Normal myocardial function in severe right ventricular volume overload hypertrophy

Severe left ventricular volume overloading causes myocardial and cellular contractile dysfunction. Whether this is also true for severe right ventricular volume overloading was unknown. We therefore created severe tricuspid regurgitation percutaneously in seven dogs and then observed them for 3.5-4.0 yr. All five surviving operated dogs had severe tricuspid regurgitation and right heart failure, including massive ascites, but they did not have left heart failure. Right ventricular cardiocytes were isolated from these and from normal dogs, and sarcomere mechanics were assessed via laser diffraction. Right ventricular cardiocytes from the tricuspid regurgitation dogs were 20% longer than control cells, but neither the extent (0.171 +/- 0.005 microm) nor the velocity (2.92 +/- 0.12 microm/s) of sarcomere shortening differed from controls (0.179 +/- 0.005 microm and 3.09 +/- 0.11 microm/s, respectively). Thus, despite massive tricuspid regurgitation causing overt right heart failure, intrinsic right ventricular contractile function was normal. This finding for the severely volume-overloaded right ventricle stands in distinct contrast to our finding for the left ventricle severely volume overloaded by mitral regurgitation, wherein intrinsic contractile function is depressed.     

Sarcomere shortening in pressure overload hypertrophy

Sarcomere shortening during contraction was measured by using laser diffraction, in thin, rabbit right ventricular (RV) trabeculae from normal hearts (N) (n = 5) and from hearts subjected to RV pressure overload by pulmonary banding (H) (n = 5). Banding resulted in substantial RV hypertrophy after 2 wk. Hypertrophied preparations had the same resting muscle length (H = 3.15 +/- 0.29 mm) and resting sarcomere lengths (H = 2.16 +/- 0.005 micron) as the normal preparations (3.10 +/- 0.37 mm, 2.16 +/- 0.008 micron, respectively). Total tension at the peak of isometric twitches was the same as normal in the hypertrophied muscles (N = 8.06 +/- 1.20, H = 8.51 +/- 1.95 g/mm2). However, the amount of auxotonic sarcomere shortening was much less than normal in the hypertrophied preparations (N = 0.39 +/- 0.028, H = 0.19 +/- 0.034 micron; P less than 0.001). In isotonic contractions in which the ratio of muscle shortening to resting muscle length was the same in both the normal and hypertrophied muscles (ratio of 0.05 in both groups), the extent of sarcomere shortening relative to resting sarcomere length was less in the hypertrophied muscles than in the normal preparations (N = 0.14 +/- 0.01), H = 0.07 +/- 0.01; P less than 0.01). Series elasticity was the same as normal in the hypertrophied muscle P less than 0.05). Less auxotonic sarcomere shortening for a given level of isometric tension development and less isotonic sarcomere shortening per unit muscle shortening indicate that there is less than normal work per sarcomere during contraction in hypertrophied myocardium. These findings may have important implications for intracellular compensatory adaptation in pressure overload cardiac hypertrophy.     

Absence of pressure overload induced myocardial hypertrophy after conditional inactivation of Galphaq/Galpha11 in cardiomyocytes.

Myocardial hypertrophy is an adaptational response of the heart to increased work load, but it is also associated with a high risk of cardiac mortality due to its established role in the development of cardiac failure, one of the leading causes of death in developed countries. Multiple growth factors and various downstream signaling pathways involving, for example, ras, gp-130 (ref. 4), JNK/p38 (refs. 5,6) and calcineurin/NFAT/CaM-kinase have been implicated in the hypertrophic response. However, there is evidence that the initial phase in the development of myocardial hypertrophy involves the formation of cardiac para- and/or autocrine factors like endothelin-1, norepinephrine or angiotensin II (refs. 7,8), the receptors of which are coupled to G-proteins of the Gq/11-, G12/13- and Gi/o-families. Cardiomyocyte-specific transgenic overexpression of alpha1-adrenergic or angiotensin (AT1)-receptors as well as of the Gq alpha-subunit, Galphaq, results in myocardial hypertrophy. These data demonstrate that chronic activation of the Gq/G11-family is sufficient to induce myocardial hypertrophy. In order to test whether Gq/G11 mediate the physiological hypertrophy response to pressure overload, we generated a mouse line lacking both Galphaq and Galpha11 in cardiomyocytes. These mice showed no detectable ventricular hypertrophy in response to pressure-overload induced by aortic constriction. The complete lack of a hypertrophic response proves that the Gq/G11-mediated pathway is essential for cardiac hypertrophy induced by pressure overload and makes this signaling process an interesting target for interventions to prevent myocardial hypertrophy.     

