Melatonin ameliorates pressure overload-induced cardiac hypertrophy by attenuating Atg5-dependent autophagy and activating the Akt/mTOR pathway

Cardiac hypertrophy, including hypertension and valvular dysfunction, is a pathological feature of many cardiac diseases that ultimately leads to heart failure. Melatonin confers a protective role against pathological cardiac hypertrophy, but the underlying mechanisms remain elusive. In the present study, we hypothesized that melatonin protects against pressure overload-induced cardiac hypertrophy by attenuating Atg5-dependent autophagy and activating the Akt/mTOR pathway. Male C57BL/6 mice that received adenovirus carrying cardiac-specific Atg5 (under the cTNT promoter; Ad-cTNT-Atg5) underwent transverse aortic constriction (TAC) or sham operation and received an intraperitoneal injection of melatonin (10 mg/kg/d), vehicle or LY294002 (10 mg/kg/d) for 8 weeks. Melatonin treatment for 8 weeks markedly attenuated cardiac hypertrophy and restored impaired cardiac function, as indicated by a decreased HW/BW ratio, reduced cell cross-sectional area and fibrosis, downregulated the mRNA levels of ANP, BNP, and β-MHC and ameliorated adverse effects on the LVEF and LVFS. Melatonin treatment also inhibited apoptosis and alleviated autophagy dysfunction. Furthermore, melatonin inhibited Akt/mTOR pathway activation, while these effects were blocked by LY294002. In addition, the effect of melatonin regulation on TAC-induced autophagy dysfunction was inhibited by LY294002 or cardiac-specific Atg5 overexpression. As expected, Akt/mTOR pathway inhibition or cardiac-specific Atg5 overexpression restrained melatonin alleviation of pressure overload-induced cardiac hypertrophy. These results demonstrated that melatonin ameliorated pressure overload-induced cardiac hypertrophy by attenuating Atg5-dependent autophagy and activating the Akt/mTOR pathway.     

Thyroid hormone inhibits ERK phosphorylation in pressure overload-induced hypertrophied mouse hearts through a receptor-mediated mechanism

Pressure overload-induced cardiac hypertrophy results in a pathological type of hypertrophy with activation of signaling cascades like the extracellular signal-regulated kinase (ERK) pathway, which promotes negative cardiac remodeling and decreased contractile function. In contrast, thyroid hormone mediates a physiological type of hypertrophy resulting in enhanced contractile function. In addition, thyroid hormone action is diminished in pressure overload-induced cardiac hypertrophy. We hypothesized that thyroid hormone status modulates ERK activity and that administration of thyroid hormone could alter the activity of this kinase in cardiac hypertrophy induced by pressure overload. ERK is activated by phosphorylation; accordingly, we investigated phosphorylation of ERK in hearts of control, hypothyroid, and hyperthyroid mice. In addition, the effect of T3 treatment on ERK phosphorylation in hypertrophied hearts from transverse aortic-constricted (TAC) mice was investigated. Results showed that phosphorylated ERK (p-ERK) was decreased by 25% in hyperthyroid mice. In contrast, hypothyroid mice presented increased p-ERK by 80%. TAC mice presented a greater than fourfold increase of p-ERK compared with control mice. Interestingly, T3 administration dramatically canceled TAC-induced ERK phosphorylation (36% lower compared with control). Raf-1 is upstream of the ERK pathway. TAC mice presented a 45% increase in phospho-Raf-1 (Ser338). T3 treatment inhibited this effect of pressure overload and further decreased p-Raf-1 (Ser338) by 37%, compared with control. Overexpression of thyroid hormone receptor-α in cultured cardiomyocytes potentiated the inhibitory effect of T3 on ERK phosphorylation. We concluded that thyroid hormone has an inhibitory effect on the Raf-1/ERK pathway. Furthermore, treatment of TAC mice with T3 inhibited Raf-1/ERK pathway by a thyroid hormone receptor-dependent mechanism.     

