Isorhapontigenin, a new resveratrol analog, attenuates cardiac hypertrophy via blocking signaling transduction pathways

Cardiac hypertrophy is a major cause of morbidity and mortality worldwide. The hypertrophic process is mediated, in part, by oxidative stress-mediated signaling pathways. We hypothesized that isorhapontigenin (ISO), a new resveratrol analog, inhibits cardiac hypertrophy by blocking oxidative stress and oxidative stress-mediated signaling pathways. We treated cardiomyocytes with angiotensin II (Ang II) with or without ISO and found that ISO inhibited Ang II-induced cardiac hypertrophy. These effects were associated with a decrease in the levels of reactive oxygen species and H2O2 and the content of intracellular malonaldehyde and an increase in the activities of superoxide dismutase and glutathione peroxidase. Ang II induced the phosphorylation of PKC, Erk1/2, JNK, and p38 in cardiomyocytes and such phosphorylation was inhibited by ISO. ISO also blocked the PKC-dependent PI3K-Akt-GSK3beta/p70S6K pathway. These effects lead to direct or indirect inhibition of NF-kappaB and AP-1 activation. Our results revealed that pretreatment with ISO significantly inhibited Ang II-mediated NF-kappaB through affecting the degradation and phosphorylation of IkappaBalpha and the activity of IKKbeta and AP-1 activation by influencing the expression of c-Fos and c-Jun proteins. In addition, we also established the molecular link between activation of PKC and MAPKs and activation of NF-kappaB and AP-1 in cardiomyocytes. We also found that ISO treatment significantly attenuated heart weight/body weight ratio by approximately 25%, decreased posterior wall thickness and left ventricle diastolic and systolic diameters, and increased 10% fractional shortening in an aortic-banded rat model. Furthermore, treatment with ISO significantly decreased cardiac myocyte size and systolic blood pressure. These findings suggest that ISO prevents the development of cardiac hypertrophy through an antioxidant mechanism involving inhibition of different intracellular signaling transduction pathways.     

Endothelin-1- and isoproterenol-induced differential protein expression and signaling pathway in HL-1 cardiomyocytes

It is well known that the two chemical compounds endothelin-1 (ET-1) and isoproterenol (ISO) can individually induce cardiac hypertrophy through G protein-coupled receptors in cardiomyocytes. However, the cardiac hypertrophy signaling pathway activated by ET-1 and ISO is not well defined. Therefore, we investigated the protein expression profile and signaling transduction in HL-l cardiomyocyte cells treated with ET-1 and ISO. Following separation of the cell lysates by using 2-DE and silver staining, we identified 16 protein spots that were differentially expressed as compared to the controls. Of these 16 spots, three changed only after treatment with ET-1, whereas four changed only after treatment with ISO, suggesting that these two stimuli could induce different signaling pathways. In order to reveal the differences between ET-1- and ISO-induced signaling, we studied the different events that occur at each step of the signaling pathways, when selected biocomponents were blocked by inhibitors. Our results indicated that ET-1 and ISO used different pathways for phosphorylation of glycogen synthase kinase-3β (GSK3β). ET-1 mainly used the mitogen-activated protein kinase and phosphatidylinositol-3-kinase/AKT pathways to activate GSK3β, whereas under ISO stimulation, only the phosphatidylinositol-3-kinase/AKT pathway was required to trigger the GSK3β pathway. Furthermore, the strength of the GSK3β signal in ISO-induced cardiac hypertrophy was stronger than that in ET-1-induced cardiac hypertrophy. We found that these two agonists brought about different changes in the protein expression of HL-1 cardiomyocytes through distinct signaling pathways even though the destination of the two signaling pathways was the same.     

Reciprocal regulation of miR-23a and lysophosphatidic acid receptor signaling in cardiomyocyte hypertrophy

