Piceatannol attenuates cardiac hypertrophy in an animal model through regulation of the expression and binding of the transcription factor GATA binding factor 6

Piceatannol is found in grapes, passion fruit, and Japanese knotweed. Piceatannol pretreatment suppresses cardiac hypertrophy induced by isoproterenol as assessed by heart weight/body weight ratio, cross-sectional area, and expression of hypertrophic markers. The anti-hypertrophic effect of piceatannol in rat neonatal cardiomyocytes is the same as that in vivo. Piceatannol inhibits lentiviral-GATA6-induced cardiomyocyte hypertrophy. Furthermore, piceatannol reduces the interaction between GATA4 and GATA6 as well as the DNA-binding activity of endogenous GATA6 in the ANP promoter. Our results suggest that piceatannol may be a novel therapeutic agent for the prevention of cardiac hypertrophy.Structured summary of protein interactionsGATA6 physically interacts with GATA4 by anti V5 tag coimmunoprecipitation (View interaction)     

The histone acetyltransferase MOF overexpression blunts cardiac hypertrophy by targeting ROS in mice

Imbalance between histone acetylation/deacetylation critically participates in the expression of hypertrophic fetal genes and development of cardiac hypertrophy. While histone deacetylases play dual roles in hypertrophy, current evidence reveals that histone acetyltransferase such as p300 and PCAF act as pro-hypertrophic factors. However, it remains elusive whether some histone acetyltransferases can prevent the development of hypertrophy. Males absent on the first (MOF) is a histone acetyltransferase belonging to the MYST (MOZ, Ybf2/Sas3, Sas2 and TIP60) family. Here in this study, we reported that MOF expression was down-regulated in failing human hearts and hypertrophic murine hearts at protein and mRNA levels. To evaluate the roles of MOF in cardiac hypertrophy, we generated cardiac-specific MOF transgenic mice. MOF transgenic mice did not show any differences from their wide-type littermates at baseline. However, cardiac-specific MOF overexpression protected mice from transverse aortic constriction (TAC)-induced cardiac hypertrophy, with reduced radios of heart weight (HW)/body weight (BW), lung weight/BW and HW/tibia length, decreased left ventricular wall thickness and increased fractional shortening. We also observed lower expression of hypertrophic fetal genes in TAC-challenged MOF transgenic mice compared with that of wide-type mice. Mechanically, MOF overexpression increased the expression of Catalase and MnSOD, which blocked TAC-induced ROS and ROS downstream c-Raf-MEK-ERK pathway that promotes hypertrophy. Taken together, our findings identify a novel anti-hypertrophic role of MOF, and MOF is the first reported anti-hypertrophic histone acetyltransferase.     

Antihypertrophic effects of adiponectin on cardiomyocytes are associated with the inhibition of heparin-binding epidermal growth factor signaling

This study was aimed to investigate whether the antihypertrophic effects of adiponectin in murine hearts are associated with the modulation of HB-EGF signaling. We determined the myocardial expressions of adiponectin and adiponectin receptors, brain natriuretic peptide (BNP), and HB-EGF in normal and hypertrophied hearts of adiponectin knockout mice or wild-type mice with transverse aortic constriction (TAC). Then, we observed the effects of adiponectin on cardiac hypertrophy and HB-EGF signaling in cultured neonatal rat cardiomyocytes and whole hearts of adiponectin-null mice. The myocardial mRNA and protein expressions of adiponectin in the hypertrophied hearts were significantly downregulated, and the mRNA expression of adiponectin was inversely correlated with the heart-to-body weight ratio, BNP, and HB-EGF. The TAC-induced cardiac hypertrophy and EGF receptor (EGFR) activation in the adiponectin knockout mice were significantly greater than those in the wild-type mice. Furthermore, in vitro experiments revealed that adiponectin inhibited HB-EGF-stimulated protein synthesis, HB-EGF shedding, and EGFR phosphorylation. We conclude that the inhibition of HB-EGF mediated EGFR activation is one of the alternative mechanisms for the antihypertrophic action of adiponectin.     

