Atrial Natriuretic Factor Expression Research Articles

Stem cell-derived extracellular vesicles reduce the expression of molecules involved in cardiac hypertrophy-In a model of human-induced pluripotent stem cell-derived cardiomyocytes.

Cardiac pathological hypertrophy is the major risk factor that usually progresses to heart failure. We hypothesized that extracellular vesicles (EVs), known to act as important mediators in regulating physiological and pathological functions, could have the potential to reduce the cardiac hypertrophy and the ensuing cardiovascular diseases. Herein, the effects of mesenchymal stem cell-derived extracellular vesicles (EV-MSCs) on cardiac hypertrophy were investigated. EVs were isolated from the secretome of human adipose tissue-derived stem cells (EV-ADSCs) or bone marrow-derived stem cells (EV-BMMSCs). Human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) were stimulated with AngII and TGF-β1, in absence or presence of EVs. The results showed that exposure of hiPSC-CMs to AngII and TGF-β1 generated in vitro model of hypertrophic cardiomyocytes characterized by increases in surface area, reactive oxygen species production, protein expression of cardiac-specific biomarkers atrial natriuretic factor, migration inhibitory factor, cTnI, COL1A1, Cx43, α-SMA and signalling molecules SMAD2 and NF-kBp50. The presence of EV-ADSCs or EV-BMMSCs in the hiPSC-CM culture along with hypertrophic stimuli reduced the protein expressions of hypertrophic specific markers (ANF, MIF, cTnI, COL1A1) and the gene expressions of IL-6 molecule involved in inflammatory process associated with cardiac hypertrophy and transcription factors SMAD2, SMAD3, cJUN, cFOS with role in cardiomyocyte hypertrophic response induced by AngII and TGF-β1. The EV-ADSCs were more effective in reducing the protein expressions of hypertrophic and inflammatory markers, while EV-BMMSCs in reducing the gene expressions of transcription factors. Notably, neither EV-ADSCs nor EV-BMMSCs induced significant changes in cardiac biomarkers Cx43, α-SMA and fibronectin. These different effects of stem cell-derived EVs could be attributed to their miRNA content: some miRNAs (miR-126-3p, miR-222-3p, miR-30e-5p, miR-181b-5p, miR-124-3p, miR-155-5p, miR-210-3p hsa-miR-221-3p) were expressed in both types of EVs and others only in EV-ADSCs (miR-181a-5p, miR-185-5p, miR-21-5p) or in EV-BMMSCs (miR-143-3p, miR-146a-5p, miR-93-5p), some of these attenuating the cardiac hypertrophy while others enhance it. In conclusion, in hiPSC-CMs the stem cell-derived EVs through their cargo reduced the expression of hypertrophic specific markers and molecules involved in inflammatory process associated with cardiac hypertrophy. The data suggest the EV potential to act as therapeutic mediators to reduce cardiac hypertrophy and possibly the subsequent cardiovascular events.

Omentin inhibits the resistin-induced hypertrophy of H9c2 cardiomyoblasts by inhibiting the TLR4/MyD88/NF-κB signaling pathway.

The recently identified adipocytokine omentin was previously found to be expressed mainly in human omental and visceral adipose tissues. As such, reduced plasma concentrations of omentin were revealed to be associated with increased risks of cardiovascular diseases. Omentin has also been previously demonstrated to exert anti-inflammatory effects. By contrast, resistin is a protein that has been associated with obesity and type-2 diabetes mellitus, and the serum concentration of resistin is increased significantly in these populations. Resistin is involved in mediating inflammation development, where they can promote cardiac hypertrophy in humans through toll-like receptor 4 (TLR4)-related signaling. In the present study, the potential effects of omentin on resistin-induced hypertrophy in H9c2 cardiomyoblasts were investigated. In the absence/presence of omentin, H9c2 cardiomyoblasts were treated with resistin. Omentin was found to significantly inhibit resistin-induced increases in the surface area of H9c2 cardiomyoblasts as determined by immunofluorescence. In addition, omentin significantly inhibited resistin-induced increases in the mRNA expression of atrial natriuretic factor, brain natriuretic peptide, β-myosin heavy chain (which is a characteristic feature of cardiac hypertrophy) and TLR4, which was determined using reverse-transcription-quantitative PCR. According to western blotting results, omentin significantly inhibited resistin-induced ERK phosphorylation, which is an important mediator of cardiomyoblast hypertrophy. Furthermore, omentin significantly inhibited resistin-induced protein expression of TLR4, myeloid differentiation factor 88 (MyD88) and NF-κB phosphorylation, both of which are important members of inflammatory signaling. To conclude, data from the present study suggest that omentin can inhibit resistin-induced H9c2 cardiomyoblast hypertrophy through inhibition of the TLR4/MyD88/NF-κB signaling pathway. Therefore, omentin serve as an attractive therapeutic target against resistin-induced cardiac hypertrophy.

