atrial-natriuretic-factor and Hypertrophy

Magnesium isoglycyrrhizinate (MgIG) is a magnesium salt of the 18-α glycyrrhizic acid stereoisomer that has exhibited hepato-protective effects and has anti-inflammatory, antioxidant, and antiviral activities. Here, we have investigated the effects and potential mechanisms of action of MgIG, with respect to myocardial fibrosis induced by isoproterenol (ISO) in mice. Mice were administered MgIG for 14days, with concurrent ISO dosing, and were sacrificed two weeks later. Lactate dehydrogenase (LDH) and creatine kinase (CK) concentrations were measured in the blood. Pathological changes in the myocardium were observed via light microscopy. In addition, the expression of the Bax and Bcl-2 genes, and the basic fibroblast growth factor (bFGF) protein were measured via an immunohistochemical method. The RNA expression of atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), c-fos, and c-jun mRNA were quantified by reverse transcription-polymerase chain reaction (RT-PCR) in the myocardial tissue. The protein expression of toll-like receptor (TLR) 4, and nuclear factor kappa B (NF-κB) (p65) were measured using Western blot assays. Compared with the control group, the ISO group showed significant increases in bFGF, Bax, Bcl-2, TLR4, and NF-κB (p65) expressions, as well as increased serum levels of LDH and CK. MgIG had a protective effect on ISO-induced myocardial fibrosis, which might be ascribed, at least in part, to the inhibition of the TLR4/NF-κB (p65) signaling pathway.

Topics: Animals; Anti-Inflammatory Agents; Atrial Natriuretic Factor; bcl-2-Associated X Protein; Creatine Kinase; Disease Models, Animal; Fibroblast Growth Factor 2; Fibrosis; Heart Diseases; Humans; Hypertrophy; Isoproterenol; L-Lactate Dehydrogenase; Mice; Mice, Inbred Strains; Myocardium; Natriuretic Peptide, Brain; NF-kappa B; Proto-Oncogene Proteins c-bcl-2; Saponins; Toll-Like Receptor 4; Triterpenes

The natriuretic peptides (NPs), through their diuretic, vasodilatory and anti-mitogenic properties, play an important role in the regulation of cardiovascular, renal and endocrine homeostasis. Recent studies suggest that they have utility in both the diagnosis and management of heart failure. Plasma brain NP (BNP) levels have been used to establish prognosis in patients with heart failure and those at risk for heart failure post-myocardial infarction. They have been used to establish a cardiac etiology for acute shortness of breath, and to guide and assess the efficacy of therapy in patients with established heart failure. BNP is also approved for use in the management of acute decompensated heart failure. Of note, recent studies suggest that cardiac NPs suppress myocyte hypertrophy and interstitial fibrosis in the heart, arguing for an important autocrine-paracrine role of these peptides in controlling the cardiac response during hypertrophy. Therefore, the existing evidence supports a role for BNP as both a marker and a modulator of hypertrophy.

Topics: Atrial Natriuretic Factor; Biomarkers; Cardiomyopathy, Dilated; Gene Expression Regulation; Humans; Hypertrophy; Myocardium; Natriuretic Peptide, Brain; Natriuretic Peptides

The natriuretic peptides (NPs) constitute a family of polypeptide hormones that regulate mammalian blood volume and blood pressure. The ability of the NPs to modulate cardiac hypertrophy and cell proliferation as well is now beginning to be recognized. The NPs interact with three membrane-bound receptors, all of which contain a well-characterized extracellular ligand-binding domain. The R1 subclass of NP receptors (NPR-A and NPR-B) contains a C-terminal guanylyl cyclase domain and is responsible for most of the NPs downstream actions through their ability to generate cGMP. The R2 subclass lacks an obvious catalytic domain and functions primarily as a clearance receptor. This review focuses on the signal transduction pathways initiated by ligand binding and other factors that help to determine signalling specificities, including allosteric factors modulating cGMP generation, receptor desensitization, the activation and function of cGMP-dependent protein kinase (PKG), and identification of potential nuclear or cytoplasmic targets such as the mitogen-activated protein kinase signalling (MAPK) cascade. The inhibition of cardiac growth and hypertrophy may be an important but underappreciated action of the NP signalling system.

Topics: Amino Acid Sequence; Animals; Atrial Natriuretic Factor; Catalytic Domain; Cell Division; Cyclic GMP; Cyclic GMP-Dependent Protein Kinases; Cytoplasm; Guanylate Cyclase; Humans; Hypertrophy; Ligands; MAP Kinase Signaling System; Models, Biological; Molecular Sequence Data; Natriuretic Peptide, Brain; Protein Binding; Protein Structure, Tertiary; Receptors, Atrial Natriuretic Factor; Signal Transduction

Topics: Animals; Atrial Natriuretic Factor; Autophagy; Hypertrophy; Mice; Mice, Knockout; Myocytes, Cardiac

Diabetic cardiomyopathy (DCM) is a chronic complication of diabetes that emphasizes the urgency of developing new drug therapies. With an illustrious history in traditional medicine to improve diabetes, cinnamon has been shown to possess blood lipids lowering effects and antioxidative and anti-inflammatory properties. However, the extent to which it protects the diabetic heart has yet to be determined. Forty-eight rats were administered in the study and grouped as: control; diabetic; diabetic rats given 100, 200, or 400 mg/kg cinnamon extract, metformin (300 mg/kg), valsartan (30 mg/kg), or met/val (combination of both drugs), via gavage for six weeks. Fasting blood sugar (FBS) and markers of cardiac injury including creatine kinase-muscle/brain (CK-MB), aspartate aminotransferase (AST), and lactate dehydrogenase (LDH) were evaluated in blood samples. Malondialdehyde (MDA) levels, the total contents of thiol, superoxide dismutase (SOD), and catalase (CAT) activities were measured. Histopathology study and gene expression measurement of angiotensin II type 1 receptor (AT1), atrial natriuretic peptide (ANP), beta-myosin heavy chain (β-MHC), and brain natriuretic peptide (BNP) were done on cardiac tissue. FBS and cardiac enzyme indicators were reduced in all treated groups. A reduction in MDA level and enhancement in thiol content alongside with increase of SOD and CAT activities were observed in extract groups. The decrease of inflammation and fibrosis was obvious in treated groups, notably in the high-dose extract group. Furthermore, all treated diabetic groups showed a lowering trend in AT1, ANP, β-MHC, and BNP gene expression. Cinnamon extract, in addition to its hypoglycemic and antioxidant properties, can prevent diabetic heart damage by alleviating cardiac inflammation and fibrosis. PRACTICAL APPLICATIONS: This study found that cinnamon extract might protect diabetic heart damage by reducing inflammation and fibrosis in cardiac tissue, in addition to lowering blood glucose levels and increasing antioxidant activity. Our data imply that including cinnamon in diabetic participants' diets may help to reduce risk factors of cardiovascular diseases.

Topics: Animals; Antioxidants; Atrial Natriuretic Factor; Cinnamomum zeylanicum; Diabetes Mellitus, Experimental; Diabetic Cardiomyopathies; Fibrosis; Heart Injuries; Humans; Hypertrophy; Inflammation; Plant Extracts; Rats; Sulfhydryl Compounds; Superoxide Dismutase

Atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) are cardiac hormones located in atria granules. Both peptides respond to cardiac pressure and volume dynamics and accordingly serve as translation biomarkers for the clinical treatment of heart failure. Serum ANP and BNP play central secretary roles in blood pressure and cardiac output regulation and have proven utility as differential biomarkers of cardiovascular proficiency and drug-induced maladaptation, yet both peptides are impervious to exercise-induced hypertrophy. We employed immunoelectron microscopy to examine the effects of 28 days of chronic swim exercise or administration of a PPARγ agonist on atrial granules and their stored natriuretic peptides in Sprague Dawley rats. Chronic swimming and drug treatment both resulted in a 15% increase in heart weight compared with controls, with no treatment effects on perinuclear granule area in the left atria (LAs). Drug treatment resulted in larger size granules with greater BNP density in the right atria. Comparing swimming and PPARγ agonist treatment effects on ANP:BNP granule density ratios between atrial chambers revealed a shift toward a greater proportion of ANP than BNP in LAs of swim-trained rats. These data suggest a distinction in the population of ANP and BNP after chronic swim or PPARγ that makes it a novel metric for the differentiation of pathological and physiological hypertrophy.

Topics: Animals; Atrial Natriuretic Factor; Biomarkers; Heart Atria; Hypertrophy; Natriuretic Peptide, Brain; Peptides; PPAR gamma; Rats; Rats, Sprague-Dawley

Clinical studies have indicated that 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors, also known as statins, can potentially inhibit chronic heart failure. In the Stat-LVDF study, a difference was noted in terms of the effect of lipophilic pitavastatin (PTV) and hydrophilic rosuvastatin (RSV) on plasma BNP, suggesting that statin lipophilicity and pharmacokinetics change the pleiotropic effect on heart failure in humans. Therefore, we assessed the beneficial effects of PTV on hypertrophy in cardiac myocytes compared with RSV at clinically used doses. Cultured cardiomyocytes were stimulated with 30 μM phenylephrine (PE) in the presence of PTV (250 nM) or RSV (50 nM). These doses were calculated based on the maximum blood concentration of statins used in clinical situations in Japan. The results showed that PTV, but not RSV, significantly inhibits the PE-induced increase in cell size and leucine incorporation without causing cell toxicity. In addition, PTV significantly suppressed PE-induced mRNA expression of hypertrophic response genes. PE-induced ERK phosphorylation was inhibited by PTV, but not by RSV. Furthermore, PTV significantly suppressed the angiotensin-II-induced proline incorporation in primary cultured cardiac fibroblasts. In conclusion, a clinical dose of PTV was noted to directly inhibit cardiomyocyte hypertrophy and cardiac fibrosis, suggesting that lipophilic PTV can be a potential drug candidate against chronic heart failure.

Topics: Actins; Animals; Atrial Natriuretic Factor; Cells, Cultured; Extracellular Signal-Regulated MAP Kinases; Gene Expression; Hydroxymethylglutaryl-CoA Reductase Inhibitors; Hypertrophy; Leucine; Myocytes, Cardiac; Natriuretic Peptide, Brain; Phosphorylation; Quinolines; Rats, Sprague-Dawley; RNA, Messenger; Rosuvastatin Calcium

Angiotensin II (Ang II) is an important bioactive peptide in the renin‑angiotensin system, and it can contribute to cell proliferation and cardiac hypertrophy. Dysfunctions in transient receptor potential canonical (TRPC) channels are involved in many types of cardiovascular diseases. The aim of the present study was to investigate the role of the TRPC channel inhibitor SKF‑96365 in cardiomyocyte hypertrophy induced by Ang II and the potential mechanisms of SKF‑96365. H9c2 cells were treated with different concentrations of Ang II. The expression levels of cardiomyocyte hypertrophy markers and TRPC channel‑related proteins were also determined. The morphology and surface area of the H9c2 cells, the expression of hypertrophic markers and TRPC channel‑related proteins and the [3H] leucine incorporation rate were detected in the Ang II‑treated H9c2 cells following treatment with the TRPC channel inhibitor SKF‑96365. The intracellular Ca2+ concentration was tested by flow cytometry. The present results suggested that the surface area of H9c2 cells treated with Ang II was significantly increased compared with untreated H9c2 cells. The fluorescence intensity of α‑actinin, the expression of hypertrophic markers and TRPC‑related proteins, the [3H] leucine incorporation rate and the intracellular Ca2+ concentration were all markedly increased in the Ang II‑treated H9c2 cells but decreased following SKF‑96365 treatment. The present results suggested that Ang II induced cardiomyocyte hypertrophy in H9c2 cells and that the TRPC pathway may be involved in this process. Therefore, SKF‑96365 can inhibit cardiomyocyte hypertrophy induced by Ang II by suppressing the TRPC pathway. The present results indicated that TRPC may be a therapeutic target for the development of novel drugs to treat cardiac hypertrophy.

Topics: Actinin; Angiotensin II; Animals; Atrial Natriuretic Factor; Calcium; Cell Line; Cell Membrane; Cell Shape; Fluorescence; Gene Expression Regulation; Hypertrophy; Imidazoles; Myocytes, Cardiac; Rats; TRPC Cation Channels

Luteolin (LTL) serves essential roles in a wide variety of biological processes. Lipopolysaccharide (LPS) can lead to myocardial hypertrophy and autophagy. However, the roles of LTL on LPS‑induced cardiomyocyte hypertrophy and autophagy in rat cardiomyocytes have not yet been fully elucidated. In the present study, the morphology of cultured rat cardiomyocytes was observed under an inverted microscope. Cell viability was detected by MTT assay. α‑Actinin and microtubule‑associated protein 1 light chain 3 (LC3) expression levels were measured by immunofluorescence assay. In addition, the expression levels of atrial natriuretic peptide/brain natriuretic peptide (ANP/BNP), LC3, and autophagy‑ and Wnt signaling pathway‑associated genes were analyzed by reverse transcription‑quantitative polymerase chain reaction or western blot assays. The results indicated that LTL increased the cell viability of cardiomyocytes treated with LPS. LTL decreased the expression of cardiac hypertrophy associated markers (ANP and BNP). LTL decreased α‑actinin and LC3 expression levels in LPS‑treated cardiomyocytes. It was also demonstrated that LTL suppressed the mRNA and protein expression levels of LPS‑mediated autophagy and Wnt signaling pathway‑associated genes. In addition, it was demonstrated that silencing of β‑catenin inhibited LPS‑induced cardiomyocyte hypertrophy and the formation of autophagosomes. Thus, the present study suggested that LTL protected against LPS‑induced cardiomyocyte hypertrophy and autophagy in rat cardiomyocytes.

