Cardiac-specific overexpression of catalase rescues ventricular myocytes from ethanol-induced cardiac contractile defect

Oxidative stress is intimately involved in alcoholic cardiomyopathy. Catalase is responsible for detoxification of hydrogen peroxide (H(2)O(2)) and may interfere with ethanol-induced cardiac toxicity. To test this hypothesis, a transgenic mouse line was produced to overexpress catalase (~50-fold) in the heart, ranging from sarcoplasm, the nucleus and peroxisomes within myocytes. Mechanical and intracellular Ca(2+) properties were evaluated in ventricular myocytes from catalase transgenic (CAT) and wild-type FVB mice. Protein abundance of sarco (endo) plasmic reticulum Ca(2+)-ATPase (SERCA), phospholamban (PLB), Na(+)/Ca(2+) exchanger (NCX), dihydropyridine Ca(2+) receptor (DHPR), ryanodine receptor (RyR), Akt and phosphorylated Akt (pAkt) were measured by western blot. CAT itself did not alter body and organ weights, as well as myocyte contractile properties. Acute exposure of ethanol elicited a concentration-dependent depression in cell shortening and intracellular Ca(2+) in FVB mice with maximal inhibitions of 65.4% and 35.8%, respectively. The ethanol-induced cardiac depression was significantly attenuated in myocytes from CAT with maximal inhibitions of 42.4% and 27.3%. CAT also abrogated the ethanol-induced inhibition of maximal velocity of shortening/relengthening, prolongation of relengthening duration and intracellular Ca(2+) clearing time. Cell shortening at different extracellular Ca(2+) revealed stronger myocyte-shortening amplitude under lower (0.5 mM) Ca(2+) in CAT mice. Protein expression of NCX, RyR, Akt and pAkt were elevated in myocytes from CAT mice, while those of SERCA, PLB and DHPR were not affected. In conclusion, our data suggest that catalase overexpression may protect cardiac myocytes from ethanol-induced contractile defect, partially through improved intracellular Ca(2+) handling and Akt signaling.     

Cardiac-specific overexpression of Galphaq alters excitation-contraction coupling in isolated cardiac myocytes.

Transgenic mice with cardiac-specific overexpression of G alpha q exhibit a biochemical and physiological phenotype of load-independent cardiac hypertrophy with contractile dysfunction. To elucidate the cellular basis for altered contractility, we measured cellular contraction, Ca(2+)transients, and l -type Ca(2+)channel currents (I(Ca)) in left ventricular (LV) myocytes isolated from non transgenic (NT) controls or G alpha q hearts. Although baseline contractile function (% shortening) and the amplitude of Ca(2+)transients in G alpha q myocytes were similar to NT myocytes, the rates of cellular shortening and relengthening and the duration of Ca(2+)transients were prolonged in G alpha q myocytes. Myocytes from G alpha q hearts had larger cell capacitance but no change in I(Ca)density, voltage-dependence of activation and inactivation. The responses of I(Ca)to dihydropyridine drugs and a membrane permeable cAMP analog, 8-(4-chlorophenylthio) cAMP, were not altered; however, the time course of I(Ca)inactivation was significantly slower in G alpha q myocytes compared to NT myocytes. The kinetic difference in inactivation was abolished when Ba(2+)was used as the charge carrier or when the sarcoplasmic reticulum (SR) Ca(2+)was depleted by ryanodine, suggesting that Ca(2+)-dependent inactivation is reduced in G alpha q myocytes due to altered SR Ca(2+)release. Consistent with this hypothesis, the function of SR as assessed by the maximal Ca(2+)uptake rates and the apparent affinity of SR Ca(2+)-ATPase for Ca(2+)was reduced in ventricles of G alpha q heart. These results suggest that the reduced SR function contributes to the depressed contractility associated with this form of cardiac hypertrophy.     