Estrogen receptor-beta mediates male-female differences in the development of pressure overload hypertrophy

The goal of this study was to determine the role of estrogen receptor subtypes in the development of pressure overload hypertrophy in mice. Epidemiological studies have suggested gender differences in the development of hypertrophy and heart disease, but the mechanism and the role of estrogen receptor subtypes are not established. We performed transverse aortic constriction (TAC) and sham operations in male and female wild-type (WT) mice and mice lacking functional estrogen receptor-alpha [alpha-estrogen receptor knockout (alpha-ERKO)] and mice lacking estrogen receptor-beta (beta-ERKO). Body, heart, and lung weights were measured 2 wk postsurgery. WT male mice subjected to TAC showed a 64% increase in the heart weight-to-body weight ratio (HW/BW) compared with sham, and WT males have increased lung weight at 2 wk. WT female mice subjected to TAC showed a 31% increase in HW/BW compared with sham, which was significantly less than their male counterparts and with no evidence of heart failure. alpha-ERKO females developed HW/BW nearly identical to that seen in WT littermate females in response to TAC, indicating that estrogen receptor-alpha is not essential for the attenuation of hypertrophy observed in WT females. In contrast, beta-ERKO females responded to TAC with a significantly greater increase in HW/BW than WT littermate females. beta-ERKO females have lower expression of lipoprotein lipase at baseline than WT or alpha-ERKO females. These data suggest an important role for estrogen receptor-beta in attenuating the hypertrophic response to pressure overload in females.     

Apoptosis in severe, compensated pressure overload predominates in nonmyocytes and is related to the hypertrophy but not function

It is widely held that myocyte apoptosis in left ventricular hypertrophy (LVH) contributes to left ventricle (LV) dysfunction and heart failure. The main goal of this investigation was to determine if there is a statistical relationship among LV hypertrophy, apoptosis and LV function, and importantly whether the apoptosis occurs in myocytes or nonmyocytes in the heart. We used both rat and canine models of severe LVH induced by chronic thoracic aortic banding with resultant LV-aortic pressure gradients 145-155 mmHg and increases in LV/body weight of 58 and 70%. These models also provided the ability to examine transmural apoptosis in LVH. In both models, the overwhelming majority (88%) of apoptotic cells were nonmyocytes. The regressions for apoptosis vs. LVH were stronger for nonmyocytes than myocytes and also stronger in the subendocardium than the subepicardium. Importantly, LV systolic and diastolic wall stresses were normal, indicating that the apoptosis could not be attributed to LV stretch or heart failure. In addition, there was no relationship between the extent of apoptosis and LV ejection fraction, which actually increased (P < 0.05), in the face of elevated LV systolic pressure, indicating that greater apoptosis did not result in a decrease in LV function. Thus, in response to chronic, severe pressure overload, LVH in the absence of LV dilation, and elevated LV wall stress, apoptosis occurred predominantly in nonmyocytes in the myocardial interstitium, more in the subendocardium than the subepicardium. The extent of apoptosis was linearly related to the amount of LV hypertrophy, but not to LV function.     