Topics: Animals; Atrial Natriuretic Factor; Autophagy; Disease Models, Animal; Gene Expression; Humans; Hypertrophy; Lipopolysaccharides; Luteolin; Microtubule-Associated Proteins; Myocytes, Cardiac; Natriuretic Peptide, Brain; Rats; RNA, Messenger; Signal Transduction

This paper was aimed to investigate the inhibitory effect of aconitine(AC) on angiotensin Ⅱ(Ang Ⅱ)-induced H9 c2 cell hypertrophy and explore its mechanism of action. The model of hypertrophy was induced by Ang Ⅱ(1×10-6 mol·L-1),and cardiomyocytes were incubated with different concentrations of AC. Western blot was used to quantify the protein expression levels of atrial natriuretic peptide(ANP),brain natriuretic peptide(BNP),β-myosin heavy chain(β-MHC),and α-smooth muscle actin(α-SMA). Real-time quantitative PCR(qRT-PCR) was used to quantify the mRNA expression levels of cardiac hypertrophic markers ANP,BNP and β-MHC. In addition,the fluorescence intensity of the F-actin marker,an important component of myofibrils,was detected by using laser confocal microscope. AC could significantly reverse the increase of total protein content in H9 c2 cells induced by Ang Ⅱ; qRT-PCR results showed that AC could significantly inhibit the ANP,BNP and β-MHC mRNA up-regulation induced by AngⅡ. Western blot results showed that AC could significantly inhibit the ANP,BNP and β-MHC protein up-regulation induced by AngⅡ. In addition,F-actin expression induced by Ang Ⅱ could be inhibited by AC,and multiple indicators of cardiomyocyte hypertrophy induced by Ang Ⅱ could be down-regulated,indicating that AC may inhibit cardiac hypertrophy by inhibiting the expression of hypertrophic factors,providing new clues for exploring the cardiovascular protection of AC.

Topics: Aconitine; Actins; Angiotensin II; Atrial Natriuretic Factor; Cardiac Myosins; Cardiomegaly; Cells, Cultured; Humans; Hypertrophy; Myocytes, Cardiac; Myosin Heavy Chains; Natriuretic Peptide, Brain

In response to assorted stimuli, the heart will develop into cardiomyocyte hypertrophy, but sustained cardiomyocyte hypertrophy will finally lead to heart failure. This research is aimed to examine the effect of VEGFB on cardiomyocyte hypertrophy by using the cardiomyocyte-derived cell line H9C2 of cultured rates. It turns out that VEGFB can positively prevent the Ang II-induced rising in the size of cardiomyocyte as well as reduce Ang II-induced mRNA and protein levels of β-MHC (β-myosin heavy chain), BNP (brain natriuretic peptide), and ANP (atrial natriuretic peptide). Moreover, VEGFB can regulate the decline of the Ang II-induced rising in Ca

Topics: Angiotensin II; Animals; Atrial Natriuretic Factor; Calcium; Cell Line; Cell Size; Cyclic GMP-Dependent Protein Kinase Type I; Gene Knockdown Techniques; Hypertrophy; Myocytes, Cardiac; Myosin Heavy Chains; Natriuretic Peptide, Brain; Rats; Signal Transduction; Vascular Endothelial Growth Factor B; Vascular Endothelial Growth Factor Receptor-1

This study aimed to determine the effects of levosimendan, a calcium sensitizer, on atrial contractility and atrial natriuretic peptide (ANP) secretion and its modification in hypertrophied atria. Isolated perfused beating rat atria were used from control and isoproterenol-treated rats. Levosimendan and its metabolite OR-1896 caused a positive inotropic effect and suppressed ANP secretion in rat atria. Similar to levosimendan, the selective phosphodiesterase 3 (PDE3) or PDE4 inhibitor also suppressed ANP secretion. Suppression of ANP secretion by 1 µM levosimendan was abolished by PDE3 inhibitor, but reversed by PDE4 inhibitor. Levosimendan-induced suppression of ANP secretion was potentiated by K

Topics: Animals; Atrial Function; Atrial Natriuretic Factor; Atrial Pressure; Calcium; Extracellular Space; Heart Atria; Hemodynamics; Hydrazones; Hypertrophy; Male; Pyridazines; Rats; Rats, Sprague-Dawley; Simendan

Many clinical and experimental studies have shown that treatment with statins could prevent myocardial hypertrophy and remodeling induced by hypertension and myocardial infarction. But the molecular mechanism was not clear. We aimed to investigate the beneficial effects of atorvastatin on hypertension-induced myocardial hypertrophy and remodeling in spontaneously hypertensive rats (SHR) with the hope of revealing other potential mechanisms or target pathways to interpret the pleiotropic effects of atorvastatin on myocardial hypertrophy.. The male and age-matched animals were randomly divided into three groups: control group (8 WKY), SHR (8 rats) and intervention group (8 SHR). The SHR in intervention group were administered by oral gavage with atorvastatin (suspension in distilled water, 10 mg/Kg once a day) for 6 weeks, and the other two groups were administered by gavage with equal quantity distilled water. Blood pressure of rats was measured every weeks using a standard tail cuff sphygmomanometer. Left ventricular (LV) dimensions were measured from short-axis views of LV under M-mode tracings using Doppler echocardiograph. Cardiomyocyte apoptosis was assessed by the TUNEL assay. The protein expression of C/EBPβ, PGC-1α and UCP3 were detected by immunohistochemistry or Western blot analysis.. At the age of 16 weeks, the mean arterial pressure of rats in three groups were 103.6±6.1, 151.8±12.5 and 159.1±6.2 mmHg respectively, and there wasn't statistically significant difference between the SHR and intervention groups. Staining with Masson's trichrome demonstrated that the increased interstitial fibrosis of LV and ventricular remodeling in the SHR group were attenuated by atorvastatin treatment. Echocardiography examination exhibited that SHR with atorvastatin treatment showed an LV wall thickness that was obviously lower than that of water-treated SHR. In hypertrophic myocardium, accompanied by increasing C/EBPβ expression and the percentage of TUNEL-positive cells, the expression of Bcl-2/Bax ratio, PGC-1α and UCP3 were reduced, all of which could be abrogated by treatment with atorvastatin for 6 weeks.. This study further confirmed that atorvastatin could attenuate myocardial hypertrophy and remodeling in SHR by inhibiting apoptosis and reversing changes in mitochondrial metabolism. The C/EBPβ/PGC-1α/UCP3 signaling pathway might also be important for elucidating the beneficial pleiotropic effects of atorvastatin on myocardial hypertrophy.

Topics: Animals; Apoptosis; Atorvastatin; Atrial Natriuretic Factor; Blood Pressure; CCAAT-Enhancer-Binding Protein-beta; Echocardiography; Hypertension; Hypertrophy; Male; Myocardium; Myocytes, Cardiac; Natriuretic Peptide, Brain; Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha; Rats; Rats, Inbred SHR; Rats, Inbred WKY; Signal Transduction; Uncoupling Protein 3; Up-Regulation; Ventricular Remodeling

Cardiac hypertrophy is a one of common type of CHD, responsible for cardiac mortality worldwide. The present study designed to investigate the effect of muscarinic receptors agonist in the rat model of cardiac hypertrophy. A total of 30 male adult Wistar rats having body weight 300-400 gram were equally distributed in two groups (Test group: Rats with Angiotensin II + M3 receptor agonist [acetylcholine]; Reference group: Rats with cardiac hypertrophy induced by Angiotensin II). Rat model of cardiac hypertrophy were induced by Angiotensin II. Effect of M3 receptor agonist on cardiac hypertrophy was evaluated by electrocardiography, hemodynamic and histological assessment. Also, expression of M3 receptor was analyzed using by real-time-PCR and Western blot analysis. Also, vital signs such as pulse rate, and blood pressure were measured. Echocardiographic related variable including ejection fraction were also assessed in both the groups. The results of this study showed acetylcholine attenuates the hypertrophic response triggered by Angiotensin II, by upregulation of M3 receptor. Upregulation of M3 receptor after administration of acetylcholine ameliorates hypertrophic responses induced by angiotensin II. Also acetylcholine treatment prevents Angiotensin II induced increase in level of ANP and β-myosin, which are responsible for inducing cardiac hypertrophic responses. Moreover, acetylcholine ameliorates Angiotensin II induced cell enlargement by reducing the surface area of cells. Overall finding suggested that acetylcholine improves left ventricle hypertrophy and ejection fraction by activating M3 receptor in heart. The finding of this study gives the new vision to cardiovascular researchers to develop anti- hypertrophy therapy based on M3 receptor.

Topics: Acetylcholine; Angiotensin II; Animals; Atrial Natriuretic Factor; Cardiomegaly; Cells, Cultured; Electrocardiography; Hemodynamics; Hypertrophy; Male; Muscarinic Agonists; Rats; Receptor, Muscarinic M3; Ventricular Function, Left; Ventricular Myosins

We investigated the effect of ginsenoside Rb1 on cardiac function and remodeling in heart failure (HF). Four weeks after HF induction, the rats were administrated with ginsenoside Rb1 (35 and 70 mg/kg) and losartan (4.5 mg/kg) for 8 weeks. Losartan was used as a positive control. Cardiac function was assessed by measuring hemodynamic parameters. Histological changes were analyzed by HE and Masson's trichrome staining. Cardiac hypertrophy, fibrosis, mitochondrial membrane potential and glucose transporter type 4 (GLUT4) levels were evaluated. In the present study, high dose of (H-) ginsenoside Rb1 decreased heart rate, improved cardiac function and alleviated histological changes induced by HF. H-ginsenoside Rb1 attenuated cardiac hypertrophy and myocardial fibrosis by decreasing left ventricular (LV) weight/heart weight ratio and cardiomyocyte cross-sectional area and reducing the levels of atrial natriuretic factor (ANF), β-myosin heavy chain (β-MHC), periostin, collagen I, Angiotensin II (Ang II), Angiotensin converting enzyme (ACE) and Ang II type 1 (AT1) receptor. Moreover, H-ginsenoside Rb1 decreased mitochondrial membrane potential and enhanced the translocation of GLUT4 to plasma membrane. The TGF-β1/Smad and ERK signaling pathways were inhibited and the Akt pathway was activated. These findings suggest that ginsenoside Rb1 might restore cardiac/mitochondrial function, increase glucose uptake and protect against cardiac remodeling via the TGF-β1/Smad, ERK and Akt signaling pathways.

Topics: Animals; Aortic Coarctation; Atrial Natriuretic Factor; Disease Models, Animal; Dose-Response Relationship, Drug; Fibrosis; Ginsenosides; Glucose Transporter Type 4; Heart Failure; Hypertrophy; Male; Membrane Potential, Mitochondrial; Myocardium; Phytotherapy; Rats, Sprague-Dawley; Ventricular Remodeling

The present study was aimed to determine whether stimulating Npr1 gene activity using Valporic acid (VA), a small short chain fatty acid molecule can enhance ANP mediated anti-hypertrophic activity in isoproterenol (ISO) - treated H9c2 cells in vitro. H9c2 cells were treated with ISO (10

Topics: Animals; Atrial Natriuretic Factor; Biomarkers; Cell Line; Cyclic GMP; Drug Synergism; Gene Expression Regulation; Histone Deacetylase 1; Histone Deacetylase 2; Hypertrophy; Myocardium; Rats; Receptors, Atrial Natriuretic Factor; RNA, Messenger; Transcription, Genetic; Valproic Acid

UbiA prenyltransferase domain containing 1 (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.

Topics: Angiotensin II; Apoptosis; Atrial Natriuretic Factor; Caspase 3; Cell Line; Dimethylallyltranstransferase; Gene Expression; Humans; Hypertrophy; Myocytes, Cardiac; Nitric Oxide; Nitric Oxide Synthase Type III; RNA Interference; RNA, Small Interfering; Ubiquinone

We sought to determine the effect of chronic activation of β-adrenergic receptor (β-AR) on the left ventricular (LV) expression profile of Npr1 and Npr2 (coding for NPR-A and NPR-B, respectively) genes, and the functional activity of these receptors in adult Wistar rat hearts. The Npr1 gene expression was markedly reduced (3.5-fold), while the Npr2 gene expression was up regulated (4-fold) in Isoproterenol (ISO)-treated heart as compared with controls. A gradual reduction in NPR-A protein (3-fold), cGMP levels (75%) and a steady increased expression of NPR-B protein (4-fold), were noticed in ISO hearts. Further, in-vitro membranes assay shows that NPR-A dependent guanylyl cyclase (GC) activity was down-regulated (2-fold), whereas NPR-B dependent GC activity was increased (5-fold) in ISO treated hearts. Atenolol treatment normalized the altered expression of Npr1 and Npr2 genes. In conclusion, the chronic β-AR activation differentially regulates Npr1 and Npr2 genes in the heart. Npr1 down regulation is positively associated with the development of left ventricular hypertrophy (LVH) in ISO rats.