Activated protein C protects against pressure overload-induced hypertrophy through AMPK signaling

We found that the anticoagulant plasma protease, activated protein C (APC), stimulates the energy sensor kinase, AMPK, in the stressed heart by activating protease-activated receptor 1 (PAR1) on cardiomyocytes. Wild-type (WT) and AMPK-kinase dead (KD) transgenic mice were subjected to transverse aortic constriction (TAC) surgery. The results demonstrated that while no phenotypic differences can be observed between WT and AMPK-KD mice under normal physiological conditions, AMPK-KD mice exhibit significantly larger hearts after 4 weeks of TAC surgery. Analysis by echocardiography suggested that the impairment in the cardiac function of AMPK-KD hearts is significantly greater than that of WT hearts. Immunohistochemical staining revealed increased macrophage infiltration and ROS generation in AMPK-KD hearts after 4 weeks of TAC surgery. Immunoblotting results demonstrated that the redox markers, pShc⁶⁶, 4-hydroxynonenal and ERK, were all up-regulated at a higher extent in AMPK-KD hearts after 4 weeks of TAC surgery. Administration of APC-WT and the signaling selective APC-2Cys mutant, but not the anticoagulant selective APC-E170A mutant, significantly attenuated pressure overload-induced hypertrophy and fibrosis. Macrophage infiltration and pShc⁶⁶ activation caused by pressure overload were also inhibited by APC and APC-2Cys but not by APC-E170A. Therefore, the cardiac AMPK protects against pressure overload-induced hypertrophy and the signaling selective APC-2Cys may have therapeutic potential for treating hypertension-related hypertrophy without increasing the risk of bleeding.     

Grape polyphenols and exercise training have distinct molecular effects on cardiac hypertrophy in a model of obese insulin-resistant rats

Obesity and exercise lead to structural changes in heart such as cardiac hypertrophy. The underlying signaling pathways vary according to the source of the overload, be it physiological (exercise) or pathologic (obesity). The physiological pathway relies more on PI3K-Akt signaling while the pathologic pathway involves calcineurin-Nuclear factor of activated T-cells activation and fibrosis accumulation. Independently, exercise and polyphenols have demonstrated to prevent pathologic cardiac hypertrophy. Therefore, we investigated the molecular adaptations of the combination of exercise training and grape polyphenols supplementation (EXOPP) in obese high-fat fed rats on heart adaptation in comparison to exercise (EXO), polyphenols supplementation (PP) and high-fat fed rats (HF), alone. Exercised and PP rats presented a higher heart weight/body weight ratio compared to HF rats. EXO and EXOPP depicted an increase in cell-surface area, P-Akt/Akt, P-AMPK/AMPK ratios with a decreased fibrosis and calcineurin expression, illustrating an activation of the physiological pathway, but no additional benefit of the combination. In contrast, neither cell-surface area nor Akt signaling increased in PP rats; but markedly decreased fibrosis, calcineurin expression, systolic blood pressure, higher SERCA and P-Phospholamdan/Phospholamdan levels were observed. These data suggest that PP rats have a shift from pathologic toward physiological hypertrophy. Our study demonstrates that polyphenols supplementation has physical-activity-status-specific effects; it appears to be more protective in sedentary obese insulin-resistant rats than in the exercised ones. Exercise training improved metabolic and cardiac alterations without a synergistic effect of polyphenols supplementation. These data highlight a greater effect of exercise than polyphenols supplementation for the treatment of cardiac alterations in obese insulin-resistant rats.     

Exercise training reduces insulin resistance and upregulates the mTOR/p70S6k pathway in cardiac muscle of diet‐induced obesity rats