Earlier, our study demonstrated that lysophosphatidic acid (LPA) receptor mediated cardiomyocyte hypertrophy. However, the subtype-specific functions for LPA1 and LPA3 receptors in LPA-induced hypertrophy have not been distinguished. Growing evidence indicates that microRNAs (miRNAs) are involved in the pathogenesis of cardiac hypertrophy by down-regulating target molecules. The present work therefore aimed at elucidating the functions mediated by different subtypes of LPA receptors and investigating the modulatory role of miRNAs during LPA induced hypertrophy. Experiments were done with cultured neonatal rat cardiomyocytes (NRCMs) exposed to LPA and we showed that knockdown of LPA1 by small interfering RNA (siRNA) enhanced LPA-induced cardiomyocyte hypertrophy, whereas LPA3 silencing repressed hypertrophy. miR-23a, a pro-hypertrophic miRNA, was up-regulated by LPA in cardiomyocytes and its down-regulation reduced LPA-induced cardiomyocyte hypertrophy. Importantly, luciferase reporter assay confirmed LPA1 to be a target of miR-23a, indicating that miR-23a is involved in mediating the LPA-induced cardiomyocyte hypertrophy by targeting LPA1. In addition, knockdown of LPA3, but not LPA1, eliminated miR-23a elevation induced by LPA. And PI3K inhibitor, LY294002, effectively prevented LPA-induced miR-23a expression in cardiomyocytes, suggesting that LPA might induce miR-23a elevation by activating LPA3 and PI3K/AKT pathway. These findings identified opposite subtype-specific functions for LPA1 and LPA3 in mediating cardiomyocyte hypertrophy and indicated LPA1 to be a target of miR-23a, which discloses a link between miR-23a and the LPA receptor signaling in cardiomyocyte hypertrophy.     

Tanshinone IIA protects against cardiac hypertrophy via inhibiting calcineurin/NFATc3 pathway

Pathological cardiac hypertrophy induced by adrenergic overactivation can subsequently develop to heart failure which remains as a leading cause of mortality worldwide. Tanshinone IIA is a lipid-soluble pharmacologically active compound extracted from the rhizome of the Chinese herb Salvia miltiorrhiza, a well-known traditional Chinese medicine used for the treatment of cardiovascular disorders. However, little is know about the effect of Tanshinone IIA on cardiac hypertrophy. The present study was aimed to investigate whether Tanshinone IIA prevents cardiac hypertrophy induced by isoproterenol (ISO) and to clarify its possible mechanisms. Cardiomyocytes hypertrophy was induced by ISO 10 μM for 48 h with or without Tanshinone IIA 10, 30, 100 μM pretreatment, and evaluated by determining the cell size and the expression of ANP, BNP, β-MHC, Calcineurin, and NFATc3 by real-time PCR and western blot. We found that Tanshinone IIA pretreatment attenuated the enlargement of cell surface area induced by ISO in cultured cardiomyocytes. The mRNA level of ANP, BNP and β-MHC was obviously elevated in ISO-treated cardiac cells, which was effectively inhibited by Tanshinone IIA. Moreover, we found that Tanshinone IIA pretreatment could prevent the augment of intracellular calcium transient in ISO-treated cardiomyocytes. The further study revealed that Calcineurin, NFATc3, ANP, BNP and β-MHC proteins were upregulated by ISO in ventricular myocytes, and Tanshinone IIA pretreatment significantly attenuate the increased expression of Calcineurin, NFATc3, ANP, BNP and β-MHC proteins. In summary, Tanshinone IIA attenuated cardiomyocyte hypertrophy induced by ISO through inhibiting Calcineurin/NFATc3 pathway, which provides new insights into the pharmacological role and therapeutic mechanism of Tanshinone IIA in heart diseases.     

心脏中不同β肾上腺素受体亚型的信号体系及其病理生理意义

生理情况下,β肾上腺素受体(βAR)对心肌收缩和舒张活动起至关重要的作用;病理情况下,长期激动βAR可以诱发心肌细胞肥大、凋亡以及细胞坏死等心肌重塑性活动,从而参与了慢性心衰的发病过程。近十年以来,许多资料表明β1和β2肾上腺素受体亚型(β1AR和β2AR)共存于心脏中,且激动不同信号系统。短时间激动β1AR,使Gs蛋白-腺苷酸环化酶-环苷腺酸-蛋白激酶A(Gs-adenyly cyclase-cAMP-PKA)信号体系激活并广布于细胞内,而激动βAR则同时激活G1蛋白而产生空间及功能局限的cAMP信号;长时间激动β1AR和β2AR则对心肌细胞的命运产生不同影响：β1AR诱导细胞肥大和凋亡,β2AR促使细胞存活。β2AR的心肌保护作用是通过激活Gi蛋白-Gβγ-PI3K-Akt途径介导。但出乎意料,β1AR的心肌肥厚和凋亡效应并不依赖于经典的cAMP/PKA信号途径,而是激活钙,钙调素依赖性蛋白激酶Ⅱ(caMK Ⅱ)途径。用心肌特异性表达βAR亚型的转基因小鼠进行实验,进一步证实不同βAR亚型在调节心肌重塑和功能方面作用各异。βAR亚型作用不同的新观点不仅为β阻滞剂治疗慢性心衰提供了分子和细胞机制的依据,而且提出了选择性β1AR阻滞和β2AR激动联合治疗慢性心衰的新的治疗思路。     

