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Given the importance of the aggregation of advanced glycation end products (AGEs) and cardiac inflammation in the onset and progression of diabetic cardiomyopathy (DCM), our objective in this study was to demonstrate the cardioprotective effect of mangiferin, an antidiabetic and anti-inflammatory agent, on diabetic rat model. The DCM model was established by a high-fat diet and a low dose of streptozotocin. DCM rats were treated orally with mangiferin (20 mg/kg) for 16 weeks. Serum and left ventricular myocardium were collected for determination of inflammatory cytokines. AGEs mRNA and protein expression of nuclear factor kappa B (NF-κB) and receptor for AGEs (RAGE) in myocardium were assayed by real-time PCR and Western blot. ROS levels were measured by dihydroethidium fluorescence staining. NF-κB binding activity was assayed by TransAM NF-κB p65 ELISA kit. Chronic treatment with mangiferin decreased the levels of myocardial enzymes (CK-MB, LDH) and inflammatory mediators (TNF-α, IL-1β). Meanwhile, NF-κB is inhibited by the reduction of nuclear translocation of p65 subunit, and mangiferin reduced AGE production and decreased the mRNA and protein expression of RAGE in DCM rats. Our data indicated that mangiferin could significantly ameliorate DCM by preventing the release of inflammatory cytokines, and inhibiting ROS accumulation, AGE/RAGE production, and NF-κB nuclear translocation, suggesting that mangiferin treatment might be beneficial in DCM. Key words: advanced glycation end products, diabetic cardiomyopathy, inflammatory cytokines, NF-κB, mangiferin. Etant donne l'importance du role de l'agregation des produits finaux de glycation avancee (AGE) et de l'inflammation du muscle cardiaque dans l'apparition et revolution de la cardiomyopathie diabetique (CMD), la presente etude avait pour objectif de montrer l'effet cardioprotecteur de la mangiferine, un agent antidiabetique et anti-inflammatoire, dans un modele de rat diabetique. Le modele de CMD etait obtenu en administrant un regime a haute teneur en matieres grasses et de la streptozotocine a faible dose. Nous avons traite les rats atteints de CMD en administrant de la mangiferine (20 mg/kg) par voie orale pendant 16 semaines. Nous avons ensuite preleve du tissu myocardique du ventricule gauche et du serum en vue de doser certaines cytokines inflammatoires. A l'aide des techniques d'amplification en chaine par polymerase (PCR) en temps et reel et d'electrobuvardage de Western, nous avons mesure l'expression des AGE (ARNm et proteines) induite par l'activation du facteur nucleaire kappa (NF-κB) et les taux de recepteurs des AGE (RAGE) dans le tissu myocardique. Nous avons aussi quantifie les taux de derives reactifs de l'oxygene (ROS) a l'aide de la technique de coloration fluorescente par le dihydroethidium et mesure l'activite de la liaison du NF-κB a l'aide du kit ELISA (essai immuno- enzymatique) TransAM NF-κB p65. Nous avons observe que l'administration chronique de mangiferine permettait d'abaisser les taux d'enzymes myocardiques (CK-MB et LDH) et de mediateurs inflammatoires (TNF-α et IL-1β). Tandis que l'effet du NF- κB etait inhibe par la diminution de la translocation nucleaire de la sous-unite p65, et l'administration de mangiferine permettait de diminuer la production des AGE ainsi que l'expression des RAGE (ARNm et proteines) chez les rats atteints de CMD. En somme, nos resultats laissent entendre que la mangiferine pourrait permettre de contrecarrer la CMD de facon appreciable par les mecanismes suivants : inhibition de la liberation de cytokines inflammatoires, reduction de l'accumulation de ROS, reduction de la production d'AGE et de RAGE et reduction de la translocation nucleaire du NF-κB. Il est donc possible que l'administration de mangiferine puisse etre utile dans le traitement de la CMD. [Traduit par la Redaction] Mots-cles: produits finaux de glycation avancee, cardiomyopathie diabetique, cytokines inflammatoires, NF-κB, mangiferine., Introduction Diabetic cardiomyopathy (DCM), as an early complication of diabetes, is considered a risk factor for acute myocardial infarction and death due to coronary artery disease. It carries increased mortality [...]