Obesity and insulin resistance are rapidly expanding public health problems. These disturbances are related to many diseases, including heart pathology. Acting through the Akt/mTOR pathway, insulin has numerous and important physiological functions, such as the induction of growth and survival of many cell types and cardiac hypertrophy. However, obesity and insulin resistance can alter mTOR/p70S6k. Exercise training is known to induce this pathway, but never in the heart of diet‐induced obesity subjects. To evaluate the effect of exercise training on mTOR/p70S6k in the heart of obese Wistar rats, we analyzed the effects of 12 weeks of swimming on obese rats, induced by a high‐fat diet. Exercise training reduced epididymal fat, fasting serum insulin and plasma glucose disappearance. Western blot analyses showed that exercise training increased the ability of insulin to phosphorylate intracellular molecules such as Akt (2.3‐fold) and Foxo1 (1.7‐fold). Moreover, reduced activities and expressions of proteins, induced by the high‐fat diet in rats, such as phospho‐JNK (1.9‐fold), NF‐kB (1.6‐fold) and PTP‐1B (1.5‐fold), were observed. Finally, exercise training increased the activities of the transduction pathways of insulin‐dependent protein synthesis, as shown by increases in Raptor phosphorylation (1.7‐fold), p70S6k phosphorylation (1.9‐fold), and 4E‐BP1 phosphorylation (1.4‐fold) and a reduction in atrogin‐1 expression (2.1‐fold). Results demonstrate a pivotal regulatory role of exercise training on the Akt/mTOR pathway, in turn, promoting protein synthesis and antagonizing protein degradation. J. Cell. Physiol. 226: 666–674, 2011. © 2010 Wiley‐Liss, Inc.     

Exercise Protects against Chronic β-Adrenergic Remodeling of the Heart by Activation of Endothelial Nitric Oxide Synthase

Extensive data have shown that exercise training can provide cardio-protection against pathological cardiac hypertrophy. However, how long the heart can retain cardio-protective phenotype after the cessation of exercise is currently unknown. In this study, we investigated the time course of the loss of cardio-protection after cessation of exercise and the signaling molecules that are responsible for the possible sustained protection. Mice were made to run on a treadmill six times a week for 4 weeks and then rested for a period of 0, 1, 2 and 4 weeks followed by isoproterenol injection for 8 days. Morphological, echocardiographic and hemodynamic changes were measured, gene reactivation was determined by real-time PCR, and the expression and phosphorylation status of several cardio-protective signaling molecules were analyzed by Western-blot. HW/BW, HW/TL and LW/BW decreased significantly in exercise training (ER) mice. The less necrosis and lower fetal gene reactivation induced by isoproterenol injection were also found in ER mice. The echocardiographic and hemodynamic changes induced by β-adrenergic overload were also attenuated in ER mice. The protective effects can be sustained for at least 2 weeks after the cessation of the training. Western-blot analysis showed that the alterations in the phosphorylation status of endothelial nitric oxide synthase (eNOS) (increase in serine 1177 and decrease in threonine 495) continued for 2 weeks after the cessation of the training whereas increases of the phosphorylation of Akt and mTOR disappeared. Further study showed that L-NG-Nitroarginine methyl ester (L-NAME) treatment abolished the cardio-protective effects of ER. Our findings demonstrate that stimulation of eNOS in mice through exercise training provides acute and sustained cardioprotection against cardiac hypertrophy.     

A novel compound,ferulic acid-bound resveratrol,induces the tumor suppressor gene p15 and inhibits the three-dimensional proliferation of colorectal cancer cells

This study aimed to investigate the effects and molecular mechanisms of ivabradine in preventing cardiac hypertrophy in an established transverse aortic constriction (TAC) mouse model. A total of 56 male C57BL/6 mice were randomly assigned into the following seven groups (8 mice per group): sham, TAC model, Iva-10 (10 mg/kg/day ivabradine), Iva-20 (20 mg/kg/day ivabradine), Iva-40 (40 mg/kg/day ivabradine), Iva-80 (80 mg/kg/day ivabradine), and Rap (rapamycin, a positive control). Echocardiography and left ventricular hemodynamics were performed. Hematoxylin-eosin (H&E), Masson’s trichome staining, and TUNEL assays were conducted to evaluate cardiac hypertrophy, fibrosis, and apoptosis, respectively. Western blotting was performed to detect the expression of proteins related to the PI3K/Akt/mTOR/p70S6K pathway. Ivabradine could effectively improve left ventricular dysfunction and hypertrophy induced by TAC in a dose-independent manner. Moreover, no obvious change in heart rate (HR) was observed in the TAC and Rap groups, whereas a significant decrease in HR was found after ivabradine treatment (P?<?0.05). Cardiac hypertrophy, fibrosis, and apoptosis induced by TAC were notably suppressed after either rapamycin or ivabradine treatment (P?<?0.05). Ivabradine and rapamycin also decreased the expression of PI3K/Akt and mTOR induced by TAC. Ivabradine improved cardiac hypertrophy and fibrosis as well as reduced cardiomyocyte apoptosis via the PI3K/Akt/mTOR/p70S6K pathway in TAC model mice.