Gbetagamma-dependent phosphoinositide 3-kinase activation in hearts with in vivo pressure overload hypertrophy

Activation of phosphoinositide 3-kinases is coupled to both phosphotyrosine/growth factor and G protein-coupled receptors. We explored the role of phosphoinositide 3-kinase activation in myocardium during in vivo pressure overload hypertrophy in mice. Cytosolic extracts from wild type hypertrophied hearts showed a selective increase in the phosphoinositide 3-kinase gamma isoform. To address the role of G protein-coupled receptor-mediated activation of phosphoinositide 3-kinase, we used transgenic mice with cardiac-specific overexpression of a Gbetagamma sequestering peptide. Extracts from hypertrophied transgenic hearts showed complete loss of phosphoinositide 3-kinase activation, indicating a Gbetagamma-dependent process. To determine the class of G proteins that contribute Gbetagamma dimers for in vivo phosphoinositide 3-kinase activation, two strategies were used: 1) transgenic mice with cardiac-specific overexpression of a G(q) inhibitor peptide and 2) pertussis toxin treatment prior to pressure overload in wild type mice. Pressure overloaded G(q) inhibitor transgenic mice showed a complete absence of phosphoinositide 3-kinase activation, whereas pretreatment with pertussis toxin showed robust phosphoinositide 3-kinase activation. Taken together, these data demonstrate that activation of the phosphoinositide 3-kinase during in vivo pressure overload hypertrophy is Gbetagamma-dependent and the Gbetagamma dimers arise from stimulation of G(q)-coupled receptors.     

Interleukin-18 knockout mice display maladaptive cardiac hypertrophy in response to pressure overload

Interleukin (IL)-18 is a cardiotropic proinflammatory cytokine chronically elevated in the serum of patients with cardiac hypertrophy (LVH). The purpose of this study was to examine the role of IL-18 in pressure-overload hypertrophy using wild type (WT) and IL-18 -/- (null) mice. Adult male C57Bl/6 mice underwent transaortic constriction (TAC) for 7days or sham surgery. Heart weight/body weight ratios showed blunted hypertrophy in IL-18 null TAC mice compared to WT TAC animals. Microarray analyses indicated differential expression of hypertrophy-related genes in WT versus IL-18 nulls. Northern, Western, and EMSA analyses showed Akt and GATA4 were increased in WT but unchanged in IL-18 null mice. Our results demonstrate blunted hypertrophy with reduced expression of contractile-, hypertrophy-, and remodeling-associated genes following pressure overload in IL-18 null mice, and suggest that IL-18 plays a critical role in the hypertrophic response.     

Basis for MAP4 dephosphorylation-related microtubule network densification in pressure overload cardiac hypertrophy

Increased activity of Ser/Thr protein phosphatases types 1 (PP1) and 2A (PP2A) during maladaptive cardiac hypertrophy contributes to cardiac dysfunction and eventual failure, partly through effects on calcium metabolism. A second maladaptive feature of pressure overload cardiac hypertrophy that instead leads to heart failure by interfering with cardiac contraction and intracellular transport is a dense microtubule network stabilized by decoration with microtubule-associated protein 4 (MAP4). In an earlier study we showed that the major determinant of MAP4-microtubule affinity, and thus microtubule network density and stability, is site-specific MAP4 dephosphorylation at Ser-924 and to a lesser extent at Ser-1056; this was found to be prominent in hypertrophied myocardium. Therefore, in seeking the etiology of this MAP4 dephosphorylation, we looked here at PP2A and PP1, as well as the upstream p21-activated kinase 1, in maladaptive pressure overload cardiac hypertrophy. The activity of each was increased persistently during maladaptive hypertrophy, and overexpression of PP2A or PP1 in normal hearts reproduced both the microtubule network phenotype and the dephosphorylation of MAP4 Ser-924 and Ser-1056 seen in hypertrophy. Given the major microtubule-based abnormalities of contractile and transport function in maladaptive hypertrophy, these findings constitute a second important mechanism for phosphatase-dependent pathology in the hypertrophied and failing heart.     