Topics: Animals; Atrial Natriuretic Factor; Cyclic GMP; Down-Regulation; Guanylate Cyclase; Heart; Hypertrophy; Male; Rats; Rats, Wistar; Receptors, Adrenergic, beta; Receptors, Atrial Natriuretic Factor; Signal Transduction; Up-Regulation

Aldosterone acts on its target tissue through a classical mechanism or through the rapid pathway through a putative membrane-bound receptor. Our goal here was to better understand the molecular and biochemical rapid mechanisms responsible for aldosterone-induced cardiomyocyte hypertrophy. We have evaluated the hypertrophic process through the levels of ANP, which was confirmed by the analysis of the superficial area of cardiomyocytes. Aldosterone increased the levels of ANP and the cellular area of the cardiomyocytes; spironolactone reduced the aldosterone-increased ANP level and cellular area of cardiomyocytes. Aldosterone or spironolactone alone did not increase the level of cyclic 3',5'-adenosine monophosphate (cAMP), but aldosterone plus spironolactone led to increased cAMP level; the treatment with aldosterone + spironolactone + BAPTA-AM reduced the levels of cAMP. These data suggest that aldosterone-induced cAMP increase is independent of mineralocorticoid receptor (MR) and dependent on Ca(2+). Next, we have evaluated the role of A-kinase anchor proteins (AKAP) in the aldosterone-induced hypertrophic response. We have found that St-Ht31 (AKAP inhibitor) reduced the increased level of ANP which was induced by aldosterone; in addition, we have found an increase on protein kinase C (PKC) and extracellular signal-regulated kinase 5 (ERK5) activity when cells were treated with aldosterone alone, spironolactone alone and with a combination of both. Our data suggest that PKC could be responsible for ERK5 aldosterone-induced phosphorylation. Our study suggests that the aldosterone through its rapid effects promotes a hypertrophic response in cardiomyocytes that is controlled by an AKAP, being dependent on ERK5 and PKC, but not on cAMP/cAMP-dependent protein kinase signaling pathways. Lastly, we provide evidence that the targeting of AKAPs could be relevant in patients with aldosterone-induced cardiac hypertrophy and heart failure.

Topics: A Kinase Anchor Proteins; Aldosterone; Animals; Atrial Natriuretic Factor; Cyclic AMP; Egtazic Acid; Heart Failure; Humans; Hypertrophy; Mitogen-Activated Protein Kinase 7; Myocytes, Cardiac; Phosphorylation; Primary Cell Culture; Protein Kinase C; Rats; Receptors, Mineralocorticoid; Signal Transduction; Spironolactone

To investigate the potential effect of curcumin on cardiomyocyte hypertrophy and a possible mechanism involving the PPARγ/Akt/NO signaling pathway in diabetes.. The cardiomyocyte hypertrophy induced by high glucose (25.5mmol/L) and insulin (0.1μmol/L) (HGI) and the antihypertrophic effect of curcumin were evaluated in primary culture by measuring the cell surface area, protein content and atrial natriuretic factor (ANF) mRNA expression. The mRNA and protein expressions were assayed by reverse transcription PCR and Western blotting, whereas the NO concentration and endothelial NO synthase (eNOS) activity were determined using nitrate reduction and ELISA methods, respectively.. The cardiomyocyte hypertrophy induced by HGI was characterized by increasing ANF mRNA expression, total protein content, and cell surface area, with downregulated mRNA and protein expressions of both PPARγ and Akt, which paralleled the declining eNOS mRNA expression, eNOS content, and NO concentration. The effects of HGI were inhibited by curcumin (1, 3, 10μmol/L) in a concentration-dependent manner. GW9662 (10μmol/L), a selective PPARγ antagonist, could abolish the effects of curcumin. LY294002 (20μmol/L), an Akt blocker, and N(G)-nitro-l-arginine-methyl ester (100μmol/L), a NOS inhibitor, could also diminish the effects of curcumin.. The results suggested that curcumin supplementation can improve HGI-induced cardiomyocytes hypertrophy in vitro through the activation of PPARγ/Akt/NO signaling pathway.

Topics: Anilides; Animals; Atrial Natriuretic Factor; Cells, Cultured; Chromones; Curcumin; Glucose; Hypertrophy; Insulin; Morpholines; Myocytes, Cardiac; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase Type III; PPAR gamma; Proto-Oncogene Proteins c-akt; Rats; Rats, Sprague-Dawley; RNA, Messenger; Signal Transduction

Proteinuria is a hallmark of chronic kidney disease (CKD) and cardiovascular disease (CVD), and a good predictor of clinical outcome. Selective endothelin A (ETA) receptor antagonist used with renin-angiotensin system (RAS) inhibitors prevents development of proteinuria in CKD. However, whether the improvement in proteinuria would have beneficial effects on CVD, independent of RAS inhibition, is not well understood. In this study, we investigated whether atrasentan, an ETA receptor antagonist, has renal and cardiovascular effects independent of RAS inhibition. Male Dahl salt sensitive (DSS) rats, at six weeks of age, received water with or without different doses of atrasentan and/or enalapril under high salt (HS) diet or normal diet (ND) for 6 weeks. At the end of 12th week, atrasentan at a moderate dose significantly attenuated proteinuria and serum creatinine without reducing mean arterial pressure (MAP), thereby preventing cardiac hypertrophy and improving cardiac function. ACE inhibitor enalapril at a dose that did not significantly lowered BP, attenuated cardiac hypertrophy while moderately improving cardiac function without reducing proteinuria and serum creatinine level. Nonetheless, combined therapy of atrasentan and enalapril that does not altering BP exerted additional cardioprotective effect. Based on these findings, we conclude that BP independent monotherapy of ETA receptor antagonist attenuates the progression of CKD and significantly mitigates CVD independent of RAS inhibition.

Topics: Animals; Atrasentan; Atrial Natriuretic Factor; Blood Pressure; Cardiomegaly; Disease Models, Animal; Echocardiography; Endothelin A Receptor Antagonists; Gene Expression; Heart Diseases; Hemodynamics; Hypertension; Hypertrophy; Kidney Function Tests; Male; Myocytes, Cardiac; Phenylephrine; Pyrrolidines; Rats; Receptor, Endothelin A; Renal Insufficiency, Chronic

Anabolic-androgenic steroids are misused, including women, but little is known about the cardiovascular effects of these drugs on females.. Evaluated the effects of nandrolone decanoate (ND), physical exercise and estrogen deficiency on female rats.. Female Wistar rats were divided into 8 groups: S and OVX: (SHAM: sham surgery; OVX: ovariectomy, vehicle), SE and OVXE (resistance exercise 5 times a week, vehicle), SD and OVXD (treated with ND, 20 mg/kg/week for 4 weeks); SDE and OVXDE. Treatments were initiated 21 days after surgery. The Bezold–Jarisch reflex was assessed by Phenylbiguanide administration. The right atrium, kidney, and serum were collected for molecular analyses by RT-PCR of atrial natriuretic peptide (ANP), A-type natriuretic peptide receptor (NPR-A) and NPR-C. ELISA assay to estradiol and testosterone concentrations. The gastrocnemius muscle, heart and kidney weights/tibia length were measured.Morphometric analysis of heart was made (H/E) and collagen content of heart and kidney were evaluated using Pirossirius Red.. ND treatment increased ANP expression on atrium and decreased NPR-A expression in kidney. Physical exercise and ovariectomy did not alter this parameter. NPR-C level was reduced in the SDE and OVXDE. Renal and cardiac hypertrophy was observed after ND treatment, with collagen deposition. Plasma estrogen concentrations were reduced and serum testosterone concentrations were increased after ND treatment.. ANP has an important role in modulating the cardiovascular effects of ND in females. Thismodulating may have occurred by the increasing ANP expression, reducing NPR-A and NPR-C expression levels, and changing sex hormone levels.

Topics: Anabolic Agents; Animals; Arterial Pressure; Atrial Natriuretic Factor; Baroreflex; Biguanides; Collagen; Estradiol; Estrogens; Female; Gene Expression; Heart; Heart Atria; Heart Rate; Hypertrophy; Kidney; Muscle, Skeletal; Myocardium; Nandrolone; Nandrolone Decanoate; Natriuretic Peptide, C-Type; Organ Size; Ovariectomy; Physical Conditioning, Animal; Rats; Receptors, Atrial Natriuretic Factor; Testosterone; Tibia

Oxidative stress has been implicated in cardiac remodeling (cardiac fibrosis and hypertrophy), which impairs cardiac function and metabolism; therefore, it is anticipated antioxidative compounds will have protective properties against cardiac remodeling. Luteolin (3',4',5,7-tetrahydroxyflavone), a widely distributed flavonoid found in many herbal extracts including celery, green pepper, perilla leaves and seeds, and chamomile, is a known to be a potent antioxidant and was previously demonstrated to exert an antifibrotic effect in the lungs and the liver. In this study, we clearly demonstrate that oral pretreatment with the higher-luteolin diet (0.035% (wt/wt)) protected against cardiac fibrosis and hypertrophy as well as a hyperoxidative state in Ang II-infused rats. In cardiac tissue, increased gene expression levels of TGFβ1, CTGF, Nox2, Nox4, ANP, and BNP induced by Ang II were restored by oral pretreatment of this high-luteolin diet. In cultured rat cardiac fibroblasts, H2O2-induced TGFβ1 expression and the phosphorylation of JNK were suppressed by luteolin pretreatment. In conclusion, food-derived luteolin has protective actions against Ang II-induced cardiac remodeling, which could be mediated through attenuation of oxidative stress.

Topics: Angiotensin II; Animals; Antioxidants; Atrial Natriuretic Factor; Connective Tissue Growth Factor; Diet; Fibroblasts; Fibrosis; Flavonoids; Food; Heart; Hydrogen Peroxide; Hypertrophy; Luteolin; Male; Myocardium; Oxidative Stress; Phosphorylation; Rats; Rats, Sprague-Dawley; Signal Transduction; Transforming Growth Factor beta1; Ventricular Remodeling

N-terminal segment of B-type natriuretic peptide prohormone (NT-proBNP) is elevated in patients with acute myocardial infarction (AMI) thus providing both diagnostic information and prognostic information. The aim of the present study was to determine the time course of NT-proBNP release in patients undergoing transcoronary ablation of septal hypertrophy (TASH) a procedure mimicking AMI.. We analyzed the release kinetics of NT-proBNP in 18 consecutive patients with hypertrophic obstructive cardiomyopathy undergoing TASH. Serum samples were collected prior to and at 15, 30, 45, 60, 75, 90, and 105 min, and 2, 4, 8, and 24h after TASH.. NT-proBNP concentrations showed a continuous increase during the first 75 min with a significant percent change compared to baseline value already 15 min after TASH (105.6% [IQR 102.2-112.7]; P<0.001). All patients had a significant increase of NT-proBNP at 45 min (range of percent increase [min-max]: 103.5-137.2%; range of absolute increase [min-max]: 23.5-304.0 ng/L). NT-proBNP concentrations decreased below the baseline value until the 8th h after initiation of myocardial infarction.. NT-proBNP concentration increases immediately after induction of myocardial infarction proving early evidence of myocardial injury despite the decrease of the left ventricular wall stress due to the TASH related reduction of the left ventricular outflow gradient.

Topics: Ablation Techniques; Acute Disease; Atrial Natriuretic Factor; Female; Heart Septum; Humans; Hypertrophy; Kinetics; Male; Middle Aged; Myocardial Infarction; Protein Precursors

The precise molecular mechanisms leading to disturbance of Ca(2+)/calmodulin-dependent intracellular signalling in cardiac hypertrophy remains unclear. As an endogenous calmodulin regulator protein, the pathophysiology role of PEP-19 during cardiac hypertrophy was investigated in the present study. We here demonstrated that PEP-19 protein levels are significantly elevated in the aortic banding model in vivo and angiotensin II-induced cardiomyocyte hypertrophy in vitro. Consistent with inhibitory actions of PEP-19 on cardiomyocyte hypertrophy, induction of CaMKII and calcineurin activation as well as hypertrophy-related genes including atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) was significantly inhibited by PEP-19 transfection. Moreover, PEP-19 partially ameliorates angiotensin II-induced elevation of phospho-phospholamban (Thr-17) and sarcoplasmic reticulum Ca(2+) release in cardiomyocytes. Together, our results suggest that PEP-19 attenuates angiotensin II-induced cardiomyocyte hypertrophy via suppressing the disturbance of CaMKII and calcineurin signaling.