RGS2 is upregulated by and attenuates the hypertrophic effect of alpha1-adrenergic activation in cultured ventricular myocytes

Regulator of G protein signaling (RGS) proteins counter the effects of G protein-coupled receptors (GPCRs) by limiting the abilities of G proteins to propagate signals, although little is known concerning their role in cardiac pathophysiology. We investigated the potential role of RGS proteins on alpha1-adrenergic receptor signals associated with hypertrophy in primary cultures of neonatal rat cardiomyocytes. Levels of mRNA encoding RGS proteins 1-5 were examined, and the alpha1-adrenergic agonist phenylephrine (PE) significantly increased RGS2 gene expression but had little or no effect on the others. The greatest changes in RGS2 mRNA occurred within the first hour of agonist addition. We next investigated the effects of RGS2 overexpression produced by infecting cells with an adenovirus encoding RGS2-cDNA on cardiomyocyte responses to PE. As expected, PE increased cardiomyocyte size and also significantly upregulated alpha-skeletal actin and ANP expression, the markers of hypertrophy, as well as the Na-H exchanger 1 isoform. These effects were blocked in cells infected with the adenovirus expressing RGS2. We also examined hypertrophy-associated MAP kinase pathways, and RGS2 overexpression completely prevented the activation of ERK by PE. In contrast, the activation of both JNK and p38 unexpectedly were increased by RGS2, although the ability of PE to further activate the p38 pathway was reduced. These results indicate that RGS2 is an important negative-regulatory factor in cardiac hypertrophy produced by alpha1-adrenergic receptor stimulation through complex mechanisms involving the modulation of mitogen-activated protein kinase signaling pathways.     

A role for focal adhesion kinase in phenylephrine-induced hypertrophy of rat ventricular cardiomyocytes

A variety of agonists including phenylephrine (PE) induce hypertrophy in neonatal ventricular cardiomyocytes. Here we report that signals provided by extracellular matrix proteins (ECM) augment the PE-induced hypertrophic response of cardiomyocytes and provide evidence that ECM-dependent signaling is mediated in part by the protein tyrosine kinase, focal adhesion kinase (FAK). Addition of PE to cultured neonatal cardiomyocytes stimulated sarcomeric organization, increased cell size, and induced atrial natriuretic factor in cardiomyocytes plated on the ECM protein laminin or fibronectin. In contrast, cardiomyocytes plated on the non-adhesive substrate gelatin exhibited a reduced capacity to undergo these PE-stimulated hypertrophic changes. In cardiomyocytes cultured on ECM, PE stimulated a rapid increase in tyrosine phosphorylation of focal adhesion proteins including FAK, paxillin, and p130 Crk-associated substrate and subsequent formation of peripheral focal complexes. Inhibition of the PE-induced hypertrophic response by genistein and herbimycin-A indicated a requirement for protein tyrosine kinases in PE signaling. To determine whether activation of FAK is required for PE-induced hypertrophy, a dominant-interfering mutant form of FAK, termed FRNK (FAK-related non-kinase), was ectopically expressed in cardiomyocytes using a replication-defective adenovirus expression system. FRNK expression attenuated PE-stimulated hypertrophy as assessed by cell size, sarcomeric organization, and induction of atrial natriuretic factor. These data indicate that the signal transduction pathways leading to cardiomyocyte hypertrophy are strongly influenced by and/or dependent upon an integrin-mediated signaling process requiring FAK.     