Mice lacking functional TRPV1 are protected from pressure overload cardiac hypertrophy

TRPV1 (transient receptor potential cation channel, subfamily V, member 1) is best studied in peripheral sensory neurons as a pain receptor; however TRPV1 is expressed in numerous tissues and cell types including those of the cardiovascular system. TRPV1 expression is upregulated in the hypertrophic heart, and the channel is positioned to receive stimulatory signals in the hypertrophic heart. We hypothesized that TRPV1 has a role in regulating cardiac hypertrophy. Using transverse aortic constriction to model pressure overload cardiac hypertrophy we show that mice lacking functional TRPV1, compared to wild type, have improved heart function, and reduced hypertrophic, fibrotic and apoptotic markers. This suggests that TRPV1 plays a role in the progression of cardiac hypertrophy, and presents a possible therapeutic target for the treatment of cardiac hypertrophy and heart failure.     

Mice lacking functional TRPV1 are protected from pressure overload cardiac hypertrophy

TRPV1 (transient receptor potential cation channel, subfamily V, member 1) is best studied in peripheral sensory neurons as a pain receptor; however TRPV1 is expressed in numerous tissues and cell types including those of the cardiovascular system. TRPV1 expression is upregulated in the hypertrophic heart, and the channel is positioned to receive stimulatory signals in the hypertrophic heart. We hypothesized that TRPV1 has a role in regulating cardiac hypertrophy. Using transverse aortic constriction to model pressure overload cardiac hypertrophy we show that mice lacking functional TRPV1, compared to wild type, have improved heart function, and reduced hypertrophic, fibrotic and apoptotic markers. This suggests that TRPV1 plays a role in the progression of cardiac hypertrophy, and presents a possible therapeutic target for the treatment of cardiac hypertrophy and heart failure.     

Enhanced activity of ventricular Na+-HCO3- cotransport in pressure overload hypertrophy

The Na(+)-HCO(3)(-) cotransporter (NBC) plays a key role in intracellular pH (pH(i)) regulation in normal ventricular muscle. However, the state of NBC in nonischemic hypertrophied hearts is unresolved. In this study, we examined functional and molecular properties of NBC in adult rat ventricular myocytes. The cells were enzymatically isolated from both normal and hypertrophied hearts. Ventricular hypertrophy was induced by pressure overload created by suprarenal abdominal aortic constriction of 50% for 7 wk. pH(i) was measured in single cells using the fluorescent pH indicator 2',7'-bis(2-carboxyethyl)5-(6)carboxyfluorescein. Real-time PCR analysis was used to quantitatively assess expression of NBC-encoding mRNA, including SLC4A4 (encoding electrogenic NBC, NBCe1) and SLC4A7 (electroneutral NBC, NBCn1). Our results demonstrate that: 1) mRNA levels of both the electrogenic NBCe1 (SLC4A4) and electroneutral NBCn1 (SLC4A7) forms of NBC were increased by aortic constriction, 2) the onset of NBC upregulation occurred within 3 days after constriction, 3) normal and hypertrophied ventricles displayed regional differences in NBC expression, 4) acid extrusion via NBC (J(NBC)) was increased significantly in hypertrophied myocytes, 5) although acid extrusion via Na(+)/H(+) exchange was also increased in hypertrophied myocytes, the relative enhancement of J(NBC) was larger, 6) membrane depolarization markedly increased J(NBC) in hypertrophied myocytes, and 7) losartan, an ANG II AT(1) receptor antagonist, significantly attenuated the upregulation of both NBCs induced by 3 wk of aortic constriction. Enhanced NBC activity during hypertrophic development provides a mechanism for intracellular Na(+) overload, which may render the ventricles more vulnerable to Ca(2+) overload during ischemia-reperfusion.     

Right ventricular diastolic function in canine models of pressure overload,volume overload,and ischemia

By limiting filling, abnormalities of right ventricular (RV) diastolic function may impair systolic function and affect adaptation to disease. To quantify diastolic RV pressure-volume relations and myocardial compliance (MC), a new sigmoidal model was developed. RV micromanometric and sonomicrometric data in alert dogs at control (n = 16) and under surgically induced subacute (2-5 wk) RV pressure overload (n = 6), volume overload (n = 7), and ischemia (n = 6) were analyzed. The conventional exponential model detected no changes from control in the passive filling pressure-volume (P(pf)-V) relations. The new sigmoidal model revealed significant quantifiable changes in P(pf)-V relations. Maximum RV MC (MC(max)), attained during early filling, is reduced from control in pressure overload (P = 0.0016), whereas filling pressure at maximum MC (P(MCmax)) is increased (P = 0.0001). End-diastolic RV MC increases significantly in volume overload (P = 0.0131), whereas end-diastolic pressure is unchanged. In ischemia, MC(max) is decreased (P = 0.0102), with no change in P(MCmax). We conclude that the sigmoidal model quantifies important changes in RV diastolic function in alert dog models of pressure overload, volume overload, and ischemia.     