Cardiac contractile function is enhanced in isolated ventricular myocytes from growth hormone transgenic mice

Growth hormone (GH) plays a key role in cardiac growth and function. However, excessive levels of GH often result in cardiac dysfunction, which is the major cause of death in acromegalic patients. Transgenic mice with GH over-expression serve as useful models for acromegaly and exhibit impaired cardiac functions using echocardiography, similar to those of human acromegaly. However, the mechanism underscoring the impaired ventricular function has not been well defined. This study was designed to evaluate the cardiac excitation-contraction coupling in GH over-expressing transgenic mice at the single ventricular myocyte level. Myocytes were isolated from GH and age-matched wild-type mouse hearts. Mechanical properties were evaluated using an IonOptix MyoCam system. The contractile properties analyzed included peak shortening (PS), time-to-peak shortening (TPS) and time-to-90% relengthening (TR(90)), and maximal velocities of shortening/relengthening (+/-dL/dt). Intracellular Ca2+ properties were evaluated by fura-2. GH transgenic mice exhibited significantly increased body weights and enlarged heart and myocyte size. Myocytes from GH transgenic mice displayed significantly enhanced PS and+/-dL/dt associated with similar TPS and TR(90) compared with the wild-type littermates. Myocytes from GH transgenic mice displayed a similar resting intracellular Ca2+ level and Ca2+ removal rate but exhibited an elevated peak intracellular Ca2+ level compared with the wild-type group. Myocytes from both groups were equally responsive to increases in extracellular Ca2+ concentration and stimulating frequency. These results suggest that GH over-expression is associated with enhanced contractile function in isolated myocytes and that the impaired cardiac function observed in whole hearts may not be due to defects at the myocyte level.     

Increased Contraction of Myocytes Isolated From the Young Spontaneously Hypertensive Rat: Relationship Between Systolic and Diastolic Function

This study was designed to assess heart performance in young (10-week-old) spontaneously hypertensive rats (SHR) and Wistar-Kyoto (WKY) rats, in terms of whole heart function in vivo and mechanics of isolated ventricular myocytes in vitro. The data suggest that left ventricular pressure (LVP) generation is greater, and the maximal velocities of LVP generation and decline are faster in SHR than in WKY. Two-dimensional morphologic measurements show that SHR myocytes are hypertrophied and that augmented contractile function is also present in isolated cells as determined by the extent of shortening and velocity of shortening. Relaxation is also faster at the myocyte level as determined by velocity of relengthening. However, the slope of the relationship between myocyte peak shortening and velocity of relaxation was similar in both groups. These results suggest that hyperdynamic myocyte relengthening may reflect changes in elastic recoil from increased shortening rather than intrinsic changes in cellular mechanisms, which are independent of shortening.     

Cardiac overexpression of alcohol dehydrogenase exacerbates cardiac contractile dysfunction,lipid peroxidation,and protein damage after chronic ethanol ingestion

BACKGROUND: Alcoholic cardiomyopathy is manifested as ventricular dysfunction, although its specific toxic mechanism remains obscure. This study was designed to examine the impact of enhanced acetaldehyde exposure on cardiac function via cardiac-specific overexpression of alcohol dehydrogenase (ADH) after alcohol intake. METHODS: ADH transgenic and wild-type FVB mice were placed on a 4% alcohol or control diet for 8 weeks. Mechanical and intracellular Ca2+ properties were evaluated in cardiac myocytes. Levels of acetaldehyde, lipid peroxidation, and protein carbonyl formation were determined. RESULTS: FVB and ADH mice consuming ethanol exhibited elevated blood ethanol/acetaldehyde, cardiac acetaldehyde, and cardiac hypertrophy compared with non-ethanol-consuming mice. However, the levels of cardiac acetaldehyde and hypertrophy were significantly greater in ADH ethanol-fed mice than FVB ethanol-fed mice. ADH transgene itself did not affect mechanical and intracellular Ca2+ properties with the exception of reduced resting intracellular Ca2+ and Ca2+ re-sequestration at low pace frequency. Myocytes from ethanol-fed mice showed significantly depressed peak shortening, velocity of shortening/relengthening, rise of intracellular Ca2+ transients, and sarco(endo)plasmic reticulum Ca2+ load associated with similar duration of shortening/relengthening compared with myocytes from control mice. Strikingly, the ethanol-induced mechanical and intracellular Ca2+ defects were exacerbated in ADH myocytes compared with the FVB group except velocity of shortening/relengthening. The lipid peroxidation end products malondialdehyde and protein carbonyl formation were significantly elevated in both livers and hearts after chronic ethanol consumption, with the cardiac lipid and protein damage being exaggerated by ADH transgene. CONCLUSION: These data suggest that increased cardiac acetaldehyde exposure due to ADH transgene may play an important role in cardiac contractile dysfunctions associated with lipid and protein damage after alcohol intake.     