Metformin suppresses phenylephrine-induced hypertrophic responses by inhibiting p300-HAT activity in cardiomyocytes

IntroductionHeart failure is the final pathway for a wide spectrum of myocardial stress, including hypertension and myocardial infarction. However, the potential effects of metformin on cardiac hypertrophy are still unclear. PurposeThe purpose of this study was to investigate whether metformin leads to suppression of hypertrophic responses in cardiomyocytes. Methods and resultsTo investigate whether metformin inhibited p300-histone acetyltransferase (HAT), we performed an in vitro HAT assay. Metformin directly inhibited p300-mediated acetylation of histone-H3K9. To examine the effects of metformin on hypertrophic responses, cardiomyocytes prepared from neonatal rats were treated with metformin and stimulated with saline or phenylephrine (PE), a α1-adrenergic agonist for 48 h. PE stimulus showed an increase in cell size, myofibrillar organization, expression of the endogenous atrial natriuretic factor and brain natriuretic peptide genes, and acetylation of histone-H3K9 compared with saline-treated cells. These PE-induced changes were inhibited by metformin. Next, to examine the effect of metformin on p300-mediated hypertrophy, cardiomyocytes were transfected with expression vector of p300. Metformin significantly suppressed p300-induced hypertrophic responses and acetylation of histone-H3K9. ConclusionsThe study demonstrates that metformin can suppress PE-induced and p300-mediated hypertrophic responses. Metformin may be useful for the treatment of patients with diabetes and heart failure.

BioID‐Based Network Analysis to Probe Alterations and Functional Consequences of Adenylyl Cyclase‐Containing Macromolecular Complexes in Cardiomyocytes

Numerous cardiac functions rely on the proper formation of macromolecular complexes that regulate cyclic AMP (cAMP) signaling. Localized cAMP production by adenylyl cyclase (AC) is coordinated by scaffolding proteins such as A-kinase anchoring proteins (AKAPs) that anchor AC, the cAMP effector protein kinase A (PKA), along with downstream targets of PKA and other regulators. Alteration of AKAP complexes can promote hypertrophy and changes in contractility, leading to cardiac remodeling and cardiomyopathy. However, the composition and dynamics of AC-containing complexes in heart disease remain mostly unknown. We have therefore carried out a comprehensive study of AC complexes in heart using BioID (proximity-dependent identification of near-neighbor interaction) proteomics technique to probe alterations in complex formation resulting from hypertrophic cardiomyopathy. We have fused the mini-Turbo biotin ligase with three different AC isoforms, two AKAPs, and control GFP and have expressed these fusion proteins in neonatal cardiomyocytes using adenoviruses. Neonatal cardiomyocytes were subsequently treated with β adrenergic receptor agonist norepinephrine (NE) for 48 hours, incubated with biotin for 3 hours, and harvested for streptavidin pull-down and mass spectrometry. Norepinephrine treatment induced hypertrophy, leading to a 229% increase in cell size and a 295% increase in atrial natriuretic factor expression. Clear differences in biotinylation patterns were observed with and without NE by western blot analysis for each of the mini-Turbo fusions. Biotin-labeled proteins resulting from AC or AKAP expression were sorted by Significance Analysis of INTeractome (SAINT) and bioinformatics analysis identified unique protein patterns with different AC and AKAP isoforms under physiological and hypertrophic conditions. Our comprehensive network analysis of localized complex formation is expected to provide important insights for understanding the functional changes of signalosomes and signaling pathways in disease states.

Smyd1 Orchestrates Early Heart Development Through Positive and Negative Gene Regulation.