Using the UM-X7.1 hereditary cardiomyopathic and muscular dystrophy hamsters (HCMH), we tested the effects of lifelong preventive or curative treatments during the heart failure phase with the NHE-1 inhibitor EMD 87580 (EMD) or with the angiotensin-converting enzyme inhibitor cilazapril on the intracellular [Na.sup.+] and [Ca.sup.2+] overloads, elevated level of NHE-1, necrosis, hypertrophy, heart failure, and early death. Our results showed that 310-day pretreatment of 30-day-old HCMHs with EMD significantly prevented cardiac necrosis, cardiomyocyte hypertrophy, and reduced the heart to body mass ratio. This treatment significantly prevented [Na.sup.+] and [Ca.sup.2+] overloads and the increase in NHE-1 protein level observed in HCMHs. Importantly, this lifelong preventive treatment significantly decreased the levels of creatine kinase and prevented early death of HCMHs. Curative 0treatment of hypertrophic 275-day-old HCMHs for 85 days with EMD significantly prevented hypertrophy and early death of HCMHs. However, treatments with cilazapril did not have any significant effects on the cardiac parameters studied or on early death of HCMHs. Our results suggest that the increase in the NHE-1 level and the consequent [Na.sup.+] and [Ca.sup.2+] overloads are implicated in the pathological process leading to heart failure and early death in HCMHs, and treatment with the NHE-1 inhibitor is promising for preventing early death in hereditary cardiomyopathy. Key words: cardiomyopathy, early death, heart failure, NHE-1, NHE-1 inhibitor EMD 87580, ACE inhibitor. A l'aide d'un modele de hamster atteint de cardiomyopathie et de dystrophie musculaire hereditaires (souche UM-X7.1), nous avons etudie les effets de traitements preventifs a vie et de traitements curatifs pendant la phase d'insuffisance cardiaque par l'administration de l'EMD 87580, un inhibiteur de la NHE-1 (isoforme 1 de la proteine echangeuse d'ions sodium-proton), ou de cilazapril, un IECA (inhibiteur de l'enzyme de conversion de l'angiotensine), sur l'etablissement de surcharges intracellulaires en [Na.sup.+] et en [Ca.sup.2+], l'augmentation des taux de NHE-1, la necrose, l'hypertrophie, l'insuffisance cardiaque et la mort prematuree. Nos resultats ont montre qu'un pretraitement des hamsters UM-X7.1 par l'EMD 87580 pendant 310 jours permettait manifestement de prevenir la necrose cardiaque, l'hypertrophie des cardiomyocytes et l'augmentation du rapport entre le poids du creur et le poids corporel total. Ce traitement permettait de prevenir l'etablissement de surcharges en [Na.sup.+] et en [Ca.sup.2+] et d'augmenter les taux de proteine NHE-1 observes dans ce modele. Mais avant tout, administre a vie de maniere preventive, ce traitement permettait de diminuer les taux de creatine kinase et de prevenir la mort prematuree de maniere importante chez les hamsters UM-X7.1. Un traitement curatif par l'EMD 87580 pendant 85 jours administre a des hamsters UM-X7.1ages de 275 jours permettait de prevenir l'hypertrophie et la mort prematuree dans ce modele. Cependant, les traitements par le cilazapril n'avaient aucun effet notable sur les parametres cardiaques etudies ni sur la mort prematuree chez les hamsters UM-X7.1. Nos resultats indiquent que l'augmentation des taux de NHE-1 et les surcharges en [Na.sup.+] et en [Ca.sup.2+] qui en decoulent joueraient un role dans les processus pathologiques entrainant l'insuffisance cardiaque et la mort prematuree chez les hamsters UM-X7.1. En fait, l'administration d'inhibiteurs de la NHE-1 pourrait etre une avenue therapeutique prometteuse pour la prevention de la mort prematuree en cas de cardiomyopathie hereditaire. [Traduit par la Redaction] Mots-cles: cardiomyopathie, mort prematuree, insuffisance cardiaque, NHE-1, inhibiteur de la NHE-1 EMD 87580, IECA., Introduction An important characteristic of several types of hypertrophy and congestive heart failure (HF) is a disturbed intracellular calcium ([[[Ca.sup.2+]].sub.i]) handling in heart muscles (Jasmin and Proschek 1984, 1994; Bkaily [...]

Type 1 diabetes is associated with a unique form of cardiomyopathy that is present without atherosclerosis. Redox imbalance and/or changes in vascular endothelial growth factor (VEGF) expression have been associated with diabetes-related cardiomyopathy. However, the mechanisms of these changes and their interrelationships remain unclear. Using a murine type 1 diabetes model, we tested the hypothesis that alterations in cardiac performance are associated with decreased cardiac microvascular prevalence, as well as downregulation of VEGF isoforms. We also investigated oxidative stress as a contributor to regulate individual VEGF isoforms and microvascular rarefaction. Significant and rapid hyperglycemia was observed at 1 wk post-streptozotocin (STZ) and persisted throughout the 5-wk study. Left ventricular (LV) fractional shortening was reduced at week 1 and 5 post-STZ insult relative to age-matched controls. We also observed the early reduction in E/A ratio at 1 wk. Immunostaining for CD31 and digital image analysis demonstrated a 35% reduction in microvessels/myocardial area, indicative of rarefaction, which was highly correlated with fractional shortening. Furthermore, a significant increase in the prevalence of protein 3-nitrotyrosine was observed in the diabetic cardiac tissue, which was inversely associated with microvascular rarefaction. The expressions of three VEGF isoforms were significantly reduced to different extents. The reduction of VEGF164 was associated with GSSG accumulation. These data demonstrate that the mouse model of STZ-induced diabetes has hallmark features observed in humans with respect to nonischemic systolic and diastolic performance and microvascular rarefaction, which are associated with changes in VEGF isoform expression and redox imbalance in the myocardium. microvascular density; vascular endothelial growth factor; cardiomyopathy; oxidants