Temporal alterations in cardiac fibroblast function following induction of pressure overload

Increases in cardiovascular load (pressure overload) are known to elicit ventricular remodeling including cardiomyocyte hypertrophy and interstitial fibrosis. While numerous studies have focused on the mechanisms of myocyte hypertrophy, comparatively little is known regarding the response of the interstitial fibroblasts to increased cardiovascular load. Fibroblasts are the most numerous cell type in the mammalian myocardium and have long been recognized as producing the majority of the myocardial extracellular matrix. It is only now becoming appreciated that other aspects of fibroblast behavior are important to overall cardiac function. The present studies were performed to examine the temporal alterations in fibroblast activity in response to increased cardiovascular load. Rat myocardial fibroblasts were isolated at specific time-points (3, 7, 14, and 28 days) after induction of pressure overload by abdominal aortic constriction. Bioassays were performed to measure specific parameters of fibroblast function including remodeling and contraction of 3-dimensional collagen gels, migration, and proliferation. In addition, the expression of extracellular matrix receptors of the integrin family was examined. Myocardial hypertrophy and fibrosis were evident within 7 days after constriction of the abdominal aorta. Collagen gel contraction, migration, and proliferation were enhanced in fibroblasts from pressure-overloaded animals compared to fibroblasts from sham animals. Differences in fibroblast function and protein expression were evident within 7 days of aortic constriction, concurrent with the onset of hypertrophy and fibrosis of the intact myocardium. These data provide further support for the idea that rapid and dynamic changes in fibroblast phenotype accompany and contribute to the progression of cardiovascular disease.     

Effects of moderate pressure overload cardiac hypertrophy on the distribution of creatine kinase isozymes

Myocardial activities and isozyme distributions of creatine kinase (CK) and lactate dehydrogenase (LDH) were measured in rats with moderate pressure overload hypertrophy. Three weeks after aortic banding, the ratio of left ventricular (LV) weight to body weight increased by 30%. Values for enzyme activity in the hypertrophied LV were compared to values for control rats as well as to the contralateral relatively unaffected right ventricle (RV). In rats with moderate LV hypertrophy, total CK activity was unchanged. The percent MB-CK increased significantly (p less than 0.01) only in the hypertrophied LV, from 13 +/- 1% to 19 +/- 1% of total CK, while the sum of MM and mitochondrial-CK decreased from 86 +/- 3 to 80 +/- 3% (p less than 0.01). LDH activity increased (p less than 0.05) only in the hypertrophied ventricle from a control of 2.90 +/- 0.13 to 3.21 +/- 0.13 IU/mg protein, while the ratio of LDH activity at high to low substrate increased from 0.12 +/- 0.02 to 0.14 +/- 0.02 (p less than 0.05). Thus, the development of moderate pressure overload hypertrophy in the LV is associated with normal levels of total CK, but the percentage of MB-CK increases selectively in the primarily affected ventricle. Also, total LDH and LDH activity at high to low substrate concentration increases significantly in LV hypertrophy.     