SET and MYND domain-containing protein 1 (Smyd1) is a striated muscle-specific histone methyltransferase. Our previous work demonstrated that deletion of Smyd1 in either cardiomyocytes or the outflow tract (OFT) resulted in embryonic lethality at E9.5, with cardiac structural defects such as truncation of the OFT and right ventricle and impaired expansion and proliferation of the second heart field (SHF). The cardiac phenotype was accompanied by the downregulation of ISL LIM Homeobox 1 (Isl1) and upregulation of atrial natriuretic factor (ANF). However, the mechanisms of Smyd1 regulating Isl1 and ANF during embryonic heart development remain to be elucidated. Here, we employed various biochemical and molecular biological approaches including chromatin immunoprecipitation polymerase chain reaction (ChIP-PCR), pGL3 fluorescence reporter system, and co-immunoprecipitation (CoIP) and found that Smyd1 interacted with absent small homeotic-2-like protein (ASH2L) and activated the promoter of Isl1 by trimethylating H3K4. We also found that Smyd1 associated with HDAC to repress ANF expression using trichostatin A (TSA), a deacetylase inhibitor. In conclusion, Smyd1 participates in early heart development by upregulating the expression of Isl1 and downregulating the expression of ANF.

Whole-Exome Sequencing and hiPSC Cardiomyocyte Models Identify MYRIP, TRAPPC11, and SLC27A6 of Potential Importance to Left Ventricular Hypertrophy in an African Ancestry Population.

Background: Indices of left ventricular (LV) structure and geometry represent useful intermediate phenotypes related to LV hypertrophy (LVH), a predictor of cardiovascular (CV) disease (CVD) outcomes.Methods and Results: We conducted an exome-wide association study of LV mass (LVM) adjusted to height2.7, LV internal diastolic dimension (LVIDD), and relative wall thickness (RWT) among 1,364 participants of African ancestry (AAs) in the Hypertension Genetic Epidemiology Network (HyperGEN). Both single-variant and gene-based sequence kernel association tests were performed to examine whether common and rare coding variants contribute to variation in echocardiographic traits in AAs. We then used a data-driven procedure to prioritize and select genes for functional validation using a human induced pluripotent stem cell cardiomyocyte (hiPSC-CM) model. Three genes [myosin VIIA and Rab interacting protein (MYRIP), trafficking protein particle complex 11 (TRAPPC11), and solute carrier family 27 member 6 (SLC27A6)] were prioritized based on statistical significance, variant functional annotations, gene expression in the hiPSC-CM model, and prior biological evidence and were subsequently knocked down in the hiPSC-CM model. Expression profiling of hypertrophic gene markers in the knockdowns suggested a decrease in hypertrophic expression profiles. MYRIP knockdowns showed a significant decrease in atrial natriuretic factor (NPPA) and brain natriuretic peptide (NPPB) expression. Knockdowns of the heart long chain fatty acid (FA) transporter SLC27A6 resulted in downregulated caveolin 3 (CAV3) expression, which has been linked to hypertrophic phenotypes in animal models. Finally, TRAPPC11 knockdown was linked to deficient calcium handling.Conclusions: The three genes are biologically plausible candidates that provide new insight to hypertrophic pathways.

Protective effect of histone methyltransferase NSD3 on ISO-induced cardiac hypertrophy.

Nuclear receptor-binding SET domain 3 (NSD3) is a lysine methyltransferase that plays important roles in multiple biological activities; however; its potential roles in the cardiovascular system remain unknown. In this study, we found that NSD3 expression is reduced by isoproterenol (ISO) stimuli both invitro and invivo. Overexpression of NSD3 attenuates ISO-induced cardiomyocyte hypertrophy. Mechanistically, ISO treatment decreases H3K27me2/3 modifications on the atrial natriuretic factor (ANF) promoter by suppressing NSD3 and inhibits the association between NSD3 and bromodomain-containing protein 4 (BRD4), thus suppressing the BRD4-mediated H3K27ac modifications, which ultimately promote ANF transcription and cardiomyocyte hypertrophy. In conclusion, NSD3 decreases ANF expression and, thereby, attenuates ISO-induced cardiomyocyte hypertrophy.

Sodium ferulate inhibits myocardial hypertrophy induced by abdominal coarctation in rats: Involvement of cardiac PKC and MAPK signaling pathways.