Transgenic mice were generated to express a restrictive cardiomyopathy (RCM) human cardiac troponin I (cTnI) R192H mutation in the heart ([cTnI.sup.193His] mice). The objective of this study was to assess cardiac function during the development of diastolic dysfunction and to gain insight into the pathophysiological impact of the RCM cTnI mutation. Cardiac function and pathophysiological changes were monitored in [cTnI.sup.193His] mice and wild-type littermates for a period of 12 too. It progressed gradually from abnormal relaxation to diastolic dysfunction characterized with high-resolution echocardiography by a reversed E-to-A ratio, increased deceleration time, and prolonged isovolumetric relaxation time. At the age of 12 mo, cardiac output in [cTnI.sup.193His] mice was significantly declined, and some transgenic mice showed congestive heart failure. The negative impact of [cTnI.sup.193His] on ventricular contraction and relaxation was further demonstrated in isolated mouse working heart preparations. The main morphological change in [cTnI.sup.193His] myocytes was shortened cell length. Dobutamine stimulation increased heart rate in [cTnI.sup.193His] mice but did not improve CO. The [cTnI.sup.193His] mice had a phenotype similar to that in human RCM patients carrying the cTnI mutation characterized morphologically by enlarged atria and restricted ventricles and functionally by diastolic dysfunction and diastolic heart failure. The results demonstrate a critical role of the COOH-terminal domain of cTnI in the diastolic function of cardiac muscle. troponin I; cardiac relaxation: Doppler echocardiography; working heart function

We have previously shown that mouse embryonic stem (ES) cells transplanted following myocardial infarction (MI) differentiate into the major cell types in the heart and improve cardiac function. However, the extent of regeneration was relatively meager compared with the observed functional improvement. Therefore, we hypothesize that mechanisms in addition to regeneration contribute to the functional improvement from ES cell therapy. In this study, we examined the effect of mouse ES cells transplanted post-MI on cardiac apoptosis, fibrosis, and hypertrophy. MI was produced by left coronary artery ligation in C57BL/6 mice. Two different mouse ES cell lines, expressing enhanced green fluorescent protein and [beta]-galactosidase, respectively, were tested. Post-MI intramyocardial injection of 3 x [10.sup.4] ES cells was compared with injection of medium alone. Terminal deoxynucleotidyl nick end labeling (TUNEL), immunofluorescence, and histology were used to examine the effect of transplanted ES cells on apoptosis, fibrosis, and hypertrophy. Two weeks post-MI, ES cell-transplanted hearts exhibited a significant decrease in TUNEL-stained nuclei (mean [+ or -] SE; MI+medium = 12 [+ or -] 1.5%; MI+ES cells = 6.6 [+ or -] 1%, P < 0.05). TUNEL-positive nuclei were confirmed to be apoptotic by colabeling with a caspase-3 antibody. Cardiac fibrosis was 57% less in the MI+ES cell group compared with the MI + medium group (P < 0.05) as shown with Masson's trichrome staining. Picrosirius red staining confirmed a decreased amount of collagen present in the MI+ES cell group. Cardiomyocyte hypertrophy was significantly decreased following ES cell transplantation compared with medium control animals. In conclusion, transplanted mouse ES cells in the infarcted heart inhibit apoptosis, fibrosis, and hypertrophy, thereby reducing adverse remodeling. cardiomyocytes; cell therapy; fibrosis; TIMP-1

Familial hypertrophic cardiomyopathy (FHC) is a disease caused by mutations in contractile proteins of the sarcomere. Our laboratory developed a mouse model of FHC with a mutation in the thin filament protein [alpha]-tropomyosin (TM) at amino acid 180 (Glul80Gly). The hearts of these mice exhibit dramatic systolic and diastolic dysfunction, and their myofilaments demonstrate increased calcium sensitivity. The mice also develop severe cardiac hypertrophy, with death ensuing by 6 mo. In an attempt to normalize calcium sensitivity in the cardiomyofilaments of the hypertrophic mice, we generated a chimeric [alpha]-/[beta]-TM protein that decreases calcium sensitivity in transgenic mouse cardiac myofilaments. By mating mice from these two models together, we tested the hypothesis that an attenuation of myofilament calcium sensitivity would modulate the severe physiological and pathological consequences of the FHC mutation. These double-transgenic mice 'rescue' the hypertrophic phenotype by exhibiting a normal morphology with no pathological abnormalities. Physiological analyses of these rescued mice show improved cardiac function and normal myofilament calcium sensitivity. These results demonstrate that alterations in calcium response by modification of contractile proteins can prevent the pathological and physiological effects of this disease. hypertrophy; contractile function; genetically altered mice; calcium sensitivity

I. Introduction II. Overview of Myocardial Substrate Metabolism A. Regulation of metabolic pathways in the heart B. Carbohydrate metabolism C. Fatty acid metabolism D. Ketone body metabolism E. Interregulation of [...]