Specific interactions between Epac1, β-arrestin2 and PDE4D5 regulate β-adrenergic receptor subtype differential effects on cardiac hypertrophic signaling

β1 and β2 adrenergic receptors (βARs) are highly homologous but fulfill distinct physiological and pathophysiological roles. Here we show that both βAR subtypes activate the cAMP-binding protein Epac1, but they differentially affect its signaling. The distinct effects of βARs on Epac1 downstream effectors, the small G proteins Rap1 and H-Ras, involve different modes of interaction of Epac1 with the scaffolding protein β-arrestin2 and the cAMP-specific phosphodiesterase (PDE) variant PDE4D5. We found that β-arrestin2 acts as a scaffold for Epac1 and is necessary for Epac1 coupling to H-Ras. Accordingly, knockdown of β-arrestin2 prevented Epac1-induced histone deacetylase 4 (HDAC4) nuclear export and cardiac myocyte hypertrophy upon β1AR activation. Moreover, Epac1 competed with PDE4D5 for interaction with β-arrestin2 following β2AR activation. Dissociation of the PDE4D5–β-arrestin2 complex allowed the recruitment of Epac1 to β2AR and induced a switch from β2AR non-hypertrophic signaling to a β1AR-like pro-hypertrophic signaling cascade. These findings have implications for understanding the molecular basis of cardiac myocyte remodeling and other cellular processes in which βAR subtypes exert opposing effects.     

Overexpression of microRNA-99a Attenuates Cardiac Hypertrophy

Pathological cardiomyocyte hypertrophy is associated with significantly increased risk of heart failure, one of the leading medical causes of mortality worldwide. MicroRNAs are known to be involved in pathological cardiac remodeling. However, whether miR-99a participates in the signaling cascade leading to cardiac hypertrophy is unknown. To evaluate the role of miR-99a in cardiac hypertrophy, we assessed the expression of miR-99a in hypertrophic cardiomyocytes induced by isoprenaline (ISO)/angiotensin-II (Ang II) and in mice model of cardiac hypertrophy induced by transverse aortic constriction (TAC). Expression of miR-99a was evaluated in these hypertrophic cells and hearts. We also found that miR-99a expression was highly correlated with cardiac function of mice with heart failure (8 weeks after TAC surgery). Overexpression of miR-99a attenuated cardiac hypertrophy in TAC mice and cellular hypertrophy in stimuli treated cardiomyocytes through down-regulation of expression of mammalian target of rapamycin (mTOR). These results indicate that miR-99a negatively regulates physiological hypertrophy through mTOR signaling pathway, which may provide a new therapeutic approach for pressure-overload heart failure.     

MLP参与心肌肥大发生过程与Calcineurin/NFAT信号通路有关

心肌肥大是心肌细胞面对多种病理刺激时的共同反应,以心肌细胞体积增大和胚胎期基因的重新表达为标志.心肌发育调控基因肌肉LIM蛋白(muscle LIM protein,MLP)的表达异常与心肌肥大有关.为研究MLP参与心肌肥大发生的分子机制,采用去氧肾上腺素（phenylephrine, PE）刺激大鼠原代培养心肌细胞,建立心肌细胞肥大模型,采用RNAi技术敲减MLP的表达,分析MLP与肥大信号通路钙调神经磷酸酶（calcineurin）/活化T细胞核因子（nuclear factor of activated T-cells, NFAT）的关系.结果显示, 原代培养的心肌细胞经一定浓度的PE刺激后细胞表面积增加,肥大标志蛋白ANP、BNP表达增高,并伴有MLP表达上调. RNAi方法敲减MLP的表达则明显抑制PE诱导的心肌细胞表面积增加和BNP表达增高,并且直接 影响NFAT的转录激活活性,提示MLP与心肌肥大的发生密切相关,并且可能是通过calcineurin/NFAT信号通路而参与心肌肥大的发生.     

Somatostatin receptor-2 negatively regulates β-adrenergic receptor mediated Ca dependent signaling pathways in H9c2 cells

In the present study, we report that somatostatin receptor 2 (SSTR2) plays a crucial role in modulation of β₁AR and β₂AR mediated signaling pathways that are associated with increased intracellular Ca^2 + and cardiac complications. In H9c2 cells, SSTR2 colocalizes with β₁AR or β₂AR in receptor specific manner. SSTR2 selective agonist inhibits isoproterenol and formoterol stimulated cAMP formation and PKA phosphorylation in concentration dependent manner. In the presence of SSTR2 agonist, the expression of PKCα and PKCβ was comparable to the basal condition, however SSTR2 agonist inhibits isoproterenol or formoterol induced PKCα and PKCβ expression, respectively. Furthermore, the activation of SSTR2 not only inhibits calcineurin expression and its activity, but also blocks NFAT dephosphorylation and its nuclear translocation. SSTR2 selective agonist abrogates isoproterenol mediated increase in cell size and protein content (an index of hypertrophy). Taken together, the results described here provide direct evidence in support of cardiac protective role of SSTR2 via modulation of Ca^2 + associated signaling pathways attributed to cardiac hypertrophy.     