Topics: Angiotensin II; Animals; Atrial Natriuretic Factor; Calcineurin; Calcium; Calcium Signaling; Calcium-Binding Proteins; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Calmodulin; Calmodulin-Binding Proteins; Cardiomegaly; Cells, Cultured; Gene Expression; Hypertrophy; Male; Myocytes, Cardiac; Natriuretic Peptide, Brain; Nerve Tissue Proteins; Rats, Sprague-Dawley; Sarcoplasmic Reticulum

Cardiac hypertrophy is an important compensatory mechanism in response to a pressure overload, but a sustained excessive cardiac workload may deteriorate to maladaptive hypertrophy and to increased risk of heart failure. In this study, we evaluated the effects of KS370G on left ventricular hypertrophy and function. Abdominal aortic banding was performed by constricting the abdominal aorta. Hypertrophied heart was studied at 8 weeks after the operation. After the operation, KS370G 1mg/kg (K1 group) was administered by oral gavage once a day. Left ventricular function was measured by a 1.2F pressure-volume catheter (Scisense, Canada). The levels of protein for α-SMA (smooth muscle actin), p-AKT (protein kinase B), p-GSK3β (glycogen synthase kinase 3β) and p-ERKs (extracellular signal-regulated kinases) in myocardium were analyzed by Western blot. Plasma levels of angiotensin II, atrial natriuretic peptide and lactate dehydrogenase were analyzed by commercial kits. H.E. staining and M.T. staining methods were also used to observe diameter of cardiomyocytes and collagen accumulation. Chronic oral treatment with 1mg/kg KS370G inhibited cardiac hypertrophy and improved cardiac function induced by pressure overload. KS370G also decreased the plasma levels of atrial natriuretic peptide and lactate dehydrogenase. Besides, pressure overload-induced increase of α-SMA and phosphorylation of ERK, AKT and GSK3β were significantly reduced by chronic oral treatment with KS370G. We also found that chronic oral treatment with KS370G reduced cardiac collagen accumulation. KS370G improved left ventricular function and inhibited cardiac hypertrophy through the decrease of the phosphorylation of ERK, AKT and GSK3β in pressure-overload mice heart.

Topics: Actins; Angiotensin II; Animals; Aorta; Atrial Natriuretic Factor; Caffeic Acids; Cardiotonic Agents; Collagen; Constriction; Extracellular Signal-Regulated MAP Kinases; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; Heart Ventricles; Hypertrophy; L-Lactate Dehydrogenase; Male; Mice; Mice, Inbred ICR; Myocytes, Cardiac; Phosphoproteins; Pressure; Proto-Oncogene Proteins c-akt; Time Factors

Human cytomegalovirus immediate early proteins (CMV IEs) are involved in transcriptional activities of both host and virus gene expression. This study shows that the transcriptional activity of myocardin in regulating cardiomyocyte hypertrophy is enhanced by co-expressing CMV IE2. Forced expression of IE2 increases the augmented cell size of neonatal rat cardiac myocytes induced by myocardin, as well as the mRNA and protein levels of hypertrophic genes, whereas deletion of CArG boxes in the atrial natriuretic factor (ANF) promoter attenuates the effect of CMV IE2 with myocardin. In conclusion, CMV IE2 synergistically stimulates myocardin transactivity in the hypertrophic marker gene ANF in a CArG box-dependent manner. Our study indicates that the association of CMV IEs with myocardin-induced transcription may be involved in myocardin-mediated cardiac hypertrophy.

Topics: Actinin; Animals; AT Rich Sequence; Atrial Natriuretic Factor; Chlorocebus aethiops; COS Cells; Hypertrophy; Immediate-Early Proteins; Myocytes, Cardiac; Nuclear Proteins; Promoter Regions, Genetic; Rats; Rats, Sprague-Dawley; RNA, Messenger; Serum Response Factor; Trans-Activators; Transcription, Genetic; Transcriptional Activation

Cardiac disease is frequently reported in farmed animals, and stress has been implicated as a factor for myocardial dysfunction in commercial fish rearing. Cortisol is a major stress hormone in teleosts, and this hormone has adverse effects on the myocardium. Strains of rainbow trout (Oncorhynchus mykiss) selected for divergent post-stress cortisol levels [high responsive (HR) and low responsive (LR)] have been established as a comparative model to examine how fish with contrasting stress-coping styles differ in their physiological and behavioral profiles. We show that the mean cardiosomatic index (CSI) of adult HR fish was 34% higher than in LR fish, mainly because of hypertrophy of the compact myocardium. To characterize the hypertrophy as physiological or pathological, we investigated specific cardiac markers at the transcriptional level. HR hearts had higher mRNA levels of cortisol receptors (MR, GR1 and GR2), increased RCAN1 levels [suggesting enhanced pro-hypertrophic nuclear factor of activated T-cell (NFAT) signaling] and increased VEGF gene expression (reflecting increased angiogenesis). Elevated collagen (Col1a2) expression and deposition in HR hearts supported enhanced fibrosis, whereas the heart failure markers ANP and BNP were not upregulated in HR hearts. To confirm our results outside the selection model, we investigated the effect of acute confinement stress in wild-type European brown trout, Salmo trutta. A positive correlation between post-stress cortisol levels and CSI was observed, supporting an association between enhanced cortisol response and myocardial remodeling. In conclusion, post-stress cortisol production correlates with myocardial remodeling, and coincides with several indicators of heart pathology, well-known from mammalian cardiology.

Topics: Animals; Atrial Natriuretic Factor; Biomarkers; Collagen; Female; Fibrosis; Humans; Hydrocortisone; Hypertrophy; Myocardium; Natriuretic Peptide, Brain; NFATC Transcription Factors; Receptors, Glucocorticoid; Salmonidae; Stress, Physiological; Ventricular Remodeling

Cell cycle arrest is associated with differentiation, senescence and apoptosis. We investigated alterations in the cell cycle during the development of hypertrophy induced by hydrogen peroxide (H(2)O(2)) in the H9c2 clonal myoblastic cell line. H(2)O(2) induced hypertrophy in H9c2 cells that was indicated by an increase in atrial natriuretic peptide (ANP) gene expression, a marker of cardiomyocyte hypertrophy, and a larger cell size. On induction of hypertrophy by H(2)O(2) in H9c2 cells, cell proliferation was arrested, indicated by the number of cells remaining constant during a 72-h incubation period. The cell cycle was arrested at the G1 and G2/M phases with an increase in p21 expression, a negative cell cycle regulator. Cell cycle arrest and increase in p21 expression were significantly inhibited by 1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetra (acetoxymethyl) ester (BAPTA-AM), an intracellular calcium chelator. Although ANP gene expression was induced significantly, H(2)O(2) failed to induce hypertrophy in the presence of BAPTA-AM, and the cell cycle progressed. We concluded that H(2)O(2) induced cell cycle arrest in H9c2 cells, which was related to cellular hypertrophy.

Topics: Animals; Atrial Natriuretic Factor; Biomarkers; Cell Cycle; Cell Proliferation; Cells, Cultured; Chelating Agents; Cyclin-Dependent Kinase Inhibitor p21; Egtazic Acid; Gene Expression; Hydrogen Peroxide; Hypertrophy; Myocytes, Cardiac; Rats

Topics: Animals; Atrial Natriuretic Factor; Biomarkers; Blood Pressure; Humans; Hypertension; Hypertrophy; Leukocytes; Mice; Mitogen-Activated Protein Kinases; Models, Biological; Myocardium; Phosphorylation; Pressure

Pressure overload has been shown to induce mitogen activated protein kinases (MAPKs) and reactivate the atrial natriuretic factor in the heart. To test the sensitivity of these signals to pressure overload, we assayed the activity of MAPKs extracellular signal-regulated kinase, c-Jun N-terminal kinase 1, and p38 in protein lysates from the left ventricle (LV) or white blood cells (WBC) isolated from aortic banded mice with varying levels of pressure overload. In separated mice we measured atrial natriuretic factor mRNA levels by Northern blotting. As expected, a significant induction of atrial natriuretic factor mRNA levels was observed after aortic banding, and it significantly correlated with the trans-stenotic systolic pressure gradient but not with the LV weight:body weight ratio. In contrast, a significant correlation with systolic pressure gradient or LV weight:body weight ratio was observed for all of the MAPK activity detected in LV samples or WBCs. Importantly, LV activation of MAPKs significantly correlated with their activation in WBCs from the same animal. To test whether MAPK activation in WBCs might reflect uncontrolled blood pressure levels in humans, we assayed extracellular signal-regulated kinase, c-Jun N-terminal kinase 1, and p38 activation in WBCs isolated from normotensive volunteers, hypertensive patients with controlled blood pressure values, or hypertensive patients with uncontrolled blood pressure values. Interestingly, in hypertensive patients with controlled blood pressure values, LV mass and extracellular signal-regulated kinase phosphorylation were significantly reduced compared with those in hypertensive patients with uncontrolled blood pressure values. These results suggest that MAPKs are sensors of pressure overload and that extracellular signal-regulated kinase activation in WBCs might be used as a novel surrogate biomarker of uncontrolled human hypertension.

Topics: Animals; Aorta; Atrial Natriuretic Factor; Blood Pressure; Blotting, Northern; Blotting, Western; Constriction, Pathologic; Enzyme Activation; Female; Gene Expression; Humans; Hypertension; Hypertrophy; Leukocytes; Male; MAP Kinase Signaling System; Mice; Middle Aged; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 8; Mitogen-Activated Protein Kinases; Myocardium; p38 Mitogen-Activated Protein Kinases; Phosphorylation; Pressure

Aldosterone exerts its effect by binding to specific mineralocorticoid receptors (MRs). Spironolactone blocks the aldosterone system, which ameliorates heart failure in humans, but the precise molecular mechanisms of MR blockade are unclear.. Neonatal rat cardiomyocytes were stimulated with phenylephrine (PE), aldosterone, and/or spironolactone. The association of the MR with p300, a transcriptional coactivator of GATA4 required for hypertrophic responses, was examined. MR and p300 synergistically activated GATA4-dependent atrial natriuretic factor (ANF) promoter activities. The stimulation of cardiomyocytes with PE induced translocation of the MRs into the nuclei and markedly increased the association of MRs with p300. Compatible with the synergistic activation by the MR and p300, aldosterone further augmented the PE-induced increase in cell size and induction of ANF gene transcription. Blockade of MR activation by spironolactone inhibited the PE-induced nuclear translocation of MRs and hypertrophic responses.. For the first time it has been demonstrated that the aldosterone/MR system associates with the p300/GATA4 transcriptional pathway during the hypertrophic response of cardiomyocytes, and may provide a mechanism of the beneficial effects of aldosterone-blocking agents in heart failure therapy in humans. (Circ J 2010; 74: 156 - 162).

Topics: Aldosterone; Animals; Atrial Natriuretic Factor; Chlorocebus aethiops; COS Cells; Disease Models, Animal; E1A-Associated p300 Protein; GATA4 Transcription Factor; Hypertrophy; Mice; Myocytes, Cardiac; NIH 3T3 Cells; Phenylephrine; Rats; Receptors, Mineralocorticoid; Signal Transduction; Spironolactone; Transfection

The mixed-lineage kinases (MLKs) act upstream of mitogen-activated protein kinases, but their role in cardiac biology and pathology is largely unknown.. We investigated the effect of a MLK1-3 inhibitor CEP-11004 on G protein-coupled receptor agonist-induced stress response in neonatal rat cardiac myocytes in culture.. CEP-11004 administration dose-dependently attenuated phenylephrine and endothelin-1 (ET-1)-induced c-Jun N-terminal kinase activation. MLK inhibition also reduced ET-1- and phenylephrine-induced phosphorylation of p38 mitogen-activated protein kinase. In contrast, phenylephrine-induced extracellular signal-regulated kinase phosphorylation was further up-regulated by CEP-11004. ET-1 increased activator protein-1 binding activity 3.5-fold and GATA-binding protein 4 (GATA-4) binding activity 1.8-fold, both of which were attenuated with CEP-11004 administration by 59% and 63% respectively. Phenylephrine induced activator protein-1 binding activity by 2.6-fold, which was decreased by 81% with CEP-11004 administration. Phenylephrine also induced a 3.7-fold increase in the transcriptional activity of B-type natriuretic peptide (BNP), which was attenuated by 41% with CEP-11004 administration. In agreement, MLK inhibition also reduced hypertrophic agonist-induced secretion of immunoreactive atrial natriuretic peptide and BNP.. These results showed that inhibition of the MLK1-3 signalling pathway was sufficient for suppressing the activity of key nuclear effectors (GATA-4 and activator protein-1 transcription factors) in cardiac hypertrophy, and attenuated the agonist-induced atrial natriuretic peptide secretion and activation of BNP gene transcription.