Hearts with overexpression of anchored lipoprotein lipase (LpL) by cardiomyocytes (hLp[L.sup.GPI] mice) develop a lipotoxic cardiomyopathy. To characterize cardiac fatty acid (FA) and triglyceride (TG) metabolism in these mice and to determine whether changes in lipid metabolism precede cardiac dysfunction, hearts from young mice were perfused in Langendorff mode with [[sup.14]C]palmitate. In hLp[L.sup.GPI] hearts, FA uptake and oxidation were decreased by 59 and 82%, respectively. This suggests reliance on an alternative energy source, such as TG. Indeed, these hearts oxidized 88% more TG. Hearts from young hLp[L.sup.GPI] mice also had greater uptake of intravenously injected cholesteryl ester-labeled Intralipid and VLDL. To determine whether perfusion of normal hearts would mimic the metabolic alterations found in hLp[L.sup.GPI] mouse hearts, wild-type hearts were perfused with [[sup.14]C] palmitate and either human VLDL or Intralipid (0.4 mM TG). Both sources of TG reduced [[sup.14]C] palmitate uptake (48% with VLDL and 45% with Intralipid) and FA oxidation (71% with VLDL and 65% with Intralipid). Addition of either heparin or LpL inhibitor P407 to Intralipid-containing perfusate restored [[14.sup.C]]palmitate uptake and confirmed that Intralipid inhibition requires local LpL. Our data demonstrate that reduced FA uptake and oxidation occur before mechanical dysfunction in hLp[L.sup.GPI] lipotoxicity. This physiology is reproduced with perfusion of hearts with TG-containing particles. Together, the results demonstrate that cardiac uptake of TG-derived FA reduces utilization of albumin-FA. lipotoxicity; triglyceride; fatty acid metabolism; lipoprotein lipase

Myocardial cell death is an important contributor to the development of diabetic cardiomyopathy. It has been proposed that diabetes-mediated upregulation of the renin-angiotensin system leads to oxidative stress, the trigger for cardiomyocyte death and contractile dysfunction. However, the adverse effect of ANG II on the diabetic heart may extend beyond the development of the cardiomyopathy. ANG II also alters specific modulators of ischemic injury, such as PKC and calcium transport. Therefore, the present study examined the effect of ANG II on hyperglycemic preconditioning, a glucose-mediated condition associated with the elevation of PKC activity and alterations in calcium transport that render the cell resistant to hypoxia. Exposure of the glucose-treated cell to ANG II during the prehypoxic period blocked glucose-mediated cardioprotection. The reversal of hyperglycemic preconditioning was associated with enhanced accumulation of [Ca.sup.2+] during hypoxia, an effect prevented by inhibition of the [Na.sup.+]/[H.sup.+] exchanger and the T-type [Ca.sup.2+] channel. The inhibitors of hypoxiamediated [Ca.sup.2+] accumulation also blocked the reversal of hyperglycemic preconditioning by ANG II. Thus ANG II and glucose treatment exert opposite actions on the [Na.sup.+]/[H.sup.+] exchanger and the T-type [Ca.sup.2+] channel. Because those transporters are involved in hypoxiamediated apoptosis, they are logical candidates for the beneficial effects of high glucose and the adverse effects of ANG II on the hypoxic cardiomyocyte. sodium-hydrogen exchanger; T-type calcium channel; protein kinase C; hyperglycemia; hypoxia; apoptosis; calcium overload

The current study investigated the phosphorylation of mitogen-activated protein kinases (MAPKs) as well as pro- and anti-apoptotic proteins in adriamycin (ADR)-induced cardiomyopathy (AIC) and heart failure in rats. Modulatory effects of antioxidant probucol on the activation of MAPKs were also examined. Male rats were administered with ADR (15 mg/kg body wt ip, over 2 wk) with and without probucol (120 mg/kg body wt for 4 wk ip). Hearts from these animals were studied at 1- to 24-h as well as at 3-wk posttreatment durations. In the 3-wk group, ADR depressed cardiac function, increased left ventricular end-diastolic pressure (LVEDP), and caused dyspnea and mortality. These changes were prevented by probucol. Phosphorylation of extracellular signal-regulated kinase (ERK) 1/2, in the early stage of AIC, showed a biphasic response, with a maximum increase to 513% seen at 4 h, followed by a decrease to 66.8% at 3 wk after the last injection of ADR. Phosphorylation of p38 and c-Jun N[H.sub.2]-terminal kinases (JNKs) showed a steady increase through 2, 4, and 24 h and 3 wk (116% to 148%). In gene microarray analysis at 3 wk (heart failure stage), mRNA expression for both ERK1/2 and p38 kinases was decreased, whereas JNK mRNA was undetectable. Probucol completely prevented these MAPK changes. Activation of caspase-3 as well as the increase in the ratio of Bax to Bcl-xl were seen at early time points (1-24 h) as well as in the heart failure stage (3 wk). It is suggested that a transient increase in ERK1/2 at a shorter interval indicate an early adaptive response, and failure of this response corresponded with heart failure. In contrast, a gradual and persistent increase in p38 and JNK MAPKs as well as in caspase-3 and the Bax-to-Bcl-xl ratio may contribute in the initiation of apoptosis and progression of heart failure. Because probucol modulated changes in cellular signaling pathways and cardiac function, it is likely that oxidative stress plays a key role in AIC and heart failure. dilated cardiomyopathy; oxidative stress and apoptosis