LncRNA SNHG1 exerts a protective role in cardiomyocytes hypertrophy via targeting miR‐15a‐5p/HMGA1 axis

Heart failure preceded by pathological cardiac hypertrophy is a leading cause of death. Long noncoding RNA small nucleolar RNA host gene 1 (SNHG1) was reported to inhibit cardiomyocytes apoptosis, but the role and underlying mechanism of SNHG1 in pathological cardiac hypertrophy have not yet been understood. This study was designed to investigate the role and molecular mechanism of SNHG1 in regulating cardiac hypertrophy. We found that SNHG1 was upregulated during cardiac hypertrophy both in vivo (transverse aortic constriction treatment) and in vitro (phenylephrine [PE] treatment). SNHG1 overexpression attenuated the cardiomyocytes hypertrophy induced by PE, while SNHG1 inhibition promoted hypertrophic response of cardiomyocytes. Furthermore, SNHG1 and high‐mobility group AT‐hook 1 (HMGA1) were confirmed to be targets of miR‐15a‐5p. SNHG1 promoted HMGA1 expression by sponging miR‐15a‐5p, eventually attenuating cardiomyocytes hypertrophy. There data revealed a novel protective mechanism of SNHG1 in cardiomyocytes hypertrophy. Thus, targeting of SNHG1‐related pathway may be therapeutically harnessed to treat cardiac hypertrophy.     

Beta-adrenergic receptor-mediated DNA synthesis in cardiac fibroblasts is dependent on transactivation of the epidermal growth factor receptor and subsequent activation of extracellular signal-regulated kinases

Cardiac hypertrophy often leads to heart failure and is associated with abnormal myocardial adrenergic signaling. This enlargement of myocardial mass can involve not only an increase in cardiomyocyte size, but increased proliferation of cardiac fibroblasts. A potential key player in the cardiac hypertrophic response is the ERK family of MAPKs. To gain mechanistic insight into adrenergic regulation of myocardial mitogenic signaling, we examined beta-adrenergic receptor (beta-AR) stimulation of ERK activation and DNA synthesis in cultured adult rat cardiac fibroblasts, including the involvement of tyrosine kinases in this signaling pathway. Addition of the beta-AR agonist isoproterenol (ISO) to serum-starved cells induced DNA synthesis in a dose-dependent manner, and this was inhibited by selective inhibitors of the epidermal growth factor receptor (EGFR). Importantly and in agreement with the involvement of MAPKs and the EGFR in this response in cardiac fibroblasts, the EGFR inhibitor AG1478 attenuated ISO-induced ERK phosphorylation. Moreover, pretreatment with PP2, a selective inhibitor of the Src tyrosine kinase, attenuated both ISO-mediated EGFR phosphorylation and ERK activation. Furthermore, studies in these cardiac fibroblasts showed that phosphatidylinositol 3-kinase contributed to beta-AR-mediated ERK activation, but not to EGFR activation. Finally, studies using selective inhibitors of matrix metalloproteases indicated that they and heparin-bound EGF shedding were involved in beta-AR-induced ERK activation and subsequent DNA synthesis in cardiac fibroblasts. Because these cells primarily express the beta(2)-AR subtype, our findings indicate that beta(2)-AR-mediated EGFR transactivation of intracellular tyrosine kinase signaling pathways is the major signaling pathway responsible for the adrenergic stimulation of mitogenesis of cardiac fibroblasts.     