     

举重运动员的卫星细胞活化和信号通路对阻力或联合运动的反应

为了考察单一抗阻运动模式和联合运动模式对举重运动员的卫星细胞活化和PI3K/Akt/mTOR信号通路的影响。本研究以30名男性举重运动员为研究对象,将受试者随机分为抗阻运动组和联合运动组,抗阻运动组接受60%最大重复次数(1 RM)的负重蹲起训练,联合运动组接受60%1 RM的负重蹲起和卧推训练。运动前和运动后3 h立即获得肌肉活检样品,采用双重免疫荧光染色检测活化的卫星细胞数(Pax7+/MyoD+)。采用蛋白质印迹法检测肌肉组织中Akt、mTOR、p70S6K和4E-BP1的磷酸化情况。研究发现,运动后,联合运动组活化的卫星细胞数显著高于抗阻运动组(35.14 vs 29.86个,p=0.011)。运动后联合运动组的Akt、mTOR、p70S6K和4E-BP1的磷酸化水平显著高于抗阻运动组(p<0.05)。本研究表明,与单一抗阻运动模式相比,联合运动模式更有助于肌肉卫星细胞的活化和PI3K/Akt/mTOR信号通路的活化,从而改善肌肉功能。     

Long-term activation of adenosine monophosphate-activated protein kinase attenuates pressure-overload-induced cardiac hypertrophy

Recent in vitro studies suggest that adenosine monophosphate (AMP)-activated protein kinase (AMPK) exerts inhibitory effects on cardiac hypertrophy. However, it is unclear whether long-term activation of AMPK will affect cardiac hypertrophy in vivo. In these reports, we investigate the in vivo effects of long-term AMPK activation on cardiac hypertrophy and the related molecular mechanisms. To examine the effects of AMPK activation in the development of pressure overload-induced cardiac hypertrophy, we administered 5-aminoimidazole 1 carboxamide ribonucleoside (AICAR, 0.5 mg/g body wt), a specific activator of AMPK, to rats with transaortic constriction (TAC) for 7 weeks. We found that long-term AMPK activation attenuated cardiac hypertrophy, and improved cardiac function in rats subjected to TAC. Furthermore, long-term AMPK activation attenuated protein synthesis, diminished calcineurin-nuclear factor of activated T cells (NFAT) and nuclear factor kappaB (NF-kappaB) signaling in pressure overload-induced hypertrophic hearts. Our in vitro experiments further proved that activation of AMPK by infection of AdAMPK blocked cardiac hypertrophy and NFAT, NF-kappaB, and MAPK signal pathways. The present study demonstrates for the first time that pharmacological activation of AMPK inhibits cardiac hypertrophy in through blocking signaling transduction pathways that are involved in cardiac growth. It presents a potential therapy strategy to inhibit pathological cardiac hypertrophy by increasing the activity of AMPK.     