Chagasic (Ch) and nonchagasic (NCh) IgG fraction (20 [micro]g/ml) effects on cardiac performance of adult Wistar rat ventricles were studied with a novel approach applying a microcalorimetric technique. Resting heat ([H.sub.r]) was significantly decreased by Ch antibodies ([DELTA][H.sub.rCh] = 4.8 [+ or -] 0.9 mW/g). Although the [H.sub.r] decrease can be associated with diminished activity of the [Na.sup.+]/[K.sup.+] pump, the magnitude of the effect (25% of control [H.sub.r]) indicates that additional processes may also be affected. Ch antibodies induced an initial increase in developed pressure (P), which was associated with a decreased contractile economy. However, after 30 min of Ch antibody perfusion, P reached a significantly lower level ([DELTA][P.sub.Ch] = 3.8 [+ or -] 1.2 mN/[mm.sup.2]) without changes in active heat per beat ([H'.sub.a]). Consequently, [H'.sub.A]/P ratio increased, indicating that the energetic cost per unit of P was higher. In contrast, P and [H'.sub.a] were both significantly and reversibly decreased by NCh antibodies ([DELTA][P.sub.NCh] = 4.4 [+ or -] 1.2 mN/[mm.sup.2]; [DELTA][H'.sub.aNCh] = 9.7 [+ or -] 2.2 mJ/g), but [H'.sub.a]/P remained unaffected. According to these data, normal hearts exposed to Ch antibodies present a biphasic mechanical response: 1) an initial period of increased contractility (and decreased global muscle economy) consistent with antibodies with [[beta].sub.1]-adrenergic activity, such as those used in the present study, and 2) a decrease in P at 30 min of Ch antibody perfusion, which suggests that another [Ca.sup.2+]-related mechanism is compromised. These data contribute to redefine the role of antibody-mediated responses in the pathophysiology of chronic chagasic cardiomyopathy as agents of myocardial failure. cardiomyopathy; energy metabolism; ventricular function; muscle economy; excitation-contraction coupling

The P2[X.sub.4] purinergic receptor (P2[X.sub.4]R) is a ligand-gated ion channel. Its activation by extracellular ATP results in [Ca.sub.2+] influx. Transgenic cardiac overexpression of the human P2[X.sub.4] receptor showed an in vitro phenotype of enhanced basal contractility. The objective here was to determine the in vivo cardiac physiological role of this receptor. Specifically, we tested the hypothesis that this receptor plays an important role in modulating heart failure progression. Transgenic cardiac overexpression of canine calsequestrin (CSQ) showed hypertrophy, heart failure, and premature death. Crossing the P2[X.sub.4]R mouse with the CSQ mouse more than doubled the lifespan (182 [+ or -] 91 days for the binary CSQ/P2[X.sub.4]R mouse, n = 35) of the CSQ mouse (71.3 [+ or -] 25.4 days, n = 50, P < 0.0001). The prolonged survival in the binary CSQ/P2[X.sub.4]R mouse was associated with an improved left ventricular weight-to-body weight ratio and a restored [beta]-adrenergic responsiveness. The beneficial phenotype of the binary mouse was not associated with any downregulation of the CSQ level but correlated with improved left ventricular developed pressure and [+ or -]dP/dt. The enhanced cardiac performance was manifested in young binary animals and persisted in older animals. The increased contractility likely underlies the survival benefit from P2[X.sub.4] receptor overexpression. An increased expression or activation of this receptor may represent a new approach in the therapy of heart failure. cardiac failure; contraction; isoproterenol; purines; adenine nucleotide

In contrast to skeletal muscle isoforms of myosin binding protein C (MyBP-C), the cardiac isoform has 11 rather than 10 fibronectin or Ig modules (modules are identified as CO to C10, N[H.sub.2] to COOH terminus), 3 phosphorylation sites between modules C1 and C2, and 28 additional amino acids rich in proline in C5. Phosphorylation between C1 and C2 increases maximum Ca-activated force (Fmax), alters thick filament structure, and increases the probability of myosin heads on the thick filament binding to actin on the thin filament. Unphosphorylated C1C2 fragment binds to myosin, but phosphorylation inhibits the binding. MyBP-C also binds to actin. Using two types of immunoprecipitation and cosedimentation, we show that fragments of MyBP-C containing CO bind to actin. In low concentrations CO-containing fragments bind to skinned fibers when the N[H.sub.2] terminus of endogenous MyBP-C is bound to myosin, but not when MyBP-C is bound to actin. C1C2 fragments bind to skinned fibers when endogenous MyBP-C is bound to actin but not to myosin. Disruption of interactions of endogenous CO with a high concentration of added COC2 fragments produces the same effect on contractility as extraction of MyBP-C, namely decrease in Fmax and increase in Ca sensitivity. These results suggest that cardiac contractility can be regulated by shifting the binding of the N[H.sub.2] terminus of MyBP-C between actin and myosin. This mechanism may have an effect on diastolic filling of the heart. KEY WORDS: C1C2 * CO * cardiomyopathy * relaxation * phosphorylation