Topics: Animals; Animals, Newborn; Atrial Natriuretic Factor; Carbazoles; Cardiomegaly; Cell Nucleus; Endothelin-1; Genes, jun; Heart; Hypertrophy; JNK Mitogen-Activated Protein Kinases; MAP Kinase Kinase Kinases; Mitogen-Activated Protein Kinase Kinase Kinase 11; Mitogen-Activated Protein Kinases; Myocytes, Cardiac; Natriuretic Peptide, Brain; p38 Mitogen-Activated Protein Kinases; Phenylephrine; Phosphorylation; Rats; Rats, Sprague-Dawley; Signal Transduction; Transcription Factor AP-1; Transcription Factors

Atrial and brain natriuretic peptides (ANP and BNP, respectively) exert antihypertrophic effects in the heart via their common receptor, guanylyl cyclase (GC)-A, which catalyzes the synthesis of cGMP, leading to activation of protein kinase (PK)G. Still, much of the network of molecular mediators via which ANP/BNP-GC-A signaling inhibit cardiac hypertrophy remains to be characterized.. We investigated the effect of ANP-GC-A signaling on transient receptor potential subfamily C (TRPC)6, a receptor-operated Ca(2+) channel known to positively regulate prohypertrophic calcineurin-nuclear factor of activated T cells (NFAT) signaling.. In cardiac myocytes, ANP induced phosphorylation of TRPC6 at threonine 69, the PKG phosphorylation site, and significantly inhibited agonist-evoked NFAT activation and Ca(2+) influx, whereas in HEK293 cells, it dramatically inhibited agonist-evoked TRPC6 channel activity. These inhibitory effects of ANP were abolished in the presence of specific PKG inhibitors or by substituting an alanine for threonine 69 in TRPC6. In model mice lacking GC-A, the calcineurin-NFAT pathway is constitutively activated, and BTP2, a selective TRPC channel blocker, significantly attenuated the cardiac hypertrophy otherwise seen. Conversely, overexpression of TRPC6 in mice lacking GC-A exacerbated cardiac hypertrophy. BTP2 also significantly inhibited angiotensin II-induced cardiac hypertrophy in mice.. Collectively, these findings suggest that TRPC6 is a critical target of antihypertrophic effects elicited via the cardiac ANP/BNP-GC-A pathway and suggest TRPC6 blockade could be an effective therapeutic strategy for preventing pathological cardiac remodeling.

Topics: Anilides; Animals; Atrial Natriuretic Factor; Calcium Channels; Cells, Cultured; Cyclic GMP; Cyclic GMP-Dependent Protein Kinases; Disease Models, Animal; Humans; Hypertrophy; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Myocardium; Myocytes, Cardiac; Natriuretic Peptide, Brain; NFATC Transcription Factors; Patch-Clamp Techniques; Rats; Receptors, Atrial Natriuretic Factor; Signal Transduction; Thiadiazoles; TRPC Cation Channels; TRPC6 Cation Channel

Natriuretic peptides (NPs) inhibit cardiomyocyte hypertrophy through a cyclic GMP (cGMP)-dependent process, although these effects are associated with substantial vasodilatation. In this study, we used CU-NP, a non-vasodilatating novel NP synthesized from the ring structure of human C-type NP (CNP) and both C- and N-termini of urodilatin, and investigated whether it can directly modulate cardiomyocyte hypertrophy.. Experiments were carried out in cultured neonatal rat ventricular myocytes exposed to phenylephrine, angiotensin II, or endothelin-1 in the absence or presence of CU-NP. CU-NP produced a concentration- and time-dependent increase in intracellular cGMP levels. The hypertrophic responses to all agonists were abrogated by 10 nM CU-NP. CU-NP treatment also prevented increased activity, gene and protein expression of sodium-hydrogen exchanger-1 (NHE-1) as well as elevations in intracellular Na(+) concentrations caused by hypertrophic agents. In addition, these effects were associated with a more than two-fold increase in activity of the Ca(2+)-dependent protein phosphatase calcineurin that peaked 6 h after addition of hypertrophic stimuli. Early (1-3 h) calcineurin activation was unaffected by CU-NP, although activation at 6 and 24 h was prevented by CU-NP as was the resultant translocation of the transcriptional factor NFAT into nuclei.. Our study demonstrates a direct anti-hypertrophic effect of the chimeric peptide CU-NP via NHE-1 inhibition, thereby preventing calcineurin activation and NFAT nuclear import. Thus, CU-NP represents a novel fusion peptide of CNP and urodilatin that has the potential to be developed into a therapeutic agent to treat cardiac hypertrophy and heart failure.

Topics: Animals; Atrial Natriuretic Factor; Calcineurin; Cell Nucleus; Cells, Cultured; Cyclic GMP; Dose-Response Relationship, Drug; Hypertrophy; Mitogen-Activated Protein Kinase Kinases; Models, Animal; Myocytes, Cardiac; Peptide Fragments; Rats; Rats, Sprague-Dawley; Recombinant Fusion Proteins; Signal Transduction; Sodium; Sodium-Hydrogen Exchangers

Left atrial volume (LAV) has recently emerged as a useful biomarker for risk stratification and risk monitoring in patients with end stage renal disease. We investigated the relationship between cardiac natriuretic peptides (atrial natriuretic peptide [ANP] and brain natriuretic peptide [BNP]) and norepinephrine (NE) with LAV and LAV changes over time in 199 end stage renal disease patients. At baseline, LAV was directly related to BNP (r=0.60), ANP (r=0.59), and NE (r=0.28; P<0.001), and these relationships held true in multiple-regression models adjusting for potential confounders (P< or =0.003). In the longitudinal study (17+/-2 months), LAV increased from 9.8+/-4.6 to 10.9+/-5.4 mL/m(2.7) (+11%). In a multiple linear regression model, BNP (beta=0.28; P=0.003), ANP (beta=0.22; P=0.03), and NE (beta=0.27; P=0.003) predicted LAV changes. The area under the receiver operating characteristic curve for predicting LAV changes (>3 mL/m(2.7) per year) of a risk score on the basis of standard risk factors was 0.72. Plasma BNP (+12%; P=0.004), ANP (+8%; P=0.03), NE (+8%; P=0.05) and midwall fraction shortening (+8%; P=0.05) increased the area under the receiver operating characteristic curve to a significant extent, whereas LV mass did not (+5%; P=0.18). Predictive models, including BNP, ANP, and NE, maintained a satisfactory discriminatory power for LAV and LAV changes also when tested by a bootstrap resampling technique. BNP and ANP are strongly related to LAV in the end stage renal disease patients and predict LAV changes over time in these patients. Because an increased LAV underlies diastolic dysfunction and/or volume overload (ie, potentially modifiable risk factors), the measurement of the plasma concentration of these compounds might be useful for risk stratification and for guiding treatment in dialysis patients.

Topics: Adult; Aged; Atrial Natriuretic Factor; Biomarkers; Cardiovascular Diseases; Cohort Studies; Echocardiography; Female; Heart Atria; Humans; Hypertrophy; Kidney Failure, Chronic; Longitudinal Studies; Male; Middle Aged; Natriuretic Peptide, Brain; Norepinephrine; Predictive Value of Tests; Regression Analysis; Risk Factors; ROC Curve

Altered mechanical stress and strain in cardiac myocytes induce modifications in gene expression that affects cardiac remodeling and myocyte contractile function. To study the mechanisms of mechanotransduction in cardiomyocytes, probing alterations in mechanics and gene expression has been an effective strategy. However, previous studies are self-limited due to the general use of isolated neonatal rodent myocytes or intact animals. The main goal of this study was to develop a novel tissue culture chamber system for mouse myocardium that facilitates loading of cardiac tissue, while measuring tissue stress and deformation within a physiological environment. Intact mouse right ventricular papillary muscles were cultured in controlled conditions with superfusate at 95% O2/ 5% CO2, and 34 degrees C, such that cell to extracellular matrix adhesions as well as cell to cell adhesions were undisturbed and both passive and active mechanical properties were maintained without significant changes. The system was able to measure the induction of hypertrophic markers (BNP, ANP) in tissue after 2 hrs and 5 hrs of stretch. ANP induction was highly correlated with the diastolic load of the muscle but not with developed systolic load. Load induced ANP expression was blunted in muscles from muscle-LIM protein knockout mice, in which defective mechanotransduction pathways have been predicted.

Topics: Animals; Atrial Natriuretic Factor; Gene Expression Regulation; Hypertrophy; Male; Mice; Mice, Inbred C57BL; Myocardial Contraction; Natriuretic Peptide, Brain; Papillary Muscles; Reverse Transcriptase Polymerase Chain Reaction; Stress, Mechanical; Time Factors; Tissue Culture Techniques

Our understanding of the natriuretic peptide system continues to evolve rapidly. B-type natriuretic peptide (BNP), originally thought to be a simple volume-regulating hormone that is produced in response to cardiac stretch, has been shown to also play important roles in modulating bronchodilation, endothelial function, and cardiac remodeling. Recent data demonstrate that elevated levels of BNP in patients with heart failure do not represent a simple ratcheting up of normal production in response to increased stimulus. Instead, we now know that chronic stimulation of BNP synthesis induces a reversion to fetal gene expression, resulting in production of high molecular weight forms of BNP that are functionally deficient. Standard point-of-care BNP assays are immunoassays that will detect any molecule containing the target epitopes. Consequently, these assays cannot distinguish between defective, high molecular weight forms of BNP and normal, physiologically active BNP. In 2 separate evaluations, mass spectroscopy detected little, if any, normal BNP in patients with heart failure, despite the appearance of high circulating levels of immunoreactive BNP (iBNP) using commercial assays. Therefore, these commercial assays should be considered to be only an indication of myocardial stress. They do not measure physiologic BNP activity. This accounts for the "BNP paradox," namely, that administration of exogenous recombinant human BNP (rhBNP, nesiritide) has substantial clinical and hemodynamic impact in the presence of high levels of circulating iBNP using commercial assays. In addition to its short-term hemodynamic impact, rhBNP may have other important effects in this setting, and further investigation is warranted.

Topics: Atrial Natriuretic Factor; Creatinine; Fibrosis; Heart Failure; Humans; Hypertrophy; Immunoassay; Myocardium; Natriuretic Agents; Natriuretic Peptide, Brain; Renin-Angiotensin System

The biomechanical environment to which cells are exposed is important to their normal growth, development, interaction, and function. Accordingly, there has been much interest in studying the role of biomechanical forces in cell biology and pathophysiology. This has led to the introduction and even commercialization of many experimental devices. Many of the early devices were limited by the heterogeneity of deformation of cells cultivated in different locations of the culture plate membranes and were also attached with complicated technical/electronic efforts resulting in a restriction of the reproducibility of these devices. The objective of this study was to design and build a simple device to allow the application of dose-dependent homogeneous equibiaxial static stretch to cells cultured on flexible silicone membranes to investigate biological and biomedical questions. In addition, cultured neonatal rat atrial cardiomyocytes were stretched with the proposed device with different strain gradients. For the first time with this study we could demonstrate that stretch up to 21% caused dose-dependent changes in biological markers such as the calcineurin activity, modulatory calcineurin-interacting protein-1, voltage-gated potassium channel isoform 4.2, and voltage-gated K(+) channel-interacting proteins-2 gene expression and transient outward potassium current densities but not the protein-to-DNA ratio and atrial natriuretic peptide mRNA. With both markers mentioned last, dose-dependent stretch alterations could only be achieved with stretch up to 13%. The simple and low-cost device presented here might be applied to a wide range of experimental settings in different fields of research.

Topics: Animals; Animals, Newborn; Atrial Natriuretic Factor; Calcineurin; Cell Culture Techniques; Cell Size; Cells, Cultured; Equipment Design; Heart Atria; Hypertrophy; Intracellular Signaling Peptides and Proteins; Kv Channel-Interacting Proteins; Materials Testing; Membrane Potentials; Membranes, Artificial; Myocytes, Cardiac; Pliability; Potassium; Rats; Rats, Wistar; RNA, Messenger; Shal Potassium Channels; Silicones; Stress, Mechanical; Transcription Factors

Obstructive sleep apnea (OSA) increases cardiovascular morbidity and mortality. We have reported that chronic intermittent hypoxia (CIH), a direct consequence during OSA, leads to left ventricular (LV) remodeling and dysfunction in rats. The present study is to determine LV myocardial cellular injury that is possibly associated with LV global dysfunction. Fifty-six rats were exposed either to CIH (nadir O(2) 4-5%) or sham (handled normoxic controls, HC), 8 h/day for 6 wk. At the end of the exposure, we studied LV global function by cardiac catheterization, and LV myocardial cellular injury by in vitro analyses. Compared with HC, CIH animals demonstrated elevations in mean arterial pressure and LV end-diastolic pressure, but reductions in cardiac output (CIH 141.3 +/- 33.1 vs. HC 184.4 +/- 21.2 ml x min(-1) x kg(-1), P < 0.01), maximal rate of LV pressure rise in systole (+dP/dt), and maximal rate of LV pressure fall in diastole (-dP/dt). CIH led to significant cell injury in the left myocardium, including elevated LV myocyte size, measured by cell surface area (CIH 3,564 +/- 354 vs. HC 2,628 +/- 242 microm(2), P < 0.05) and cell length (CIH 148 +/- 23 vs. HC 115 +/- 16 microm, P < 0.05), elevated terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL)-stained positive cell number (CIH 98 +/- 45 vs. HC 15 +/- 13, P < 0.01), elevated caspase-3 activity (906 +/- 249 vs. 2,275 +/- 1,169 pmol x min(-1) x mg(-1), P < 0.05), and elevated expression of several remodeling gene markers, including c-fos, atrial natriuretic peptide, beta-myosin heavy chain, and myosin light chain-2. However, there was no difference between groups in sarcomere contractility of isolated LV myocytes, or in LV collagen deposition on trichrome-stained slices. In conclusion, CIH-mediated LV global dysfunction is associated with myocyte hypertrophy and apoptosis at the cellular level.