Plantago asiatica L. seeds extract protects against cardiomyocyte injury in isoproterenol- induced cardiac hypertrophy by inhibiting excessive autophagy and apoptosis in mice

BackgroundCardiac hypertrophy is the early stage of many heart diseases, such as coronary heart disease, hypertension, valvular dysfunction and cardiomyopathy. Cardiomyocyte autophagy and apoptosis play an important role in the process of cardiac hypertrophic response. Plantago asiatica L. seeds extract (PASE) is prepared from a traditional herbal medicine in Asia with tremendous pharmacological activities. However, whether PASE could relieve cardiac hypertrophy has not been elucidated. The present study is aimed to investigate the effect of PASE on cardiac hypertrophy and explore its potential underlying mechanism.MethodsCardiac hypertrophy was induced in C57BL/6 mice by subcutaneous injection of isoproterenol (ISO) for two weeks. Meanwhile, the mice were intraperitoneally injected with PASE at dosages of 20, 40 and 80 mg/kg/day. Cardiac hypertrophy was evaluated by echocardiographic examination, haematoxylin and eosin staining and quantitative real-time polymerase chain reaction. Expressions of proteins involved in autophagy and apoptosis such as Beclin1, p62, LC3II, Bax, Bcl-2 and Cleaved-caspase-3 were detected by western blot analysis. Western blot, transient transfection, acridine orange staining, TUNEL staining and autophagy inducer were used to observe the effect and explore the mechanism of PASE on cardiomyocyte and H9c2 cells with excessive autophagy and apoptosis induced by ISO.ResultsISO induction for two weeks disturbed the myocardial contractility and cardiac function of left ventricles of mice. PASE treated mice showed significantly improved cardiac function indexes, including EF, FS, SV and CO, compared with the ISO group. Treatment with PASE also decreased the heart weight/body weight ratio and cardiomyocyte size, and downregulated the mRNA and protein expressions of hypertrophic markers ANP, BNP, and β-MHC. Furthermore, the changes of autophagy and apoptosis markers, such as LC3II, Beclin1, p62, Bcl-2, Bax and Cleaved-caspase-3 induced by ISO were resumed by PASE treatment. Consistently, PASE demonstrated similar effects on ISO-induced H9c2 cells as it did in vivo. In addition, PASE could counteract the increased autophagy induced by the autophagy inducer, rapamycin.ConclusionPASE attenuated ISO-induced cardiac hypertrophy in mice by inhibiting excessive autophagy and apoptosis in cardiomyocytes. The novel findings may pave the way for the clinical usage of PASE for the prevention of heart diseases related with cardiac hypertrophy.     

GPR39 promotes cardiac hypertrophy by regulating the AMPK–mTOR pathway and protein synthesis

Hypertrophic growth of the cardiomyocytes is one of the core mechanisms underlying cardiac hypertrophy. However, the mechanism underlying cardiac hypertrophy remains not fully understood. Here we provided evidence that G protein-coupled receptor 39 (GPR39) promotes cardiac hypertrophy via inhibiting AMP-activated protein kinase (AMPK) signaling. GRP39 expression is overexpressed in hypertrophic hearts of humans and transverse aortic constriction (TAC)-induced cardiac hypertrophy in mice. In neonatal cardiomyocytes, adenovirus-mediated overexpression of GPR39 promoted angiotensin II-induced cardiac hypertrophy, while GPR39 knockdown repressed hypertrophic response. Adeno-associated virus 9-mediated knockdown of GPR39 suppressed TAC-induced decline in fraction shortening and ejection fraction, increase in heart weight and cardiomyocyte size, as well as overexpression of hypertrophic fetal genes. A mechanism study demonstrated that GPR39 repressed the activation of AMPK to activate the mammalian target of rapamycin (mTOR) and ribosomal protein S6 kinase β-1 (S6K1), subsequently promoted de novo protein synthesis. Inhibition of mTOR with rapamycin blocked the effects of GPR39 overexpression on protein synthesis and repressed cardiac hypertrophy. Collectively, our findings demonstrated that GPR39 promoted cardiac hypertrophy via regulating the AMPK–mTOR–S6K1 signaling pathway, and GRP39 can be targeted for the treatment of cardiac hypertrophy.     