Sodium ferulate (SF) is the sodium salt of ferulic acid which is an active ingredient of Radix Angelica Sinensis and Ligusticum chuanxiong hort. Here, we investigated SF inhibition in a rat model of myocardial hypertrophy induced by coarctation of the abdominal aorta. Following coarctation, rats were given SF (20, 40, and 80 mg/kg/day) for 25 consecutive days. We characterized myocardial hypertrophy using myocardial hypertrophic parameters, histopathology, and gene expression of atrial natriuretic factor (ANF) -a gene related to myocardial hypertrophy. We detected the levels of angiotensin II (Ang II) and endothelin-1 (ET-1), protein kinase C beta (PKC-β), Raf-1, extracellular regulated protein kinase 1/2 (ERK1/2), and mitogen-activated protein kinase phosphatase-1 (MKP-1) in myocardium. Notably, coarctation of the abdominal aorta increases myocardial hypertrophic parameters, cardiac myocyte diameter, the concentration of Ang II and ET-1 in myocardium, and gene expression of ANF. SF significantly ameliorates myocardial hypertrophy caused by coarctation of the abdominal aorta; reduces concentrations of Ang II and ET-1; suppresses the overexpression of ANF, PKC-β, Raf-1, and ERK1/2; and increases the expression of MKP-1. These results indicate that SF alleviates myocardial hypertrophy induced by coarctation of the abdominal aorta, and these protective effects could be related to the inhibition of PKC and mitogen-activated protein kinase (MAPK) signaling pathways.

GRP78 (Glucose-Regulated Protein of 78 kDa) Promotes Cardiomyocyte Growth Through Activation of GATA4 (GATA-Binding Protein 4).

The heart manifests hypertrophic growth in response to elevation of afterload pressure. Cardiac myocyte growth involves new protein synthesis and membrane expansion, of which a number of cellular quality control machineries are stimulated to maintain function and homeostasis. The unfolded protein response is potently induced during cardiac hypertrophy to enhance protein-folding capacity and eliminate terminally misfolded proteins. However, whether the unfolded protein response directly regulates cardiac myocyte growth remains to be fully determined. Here, we show that GRP78 (glucose-regulated protein of 78 kDa)-an endoplasmic reticulum-resident chaperone and a critical unfolded protein response regulator-is induced by cardiac hypertrophy. Importantly, overexpression of GRP78 in cardiomyocytes is sufficient to potentiate hypertrophic stimulus-triggered growth. At the in vivo level, TG (transgenic) hearts overexpressing GRP78 mount elevated hypertrophic growth in response to pressure overload. We went further to show that GRP78 increases GATA4 (GATA-binding protein 4) level, which may stimulate Anf (atrial natriuretic factor) expression and promote cardiac hypertrophic growth. Silencing of GATA4 in cultured neonatal rat ventricular myocytes significantly diminishes GRP78-mediated growth response. Our results, therefore, reveal that protein-folding chaperone GRP78 may directly enhance cardiomyocyte growth by stimulating cardiac-specific transcriptional factor GATA4.

Gestational diabetes influences the expression of hypertrophic genes in left ventricle of rat's offspring.

Objective(s):Gestational diabetes increases the risk of congenital heart disease in the offspring, but the molecular mechanism underlying this process remains unclear. Therefore, the current study was conducted to assess the effects of induced gestational diabetes on expression of some involved genes in cardiac hypertrophy in the offspring of diabetic rats.Materials and Methods:Diabetes was induced in 40 adult Wistar rats by intraperitoneal injection of 45 mg/kg of streptozotocin. The day of appearance of the vaginal plug was assumed as day zero of gestation for inducing diabetes. After pregnancy, the offspring was maintained until they reach the age of 12 weeks. Then, their hearts were excised and were sectioned for molecular study. We analyzed the expression pattern of some hypertrophic genes by the quantitative real-time RT-PCR.Results:The mRNA expression levels of all studied genes including c-jun, c-fos, c-myc, alpha-myosin heavy chain (α-MHC), atrial natriuretic factor (ANF) and β-MHC, which are important in cardiomyocyte hypertrophy, were higher in the offspring of the diabetic group compared to controls. Significant differences were found for β-MHC and c-myc with P<0.01 and for α-MHC and c-fos with P<0.05.Conclusion:Gestational diabetes upregulates expression of c-jun, c-fos c-myc, α-MHC, ANF and β-MHC genes that are involved in cardiac hypertrophy in the offspring of diabetic rats.

Depletion of ubiA prenyltransferase domain containing 1 expression promotes angiotensin II‑induced hypertrophic response in AC16 human myocardial cells via modulating the expression levels of coenzyme Q10 and endothelial nitric oxide synthase.