Attenuation of cardiac hypertrophy by inhibiting both mTOR and NFkappaB activation in vivo

A role for the PI3K/Akt/mTOR pathway in cardiac hypertrophy has been well documented. We reported that NFκB activation is needed for cardiac hypertrophy in vivo. To investigate whether both NFκB activation and PI3K/Akt/mTOR signaling participate in the development of cardiac hypertrophy, two models of cardiac hypertrophy, namely, induction in caAkt-transgenic mice and by aortic banding in mice, were employed. Rapamycin (2 mg/kg/daily), an inhibitor of the mammalian target of rapamycin, and the antioxidant pyrrolidine dithiocarbamate (PDTC; 120 mg/kg/daily), which can inhibit NFκB activation, were administered to caAkt mice at 8 weeks of age for 2 weeks. Both rapamycin and PDTC were also administered to the mice immediately after aortic banding for 2 weeks. Administration of either rapamycin or PDTC separately or together to caAkt mice reduced the ratio of heart weight/body weight by 21.54, 32.68, and 42.07% compared with untreated caAkt mice. PDTC administration significantly reduced cardiac NFκB activation by 46.67% and rapamycin significantly decreased the levels of p70S6K by 34.20% compared with untreated caAkt mice. Similar results were observed in aortic-banding-induced cardiac hypertrophy in mice. Our results suggest that both NFκB activation and the PI3K/Akt signaling pathway participate in the development of cardiac hypertrophy in vivo.     

Resistance training-induced muscle hypertrophy is mediated by TGF-β1-Smad signaling pathway in male Wistar rats

The TGF-β1-Smad pathway is a well-known negative regulator of muscle growth; however, its potential role in resistance training-induced muscle hypertrophy is not clear. The present study proposed to determine whether and how this pathway may be involved in resistance training-induced muscle hypertrophy. Skeletal muscle samples were collected from the control, trained (RT), control + SB431542 (CI_TGF), and trained + SB431542 (RTI_TGF) animals following 3, 5, and 8 weeks of resistance training. Inhibition of the TGF-β1-Smad pathway by SB431542 augmented muscle satellite cells activation, upregulated Akt/mTOR/S6K1 pathway, and attenuated FOXO1 and FOXO3a expression in the CI_TGF group (all p < .01), thereby causing significant muscle hypertrophy in animals from the CI_TGF. Resistance training significantly decreased muscle TGF-β1 expression and Smad3 (P-Smad3^S423/425) phosphorylation at COOH-terminal residues, augmented Smad2 (P-Smad2-L^S245/250/255) and Smad3 (P-Smad3-L^Ser208) phosphorylation levels at linker sites (all p < .01), and led to a muscle hypertrophy which was unaffected by SB431542, suggesting that the TGF-β1-Smad signaling pathway is involved in resistance training-induced muscle hypertrophy. The effects of inhibiting the TGF-β1-Smad signaling pathway were not additive to the resistance training effects on FOXO1 and FOXO3a expression, muscle satellite cells activation, and the Akt/mTOR/S6K1 pathway. Resistance training effect of satellite cell differentiation was independent of the TGF-β1-Smad signaling pathway. These results suggested that the effect of the TGF-β1-Smad signaling pathway on resistance training-induced muscle hypertrophy can be attributed mainly to its diminished inhibitory effects on satellite cell activation and protein synthesis. Suppressed P-Smad3^S423/425 and enhanced P-Smad2-L^S245/250/255 and P-Smad3-L^Ser208 are the molecular mechanisms that link the TGF-β1-Smad signaling pathway to resistance training-induced muscle hypertrophy.     

gp130 plays a critical role in pressure overload-induced cardiac hypertrophy

gp130, a common receptor for the interleukin 6 family, plays pivotal roles in growth and survival of cardiac myocytes. In the present study, we examined the role of gp130 in pressure overload-induced cardiac hypertrophy using transgenic (TG) mice, which express a dominant negative mutant of gp130 in the heart under the control of alpha myosin heavy chain promoter. TG mice were apparently healthy and fertile. There were no differences in body weight and heart weight between TG mice and littermate wild type (WT) mice. Pressure overload-induced increases in the heart weight/body weight ratio, ventricular wall thickness, and cross-sectional areas of cardiac myocytes were significantly smaller in TG mice than in WT mice. Northern blot analysis revealed that pressure overload-induced up-regulation of brain natriuretic factor gene and down-regulation of sarcoplasmic reticulum Ca(2+) ATPase 2 gene were attenuated in TG mice. Pressure overload activated ERKs and STAT3 in the heart of WT mice, whereas pressure overload-induced activation of STAT3, but not of ERKs, was suppressed in TG mice. These results suggest that gp130 plays a critical role in pressure overload-induced cardiac hypertrophy possibly through the STAT3 pathway.     