LncRNA TUG1 alleviates cardiac hypertrophy by targeting miR-34a/DKK1/Wnt-β-catenin signalling

The current study was designed to explore the role and underlying mechanism of lncRNA taurine up-regulated gene 1 (TUG1) in cardiac hypertrophy. Mice were treated by transverse aortic constriction (TAC) surgery to induce cardiac hypertrophy, and cardiomyocytes were treated by phenylephrine (PE) to induce hypertrophic phenotype. Haematoxylin-eosin (HE), wheat germ agglutinin (WGA) and immunofluorescence (IF) were used to examine morphological alterations. Real-time PCR, Western blots and IF staining were used to detect the expression of RNAs and proteins. Luciferase assay and RNA pull-down assay were used to verify the interaction. It is revealed that TUG1 was up-regulated in the hearts of mice treated by TAC surgery and in PE-induced cardiomyocytes. Functionally, overexpression of TUG1 alleviated cardiac hypertrophy both in vivo and in vitro. Mechanically, TUG1 sponged and sequestered miR-34a to increase the Dickkopf 1 (DKK1) level, which eventually inhibited the activation of Wnt/β-catenin signalling. In conclusion, the current study reported the protective role and regulatory mechanism of TUG1 in cardiac hypertrophy and suggested that TUG1 may serve as a novel molecular target for treating cardiac hypertrophy.     

The role of β‐adrenergic receptors and p38MAPK signaling pathways in physiological processes of cardiosphere‐derived cells

The effects of β adrenergic receptors (β‐ARs) and p38 mitogen‐activated protein kinases (MAPK) pathways on cardiosphere‐derived cells (CDCs) are largely unknown. This study aimed to investigate the roles of β‐ARs and p38MAPK pathways on the proliferation, apoptosis, and differentiation capacity of CDCs. The CDCs were treated with β1‐AR blocker (Met group), β2‐AR antagonist (ICI group), and p38MAPK inhibitor (SB group), non‐selective β‐AR blocker (PRO group), and β‐AR agonist (ISO group). The viability, apoptotic rate and differentiation status of CDCs were determined by MST‐1 assay, flow cytometery, and Western blot, respectively. The CDCs viability significantly reduced in ICI group (all P < 0.05), and SB group had a significant high viability after 48 h treatment (P < 0.05). Compared with control group, all treated groups had a low apoptotic rate. After treatment for 72 h, ISO treatment elevated the expression of Nkx2.5, and could partially or fully attenuate the inhibitory effects of β‐AR antagonists and/or p38MAPK inhibitor. A similar overall trend of protein expression levels among all groups could be observed between protein pairs of cTnT and β1‐AR as well as c‐Kit and β2‐AR, respectively. These results suggested that β‐ARs and p38MAPK signaling pathways play crucial roles in the proliferation and differentiation of CDCs. Our findings should be helpful for better understanding the molecular mechanism underlying the physiological processes of CDCs.     

Distinct signaling functions for Shc isoforms in the heart

Thrombin activates protease-activated receptor-1 (PAR-1) and engages signaling pathways that influence the growth and survival of cardiomyocytes as well as extracellular matrix remodeling by cardiac fibroblasts. This study examines the role of Shc proteins in PAR-1-dependent signaling pathways that influence ventricular remodeling. We show that thrombin increases p46Shc/p52Shc phosphorylation at Tyr(239)/Tyr(240) and Tyr(317) (and p66Shc-Ser(36) phosphorylation) via a pertussis toxin-insensitive epidermal growth factor receptor (EGFR) transactivation pathway in cardiac fibroblasts; p66Shc-Ser(36) phosphorylation is via a MEK-dependent mechanism. In contrast, cardiac fibroblasts express beta(2)-adrenergic receptors that activate ERK through a pertussis toxin-sensitive EGFR transactivation pathway that does not involve Shc isoforms or lead to p66Shc-Ser(36) phosphorylation. In cardiomyocytes, thrombin triggers MEK-dependent p66Shc-Ser(36) phosphorylation, but this is not via EGFR transactivation (or associated with Shc-Tyr(239)/Tyr(240) and/or Tyr(317) phosphorylation). Importantly, p66Shc protein expression is detected in neonatal, but not adult, cardiomyocytes; p66Shc expression is induced (via a mechanism that requires protein kinase C and MEK activity) by Pasteurella multocida toxin, a Galpha(q) agonist that promotes cardiomyocyte hypertrophy. These results identify novel regulation of individual Shc isoforms in receptor-dependent pathways leading to cardiac hypertrophy and the transition to heart failure. The observations that p66Shc expression is induced by a Galpha(q) agonist and that PAR-1 activation leads to p66Shc-Ser(36) phosphorylation identifies p66Shc as a novel candidate hypertrophy-induced mediator of cardiomyocyte apoptosis and heart failure.