UbiA prenyltransferase domain containing1 (UBIAD1) is closely associated with cardiovascular diseases. However, at the cellular level, little is known about how UBIAD1 is expressed and functions in cardiomyocyte hypertrophy. The aim of the present study was to investigate the expression and role of UBIAD1 in angiotensin II (Ang II)‑induced hypertrophy in AC16 cardiomyoblast cells. The loss‑of‑function approach was used to knock down UBIAD1 in vehicle‑ and Ang II‑stimulated AC16 cells. The levels of atrial natriuretic factor (ANF) and caspase-3 were measured and compared between vehicle‑ and Ang II‑treated AC16 cells pretreated with control siRNA or siRNA against UBIAD1. In addition, the levels of coenzyme Q10 (CoQ10) and endothelial nitric oxide synthase (eNOS) were evaluated and compared between these groups. Ang II induced hypertrophy and apoptosis in AC16 cells, accompanied by increased expression of ANF and caspase-3, and decreased expression of UBIAD1. These effects were potentiated by UBIAD1 knockdown. In addition, Ang II treatment suppressed the expression of CoQ10 and eNOS, as well as the production of NO, and these inhibitory effects were also enhanced by UBIAD1 knockdown. Thus, silencing of UBIAD1 expression promotes a myocardial hypertrophic response to Ang II stimulation, in part, by suppressing the expression of CoQ10 and eNOS.

Pathologic Role of MicroRNA‐365 in the Heart

Cardiac remodeling occurs in many cardiovascular diseases. It can severely impair ventricular function and increase the risk for arrhythmias. A better understanding of cardiac signaling regulatory mechanisms is critical for development of novel and effective treatment approaches. MicroRNAs (miRNAs) are important regulators in cardiac development and disease. It was reported that miRNA‐365 is 2.8‐fold up‐regulated in the left ventricular tissue from patients undergoing heart transplantation due to end‐stage heart failure (Thum T et. al, Circulation 2007). Using phenotypic screen of 230 synthetic miRNAs, miRNA‐365 was identified as one of four hypertrophy‐modulating miRNAs (Jentzsch et al., Jour Mol Cell Card 2012). However, to date, miRNA‐365 has not been functionally characterized in the heart. The goal of this on‐going study is to delineate the functional importance and regulatory mechanisms of miRNA‐365 in the heart. Using adult rats, we found increase of miRNA‐365 expression in the infarcted area than in the remote area and sham control only 7 days after myocardial infarction. MiRNA‐365 is also up‐regulated in neonatal rat ventricular myocytes stimulated with phenylephrine. Overexpression of miRNA‐365 in culture significantly increases both the size of neonatal cardiac myocytes and atrial natriuretic factor (ANF) expression. In contrast, ANF expression is markedly reduced by miRNA‐365 knockdown. We generated a transgenic mouse model with cardiac specific, α myosin heavy chain‐driven overexpression of miRNA‐365. Significant increase of ANF expression in ventricules with cardiac specific overexpression of miRNA‐365 suggests the activation of hypertrophy signaling. Our ex vivo optical mapping data suggest that ventricular tachycardia (VT) is highly inducible in the heart with cardiac specific overexpression of miRNA‐365 and spatially discordant calcium alternans is the major driven force for the VT events. Importantly, this spatially discordant calcium alternans can be eliminated by Dantrolene, suggesting uncontrolled SR calcium release during excitation‐contraction coupling may contribute to the VT induction. Our data further show that spatially discordant calcium alternans can be changed back to concordant calcium alternans by mitoTEMPO, suggesting that increased mitochondrial reactive oxygen species could be the upstream of the uncontrolled SR calcium release. These findings are supported by single cell patch clamps, in which the isolated myocytes from miRNA‐365 overexpressing hearts show increased early after depolarizations (EAD), which are majorly due to elevated cytosolic calcium concentrations. Taken together, our results suggest miRNA‐365 has a critical pathologic role in the heart and may regulate both structural and electrical cardiac remodeling, which warrants further investigation.Support or Funding InformationAmerican Heart Association (AHA) Founders Affiliate Pre‐doctoral Fellowship; COBRE for CardioPulmonary Vascular Biology

Molecular Changes in Children with Heart Failure Undergoing Left Ventricular Assist Device Therapy