Effects of Estrogen,an ERα Agonist and Raloxifene on Pressure Overload Induced Cardiac Hypertrophy

The aim of this study was to investigate the effects of 17β-estradiol (E2), the selective ERα agonist 16α-LE2, and the selective estrogen receptor modulator (SERM) raloxifene on remodeling processes during the development of myocardial hypertrophy (MH) in a mouse model of pressure overload. Myocardial hypertrophy in ovariectomized female C57Bl/6J mice was induced by transverse aortic constriction (TAC). Two weeks after TAC, placebo treated mice developed left ventricular hypertrophy and mild systolic dysfunction. Estrogen treatment, but not 16α-LE2 or raloxifene reduced TAC induced MH compared to placebo. E2, 16α-LE2 and raloxifene supported maintenance of cardiac function in comparison with placebo. Nine weeks after induction of pressure overload, MH was present in all TAC groups, most pronounced in the raloxifene treated group. Ejection fraction (EF) was decreased in all animals. However, 16α-LE2 treated animals showed a smaller reduction of EF than animals treated with placebo. E2 and 16α-LE2, but not raloxifene diminished the development of fibrosis and reduced the TGFβ and CTGF gene expression. Treatment with E2 or 16α-LE2 but not with raloxifene reduced survival rate after TAC significantly in comparison with placebo treatment. In conclusion, E2 and 16α-LE2 slowed down the progression of MH and reduced systolic dysfunction after nine weeks of pressure overload. Raloxifene did not reduce MH but improved cardiac function two weeks after TAC. However, raloxifene was not able to maintain EF in the long term period.     

Zerumbone prevents pressure overload-induced left ventricular systolic dysfunction by inhibiting cardiac hypertrophy and fibrosis

BackgroundCardiac hypertrophy and fibrosis are hallmarks of cardiac remodeling and are involved functionally in the development of heart failure (HF). However, it is unknown whether Zerumbone (Zer) prevents left ventricular (LV) systolic dysfunction by inhibiting cardiac hypertrophy and fibrosis.PurposeThis study investigated the effect of Zer on cardiac hypertrophy and fibrosis in vitro and in vivo.Study Design/methodsIn primary cultured cardiac cells from neonatal rats, the effect of Zer on phenylephrine (PE)-induced hypertrophic responses and transforming growth factor beta (TGF-β)-induced fibrotic responses was observed. To determine whether Zer prevents the development of pressure overload-induced HF in vivo, a transverse aortic constriction (TAC) mouse model was utilized. Cardiac function was evaluated by echocardiography. The changes of cardiomyocyte surface area were observed using immunofluorescence staining and histological analysis (HE and WGA staining). Collagen synthesis and fibrosis formation were measured by scintillation counter and picrosirius staining, respectively. The total mRNA levels of genes associated with hypertrophy (ANF and BNP) and fibrosis (Postn and α-SMA) were measured by qRT-PCR. The protein expressions (Akt and α-SMA) were assessed by western blotting.ResultsZer significantly suppressed PE-induced increase in cell size, mRNA levels of ANF and BNP, and Akt phosphorylation in cardiomyocytes. The TGF-β-induced increase in proline incorporation, mRNA levels of Postn and α-SMA, and protein expression of α-SMA were decreased by Zer in cultured cardiac fibroblasts. In the TAC male C57BL/6 mice, echocardiography results demonstrated that Zer improved cardiac function by increasing LV fractional shortening and reducing LV wall thickness compared with the vehicle group. ZER significantly reduced the level of phosphorylated Akt both in cultured cardiomyocytes treated with PE and in the hearts of TAC. Finally, Zer inhibited the pressure overload-induced cardiac hypertrophy and cardiac fibrosis.ConclusionZer ameliorates pressure overload-induced LV dysfunction, at least in part by suppressing both cardiac hypertrophy and fibrosis.     