Lats2 is a negative regulator of myocyte size in the heart

Mammalian sterile 20-like kinase (Mst)1 plays an important role in mediating apoptosis and inhibiting hypertrophy in the heart. Because Hippo, a Drosophila homolog of Mst1, forms a signaling complex with Warts, a serine/threonine kinase, which in turn stimulates cell death and inhibits cell proliferation, mammalian homologs of Warts, termed Lats1 and Lats2, may mediate the function of Mst1. We here show that Lats2, but not Lats1, dose-dependently increased apoptosis in cultured cardiac myocytes. Lats2 also dose-dependently reduced [(3)H]phenylalanine incorporation and cardiac myocyte size, whereas dominant negative Lats2 (DN-Lats2) increased them, suggesting that endogenous Lats2 negatively regulates myocyte growth. DN-Lats2 significantly attenuated induction of apoptosis and inhibition of hypertrophy by Mst1, indicating that Lats2 mediates the function of Mst1 in cardiac myocytes. Cardiac specific overexpression of Lats2 in transgenic mice significantly reduced the size of left and right ventricles, whereas that of DN-Lats2 caused hypertrophy in both ventricles. Overexpression of Lats2 reduced left ventricular systolic and diastolic function without affecting baseline levels of myocardial apoptosis. Expression of endogenous Lats2 was significantly upregulated in response to transverse aortic constriction. Overexpression of DN-Lats2 significantly enhanced cardiac hypertrophy and inhibited cardiac myocyte apoptosis induced by transverse aortic constriction. These results suggest that Lats2 is necessary and sufficient for negatively regulating ventricular mass in the heart. Although Lats2 is required for cardiac myocyte apoptosis in response to pressure overload, it was not sufficient to induce apoptosis at baseline. In conclusion, Lats2 affects both growth and death of cardiac myocytes, but it primarily regulates the size of the heart and acts as an endogenous negative regulator of cardiac hypertrophy.     

Beta1 integrins modulate beta-adrenergic receptor-stimulated cardiac myocyte apoptosis and myocardial remodeling

Sympathetic nerve activity increases in the heart during cardiac failure. Here, we hypothesized that beta1 integrins play a protective role in chronic beta-adrenergic receptor-stimulated cardiac myocyte apoptosis and heart failure. L-isoproterenol (iso; 400 microg/kg per hour) was infused in a group of wild-type (WT) and beta1 integrin heterozygous knockout (hKO) mice. Left ventricular structural and functional remodeling was studied at 7 and 28 days of iso-infusion. Western blot analysis demonstrated reduced beta1 integrin levels in the myocardium of hKO-sham. Iso-infusion increased heart weight:body weight ratios in both groups. However, the increase was significantly higher in WT-iso. M-mode echocardiography indicated increased left ventricular end-diastolic diameter, percentage of fractional shortening, and ejection fraction in the WT-iso group. The percentage of fractional shortening and ejection fraction were significantly lower in hKO-iso versus hKO-sham and WT-iso. Peak left ventricular developed pressure and left ventricular end-diastolic pressure measured using Langendorff-perfusion analyses were significantly higher in the WT-iso group (P<0.05 versus WT-sham and hKO-Iso). The number of TUNEL-positive myocytes was significantly higher in hKO-iso hearts 7 and 28 days after iso-infusion. The increase in myocyte cross-sectional area and fibrosis was higher in the WT-iso group. Matrix metalloproteinase-9 protein levels were significantly higher in WT-iso, whereas matrix metalloproteinase-2 levels were increased in hKO-iso hearts. Iso-infusion increased phosphorylation of c-Jun N-terminal kinase and extracellular signal-regulated kinase 1/2 in both groups. The increase in c-Jun N-terminal kinase phosphorylation was significantly higher in hKO-iso (P<0.001 versus WT-iso). Thus, beta1 integrins play a crucial role in beta-adrenergic receptor-stimulated myocardial remodeling with effects on cardiac myocyte hypertrophy, apoptosis, and left ventricular function.     