To determine whether left ventricular assist device (LVAD) treatment in children with heart failure would result in the modification of molecular pathways involved in heart failure pathophysiology. Forty-seven explanted hearts from children were studied (16 nonfailing control, 20 failing, and 11 failing post-LVAD implantation [F-LVAD]). Protein expression and phosphorylation states were determined by receptor binding assays and Western blots. mRNA expression was measured with real-time quantitative polymerase chain reaction. To evaluate for interactions and identify correlations, 2-way ANOVA and regression analysis were performed. Treatment with LVAD resulted in recovery of total β-adrenergic receptor expression and β1-adrenergic receptor (β1-AR) in failing hearts to normal levels (β-adrenergic receptor expression : 67.2 ± 11.5 fmol/mg failing vs 99.5 ± 27.7 fmol/mg nonfailing, 104 ± 38.7 fmol/mg F-LVAD, P ≤ .01; β1-AR: 52.2 ± 10.3 fmol/mg failing vs 83.0 ± 23 fmol/mg non-failing, 76.5 ± 32.1 fmol/mg F-LVAD P ≤ .03). The high levels of G protein-coupled receptor kinase-2 were returned to nonfailing levels after LVAD treatment (5.6 ± 9.0 failing vs 1.0 ± 0.493 nonfailing, 1.0 ± 1.3 F-LVAD). Interestingly, β2-adrenergic receptor expression was significantly greater in F-LVAD (27.5 ± 12; P < .005) hearts compared with nonfailing (16.4 ± 6.1) and failing (15.1 ± 4.2) hearts. Phospholamban phosphorylation at serine 16 was significantly greater in F-LVAD (7.7 ± 11.7) hearts compared with nonfailing (1.0 ± 1.2, P = .02) and failing (0.8 ± 1.0, P = .01) hearts. Also, atrial natriuretic factor (0.6 ± 0.8) and brain natriuretic peptide (0.1 ± 0.1) expression in F-LVAD was significantly lower compared with failing hearts (2.8 ± 3.6, P = .01 and 0.6 ± 0.7, P = .02). LVAD treatment in children with heart failure results in reversal of several pathologic myocellular processes, and G protein-coupled receptor kinase-2 may regulate β1-AR but not β2-adrenergic receptor expression in children with heart failure.

Left-Ventricular Plication Reduces Wall Stress and Cardiomyocyte Hypertrophy in a Rat Model of Ischemic Cardiomyopathy

Background: The indications of left-ventricular plication (LVP) are controversial, although several studies have reported favorable outcomes in heart failure patients. The aim of this study was to assess left-ventricular (LV) wall stress and myocardial remodeling after LVP in a rat model of myocardial infarction (MI). Methods: Sixteen rats underwent LVP by excluding the LV anterior wall scar 4 weeks after ligation of the left anterior descending artery. After 4 weeks, LV wall stress was assessed using transthoracic echocardiography and an LV catheter. Gene expression of the wall stress markers, atrial natriuretic factor (ANF) and brain natriuretic peptide (BNP), were evaluated via reverse transcription polymerase chain reaction. Cardiomyocyte area and myocardial fibrosis were also examined through histological examinations. These parameters were compared to those in 16 rats that underwent coronary artery ligation but not LVP. Results: We noted that the LV end-diastolic dimension was smaller (9.9 ± 0.3 vs. 11.2 ± 0.2 mm, p < 0.05) and fractional shortening was greater (25 ± 2 vs. 15 ± 1%, p < 0.05) in LVP rats than in sham rats. Moreover, systolic wall stress was lower in LVP rats (71 ± 7 vs. 111 ± 9 × 10³ dyn/cm², p < 0.05). Myocardial ANF and BNP expression levels were lower in LVP rats (2.6 ± 0.3 vs. 4.4 ± 0.5 and 1.0 ± 0.1 vs. 1.5 ± 0.2 arbitrary units, respectively; p < 0.05). Cardiomyocyte area was significantly decreased in LVP rats (556 ± 15 vs. 670 ± 28 μm², p = 0.003) and was correlated with LV wall stress (r = 0.669, p = 0.002). The reduction in myocardial fibrosis after LVP was not significant. Conclusion: LVP reduced LV wall stress and cardiomyocyte hypertrophy in a rat model of MI.