The protective effect of isosteviol sodium on cardiac function and myocardial remodelling in transverse aortic constriction rat

Pathological hypertrophy contributes to heart failure and there is not quite effective treatment to invert this process. Isosteviol has been shown to protect the heart against ischaemia-reperfusion injury and isoproterenol-induced cardiac hypertrophy, but its effect on pressure overload-induced cardiac hypertrophy is still unknown. Pressure overload induced by transverse aortic constriction (TAC) causes cardiac hypertrophy in rats to mimic the pathological condition in human. This study examined the effects of isosteviol sodium (STVNa) on cardiac hypertrophy by the TAC model and cellular assays in vitro. Cardiac function test, electrocardiogram analysis and histological analysis were conducted. The effects of STVNa on calcium transient of the adult rat ventricular cells and the proliferation of neonatal rat cardiac fibroblasts were also studied in vitro. Cardiac hypertrophy was observed after 3-week TAC while the extensive cardiac dysfunction and electronic remodelling were observed after 9-week TAC. Both STVNa and sildenafil (positive drug) treatment reversed the two process, but STVNa appeared to be more superior in some aspects and did not change calcium transient considerably. STVNa also reversed TAC-induced cardiac fibrosis in vivo and TGF-β1-induced fibroblast proliferation in vitro. Moreover, STVNa, but not sildenafil, reversed impairment of the autonomic nervous system induced by 9-week TAC.     

Effects of different doses of leucine ingestion following eight weeks of resistance exercise on protein synthesis and hypertrophy of skeletal muscle in rats

[Purpose]

This study was designed to determine the appropriate Leucine intake volume to obtain the effects of restoring damaged muscle through the synthesis of muscle proteins to increase skeletal muscle and improve exercise performance, and to achieve enhanced muscle hypertrophy.

[Methods]

To clarify the effects of leucine on skeletal muscle hypertrophy of SD rats, following eight weeks of resistance exercise (climbing ladder), the mass of the FHL (Flexor hallucis longus) was measured after extraction, after which change in the activity of muscle signaling proteins (PKB/Akt, mTOR, p70S6K, 4EBP1) was analyzed.

[Results]

The expressions of PKB/Akt, mTOR and p70S6K were increased in L5 (Leucine 50% administration group) compared with the control group (CON) and exercise group (Ex, exercise training group); EL1 (exercise + 10% leucine administration group) and EL5 (exercise + 50% Leucine administration) also exhibited increased expressions of PKB/Akt, mTOR, and p70S6K, while no difference between EL1 and EL5 were observed. No significant differences in 4EBP1 were found among any of the groups. In addition, there were no differences in FHL mass, while relative mass (FHL/body mass) was increased in the exercise group (Ex, EL1, EL5) compared with the control group. No differences were observed among the exercise groups.

[Conclusion]

The present study demonstrated that the relative body mass was increased in the EX group compared with the CON group, while no significant differences in muscle mass could be found among the groups. Even though some signaling proteins were increased, or some differences existed among groups, there were no differences in muscle mass between the leucine administration and exercise training combined with leucine administration groups in the present study.     

跑步训练诱导小鼠生理性心脏肥厚模型

目的用长期跑步训练诱导小鼠的生理性心脏肥厚模型,与主动脉缩窄手术诱导的病理性心脏肥厚模型进行比较。方法 8周龄野生型雄性C57BL/6小鼠分为跑步运动组,正常对照组,手术刺激组和假手术组。运动组跑步训练40d,手术刺激组行主动脉缩窄手术2周,从组织形态学、超声心动图、分子标志物表达等方面对模型进行全面评估。结果运动训练组小鼠心脏体重比与正常对照组相比增加27.2%(P<0.05),左心室体重比增加25.8%(P<0.01),心脏显著肥厚。超声心动图显示,与各自的对照组相比,运动组和手术组小鼠模型的左心室后壁厚度均显著增加(P<0.05),但运动组小鼠的相对室壁厚度无明显变化,而手术组小鼠相对室壁厚度显著增加50%(P<0.05),提示两种不同的心脏肥厚导致在心脏结构改变上差别显著。心脏肥厚分子标志物心房利钠肽和脑钠肽在手术组表达显著上调9.5倍和4.5倍,而在运动组下调为对照组的0.48倍和0.58倍,提示两种不同肥厚的分子机制差别迥异。结论长期跑步运动可以成功的诱导小鼠生理性心脏肥厚模型,其表型和分子机制与手术刺激的病理性肥厚差别显著。     