Topics: Animals; Apoptosis; Atrial Natriuretic Factor; Blood Pressure; Body Weight; Cardiac Myosins; Cardiac Output; Cardiomegaly; Caspase 3; Cell Size; Chronic Disease; Collagen; Disease Models, Animal; Hypertrophy; Hypoxia; Male; Myocardial Contraction; Myocardium; Myocytes, Cardiac; Myosin Heavy Chains; Myosin Light Chains; Proto-Oncogene Proteins c-fos; Rats; Rats, Sprague-Dawley; Research Design; RNA, Messenger; Ventricular Dysfunction, Left; Ventricular Pressure; Ventricular Remodeling

Chronic inhibition of nitric oxide (NO) synthesis is associated with hypertension, myocardial ischemia, oxidative stress and hypertrophy; expression of adrenomedullin (AM) and intermedin (IMD) and their receptor activity modifying proteins (RAMPs 1-3) is augmented in cardiomyocytes, indicating that the myocardial AM/ IMD system may be activated in response to pressure loading and ischemic insult. The aim was to examine effects on (i) parameters of cardiomyocyte hypertrophy and on (ii) expression of AM and IMD and their receptor components in NO-deficient cardiomyocytes of an intervention chosen specifically for ability to alleviate pressure loading and ischemic injury concurrently.. The NO synthesis inhibitor, N(G)-nitro-L-arginine methyl ester (L-NAME, 35 mg.kg(-1).day(-1)) was given to rats for 8 weeks, with/ without concurrent administration of beta-adrenoceptor antagonist, atenolol (25 mg.kg(-1).day(-1)) / calcium channel blocker, nifedipine (20mg.kg(-1).day(-1)).. In L-NAME treated rats, atenolol / nifedipine abolished increases in systolic blood pressure and plasma AM and IMD levels and in left ventricular cardiomyocytes: (i) normalized increased cell width and mRNA expression of hypertrophic (sk-alpha-actin) and cardio-endocrine (ANP, BNP, ET) genes; (ii) normalized augmented membrane protein oxidation; (iii) normalized mRNA expression of AM, IMD, RAMP1, RAMP2 and RAMP3.. normalization of blood pressure and membrane oxidant status together with prevention of hypertrophy and normalization of the augmented expression of AM, IMD and their receptor components in NO-deficient cardiomyocytes by atenolol / nifedipine supports involvement of both pressure loading and ischemic insult in stimulating cardiomyocyte hypertrophy and induction of these counter-regulatory peptides and their receptor components. Attenuation of augmented expression of IMD in this model cannot however be explained simply by prevention of cardiomyocyte hypertrophy.

Topics: Adrenomedullin; Animals; Atenolol; Atrial Natriuretic Factor; Blood Pressure; Endothelin-1; Gene Expression Regulation; Hypertrophy; Intracellular Signaling Peptides and Proteins; Male; Membrane Proteins; Myocytes, Cardiac; Natriuretic Peptide, Brain; Neuropeptides; NG-Nitroarginine Methyl Ester; Nifedipine; Nitric Oxide; Oxidative Stress; Peptides; Protein Precursors; Rats; Rats, Sprague-Dawley; Receptor Activity-Modifying Protein 1; Receptor Activity-Modifying Protein 2; Receptor Activity-Modifying Protein 3; Receptor Activity-Modifying Proteins; Receptors, Cell Surface; Systole

Evidences suggest that lipopolysaccharide (LPS) participates in the inflammatory response in the cardiovascular system; however, it is unknown if LPS is sufficient to cause the cardiac hypertrophy. In the present study, we treated H9c2 myocardiac cells with LPS to explore whether LPS causes cardiac hypertrophy, and to identify the precise molecular and cellular mechanisms behind hypertrophic responses. Here we show that LPS challenge induces pathological hypertrophic responses such as the increase in cell size, the reorganization of actin filaments, and the upregulation of hypertrophy markers including atrial natriuretic peptide (ANP) and B-type natriuretic peptide (BNP) in H9c2 cells. LPS treatment significantly promotes the activation of GATA-4 and the nuclear translocation of NFAT-3, which act as transcription factors mediating the development of cardiac hypertrophy. After administration of inhibitors including U0126 (ERK1/2 inhibitor), SB203580 (p38 MAPK inhibitor), SP600125 (JNK1/2 inhibitor), CsA (calcineurin inhibitor), FK506 (calcineurin inhibitor), and QNZ (NFkappaB inhibitor), LPS-induced hypertrophic characteristic features, such as increases in cell size, actin fibers, and levels of ANP and BNP, and the nuclear localization of NFAT-3 are markedly inhibited only by calcineurin inhibitors, CsA and FK506. Collectively, these results suggest that LPS leads to myocardiac hypertrophy through calcineurin/NFAT-3 signaling pathway in H9c2 cells. Our findings further provide a link between the LPS-induced inflammatory response and the calcineurin/NFAT-3 signaling pathway that mediates the development of cardiac hypertrophy.

Topics: Actins; Animals; Atrial Natriuretic Factor; Biomarkers; Calcineurin; Cell Nucleus; Cell Size; Fluorescent Antibody Technique; GATA4 Transcription Factor; Gene Expression Regulation; Hypertrophy; Lipopolysaccharides; Models, Biological; Myocytes, Cardiac; Natriuretic Peptide, Brain; NFATC Transcription Factors; Rats; Signal Transduction; Up-Regulation

The role played by IGF-II in signal transduction through the IGF-II/mannose-6-phosphate receptor (IGF2R) in heart tissue has been poorly understood. In our previous studies, we detected an increased expression of IGF-II and IGF2R in cardiomyocytes that had undergone pathological hypertrophy. We hypothesized that after binding with IGF-II, IGF2R may trigger intracellular signaling cascades involved in the progression of pathologically cardiac hypertrophy. In this study, we used immunohistochemical analysis of the human cardiovascular tissue array to detect expression of IGF2R. In our study of H9c2 cardiomyoblast cell cultures, we used the rhodamine phalloidin staining to measure the cell hypertrophy and western blot to measure the expression of cardiac hypertrophy markers atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) in cells treated with IGF-II. We found that a significant association between IGF2R overexpression and myocardial infarction. The treatment of H9c2 cardiomyoblast cells with IGF-II not only induced cell hypertrophy but also increased the protein level of ANP and BNP. Using Leu27IGF-II, an analog of IGF-II which interacts selectively with the IGF2R, to specifically activate IGF2R signaling cascades, we found that binding of Leu27IGF-II to IGF2R led to an increase in the phosphorylation of protein Kinase C (PKC)-alpha and calcium/calmodulin-dependent protein kinase II (CaMKII) in a Galphaq-dependent manner. By the inhibition of PKC-alpha/CaMKII activity, we found that IGF-II and Leu27IGF-II-induced cell hypertrophy and upregulation of ANP and BNP were significantly suppressed. Taken together, this study provides a new insight into the effects of the IGF2R and its downstream signaling in cardiac hypertrophy. The suppression of IGF2R signaling pathways may be a good strategy to prevent the progression of pathological hypertrophy.

Topics: Atrial Natriuretic Factor; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Cells, Cultured; GTP-Binding Protein alpha Subunits, Gq-G11; Humans; Hypertrophy; Immunohistochemistry; Insulin-Like Growth Factor II; Myocardial Infarction; Myocytes, Cardiac; Natriuretic Peptide, Brain; Protein Kinase C-alpha; Receptor, IGF Type 2; Signal Transduction

Cardiomyocyte hypertrophy and extracellular matrix remodeling, primarily mediated by inflammatory cytokine-stimulated cardiac fibroblasts, are critical cellular events in cardiac pathology. The molecular components governing these processes remain nebulous, and few genes have been linked to both hypertrophy and matrix remodeling. Here we show that p8, a small stress-inducible basic helix-loop-helix protein, is required for endothelin- and alpha-adrenergic agonist-induced cardiomyocyte hypertrophy and for tumor necrosis factor-stimulated induction, in cardiac fibroblasts, of matrix metalloproteases (MMPs) 9 and 13-MMPs linked to general inflammation and to adverse ventricular remodeling in heart failure. In a stimulus-dependent manner, p8 associates with chromatin containing c-Jun and with the cardiomyocyte atrial natriuretic factor (anf) promoter and the cardiac fibroblast mmp9 and mmp13 promoters, established activator protein 1 effectors. p8 is also induced strongly in the failing human heart by a process reversed upon therapeutic intervention. Our results identify an unexpectedly broad involvement for p8 in key cellular events linked to cardiomyocyte hypertrophy and cardiac fibroblast MMP production, both of which occur in heart failure.

Topics: Animals; Atrial Natriuretic Factor; Basic Helix-Loop-Helix Transcription Factors; Chromatin; Endothelin-1; Enzyme Induction; Fibroblasts; Gene Expression Regulation, Enzymologic; Heart Failure; HeLa Cells; Humans; Hypertrophy; Matrix Metalloproteinase 13; Matrix Metalloproteinase 9; Myocardium; Myocytes, Cardiac; Neoplasm Proteins; Phenylephrine; Promoter Regions, Genetic; Protein Binding; Proto-Oncogene Proteins c-jun; Rats; RNA, Messenger; Transcription Factor AP-1; Tumor Necrosis Factor-alpha

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.

Topics: Animals; Atrial Natriuretic Factor; Cell Size; Cells, Cultured; Enzyme Activation; Gene Expression; Heart Ventricles; Humans; Hypertrophy; MAP Kinase Signaling System; Myocytes, Cardiac; Phenylephrine; Rats; Rats, Sprague-Dawley; Receptors, Adrenergic, alpha-1; RGS Proteins; RNA, Messenger; Sodium-Hydrogen Exchanger 1; Sodium-Hydrogen Exchangers; Time Factors; Up-Regulation

The present study was performed to evaluate the effects of (2S, 3S, 4R)-N"-cyano-N-(6-amino-3, 4-dihydro-2-dimethoxymethyl-3-hydroxy-2-methyl-2H-1-benzopyran-4yl)-N'-benzylguanidine (KR-31378), a novel mitochondrial ATP-sensitive potassium channel activator, on hypertrophy of H9c2 cells and on cardiac dysfunction in rats with congestive heart failure. In rat heart-derived H9c2 cells treated with hypertrophic agonists, such as angiotensin II, phenylephrine, isoproterenol, and urotensin II, cell size was significantly increased by 27-47%. The increases in cell size induced by the hypertrophic agonists were inhibited by treatment of KR-31378 in a concentration-dependent manner. This was confirmed by the results showing that KR-31378 inhibited the angiotensin II-induced increase in cell protein content. The effect of KR-31378 on the angiotensin II-induced increase in cell size was reversed by mitochondrial ATP-sensitive potassium channel blockers, 5-hydroxydecanoate or glibenclamide. In rats with congestive heart failure, induced by permanent coronary artery occlusion for 8 weeks, KR-31378 significantly reversed the cardiac dysfunction (increase in ratios of stroke volume or cardiac output to body weight) induced by myocardial infarction without reducing infarct size. In addition, KR-31378 significantly inhibited atrial hypertrophy (decrease in ratio of right atrium to body weight) and decreased the serum pro-atrial natriuretic peptide level, a biochemical marker of heart failure. These results suggest that KR-31378 suppresses hypertrophy induced by hypertrophic agonists in H9c2 cells and improves cardiac dysfunction in rats with congestive heart failure induced by myocardial infarction, and that the effects may be mediated by the activation of mitochondrial ATP-sensitive potassium channels.