Activation of HuR downstream of p38 MAPK promotes cardiomyocyte hypertrophy

The RNA binding protein Human antigen R (HuR) interacts with specific AU-rich domains in target mRNAs and is highly expressed in many cell types, including cardiomyocytes. However, the role of HuR in cardiac physiology is largely unknown. Our results show that HuR undergoes cytoplasmic translocation, indicative of its activation, in hypertrophic cardiac myocytes. Specifically, HuR cytoplasmic translocation is significantly increased in NRVMs (neonatal rat ventricular myocytes) following treatment with phenylephrine or angiotensin II, agonists of two independent Gαq-coupled GPCRs known to induce hypertrophy. This Gq-mediated HuR activation is dependent on p38 MAP kinase, but not canonical Gq-PKC signaling. Furthermore, we show that HuR activation is necessary for Gq-mediated hypertrophic growth of NRVMs as siRNA-mediated knockdown of HuR inhibits hypertrophy as measured by cell size and expression of ANF (atrial natriuretic factor). Additionally, HuR overexpression is sufficient to induce hypertrophic cell growth. To decipher the downstream mechanisms by which HuR translocation promotes cardiomyocyte hypertrophy, we assessed the role of HuR in the transcriptional activity of NFAT (nuclear factor of activated T cells), the activation of which is a hallmark of cardiac hypertrophy. Using an NFAT-luciferase reporter assay, we show an acute inhibition of NFAT transcriptional activity following pharmacological inhibition of HuR. In conclusion, our results identify HuR as a novel mediator of cardiac hypertrophy downstream of the Gq-p38 MAPK pathway, and suggest modulation of NFAT activity as a potential mechanism.

Discovery of novel small molecule inhibitors of cardiac hypertrophy using high throughput, high content imaging

Chronic cardiac hypertrophy is maladaptive and contributes to the pathogenesis of heart failure. The objective of this study was to identify small molecule inhibitors of pathological cardiomyocyte hypertrophy. High content screening was performed with primary neonatal rat ventricular myocytes (NRVMs) cultured on 96-well plates and treated with a library of 3241 distinct small molecules. Non-toxic hit compounds that blocked hypertrophy in response to phenylephrine (PE) and phorbol myristate acetate (PMA) were identified based on their ability to reduce cell size and inhibit expression of atrial natriuretic factor (ANF), which is a biomarker of pathological cardiac hypertrophy. Many of the hit compounds are existing drugs that have not previously been evaluated for benefit in the setting of cardiovascular disease. One such compound, the anti-malarial drug artesunate, blocked left ventricular hypertrophy (LVH) and improved cardiac function in adult mice subjected to transverse aortic constriction (TAC). These findings demonstrate that phenotypic screening with primary cardiomyocytes can be used to discover anti-hypertrophic lead compounds for heart failure drug discovery. Using annotated libraries of compounds with known selectivity profiles, this screening methodology also facilitates chemical biological dissection of signaling networks that control pathological growth of the heart.

Effect and mechanism of polydatin on diabetic myocardial hypertrophy in mice

To observe the preventive effect of polydatin on diabetic myocardial hypertrophy in mice and discuss its and mechanism. The diabetic model was induced with low dose STZ (40 mg x kg(-1) x d(-1) x 5 d, ip) for five days in mice. The myocardial hypertrophy was determined by hypertrophy indexes (LVHI, left ventricular/right ventricle and septum), left ventricular/body weight (LV/BW), the histological examination and the mRNA expression of atrial natriuretic factor(ANF). The fast blood glucose(FBG), serum insulin and plasma hemoglobin A1c ( HbA1c) levels were detected, and then HOMA insulin resistance index ( HOMA. IR) was calculated. The mRNA and protein expressions were measured by qRT-PCR and western blotting, respectively. According to the results, the FBG of the model group exceeded 11.1 mmol x L(-1), with notable decrease in BW and significant increase in insulin, HbA1c and HOME. IR, suggesting the successful establishment and stability of the diabetic model. The increases in LVHI, LV/BW, cell surface and ANF mRNA indicated a myocardial hypertrophy in diabetic mice. Meanwhile, the model group showed decrease in mRNA and protein expressions of PPARβ and significant increase in NF-κB p65, COX-2 and iNOS expressions. After the preventation with PD (50, 100 mg x kg(-1) x d(-1)), diabetic mice showed increase in BW, reduction in the levels of FBG, insulin and HbA1 c, relief in insulin resistance and significant recovery in hypertrophy indexes, indicating PD has the protective effect in diabetic myocardial hypertrophy. Meanwhile, PD up-regulated the expression of PPARβ, inhibited the expressions of NF-κB p65, COX-2 and iNOS, demonstrating that PD's protective effect may be related to the activation of PPARβ and the inhibition of NF-κB, COX-2 and iNOS signaling pathways.