Compensatory hypertrophy induced by ventricular cardiomyocyte-specific COX-2 expression in mice

Cyclooxygenase-2 (COX-2) is an important mediator of inflammation in stress and disease states. Recent attention has focused on the role of COX-2 in human heart failure and diseases owing to the finding that highly specific COX-2 inhibitors (i.e., Vioxx) increased the risk of myocardial infarction and stroke in chronic users. However, the specific impact of COX-2 expression in the intact heart remains to be determined. We report here the development of a transgenic mouse model, using a loxP-Cre approach, which displays robust COX-2 overexpression and subsequent prostaglandin synthesis specifically in ventricular myocytes. Histological, functional, and molecular analyses showed that ventricular myocyte specific COX-2 overexpression led to cardiac hypertrophy and fetal gene marker activation, but with preserved cardiac function. Therefore, specific induction of COX-2 and prostaglandin in vivo is sufficient to induce compensated hypertrophy and molecular remodeling.     

Cardiac unloading alters contractility and calcium homeostasis in ventricular myocytes

Altered cardiac workload has an important effect on myocyte structure and function. Cardiac hypertrophy resulting from an increase in load has been studied extensively in the past. However, the effects of unloading and atrophy have recently become of more interest since devices for mechanical left ventricular unloading have been introduced into clinical practice for the treatment of patients with terminal heart failure, and a resulting improved cardiac and myocyte contractility have been reported. We used the heterotopic abdominal mouse heart transplant model in order to study the effects of 5 days of unloading on cell size (confocal microscopy), contractility (fractional shortening: video motion), calcium homeostasis ([Ca(2+)](i)transients, SR Ca(2+)content); and L-type Ca(2+)and sodium/calcium exchanger currents (whole cell patch clamp technique). We found unloading caused decreased cell volume consistent with atrophy. An increased fractional shortening and [Ca(2+)](i)transient were observed in myocytes from unloaded hearts as compared with controls. Transsarcolemmal I(Ca,L)and I(Na/Ca)densities, and SR Ca(2+)content were unaltered, as was membrane capacitance. A reduction in cell volume with mainteinance of internal and surface membrane areas, and/or a decrease in concentration of cellular protein Ca(2+)buffers, may contribute to the increase in the [Ca(2+)](i)transient in this model.     

Pim-1 kinase antagonizes aspects of myocardial hypertrophy and compensation to pathological pressure overload

Pim-1 kinase exerts potent cardioprotective effects in the myocardium downstream of AKT, but the participation of Pim-1 in cardiac hypertrophy requires investigation. Cardiac-specific expression of Pim-1 (Pim-WT) or the dominant-negative mutant of Pim-1 (Pim-DN) in transgenic mice together with adenoviral-mediated overexpression of these Pim-1 constructs was used to delineate the role of Pim-1 in hypertrophy. Transgenic overexpression of Pim-1 protects mice from pressure-overload-induced hypertrophy relative to wild-type controls as evidenced by improved hemodynamic function, decreased apoptosis, increases in antihypertrophic proteins, smaller myocyte size, and inhibition of hypertrophic signaling after challenge. Similarly, Pim-1 overexpression in neonatal rat cardiomyocyte cultures inhibits hypertrophy induced by endothelin-1. On the cellular level, hearts of Pim-WT mice show enhanced incorporation of BrdU into myocytes and a hypercellular phenotype compared to wild-type controls after hypertrophic challenge. In comparison, transgenic overexpression of Pim-DN leads to dilated cardiomyopathy characterized by increased apoptosis, fibrosis, and severely depressed cardiac function. Furthermore, overexpression of Pim-DN leads to reduced contractility as evidenced by reduced Ca²⁺ transient amplitude and decreased percentage of cell shortening in isolated myocytes. These data support a pivotal role for Pim-1 in modulation of hypertrophy by impacting responses on molecular, cellular, and organ levels.     