Topics: Analysis of Variance; Angiotensin II; Animals; Atrial Natriuretic Factor; Blotting, Western; Cell Enlargement; Cell Line; Dose-Response Relationship, Drug; Enzyme Activation; Guanidines; Heart; Heart Failure; Hypertrophy; Mitogen-Activated Protein Kinases; Myoblasts; Myocardial Infarction; Myocardium; Organ Size; Potassium Channels, Inwardly Rectifying; Protein Precursors; Pyrans; Rats; Rats, Sprague-Dawley

Intracellular Ca2+ plays pivotal roles in diverse cellular functions, including gene transcription that underlies cardiac remodeling during stress responses. However, the role of inositol 1,4,5-trisphosphate receptors (IP3Rs) in the mediation of cardiac intracellular Ca2+ and hypertrophic growth remains elusive. Prior work with neonatal rat ventricular myocytes suggests that activation of IP3Rs may be linked to a1 adrenergic receptor (alpha1AR) increased stereotyped Ca2+ spark occurrence and global Ca2+ oscillations. Thus, we hypothesized that Ca2+ release through IP3Rs was necessary for alpha1AR-stimulated cardiac hypertrophy.. We used myoinositol 1,4,5-trisphosphate hexakis (butyryloxymethyl) ester (IP3BM), a membrane-permeant ester of IP3, to activate IP3Rs directly, and Fluo 4/AM to measure intracellular Ca2+ signaling.. IP3BM (10 micromol x L(-1)) mimicked the effects of phenylephrine, a selective agonist of alpha1AR, in increments in local Ca2+ spark release (especially in the perinuclear area) and global Ca2+ transient frequencies. More importantly, IP3R inhibitors, 2-aminoethoxydiphenyl borate and Xestospongin C, abolished the IP3BM-induced Ca2+ responses, and significantly suppressed alpha1AR-induced cardiomyocyte hypertrophy assayed by cell size, [3H] leucine incorporation and atrial natriuretic factor gene expression, during sustained (48 h) phenylephrine stimulation.. These results, therefore, provide cellular mechanisms that link IP3R signaling to alpha1AR-stimulated gene expression and cardiomyocyte hypertrophy.

Topics: Adrenergic alpha-1 Receptor Agonists; Animals; Animals, Newborn; Atrial Natriuretic Factor; Boron Compounds; Calcium; Calcium Signaling; Cells, Cultured; Heart Ventricles; Hypertrophy; Inositol 1,4,5-Trisphosphate Receptors; Leucine; Macrocyclic Compounds; Myocytes, Cardiac; Oxazoles; Phenylephrine; Rats; Rats, Sprague-Dawley; RNA, Messenger; Signal Transduction

1. Atrial natriuretic peptide (ANP)-null mice (Nppa(-/-)) exhibit cardiac hypertrophy at baseline and adverse cardiac remodelling in response to transverse aortic constriction (TAC)-induced pressure overload stress. Previous studies have suggested that natriuretic peptides could potentially oppose mineralocorticoid signalling at several levels, including suppression of adrenal aldosterone production, inhibition of mineralocorticoid receptor (MR) activation or suppression of MR-mediated production of pro-inflammatory factors. Thus, we hypothesized that the MR blocker eplerenone would prevent the exaggerated left ventricular (LV) remodelling/fibrosis and dysfunction after TAC in Nppa(-/-). 2. In the present study, Nppa(-/-) and wild-type Nppa(+/+) mice fed eplerenone- or vehicle (oatmeal)-supplemented chow since weaning were subjected to TAC or sham operation. The daily dose of eplerenone administered was approximately 200 mg/kg. At 1 week after TAC, LV size and function were evaluated by echocardiogram and LV cross-sections were stained with picrosirius red for collagen volume measurement. Total RNA was extracted from the LV for real-time polymerase chain reaction analysis of osteopontin. 3. Eplerenone had no effect on baseline hypertrophy observed in sham-operated Nppa(-/-) compared with Nppa(+/+) mice. Eplerenone attenuated the TAC-induced increase in LV weight in both genotypes and completely prevented LV dilation, systolic dysfunction and interstitial collagen deposition seen in Nppa(-/-) mice after TAC. However, serum aldosterone levels were lower in Nppa(-/-) compared with Nppa(+/+) wild types. No interaction between eplerenone and genotype in osteopontin mRNA levels was observed. 4. Eplerenone prevents adverse cardiac remodelling related to pressure overload in ANP-deficient mice, mainly due to an antifibrotic effect. The mechanism whereby ANP deficiency leads to excess hypertrophy, fibrosis and early failure following TAC is increased profibrotic signals resulting from excess or unopposed MR activation, rather than increased levels of aldosterone.

Topics: Aldosterone; Animals; Atrial Natriuretic Factor; Blood Pressure; Cardiomegaly; Eplerenone; Heart; Hypertrophy; Male; Mice; Mice, Knockout; Myocardium; Spironolactone; Ventricular Remodeling

To examine the inhibitory effect of FAK-related nonkinase (FRNK) in cardiac hypertrophy in vitro and investigate the possible mechanisms.. A functional fragment of FRNK cDNA was amplified by reverse transcription-polymerase chain reaction and cloned into the vector pcDNA3.1. Hypertrophy in neonatal rat cardiac myocytes was established with angiotensin-II stimulation. The pcDNA3.1-FRNK or pcDNA3.1 was respectively transfected into cardiomyocytes by Lipofectamine 2000. The surface area and mRNA expression of atrial natriuretic peptide (ANP) of myocytes were employed to detect cardiac hypertrophy. NF-kappaB p65 protein in nuclear extracts, phosphorylation levels of ERK1/2 (p-ERK1/2) and AKT (p-AKT), as well as total ERK1/2, and AKT in variant treated cardiomyocytes were determined by Western blot.. Under the stimulation of angiotensin II, the surface area of myocytes and levels of ANP mRNA were significantly increased. But transient transfection with pcDNA3.1-FRNK in advance may reduce the surface area and expression of ANP mRNA of hypertrophic myocytes. The protein levels of NF-kappaB p65 in nuclear extracts and p-ERK1/2, p-AKT in FRNK treated cardiomyocytes were significantly decreased compared with that in angiotensin-II induced cardiomyocytes, while different treatments had little effect on total ERK1/2 and AKT.. FRNK may inhibit angiotensin-II-induced cardiomyocyte hypertrophy via decreasing phosphorylation levels at ERK1/2 and AKT, consequently downregulating nuclear translocation of NF-kappaB p65.

Topics: Angiotensin II; Animals; Animals, Newborn; Atrial Natriuretic Factor; Cells, Cultured; DNA, Complementary; Hypertrophy; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Myocytes, Cardiac; Protein-Tyrosine Kinases; Proto-Oncogene Proteins c-akt; Rats; Rats, Sprague-Dawley; RNA, Messenger; Transcription Factor RelA; Transfection

To investigate the role of endothelin-1 and its receptors on hypertrophy or proliferation of cultured cardial cells.. Cardiomyocytes and cardiac fibroblasts were isolated by trypsin digestion method, DNA and protein synthesis were measured by 3H-dexyribonucleotidethymine (3H-TdR) and 3H-Leucine (3H-Leu) incorporation, while protein content was measured by Bradford method. Atrial natriuretic peptide (ANP) mRNA expression of cardiomyocyte was measured by reverse transcripted-polymerase chain reaction. Selective endothelin (ET) receptor subtype antagonists BQ123 and BQ788 were used to block ET(A) receptors (ET(A)R) and ET(B)R respectively and to observe the effects of the two receptors during cardiac hypertrophy.. ET-1 significantly increased the 3H-TdR and 3H-Leu incorporation rate of cardiomyocytes and cardiac fibroblasts in a dose-dependent manner and increased protein content. Furthermore, ET-1 promoted the ANP mRNA expression of cardiomyocyte. ET(A)R antagonist remarkably blocked these effects, while ET(B)R antagonist had no obvious effect.. ET-1 can induce the hypertrophy for cardiomyocytes and the proliferation for cardiac fibroblasts. These effects are mediated by ET(A)R.

Topics: Animals; Animals, Newborn; Atrial Natriuretic Factor; Cell Proliferation; Cells, Cultured; Endothelin-1; Fibroblasts; Hypertrophy; Myocytes, Cardiac; Rats; Rats, Sprague-Dawley; Receptor, Endothelin A; RNA, Messenger

To evaluate the effects of Xinlikang (XLK) on angiotensin II (Ang II) induced hypertrophic cultured neonatal rat's cardiomyocyte (CMC).. Primary cultured neonatal rat's CMCs with the purity certified by immunohistochemical technique, were divided into three groups. Rats in the normal control group were untreated; those in the model group were established into hypertrophic models but underwent no treatment; and those in the XLK group were established to hypertrophic models and treated with XLK containing serum obtained from rats with aorta coarctation after 8 days of feeding with XLK. MTT and phase-contrast microscope were used to evaluate the effect of XLK on cell activity, pulsating rhythm and surface area; Atrial natriuretic peptide (ANP) expression was determined by radioimmunoassay; Protein content was determined by Bradford method; and DNA synthesis was detected by flow cytometric assay.. Immunohistochemistry results showed that more than 90% of the cells were alpha-sarcometin actin stained positive cells. No significant effect of XLK on normal CMC was found. Ang II could significantly induce hypertrophy in CMCs, and XLK could significantly decrease the increased surface area and the accelerated pulsating rate in them. ANP expression was 780 +/- 38 microg/L in the model group, and 430 +/- 23 microg/L in the control group, and the elevated expression of ANP in model rats was significantly decreased in the XLK group; The DNA content in the G0/G1 and G2/M phases was significantly enhanced and at the same time it was accompanied with increase of total protein content in the model rats after being stimulated by Ang II for 24 h, showing that serum-containing XLK could also significantly suppress total protein synthesis (P < 0.05).. XLK could improve Ang II mediated pathological growth of CMCs without influencing the growth of normal CMCs, suggesting that XLK is probably an effective drug for treatment of myocardial hypertrophy and heart failure.

Topics: Angiotensin II; Animals; Animals, Newborn; Atrial Natriuretic Factor; Cell Size; Cells, Cultured; DNA; Drugs, Chinese Herbal; Hypertrophy; Myocardial Contraction; Myocytes, Cardiac; Proteins; Rats; Rats, Wistar; Vasoconstrictor Agents

In cardiac hypertrophy, both excessive enlargement of cardiac myocytes (CMs) and progressive fibrosis are known to occur simultaneously. To investigate the nature of interactions between ventricular CMs and cardiac fibroblasts (CFs) in these conditions, we have established a "dedifferentiated model" of adult murine CMs in coculture with CFs. In such a model, which is recognized to study cardiac cell hypertrophy in vitro, dedifferentiated CMs in culture and in coculture were characterized by immunopositive staining to ANP (atrial natriuretic peptide) and beta-myosin heavy chain (beta-MHC). The results confirm that ANP secretion by CMs was significantly increased during the cultures. The increase size of cultured CMs was significantly higher in CM/CF cocultures than in CM cultures which was also observed when CMs were cultured with fibroblast conditioned medium (FCM). In addition, fibroblast proliferation studies showed that CMs favored fibroblast adhesion and/or growth at the beginning of the coculture and fibroblast proliferation throughout the time course of the coculture. Furthermore, a significant level of interleukin-6 (IL-6) production was detected by ELISA in CM/CF cocultures. A similar higher increase was observed when CMs were cultured in the presence of FCM. These results demonstrate that CFs enhance myocyte hypertrophy and that CMs regulate fibroblast adhesion and/or proliferation, suggesting a paracrine interaction between CMs and CFs which could involve IL-6.

Topics: Animals; Atrial Natriuretic Factor; Biomarkers; Cardiomegaly; Cell Proliferation; Cells, Cultured; Coculture Techniques; Cytokines; Fibroblasts; Hypertrophy; Mice; Myocardium; Myocytes, Cardiac; Myosin Heavy Chains; Ventricular Myosins

In patients with congestive heart failure, high serum levels of the proinflammatory cytokine interleukin (IL)-18 were reported. A positive correlation was described between serum IL-18 levels and the disease severity. IL-18 has also been shown to induce atrial natriuretic factor (ANF) gene expression in adult cardiomyocytes. Because re-expression of the fetal gene ANF is mostly associated with hypertrophy, a hallmark of heart failure, we hypothesized that IL-18 induces cardiomyocyte hypertrophy. Treatment of the cardiomyocyte cell line HL-1 with IL-18 induced hypertrophy as characterized by increases in protein synthesis, phosphorylated p70 S6 kinase, and ribosomal S6 protein levels as well as cell surface area. Furthermore, IL-18 induced ANF gene transcription in a time-dependent manner as evidenced by increased ANF secretion and ANF promoter-driven reporter gene activity. Investigation into possible signal transduction pathways mediating IL-18 effects revealed that IL-18 activates phosphoinositide 3-kinase (PI3K), an effect that was blocked by wortmannin and LY-294002. IL-18 induced Akt phosphorylation and stimulated its activity, effects that were abolished by Akt inhibitor or knockdown. IL-18 stimulated GATA4 DNA binding activity and increased transcription of a reporter gene driven by multimerized GATA4-binding DNA elements. Pharmacological inhibition or knockdown studies revealed that IL-18 induced cardiomyocyte hypertrophy and ANF gene transcription via PI3K, PDK1, Akt, and GATA4. Most importantly, IL-18 induced ANF gene transcription and hypertrophy of neonatal rat ventricular myocytes via PI3K-, Akt-, and GATA4-dependent signaling. Together these data provide the first evidence that IL-18 induces cardiomyocyte hypertrophy via PI3K-dependent signaling, defines a mechanism of IL-18-mediated ANF gene transcription, and further supports a role for IL-18 in inflammatory heart diseases including heart failure.