Treatment with hydrogen molecule attenuates cardiac dysfunction in streptozotocin-induced diabetic mice

Diabetic cardiomyopathy, a disorder of the heart muscle in diabetic patients, is one of the major causes of heart failure. The aim of present study was to investigate the therapeutic effect of hydrogen molecule on streptozotocin-induced diabetic cardiomyopathy in mice. Diabetes was induced in adult male mice by consecutive peritoneal injection of streptozotocin (50 mg/kg/day) for 5 days. Then, they were treated with hydrogen water (1.3±0.2 mg/l) for 8 weeks (four groups, n=83-88 in each group). Although treatment of diabetic mice with hydrogen water did not significantly affect blood glucose level, it significantly attenuated cardiac hypertrophy and reduced expression of atrial natriuretic factor and β-myosin heavy chain; it alleviated cardiac fibrosis and reduced expression of collagen I and III, transforming growth factor beta, alpha-smooth muscle actin, and osteopontin; it reduced cardiac caspase-3 activity and ratio of bax/bcl-2. Importantly, hydrogen water treatment improved cardiac function in streptozotocin-diabetic mice. Furthermore, it was found that hydrogen water treatment abated oxidative stress, suppressed inflammation, and attenuated endoplasmic reticulum stress in the hearts of streptozotocin-diabetic mice. In addition, hydrogen water treatment suppressed activation of Jun NH2-terminal kinase and p38 mitogen activated protein kinase signaling and nuclear factor κB signaling in the hearts of streptozotocin-diabetic mice. Treatment with hydrogen molecule attenuated cardiac dysfunction in streptozotocin-induced diabetic mice, which was independent of glycemic control. Treatment with hydrogen molecule attenuated cardiac dysfunction in streptozotocin-induced type 1 diabetic mice. Molecular hydrogen could thus be envisaged as a nutritional countermeasure for diabetic cardiomyopathy.

Nanocurcumin protects cardiomyoblasts H9c2 from hypoxia-induced hypertrophy and apoptosis by improving oxidative balance.

Hypoxia-induced cardiomyocyte hypertrophy is evident; however, the distinct molecular mechanism underlying the oxidative stress-mediated damages to cardiomyocytes remains unknown. Curcumin (diferuloylmethane) is known for anti-hypertrophic effects, but low bioavailability makes it unsuitable to exploit its pharmacological properties. We assessed the efficacy of nanotized curcumin, i.e. nanocurcumin, in ameliorating hypoxia-induced hypertrophy and apoptosis in H9c2 cardiomyoblasts and compared it to curcumin. H9c2 cardiomyoblasts were challenged with 0.5% oxygen, for 24h to assess hypoxia-induced oxidative damage, hypertrophy and consequent apoptosis. The molecular mechanism underlying the protective efficacy of nanocurcumin was evaluated in regulating Raf-1/Erk-1/2 apoptosis by caspase-3/-7 pathway and oxidative stress. Nanocurcumin ameliorated hypoxia-induced hypertrophy and apoptosis in H9c2 cells significantly (p ≤ 0.01), by downregulating atrial natriuretic factor expression, caspase-3/-7 activation, oxidative stress and stabilizing hypoxia-inducible factor-1α (HIF-1α) better than curcumin. Nanocurcumin provides insight into its use as a potential candidate in curing hypoxia-induced cardiac pathologies by restoring oxidative balance.

NF‐қB Activation is Required for Thyroid Hormone Induced Cardiomyocyte Hypertrophy

The nuclear factor NF‐қB promotes inflammatory response and mediates cardiac growth by different hypertrophic stimuli. We have previous results obtained by microarray experiments indicating that thyroid hormone (triiodotyronine, T3) up‐regulates pro‐inflammatory genes. Considering that, we investigated the influence of T3 on NF‐қB signaling and the possible participation of this nuclear factor in cardiomyocyte hypertrophy induced by T3. Primary cultures of neonatal rat ventricular cardiomyocytes were treated with T3 for 10, 30 minutes and 24 hours and the NF‐қB signaling was analyzed by western blot. Also, cardiomyocytes were treated with an inhibitor of NF‐қB (JSH‐23) followed by treatment with T3 for evaluation of mRNA expression of atrial natriuretic factor (ANF) and cell surface area analysis. Data were presented as mean ± SD and were analyzed using one‐way ANOVA and Tukey multiple comparisons test. Values of p&lt;0.05 were considered statistically significant. T3 rapidly increased the nuclear protein expression of NF‐қB without alterations on IKK and IқB expression. The inhibition of NF‐қB reduced both ANF mRNA levels and the surface area of cardiomyocytes compared to T3 treated cells. These results confirm that NF‐қB is required for development of cardiomyocyte hypertrophy by T3. The possible interactions of NF‐қB with genes associated with cardiac growth are being investigated. Financial support: Fapesp (2013/22480‐4 and 2011/23352‐4).