Down-regulation of mitofusin-2 expression in cardiac hypertrophy in vitro and in vivo

Mitofusin-2 (Mfn2) suppresses smooth muscle cell proliferation through inhibition of the Ras-extracellular signal-regulated kinases (ERK1/2) pathway. Since the ERK1/2 pathway is implicated in mediating hypertrophic signaling, we studied the changes in Mfn2 in cardiac hypertrophy using in vitro and in vivo models. Phenylephrine was used to induce hypertrophy in neonatal rat ventricular myocytes (NRVMs). In vivo hypertrophy models included spontaneously hypertensive rats (SHR), pressure-overload hypertrophy by transverse aortic constriction (TAC), hypertrophy of non-infarcted myocardium following myocardial infarction (MI), and cardiomyopathy due to cardiac-restricted overexpression of beta(2)-adrenergic receptors (beta(2)-TG). We determined hypertrophic parameters and analysed expression of atrial natriuretic peptide (ANP) and Mfn2 by real-time PCR. Phosphorylated-ERK1/2 (phospho-ERK) was measured by Western blot. Mfn2 was downregulated in phenylephrine treated NRCMs (by approximately 40%), hypertrophied hearts from SHR (by approximately 80%), mice with TAC (at 1 and 3 weeks, by approximately 50%), and beta(2)-TG mice (by approximately 20%). However, Mfn2 was not downregulated in hypertrophied hearts with 15 weeks of TAC, nor in hypertrophied non-infarcted myocardium following MI. phospho-ERK1/2 was increased in hypertrophied myocardium at 1 week post-TAC, but not in non-infarcted myocardium after MI, indicating that downregulated Mfn2 may be accompanied by an increase of phospho-ERK1/2. This study shows, for the first time, downregulated Mfn2 expression in hypertrophied hearts, which depends on the etiology and time course of hypertrophy. Further study is required to examine the causal relationship between Mfn2 and cardiac hypertrophy.     

大鼠骨髓间充质干细胞条件培养液促进心肌细胞肥大

体外模拟心肌缺血微环境,研究骨髓间充质干细胞(MSCs)的旁分泌作用对心肌细胞的影响。以大鼠MSCs各时间点的条件培养液刺激心肌细胞,观察心肌细胞蛋白含量、[3H]-Leu掺入、ANF-荧光素酶(luciferase)表达和心肌细胞面积的变化。MSCs条件培养液处理心肌细胞后,与对照组相比较6h及9h时间点的条件培养液可明显增加心肌细胞蛋白含量、[3H]-Leu掺入、ANF-荧光素酶表达以及心肌细胞面积,其中以6h时间点条件培养液的作用最为显著(P<0.01)。MSCs条件培养液能够通过旁分泌作用刺激心肌细胞肥大,此现象提示移植入心肌缺血区MSCs可能通过旁分泌作用影响心肌细胞,从而参与细胞移植后心功能的改善。     

Expression and characterization of the Na+/H+ exchanger in the mammalian myocardium

We examined two expression systems for studying the Na⁺/H⁺ exchanger in the mammalian myocardium. Mammalian NHE1 with a hemagglutinin (HA) tag and was cloned behind the alpha myosin heavy chain promoter. Transgenic mice were made with wild type NHE1 protein or with a hyperactive NHE1 protein mutated at the calmodulin-binding domain. Three lines of transgenic mice were made of each cDNA with expression levels of each type varying from high to low. Higher levels and activity of the Na⁺/H⁺ exchanger were associated with decreased long-term survival of mice, and with dilated or hypertrophic cardiomyopathy. The exogenous NHE1 protein was present in freshly made cardiomyocytes from transgenic mice, however, expression from the alpha myosin heavy chain promoter declined rapidly and little exogenous NHE1 was apparent on the fourth day after cardiomyocyte isolation. To express NHE1 protein in isolated cardiomyocytes, we transferred a mutated form of the protein into an adenoviral expression system. Infection of neonatal rat cardiomyocytes resulted in robust expression of the exogenous NHE1 protein. The mutant form of the NHE1 protein could be distinguished from the endogenous Na⁺/H⁺ exchanger by its resistance to inhibition by amiloride analogs. Our results suggest that for in vivo studies on intact hearts and animals, expression in transgenic mice is an appropriate system, however for long-term studies on cardiomyocytes, this model is inappropriate due to waning expression from the alpha myosin heavy chain promoter. Therefore, infection by adenovirus is a superior system for long-term studies on cardiomyocytes in culture.     