Byline: Matthias Behrends (1), Jurgen Peters (1) Keywords: Sepsis; Cardiomyopathy; Myocardial function; Contractility; Calcium sensitivity; Langendorff Abstract: Objective Sepsis-evoked myocardial dysfunction is possibly due to decreased myofilament calcium sensitivity, and a calcium sensitizer may thus specifically improve contractility in sepsis by enhancing myofilament calcium sensitivity. We examined whether the calcium sensitizer levosimendan mitigates myocardial dysfunction and improves contractility in hearts isolated from endotoxin-treated guinea pigs. Design and setting Prospective, controlled, randomized animal study in a university research laboratory. Subjects Guinea pig hearts isolated 4 h (n=10) or 18 h (n=8) following E. coli LPS (4 mg/kg i.p.) and hearts from sham-treated controls (n=11 and n=6). Interventions Isolated hearts were perfused at constant aortic pressure [Krebs-Henseleit buffer, heart rate: 300/min, left ventricular (LV) diastolic pressure: 6--8 mmHg], and LV developed pressure (LVdP) and LVdP/dt were continuously assessed. Levosimendan was added to the perfusate in incremental concentrations (0.03, 0.1, 0.3 AuM). Measurements and results Endotoxin resulted in a significant decrease in LVdP by 20+-6% and 43+-8%, in +LVdP/dt by 16+-5% and 44+-7%, and in -LVdP/dt by 27+-8% and 47+-8% after 4 and 18 h, respectively. In septic hearts levosimendan increased LV function concentration-dependently by 32+-4% (LVdP), 33+-5% (+LVdP/dt), and 37+-7% (-LVdP/dt) 4 h and by 31+-6% (LVdP), 33+-6% (+LVdP/dt), and 32+-7% (-LVdP/dt) 18 h after LPS. However, levosimendan increased myocardial function similarly in control hearts. Conclusions While the calcium sensitizer levosimendan markedly improved LV contractility in hearts from both endotoxic and sham animals, it failed to specifically abolish endotoxin-evoked myocardial dysfunction. Thus, decreased calcium sensitivity either does not play a major role in endotoxin-evoked cardiomyopathy or the location of its pathomechanism differs from levosimendan's site of action. Author Affiliation: (1) Klinik fur Anasthesiologie und Intensivmedizin, Universitatsklinikum Essen, Hufelandstrasse 55, 45122, Essen, Germany Article History: Received Date: 12/12/2002 Accepted Date: 27/05/2003 Online Date: 17/07/2003

To date, no study has assessed the degree of similarity between left ventricular (LV) reverse remodeling and atrophic remodeling. Stable LV hypertrophy was induced by creation of an arteriovenous fistula (AVF) in Lewis rats (32 days). LV unloading was induced by heterotopic transplantation of normal (NL-HT) and/or hypertrophic (AVF-HT) hearts (7 days). We compared indexes of remodeling in AVF, NL-HT, and AVF-HT groups with those of normal controls. LV unloading induced decreases in cardiomyocyte size in NL-HT and AVF-HT hearts. NL-HT and AVF-HT LV were both characterized by relative increases in collagen concentration that were largely a reflection of decreases in myocyte volume. NL-HT and AVF-HT LV were associated with similar increases in matrix metalloproteinase (MMP-2 and -9) zymographic activity, without change in the abundance of the tissue inhibitors of the MMPs. In contrast, AVF-HT, but not NL-HT, was associated with a dramatic increase in collagen cross-linking. Our findings suggest an overall similarity in the response of the normal and hypertrophic LV to surgical unloading. However, the dramatic increase in collagen cross-linking after just 1 wk of unloading suggests a potential difference in the dynamics of collagen metabolism between the two models. Further studies will be required to determine the precise molecular mechanisms responsible for these differences in extracellular matrix regulation. However, with respect to these and related issues, heterotopic transplantation of hypertrophied hearts will be a useful small animal model for defining mechanisms of myocyte-matrix interactions during decreased loading conditions. myocardial atrophy; reverse remodeling; extracellular matrix; volume overload

Both acute coronary occlusion and reperfusion of an infarct-related artery lead to significant myocardial cell death. Recent evidence has been presented that activation of the transcription factor nuclear factor-[kappa]B (NF-[kappa]B) plays a critical role in reperfusion injury. NF-[kappa]B is usually bound to its inhibitor, I[kappa]B, and classic activation of NF-[kappa]B occurs when the 20S proteasome degrades I[kappa]B that has been phosphorylated and ubiquitinated. In this study, activation of NF-[kappa]B was inhibited by systemic administration of a 20S proteasome inhibitor (PS-519) in a porcine model of myocardial reperfusion injury. The experimental protocol induced myocardial ischemia in the distribution of the left anterior descending coronary artery for 1 h with subsequent reperfusion for 3 h. A single systemic treatment with PS-519 reduced 20S proteasome activity; blocked activation of NF-[kappa]B induced by reperfusion; reduced creatine kinase, creatine kinase-muscle-brain fraction, and troponin I release from the myocardium; preserved regional myocardial function measured by segmental shortening; significantly reduced the size of myocardial infarction; and exhibited no acute toxicity. These data show that myocardial reperfusion injury can be inhibited by using proteasome inhibitors, which likely function through the inhibition of NF-[kappa]B activation. inflammation; myocardial contraction; multienzyme complexes; transcription factors

Hydrolysis of triacylglycerols (TG) in circulating chylomicrons by endothelium-bound lipoprotein lipase (LPL) provides a source of fatty acids (FA) for cardiac metabolism. The effect of diabetes on the metabolism of chylomicrons by perfused mouse hearts was investigated with db/db (type 2) and streptozotocin (STZ)-treated (type 1) diabetic mice. Endothelium-bound heparin-releasable LPL activity was unchanged in both type 1 and type 2 diabetic hearts. The metabolism of LPL-derived FA was examined by perfusing hearts with chylomicrons containing radiolabeled TG and by measuring [sup.3][H.sub.2]O accumulation in the perfusate (oxidation) and incorporation of radioactivity into tissue TG (esterification). Rates of LPL-derived FA oxidation and esterification were increased 2.3-fold and 1.7-fold in db/db hearts. Similarly, LPL-derived FA oxidation and esterification were increased 3.4-fold and 2.5-fold, respectively, in perfused hearts from STZ-treated mice. The oxidation and esterification of [[sup.3]H]palmitate complexed to albumin were also increased in type 1 and type 2 diabetic hearts. Therefore, diabetes may not influence the supply of LPL-derived FA, but total FA utilization (oxidation and esterification) was enhanced. diabetic cardiomyopathy; lipoprotein lipase; fatty acid metabolism

