintained by the use of any of the antihypertensive agents (including the vasodilators, if used in combination with adrenergic inhibitors and/or diuretics), there is a reasonable chance that cardiac hypertrophy can be prevented or reversed and prognosis improved.

Signals for Cardiac Muscle Hypertrophy in Hypertension

Hypertension is associated with a rise in arterial pressure and a compensatory increase in cardiac mass, which if not treated effectively, progresses to decompensated congestive heart failure. This decompensation of an initially compensatory hypertrophy has intensified interest in the factors that initiate and maintain the development of cardiac hypertrophy. The potential signals that induce the development of cardiac hypertrophy are grouped as hemodynamic, growth-promoting hormonal, vasoconstriction-promoting hormonal, and genetic factors. Growth-promoting hormones such as insulin and thyroxine appear to play a permissive, but essential, role in the development and maintenance of cardiac hypertrophy. However, changes in cardiac load, both above and below normal, result in parallel changes in cardiac mass, which will return to normal when a normal load is restored. This adaptive response of the myocardium in direct response to elevated and depressed loads demonstrates that cardiac structure, composition, and function are not fixed postneonatal cardiac properties, but instead are regulated dynamically by the cardiocyte loading environment. This adaptive response is subject to modulation by vasoconstriction-promoting hormones and genetic factors. The current thrust in this research area is to elucidate the cellular signals that transduce the physical stimulus for hypertrophy into biochemical events underlying hypertrophic cardiac growth. To remove complex systemic interactions in vivo from the experimental paradigm, several in vitro models have been used to examine three general, but distinct, cellular pathways involving protein kinase C activation, cyclic AMP formation, and increased ion fluxes. Each pathway demonstrated a stimulatory effect on general protein synthesis, which is necessary for growth in all cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Cardiac hypertrophy and arterial distensibility in essential hypertension

Echocardiographic determinations, left ventricular mass-volume ratio ( M V ), left ventricular end-systolic stress (ESS), carotidofemoral pulse wave velocity (PWV), and brachial artery compliance (BAC), deduced from pulsed Doppler measurements and from the Bramwell-Hill equation, were evaluated in 20 patients with sustained essential hypertension in comparison with 20 control subjects of the same age and sex. In hypertensive patients, M V ratio, ESS, and PWV were significantly increased while BAC was reduced. In the overall population, ESS was directly correlated with PWV ( r = 0.73), and M V ratio was significantly correlated with PWV ( r = 0.60), BAC ( r = −0.70), and systolic arterial pressure ( r = 0.71). No comparable results were observed with diastolic arterial pressure. PWV was unchanged after cadralazine, a dihydralazine-like substance, and was decreased with the same blood pressure reduction following nitrendipine, a new calcium-blocking agent. These results suggest that: (1) the distensibility of large arteries plays an important role in the maintenance of cardiac hypertrophy in hypertension, and (2) antihypertensive drugs may act differently on arterial distensibility with possible consequences on ESS and reversion of left ventricular hypertrophy.