The role of iNOS-derived NO in the antihypertrophic actions of B-type natriuretic peptide in neonatal rat cardiomyocytes

In the infarcted rat heart, the increase of NO occurs in the hypertrophied myocardium of non-infarcted areas and its antihypertrophic efficacy has been well established. As another endogenous regulator and the reliable index of heart pathology, B-type natriuretic peptide also exhibits the antihypertrophic properties in many tissues by elevating intracellular cGMP. Several studies indicate that natriuretic peptides family may exert some actions in part via a nitric oxide pathway following receptor-mediated stimulation of iNOS. Therefore, it raises our great interest to ask what role NO plays in the antihypertrophic actions of B-type natriuretic peptide in cardiomyocytes. Incubation of cardiomyocytes under mild hypoxia for 12 h caused a significant increase in cellular protein content, protein synthesis and cell surface sizes. This growth stimulation was suppressed by exogenous B-type natriuretic peptide in a concentration dependent manner. Furthermore, the generation of intracellular cGMP, the upregulation of iNOS mRNA expression, the increase of iNOS activity and subsequent nitrite generation in hypertrophic cardiomyocytes was also increased by B-type natriuretic peptide. AG, a selective iNOS inhibitor, inhibited the upregulation of iNOS expression and the increase of iNOS activity by the combination of B-type natriuretic peptide/mild hypoxia or by the combination of 8-bromo-cGMP/mild hypoxia. Rp-8-br-cGMP, cGMP dependent protein kinase inhibitor, attenuated the actions of B-type natriuretic peptide and 8-bromo-cGMP which increases intracellular cGMP independent of B-type natriuretic peptide. In conclusion, our present data suggest that B-type natriuretic peptide exerted the antihypertrophic effects in cardiomyocytes, which was partially attributed to induction of iNOS-derived NO by cGMP pathway.     

Hepatic morphological alterations,glycogen content and cytochrome P450 activities in rats treated chronically with N<Superscript>ω</Superscript>-nitro-L-arginine methyl ester (L-NAME)

Chronic treatment of rats with N(omega)-nitro-L-arginine methyl ester (L-NAME), an inhibitor of nitric oxide (NO) biosynthesis, results in hypertension mediated partly by enhanced angiotensin-I-converting enzyme (ACE) activity. We examined the influence of L-NAME on rat liver morphology, on hepatic glycogen, cholesterol, and triglyceride content, and on the activities of the cytochrome P450 isoforms CYP1A1/2, CYP2B1/2, CYP2C11, and CYP2E1. Male Wistar rats were treated with L-NAME (20 mg/rat per day via drinking water) for 2, 4, and 8 weeks, and their livers were then removed for analysis. Enzymatic induction was produced by treating rats with phenobarbital (to induce CYP2B1/2), beta-naphthoflavone (to induce CYP1A1/2), or pyrazole (to induce CYP2E1). L-NAME significantly elevated blood pressure; this was reversed by concomitant treatment with enalapril (ACE inhibitor) or losartan (angiotensin II AT(1) receptor antagonist). L-NAME caused vascular hypertrophy in hepatic arteries, with perivascular and interstitial fibrosis involving collagen deposition. Hepatic glycogen content also significantly increased. L-NAME did not affect fasting glucose levels but significantly reduced insulin levels and increased the insulin sensitivity of rats, based on an intraperitoneal glucose tolerance test. Immunoblotting experiments indicated enhanced phosphorylation of protein kinase B and of glycogen synthase kinase 3. All these changes were reversed by concomitant treatment with enalapril or losartan. L-NAME had no effect on hepatic cholesterol or triglyceride content or on the basal or drug-induced activities and protein expression of the cytochrome P450 isoforms. Thus, the chronic inhibition of NO biosynthesis produced hepatic morphological alterations and changes in glycogen metabolism mediated by the renin-angiotensin system. The increase in hepatic glycogen content probably resulted from enhanced glycogen synthase activity following the inhibition of glycogen synthase kinase 3 by phosphorylation.     

Deletion of MLIP (Muscle-enriched A-type Lamin-interacting Protein) Leads to Cardiac Hyperactivation of Akt/Mammalian Target of Rapamycin (mTOR) and Impaired Cardiac Adaptation

Aging and diseases generally result from tissue inability to maintain homeostasis through adaptation. The adult heart is particularly vulnerable to disequilibrium in homeostasis because its regenerative abilities are limited. Here, we report that MLIP (muscle enriched A-type lamin-interacting protein), a unique protein of unknown function, is required for proper cardiac adaptation. Mlip^−/− mice exhibited normal cardiac function despite myocardial metabolic abnormalities and cardiac-specific overactivation of Akt/mTOR pathways. Cardiac-specific MLIP overexpression led to an inhibition of Akt/mTOR, providing evidence of a direct impact of MLIP on these key signaling pathways. Mlip^−/− hearts showed an impaired capacity to adapt to stress (isoproterenol-induced hypertrophy), likely because of deregulated Akt/mTOR activity. Genome-wide association studies showed a genetic association between Mlip and early response to cardiac stress, supporting the role of MLIP in cardiac adaptation. Together, these results revealed that MLIP is required for normal myocardial adaptation to stress through integrated regulation of the Akt/mTOR pathways.