Age-dependent cardiomyopathy and heart failure phenotype in mice overexpressing beta(2)-adrenergic receptors in the heart

OBJECTIVE: To explore long-term cardiac phenotype in transgenic (TG) mice with 300-fold overexpression of beta(2)-adrenergic receptors (AR). METHODS: Echocardiography was performed serially on a cohort of wild-type and TG mice (n=26 each) between 4 and 15 months of age. Survival was monitored and autopsy and histological examinations were performed. RESULTS: Heart rate was higher in TG than in wild-type mice throughout the study period. The left ventricular dimensions and fractional shortening were similar between TG and wild-type groups during 4-6 months. Starting at 9 months, however, TG mice showed progressive reduction in fractional shortening and systolic wall thickening, and increase in left ventricular dimensions and left ventricular mass, indicating onset of heart failure, left ventricular hypertrophy and remodeling. Abnormal waveforms in the electrocardiogram and episodes of ventricular ectopic beats were also observed in TG mice. Death of TG mice started at 8.5 months, and the cumulative mortality was 81% by 15 months (P<0.0001 vs. 4% in wild-type mice). The majority of deaths were due to severe heart failure, indicated by cardiac dilatation, lung congestion, pleural effusion and atrial thrombus. Left ventricular sections showed widespread interstitial fibrosis, loss of myocytes and myocyte hypertrophy in TG mice. CONCLUSIONS: A high level of beta(2)AR overexpression results in cardiomyopathy and heart failure. The onset was slower and the expression levels of receptors required are much higher than previously described for the beta(1)AR overexpression.     

Cardiac stem cell and myocyte aging, heart failure, and insulin-like growth factor-1 overexpression

To determine whether cellular aging leads to a cardiomyopathy and heart failure, markers of cellular senescence, cell death, telomerase activity, telomere integrity, and cell regeneration were measured in myocytes of aging wild-type mice (WT). These parameters were similarly studied in insulin-like growth factor-1 (IGF-1) transgenic mice (TG) because IGF-1 promotes cell growth and survival and may delay cellular aging. Importantly, the consequences of aging on cardiac stem cell (CSC) growth and senescence were evaluated. Gene products implicated in growth arrest and senescence, such as p27Kip1, p53, p16INK4a, and p19ARF, were detected in myocytes of young WT mice, and their expression increased with age. IGF-1 attenuated the levels of these proteins at all ages. Telomerase activity decreased in aging WT myocytes but increased in TG, paralleling the changes in Akt phosphorylation. Reduction in nuclear phospho-Akt and telomerase resulted in telomere shortening and uncapping in WT myocytes. Senescence and death of CSCs increased with age in WT impairing the growth and turnover of cells in the heart. DNA damage and myocyte death exceeded cell formation in old WT, leading to a decreased number of myocytes and heart failure. This did not occur in TG in which CSC-mediated myocyte regeneration compensated for the extent of cell death preventing ventricular dysfunction. IGF-1 enhanced nuclear phospho-Akt and telomerase delaying cellular aging and death. The differential response of TG mice to chronological age may result from preservation of functional CSCs undergoing myocyte commitment. In conclusion, senescence of CSCs and myocytes conditions the development of an aging myopathy.     

Inhibition of protein phosphatase 1 by inhibitor-2 exacerbates progression of cardiac failure in a model with pressure overload

AIMS: The progression of human heart failure is associated with increased protein phosphatase 1 (PP1) activity, which leads to a higher dephosphorylation of cardiac regulatory proteins such as phospholamban. In this study, we tested the hypothesis whether the inhibitor-2 (I-2) of PP1 can mediate cardiac protection by inhibition of PP1 activity. METHODS AND RESULTS: We induced pressure overload by transverse aortic constriction (TAC) for 28 days in transgenic (TG) mice with heart-directed overexpression of a constitutively active form of I-2 (TG(TAC)) and wild-type littermates (WT(TAC)). Both groups were compared with sham-operated mice. TAC treatment resulted in comparable ventricular hypertrophy in both groups. However, TG(TAC) exhibited a higher atrial mass and an enhanced ventricular mRNA expression of beta-myosin heavy chain. The increased afterload was associated with the development of focal fibrosis in TG. Consistent with signs of overt heart failure, fractional shortening and diastolic function were impaired in TG(TAC) as revealed by Doppler echocardiography. The contractility was reduced in catheterized banded TG mice, which is in line with a depressed shortening of isolated myocytes. This is due to profoundly abnormal cytosolic Ca(2+) transients and a reduced stimulation of phosphorylation of phospholamban (PLB)(Ser16) after TAC in TG mice. Moreover, administration of isoproterenol was followed by a blunted contractile response in isolated myocytes of TG(TAC) mice. CONCLUSION: These results suggest that cardiac-specific overexpression of a constitutively active form of I-2 is deleterious for cardiac function under conditions of pressure overload. Thus, the long-term inhibition of PP1 by I-2 is not a therapeutic option in the treatment of heart failure.     