Development of pressure overload induced cardiac hypertrophy is unaffected by long-term treatment with losartan

Left ventricular hypertrophy with adequate wall thickness, preserved adult phenotype and extracellular matrix may be useful in the prevention of heart failure. Because activation of subtype 1 of angiotensin II (AT₁) receptors is thought to be involved in the hypertrophic response of cardiomyocytes, we tested the potential of systemic AT₁ blockade to modify the development of left ventricular hypertrophy due to pressure overload.Sham-operated rats and rats with ascending aorta constriction were treated with losartan (30 mg/kg/day) for 8 weeks. Left ventricular geometry, dynamics of isovolumic contractions, hydroxyproline concentration as well as myosin isozymes (marker of fetal phenotype) were assessed. Rats with aortic constriction exhibited a marked increase in left ventricular weight and the diastolic pressure-volume relationship was shifted to smaller volumes. An enlarged ventricular pressure-volume area and increased (p < 0.05) peak values of +dP/dtmax and -dP/dtmax demonstrated an enhanced overall ventricular performance. Signs of congestive heart failure were not apparent. In contrast, parameters of myocardial fimction (normalized length-stress area, +d/dtmax and -d/dtmax) were depressed (p < 0.05), indicating an impaired myocardial contractility. The hydroxyproline concentration remained unaltered. However, the proportion of -myosin heavy chains (NMC) was increased (p < 0.05). Administration of losartan decreased (p < 0.05) blood pressure and body weight in sham operated and pressure overloaded rats. By contrast, neither the concentric left ventricular hypertrophy or depressed myocardial function nor the increased -MHC expression were significantly altered. Thus, activation of AT₁ receptors appears not to be involved in the initial expression of the fetal phenotype of pressure overloaded heart which may be responsible for the progressive functional deterioration of the hypertrophied ventricle.     

ACE2 improves right ventricular function in a pressure overload model

Background

Right ventricular (RV) dysfunction is a complication of pulmonary hypertension and portends a poor prognosis. Pharmacological therapies targeting RV function in pulmonary hypertension may reduce symptoms, improve hemodynamics, and potentially increase survival. We hypothesize that recombinant human angiotensin-converting enzyme 2 (rhACE2) will improve RV function in a pressure overload model.

Results

rhACE2 administered at 1.8 mg/kg/day improved RV systolic and diastolic function in pulmonary artery banded mice as measured by in vivo hemodynamics. Specifically, rhACE2 increased RV ejection fraction and decreased RV end diastolic pressure and diastolic time constant (p<0.05). In addition, rhACE2 decreased RV hypertrophy as measured by RV/LV+S ratio (p<0.05). There were no significant negative effects of rhACE2 administration on LV function. rhACE2 had no significant effect on fibrosis as measured by trichrome staining and collagen1α1 expression. In pulmonary artery banded mice, rhACE2 increased Mas receptor expression and normalized connexin 37 expression.

Conclusion

In a mouse RV load-stress model of early heart failure, rhACE2 diminished RV hypertrophy and improved RV systolic and diastolic function in association with a marker of intercellular communication. rhACE2 may be a novel treatment for RV failure.     

Heart hypertrophy and function are improved by idebenone in Friedreich's ataxia

Friedreich's ataxia (FRDA) is a neuro-degenerative disease causing limb and gait ataxia and hypertrophic cardiomyopathy. It results from a triplet expansion in the first intron of the frataxin gene encoding a mitochondrial protein of yet unknown function. Cells with low frataxin content display generalized deficiency of mitochondrial iron-sulfur cluster-containing proteins, which presumably denotes overproduction of superoxide radicals in these organelles. Idebenone, a short-chain quinone, may act as a potent free radical scavenger protecting mitochondria against oxidative stress. We therefore carried out an open trial of idebenone (oral supplementation; 5mg/kg/day) in a large series of FRDA patients and followed their left ventricular mass and function. Consistent and definitive worsening being observed in the natural course of the disease and cardiac hypertrophy having no chance of spontaneous reversal and to be subject to a placebo effect, the patient's heart status before and after the treatment was used to unambiguously establish the effect of the drug. After six months, heart ultrasound revealed more than 20% reduction of left ventricular mass in about half of the patients (p < 0.001) and no significant change in the other half. Since any measurable reversion of this pathogenic trait is highly significant, this demonstrates the efficiency of idebenone in controlling heart hypertrophy in FRDA. Owing to the absence of side effects of the drug, idebenone (up to 15mg/kg/day) should be prescribed for FRDA patients continuously as early as possible.