The transcription factor nuclear factor-[kappa]B (NF-[kappa]B) regulates expression of a variety of genes involved in immune responses, inflammation, proliferation, and programmed cell death (apoptosis). Here, we show that in rat neonatal ventricular cardiomyocytes, activation of NF-[kappa]B is involved in the hypertrophic response induced by myotrophin, a hypertrophic activator identified from spontaneously hypertensive rat heart and cardiomyopathic human hearts. Myotrophin treatment stimulated NF-[kappa]B nuclear translocation and transcriptional activity, accompanied by I[kappa]B-[alpha] phosphorylation and degradation. Consistently, myotrophin-induced NF-[kappa]B activation was enhanced by wild-type I[kappa]B kinase (IKK) [beta] and abolished by the dominant-negative IKK[beta] or a general PKC inhibitor, calphostin C. Importantly, myotrophin-induced expression of two hypertrophic genes (atrial natriuretic factor [ANF] and c-myc) and also enhanced protein synthesis were partially inhibited by a potent NF-[kappa]B inhibitor, pyrrolidine dithio-carbamate (PDTC), and calphostin C. Expression of the dominantnegative form of I[kappa]B-[alpha] or IKK[beta] also partially inhibited the transcriptional activity of ANF induced by myotrophin. These findings suggest that the PKC-IKK-NF-[kappa]B pathway may play a critical role in mediating the myotrophin-induced hypertrophic response in cardiomyocytes.

We investigated 18 AIDS hearts (5 with and 13 without cardiomyopathy) by using immunocytochemistry and computerized image analysis regarding the roles of HIV-1 proteins and tumor necrosis factor ligands in HIV cardiomyopathy (HIVCM). HIVCM and cardiomyocyte apoptosis were significantly related to each other and to the expression by inflammatory cells of gp120 and tumor necrosis factor-[alpha]. In HIVCM heart, active caspase 9, a component of the mitochondrion-controlled apoptotic pathway, and the elements of the death receptor-mediated pathway, tumor necrosis factor-[alpha] and Fas ligand, were expressed strongly on macrophages and weakly on cardiomyocytes. HIVCM showed significantly greater macrophage infiltration and cardiomyocyte apoptosis rate compared with non-HIVCM. HIV-1 entered cultured neonatal rat ventricular myocytes by macropinocytosis but did not replicate. HIV-1- or gp120-induced apoptosis of rat myocytes through a mitochondrion-controlled pathway, which was inhibited by heparin, AOPRANTES, or pertussis toxin, suggesting that cardiomyocyte apoptosis is induced by signaling through chemokine receptors. In conclusion, in patients with HIVCM, cardiomyocytes die through both mitochondrion- and death receptor-controlled apoptotic pathways.

Alcoholic cardiomyopathy is characterized by impaired ventricular function although its toxic mechanism is unclear. This study examined the impact of cardiac overexpression of alcohol dehydrogenase (ADH), which oxidizes ethanol into acetaldehyde (ACA), on ethanol-induced cardiac contractile defect. Mechanical and intracellular [Ca.sup.2+] properties were evaluated in ventricular myocytes from ADH transgenic and wild-type (FVB) mice. ACA production was assessed by gas chromatography. ADH myocytes exhibited similar mechanical properties but a higher efficiency to convert ACA compared with FVB myocytes. Acute exposure to ethanol depressed cell shortening and intracellular [Ca.sup.2+] in the FVB group with maximal inhibitions of 23.3% and 23.4%, respectively. Strikingly, the ethanol-induced depression on cell shortening and intracellular [Ca.sup.2+] was significantly augmented in the ADH group, with maximal inhibitions of 43.7% and 40.6%, respectively. Pretreatment with the ADH inhibitor 4-methylpyrazole (4-MP) or the aldehyde dehydrogenase inhibitor cyanamide prevented or augmented the ethanol-induced inhibition, respectively, in the ADH but not the FVB group. The ADH transgene also substantiated the ethanol-induced inhibition of maximal velocity of shortening/relengthening and unmasked an ethanol-induced prolongation of the duration of shortening/relengthening, which was abolished by 4-MP. These data suggest that elevated cardiac ACA exposure due to enhanced ADH expression may play an important role in the development of alcoholic cardiomyopathy. transgene; acetaldehyde; shortening; intracellular calcium ion transient

The transthyretin (TTR) amyloid diseases are of keen interest, because there are >80 mutations that cause, and a few mutations that suppress, disease. The V122I variant is the most common amyloidogenic mutation worldwide, producing familial amyloidotic cardiomyopathy primarily in individuals of African descent. The substitution shifts the tetramer-folded monomer equilibrium toward monomer (lowers tetramer stability) and lowers the kinetic barrier associated with rate-limiting tetramer dissociation (pH 7; relative to wild-type TTR) required for amyloid fibril formation. Fibril formation is also accelerated because the folded monomer resulting from the tetramer-folded monomer equilibrium rapidly undergoes partial denaturation and self-assembles into amyloid (in vitro) when subjected to a mild denaturation stress (e.g., pH 4.8). Incorporation of the V122I mutation into a folded monomeric variant of transthyretin reveals that this mutation does not destabilize the tertiary structure or alter the rate of amyloidogenesis relative to the wild-type monomer. The increase in the velocity of rate-limiting tetramer dissociation coupled with the lowered tetramer stability (increasing the mol fraction of folded monomer present at equilibrium) may explain why V122I confers an apparent absolute anatomic risk for cardiac amyloid deposition.