Impaired cell shortening and relengthening with increased pacing frequency are intrinsic to the senescent mouse cardiomyocyte

Increased heart rate enhances cardiac contractility and accelerates relaxation. Both the force- and relaxation-frequency relationships are critical to myocardial function, especially during stress, and have been shown to be impaired in senescent myocardium. While senescent myocardium is characterized by decreased sarcoplasmic reticulum calcium ATPase activity, it is unclear if altered calcium regulation is directly responsible for the attenuated contractility and relaxation observed with increasing pacing frequency in aged myocardium. We examined this issue using freshly dissociated left ventricular myocytes, isolated from young adult and senescent mouse hearts. Myocytes were paced from 2 to 9 Hz at 37 degrees C, and cell shortening and [Ca(2+)](i)were simultaneously measured using video edge-detection and fura-2 fluorescence, respectively. In adult myocytes, increasing the pacing rate resulted in a progressive increase in percent cell shortening (CS) (P<0.01). This positive CS-frequency relationship was paralleled by an increase in [Ca(2+)](i)transient amplitude (P<0.05). In contrast, the CS-frequency relationship was blunted in senescent myocytes with no increase in percent CS or [Ca(2+)](i)transient amplitude with increasing pacing rate. With increased pacing, the decreases in time constants (tau) of cell relengthening and Ca(2+)transient decay were much steeper in adult compared to senescent myocytes (P<0.05). This study demonstrates that adult mouse myocytes exhibit augmented intracellular Ca(2+)transient amplitude and enhanced intracellular Ca(2+)removal with increasing pacing frequency, resulting in increased cell shortening and enhanced relengthening with frequency. In contrast, senescent mouse myocytes exhibit impaired calcium handling with increasing pacing frequency, which correlated with impairment of both cell shortening and relengthening.     

c-Flip overexpression reduces cardiac hypertrophy in response to pressure overload

OBJECTIVE: Activation of Fas signaling has been associated with the development of cardiomyocyte hypertrophy. In the present study, we investigated the effects of increased expression of c-Flip, a natural modulator of Fas receptor signaling, in a mouse model of cardiac growth response to pressure overload. METHODS: A transgenic mouse overexpressing c-Flip in the heart was generated in FVB/N strain. Echocardiographic, hemodynamic, histological and molecular analyses were carried out under basal conditions and after transverse aortic constriction (TAC)-induced pressure overload. RESULTS: Overexpression of c-Flip in ventricular heart tissue was functionally silent under basal conditions affecting neither cardiac morphology nor basal cardiac function. Transgenic mice were then subjected to pressure overload by TAC procedure. Under such conditions, c-Flip transgenic mice showed normal left ventricular function with a significantly reduced left ventricular hypertrophy compared with wild-type mice and reduced induction of the cardiac "fetal" gene programme. Further, analysis of intracellular signaling pathways indicated that c-Flip overexpression reduced phosphorylation of both the glycogen synthase kinase 3beta (GSK3 beta) and Akt as compared with controls. Finally, the reduction of the TAC-induced hypertrophy was not accompanied by significant apoptosis increase. CONCLUSION: Altogether, these findings indicate c-Flip as a key regulator of the cardiac response to ventricular pressure overload.     

Modulation of cardiac contractility by serine/threonine protein phosphatase type 5

Background

Protein phosphatase 5 (PP5) a serine/threonine phosphatase is ubiquitously expressed in mammalian tissues including the heart, but its physiological role in the heart is still unknown. Therefore, we used a transgenic mouse model to get a first insight into the cardiac role of PP5.