Diacylglycerol kinase-epsilon restores cardiac dysfunction under chronic pressure overload: a new specific regulator of Galpha(q) signaling cascade

Galpha(q) protein-coupled receptor (GPCR) signaling pathway, which includes diacylglycerol (DAG) and protein kinase C (PKC), plays a critical role in cardiac hypertrophy. DAG kinase (DGK) catalyzes DAG phosphorylation and controls cellular DAG levels, thus acting as a regulator of GPCR signaling. It has been reported that DGKepsilon acts specifically on DAG produced by inositol cycling. In this study, we examined whether DGKepsilon prevents cardiac hypertrophy and progression to heart failure under chronic pressure overload. We generated transgenic mice with cardiac-specific overexpression of DGKepsilon (DGKepsilon-TG) using an alpha-myosin heavy chain promoter. There were no differences in cardiac morphology and function between wild-type (WT) and DGKepsilon-TG mice at the basal condition. Either continuous phenylephrine infusion or thoracic transverse aortic constriction (TAC) was performed in WT and DGKepsilon-TG mice. Increases in heart weight after phenylephrine infusion and TAC were abolished in DGKepsilon-TG mice compared with WT mice. Cardiac dysfunction after TAC was prevented in DGKepsilon-TG mice, and the survival rate after TAC was higher in DGKepsilon-TG mice than in WT mice. Phenylephrine- and TAC-induced DAG accumulation, the translocation of PKC isoforms, and the induction of fetal genes were blocked in DGKepsilon-TG mouse hearts. The upregulation of transient receptor potential channel (TRPC)-6 expression after TAC was attenuated in DGKepsilon-TG mice. In conclusion, these results demonstrate the first evidence that DGKepsilon restores cardiac dysfunction and improves survival under chronic pressure overload by controlling cellular DAG levels and TRPC-6 expression. DGKepsilon may be a novel therapeutic target to prevent cardiac hypertrophy and progression to heart failure.     

Developmental changes in gene expression of Epac and its upregulation in myocardial hypertrophy

Although it has been shown that Epac1 mRNA is expressed ubiquitously and Epac2 mRNA predominantly in the brain and endocrine tissues, developmental and pathophysiological changes of these molecules have not been characterized. Developmental changes were analyzed in murine heart, brain, kidneys, and lungs by RT-PCR analysis, which revealed more drastic developmental changes of Epac2 mRNA than Epac1. Only the Epac2 mRNA in kidney showed a transient expression pattern with dramatic decline into adulthood. In addition to developmental changes, we found that Epac gene expression was upregulated in myocardial hypertrophy induced by chronic isoproterenol infusion or pressure overload by transverse aortic banding. Both Epac1 and Epac2 mRNA were upregulated in isoproterenol-induced left ventricular hypertrophy, whereas only Epac1 was increased in pressure overload-induced hypertrophy. Stimulation of H9c2, cardiac myoblast cells, with fetal calf serum, which can induce myocyte hypertrophy, upregulated Epac1 protein expression. We also demonstrated that Epac was the limiting moiety, relative to Rap, in the Epac-Rap signaling pathway in terms of stoichiometry and that Epac stimulation led to the activation of ERK1/2. Our data suggest the functional involvement of Epac in organogenesis and also in physiological as well as pathophysiological processes, such as cardiac hypertrophy. Furthermore, our results suggest the importance of the stoichiometry of Epac over that of Rap in cellular biological effects.