Nikolaidis, Lazaros A., Teresa Hentosz, Aaron Doverspike, Rhonda Huerbin, Carol Stolarski, You-Tang Shen, and Richard P. Shannon. Mechanisms whereby rapid RV pacing causes LV dysfunction: perfusion-contraction matching and NO. Am J Physiol Heart Circ Physiol 281: H2270-H2281, 2001.--Incessant tachycardia induces dilated cardiomyopathy in humans and experimental models; mechanisms are incompletely understood. We hypothesized that excessive chronotropic demands require compensatory contractility reductions to balance metabolic requirements. We studied 24 conscious dogs during rapid right ventricular (RV) pacing over 4 wk. We measured hemodynamic, coronary blood flow (CBF), myocardial [O.sub.2] consumption (M[Vo.sub.2]) responses, myocardial nitric oxide (NO) production, and substrate utilization. Early pacing (6 h) resulted in decreased heart rate (HR)-adjusted coronary blood flow (CBF), M[Vo.sub.2] (CBF/beat: 0.33 [+ or -] 0.02 to 0.19 [+ or -] 0.01 ml, P < 0.001, M[Vo.sub.2]/ beat: 0.031 [+ or -] 0.002 to 0.016 [+ or -] 0.001 ml [O.sub.2], P < 0.001), and contractility [left ventricular (LV) first derivative pressure (dP/dt)/LV end-diastolic diameter (EDD): 65 [+ or -] 4 to 44 [+ or -] 3 mmHg.[s.sup.-l].m[m.sup.-1], P < 0.01], consistent with flow-metabolism-function coupling, which persisted over the first 72 h of pacing (CBF/beat: 0.15 [+ or -] 0.01 ml, M[Vo.sub.2]/beat: 0.013 [+ or -] 0.001 ml [O.sub.2], P < 0.001). Thereafter, CBF per beat and M[Vo.sub.2] per beat increased (CBF/beat: 0.25 [+ or -] 0.01 ml, M[Vo.sub.2]/beat: 0.021 [+ or -] 0.001 ml [O.sub.2] at 28 days, P < 0.01 vs. 72 h). Contractility declined [(LV dP/dt)/LVEDD: 19 [+ or -] 2 mmHg.[s.sup.-1].m[m.sup.-1], P < 0.0001], signifying flow-function mismatch. Cardiac NO production, endothelial NO synthase expression, and fatty acid utilization decreased in late phase, whereas glycogen content and lactate uptake increased. Incessant tachycardia induces contractile, metabolic, and flow abnormalities reflecting flow-function matching early, but progresses to LV dysfunction late, despite restoration of flow and metabolism. The shift to flow-function mismatch is associated with impaired myocardial NO production. nitric oxide; cardiomyopathy; stunning; hibernation; myocardial metabolism Received 4 June 2001; accepted in final form 23 August 2001

Chronic phospholamban (PLB)-sarcoplasmic reticulum calcium ATPase interaction is discussed as the critical calcium cycling defect in dilated cardiomyopathy. A study has used a genetic complementation strategy in a well-known mouse model of dilated cardiomyopathy. It is deficient in the cytoskeletal LIM protein, which is muscle-specific. Progressive defects in excitation-contraction coupling are related to enhancement of inhibition of SER-CA2a by PLB. Inhibition of PLB expression or PLB-SERCA2a interaction can prevent heat failure.

A study was conducted to examine the effects of mutations on cardiac muscle contractile functions under physiological conditions. Reverse transcriptase polymerase chain reaction was carried out on heart mRNA to amplify human cardiac Tnt cDNA while wild-type and mutant Tnt cDNAs were determined in pET3-d and expressed in Escherichia coli BL21. Results indicated that hypertrophic cardiomyopathy related with TnT mutations is caused by a heightened sensitivity in Ca2+.

Research was conducted to study changes in protein kinase C (PKC) in early cardiomyopathy and in gracilis muscles in diabetic rats by measuring the activity and total quantity of PKC and PKC isozymes in ventricular tissues and circulating neutrophils. The rat hearts were perfused on a working heart apparatus while the transcripts for PKC isozymes were determined by reverse transcriptase-polymerase chain reaction. Results suggested that increased PKC activity and changes of the PKC isozyme profile may play a major function in myocardium transition.

Dobutamine and dopamine was administered in patients with congestive heart failure to determine their positive inotropic activity on ventricular contractility. The inotropic agents produced equivalent increases in ventricular contractility but varied properties of large vessel compliance and ventriculoarterial coupling were detected from the patients. Increased ventricular contractility was not able to induce optimal transfer of augmented ventricular contractile power to vascular circulation.