Methods and results

We generated transgenic mice with cardiac myocyte specific overexpression of PP5. Successful overexpression of PP5 was demonstrated by Western blotting, immunohistochemistry and enhanced arachidonic acid-stimulated protein phosphatase activity in transgenic hearts. Cardiac function was examined on the level of isolated cardiac myocytes, isolated organs and in intact animals. Whereas Ca²⁺ transients and cell shortening remained unchanged, L-type Ca²⁺ currents were decreased in isolated cardiac myocytes from transgenic mice. Ventricular contractility was reduced in isolated perfused hearts under basal conditions and after β-adrenergic stimulation. In intact animals, echocardiography revealed increased left ventricular diameters and decreased contractility and invasively measured hemodynamic performance by left ventricular catheterization demonstrated a reduced response to β-adrenergic stimulation in transgenic mice compared to wild type.

Conclusions

Overexpression of PP5 affected contractility and β-adrenergic signaling in the hearts of transgenic mice. Taken together, these findings are indicative of a regulatory role of PP5 in cardiac function.     

Morphological and functional alterations in ventricular myocytes from male transgenic mice with hypertrophic cardiomyopathy

Familial hypertrophic cardiomyopathy (FHC) is a human genetic disorder caused by mutations in sarcomeric proteins. It is generally characterized by cardiac hypertrophy, fibrosis, and myocyte disarray. A transgenic mouse model of FHC with mutations in the actin-binding domain of the alpha-myosin heavy chain (MyHC) gene displays many phenotypes similar to human FHC. At 4 months, male transgenic (TG) mice present with concentric cardiac hypertrophy that progresses to dilation with age. Accompanying this latter morphological change is systolic and diastolic dysfunction. Left ventricular (LV) myocytes from male TG and wild-type (WT) littermates at 5 and 12 months of age were isolated and used for morphological and functional studies. Myocytes from 5- and 12-month-old TG animals had shorter sarcomere lengths compared with WT. This sarcomere length difference was abolished in the presence of 2,3-butanedione monoxime, suggesting that the basal level of contractile element activation was increased in TG myocytes. Myocytes from 12-month-old TG mice were significantly longer than those from age-matched WT controls, and TG myocytes exhibited Z-band disorganization. When cells were paced at 0.5 Hz, TG myocyte relengthening and the fall in intracellular [Ca2+] were slowed when compared with cells from age-matched WT controls. Moreover, an increased amount of beta-myosin heavy chain protein was found in hearts from TG compared with WT. Thus, myocytes from the alpha-MyHC TG mouse model display many morphological and functional abnormalities that may help explain the LV dysfunction seen in this TG mouse model of FHC.     

Inhibition of cardiac myocyte contraction by 4-hydroxy-trans-2-nonenal

Lipid peroxidation, initiated by hydroxyl radicals, results in production of 4-Hydroxy-trans-2-nonenal (HNE) and leads to cardiac injury. However, impact of HNE on ventricular function has not been clearly defined. This study was to examine the direct effect of HNE on cardiac contractile function at cardiac myocyte level. Adult male rat ventricular myocytes were isolated and electrically stimulated to contract at 0.5 Hz. Mechanical and intracellular Ca2+ properties were evaluated using an Ionoptix Myocam system. Contractile properties analyzed included peak shortening (PS), time-PS, time-to-90% relengthening, maximal velocities of shortening and relengthening (+/-dL/dt), change of electrically stimulated intracellular Ca2+ fura-2 fluorescent intensity, and intracellular Ca2+ decay. Our results indicated that short-term incubation of HNE (10(-6) to 10(-4) M) with myocytes depressed PS, +/-dL/dt, and fura-2 fluorescent intensity; shortened time-PS; and elevated resting intracellular Ca2+ levels without affecting time-to-90% relengthening and intracellular Ca2+ decay. Interestingly, the HNE-induced cardiac mechanical effects (with the exception of shortened time-PS) were abolished by either the aldehyde dehydrogenase inhibitor cyanamide or the p38 mitogen-activated protein kinase inhibitor SB203580. These findings reveal a possible role of HNE in the lipid peroxidation-associated cardiac contractile dysfunction that is likely mediated through HNE metabolism and mitogen-activated protein kinase activation.