Topics: Androstadienes; Animals; Atrial Natriuretic Factor; Blotting, Northern; Blotting, Western; Cell Line; Cell Nucleus; Cells, Cultured; Chromones; Cytokines; DNA; DNA-Binding Proteins; Enzyme Activation; Enzyme Inhibitors; GATA4 Transcription Factor; Gene Expression Regulation; Genes, Reporter; Hypertrophy; Inflammation; Interleukin-18; Interleukin-18 Receptor alpha Subunit; Mice; Morpholines; Myocytes, Cardiac; Phenotype; Phosphatidylinositol 3-Kinases; Phosphorylation; Promoter Regions, Genetic; Protein Binding; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-akt; Rats; Receptors, Interleukin; Receptors, Interleukin-18; Ribosomal Protein S6 Kinases, 70-kDa; Signal Transduction; Time Factors; Transcription Factors; Transcription, Genetic; Transfection; Wortmannin

Cardiac hypertrophy is a complex and nonhomogenous response to various stimuli. In this study, we used high-density oligonucleotide microarray to examine gene expression profiles during physiological hypertrophy, pathological hypertrophy, and heart failure in Dahl salt-sensitive rats. There were changes in 404/3,160 and 874/3,160 genes between physiological and pathological hypertrophy and the transition from hypertrophy to heart failure, respectively. There were increases in stress response genes (e.g., heat shock proteins) and inflammation-related genes (e.g., pancreatitis-associated protein and arachidonate 12-lipoxygenase) in pathological processes but not in physiological hypertrophy. Furthermore, atrial natriuretic factor and brain natriuretic protein showed distinctive changes that are very specific to different conditions. In addition, we used a resampling-based gene score-calculating method to define significantly altered gene clusters, based on Gene Ontology classification. It revealed significant alterations in genes involved in the apoptosis pathway during pathological hypertrophy, suggesting that the apoptosis pathway may play a role during the transition to heart failure. In addition, there were significant changes in glucose/insulin signaling, protein biosynthesis, and epidermal growth factor signaling during physiological hypertrophy but not during pathological hypertrophy.

Topics: Animals; Apoptosis; Atrial Natriuretic Factor; Blotting, Northern; Cardiomegaly; Echocardiography; Epidermal Growth Factor; Gene Expression Profiling; Gene Expression Regulation; Heart Failure; Hypertrophy; Inflammation; Insulin; Natriuretic Peptide, Brain; Oligonucleotide Array Sequence Analysis; Pancreatitis-Associated Proteins; Physical Conditioning, Animal; Rats; Rats, Inbred Dahl; RNA; Signal Transduction

Atrial dilatation is an important risk factor for atrial fibrillation (AF). In the present study, we monitored the electrophysiological changes during progressive atrial dilatation in chronically instrumented goats.. In 8 goats, 2 screw-in leads with piezoelectric crystals were implanted transvenously in the right atrium. After 2 weeks, atrial diameter and effective refractory period were measured. AF paroxysms were induced by burst pacing to determine the baseline AF cycle length and stability of AF. After His-bundle ablation, the above measurements were repeated once a week. After 4 weeks of complete AV block, the free wall of the right atrium was mapped and the atrium was fixed in formalin for histological analysis. After His-bundle ablation, the ventricular rate decreased from 113.8+/-4.8 to 44.6+/-2.5 bpm. Right atrial diameter increased gradually by 13.5+/-3.9% during 4 weeks of AV block (P<0.01). The duration of induced AF paroxysms increased from 4.6 seconds to 6.4 minutes (P<0.05). Atrial effective refractory period and AF cycle length remained constant. Spontaneous paroxysms of AF were not observed. Atrial mapping during rapid pacing revealed that slow conduction (<30 cm/s) was present in 3.7+/-1.0% of the mapped area (control, 0.9+/-0.5%, P<0.05). Histological analysis showed hypertrophy without atrial fibrosis. Connexin40 and connexin43 expression was unchanged.. Chronic AV block in the goat leads to progressive atrial dilatation, prolongation of induced AF paroxysms, and local conduction delays. The increase in AF stability was not a result of a shortening of atrial refractoriness or atrial fibrosis.

Topics: Aldosterone; Angiotensin II; Animals; Atrial Fibrillation; Atrial Natriuretic Factor; Cardiac Pacing, Artificial; Cardiomegaly; Cell Size; Chronic Disease; Connexin 43; Connexins; Female; Gap Junction alpha-5 Protein; Goats; Heart Block; Heart Conduction System; Hemodynamics; Hypertrophy; Myocytes, Cardiac; Neural Conduction; Norepinephrine; Refractory Period, Electrophysiological

Diacylglycerol (DAG) is a lipid second messenger that transiently accumulates in cells stimulated by endothelin-1 (ET-1) and other Galphaq protein-coupled receptor agonists. Diacylglycerol kinase (DGK) is thought to be an enzyme that controls the cellular levels of DAG by converting it to phosphatidic acid; however, the functional role of DGK has not been examined in cardiomyocytes. Because DGK inactivates DAG, a strong activator of protein kinase C (PKC), we hypothesized that DGK inhibited ET-1-induced activation of a DAG-PKC signaling cascade and subsequent cardiomyocyte hypertrophy.. Real-time reverse transcription-polymerase chain reaction demonstrated a significant increase of DGK-zeta mRNA by ET-1 in cardiomyocytes. To determine the functional role of DGK-zeta, we overexpressed DGK-zeta in cardiomyocytes using a recombinant adenovirus encoding rat DGK-zeta (Ad-DGKzeta). ET-1-induced translocation of PKC-epsilon was blocked by Ad-DGKzeta (P<0.01). Ad-DGKzeta also inhibited ET-1-induced activation of extracellular signal-regulated kinase (P<0.01). Luciferase reporter assay revealed that ET-1-mediated increase of activator protein-1 (AP1) DNA-binding activity was significantly inhibited by DGK-zeta (P<0.01). In cardiomyocytes transfected with DGK-zeta, ET-1 failed to cause gene induction of atrial natriuretic factor, increases in [3H]-leucine uptake, and increases in cardiomyocyte surface area.. We demonstrated for the first time that DGK-zeta blocked ET-1-induced activation of the PKC-epsilon-ERK-AP1 signaling pathway, atrial natriuretic factor gene induction, and resultant cardiomyocyte hypertrophy. DGK-zeta might act as a negative regulator of hypertrophic program in response to ET-1, possibly by controlling cellular DAG levels.

Topics: Adenoviridae; Animals; Atrial Natriuretic Factor; Cell Enlargement; Cells, Cultured; Diacylglycerol Kinase; Diglycerides; Endothelin-1; Gene Expression Regulation; Hypertrophy; Myocytes, Cardiac; Protein Kinase C; Protein Kinase C-epsilon; Rats; Rats, Sprague-Dawley; RNA, Messenger; Second Messenger Systems; Transcriptional Activation; Transduction, Genetic

The crucial functions of atrial natriuretic peptide (ANP) and endothelial nitric oxide/NO in the regulation of arterial blood pressure have been emphasized by the hypertensive phenotype of mice with systemic inactivation of either the guanylyl cyclase-A receptor for ANP (GC-A-/-) or endothelial nitric-oxide synthase (eNOS-/-). Intriguingly, similar levels of arterial hypertension are accompanied by marked cardiac hypertrophy in GC-A-/-, but not in eNOS-/-, mice, suggesting that changes in local pathways regulating cardiac growth accelerate cardiac hypertrophy in the former and protect the heart of the latter. Our recent observations in mice with conditional, cardiomyocyte-restricted GC-A deletion demonstrated that ANP locally inhibits cardiomyocyte growth. Abolition of these local, protective effects may enhance the cardiac hypertrophic response of GC-A-/- mice to persistent increases in hemodynamic load. Notably, eNOS-/- mice exhibit markedly increased cardiac ANP levels, suggesting that increased activation of cardiac GC-A can prevent hypertensive heart disease. To test this hypothesis, we generated mice with systemic inactivation of eNOS and cardiomyocyte-restricted deletion of GC-A by crossing eNOS-/- and cardiomyocyte-restricted GC-A-deficient mice. Cardiac deletion of GC-A did not affect arterial hypertension but significantly exacerbated cardiac hypertrophy and fibrosis in eNOS-/- mice. This was accompanied by marked cardiac activation of both the mitogen-activated protein kinase (MAPK) ERK 1/2 and the phosphatase calcineurin. Our observations suggest that local ANP/GC-A/cyclic GMP signaling counter-regulates MAPK/ERK- and calcineurin/nuclear factor of activated T cells-dependent pathways of cardiac myocyte growth in hypertensive eNOS-/- mice.

Topics: Animals; Atrial Natriuretic Factor; Blood Pressure; Blotting, Northern; Blotting, Western; Cyclic GMP; Gene Deletion; Genotype; Heart Ventricles; Hypertension; Hypertrophy; Mice; Mice, Knockout; Mice, Transgenic; Myocardium; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Nitric Oxide Synthase Type III; Phenotype; Phosphorylation; RNA, Messenger; Signal Transduction

alpha1-Adrenergic receptors mediate several biological effects of catecholamines, including the regulation of myocyte growth and contractility and transcriptional regulation of the atrial natriuretic factor (ANF) gene whose promoter contains an alpha1-adrenergic response element. The nuclear pathways and effectors that link receptor activation to genetic changes remain poorly understood. Here, we describe the isolation by the yeast one-hybrid system of a cardiac cDNA encoding a novel nuclear zinc finger protein, Zfp260, belonging to the Krüppel family of transcriptional regulators. Zfp260 is highly expressed in the embryonic heart but is downregulated during postnatal development. Functional studies indicate that Zfp260 is a transcriptional activator of ANF and a cofactor for GATA-4, a key cardiac regulator. Knockdown of Zfp260 in cardiac cells decreases endogenous ANF gene expression and abrogates its response to alpha1-adrenergic stimulation. Interestingly, Zfp260 transcripts are induced by alpha1-adrenergic agonists and are elevated in genetic models of hypertension and cardiac hypertrophy. The data identify Zfp260 as a novel transcriptional regulator in normal and pathological heart development and a nuclear effector of alpha1-adrenergic signaling.

Topics: Adenoviridae; Amino Acid Sequence; Animals; Atrial Natriuretic Factor; Base Sequence; Blotting, Western; Cell Nucleus; Cell Proliferation; Cloning, Molecular; DNA, Complementary; Down-Regulation; Fluorescent Antibody Technique, Indirect; Gene Expression Regulation; Gene Expression Regulation, Developmental; Gene Library; Genes, Reporter; Heart; HeLa Cells; Humans; Hypertension; Hypertrophy; Immunohistochemistry; Lac Operon; Molecular Sequence Data; Myocardium; Myocytes, Cardiac; Oligonucleotides, Antisense; Plasmids; Promoter Regions, Genetic; Protein Structure, Tertiary; Rats; Rats, Inbred WKY; Rats, Sprague-Dawley; Receptors, Adrenergic, alpha-1; Recombinant Proteins; RNA; RNA, Messenger; Sequence Homology, Amino Acid; Signal Transduction; Time Factors; Trans-Activators; Transcription, Genetic; Transcriptional Activation; Zinc Fingers

Outside-in signaling from fibronectin (FN) through integrin receptors has been shown to play an important role in promoting cardiac myocyte hypertrophy and synergizes with other hypertrophic stimuli such as the alpha-adrenergic agonist phenylephrine (PE) and mechanical strain. The integrin-linked kinase (ILK) is a critical molecule involved in cell adhesion, motility and survival in nonmyocytes such as fibroblasts and epithelial cells. Its role in cardiac myocytes is unclear. In this study, we demonstrate that (1) ILK forms a complex with PINCH1 and alpha-parvin proteins (IPAP1 complex) in neonatal rat ventricular myocytes; (2) localization of IPAP1 complex proteins to costameres in cardiac myocytes is stimulated by FN, PE and synergistically by the combination of FN and PE in an integrin beta1-dependent manner; (3) a dominant-negative mutant lacking the PINCH-binding N-terminus of ILK (ILK-C) prevents costamere association of ILK and alpha-parvin, but not PINCH1; (4) FN- and PE-induced hypertrophy, measured by increased protein/DNA ratio, beating frequency and atrial natriuretic peptide expression, is stimulated by low levels of ILK-C but repressed by high ILK-C expression; and (5) overexpression of ILK-C, as well as deletion of the ILK gene in mouse neonatal ventricular myocytes, induces marked apoptosis of cardiac myocytes. These results suggest that the IPAP1 complex plays an important role in mediating integrin-signaling pathways that regulate cardiac myocyte hypertrophy and resistance to apoptosis.

Topics: Actinin; Adaptor Proteins, Signal Transducing; Adenoviridae; Adrenergic alpha-Agonists; Animals; Animals, Newborn; Atrial Natriuretic Factor; Cell Culture Techniques; Cells, Cultured; DNA-Binding Proteins; Drug Synergism; Fibronectins; Heart Ventricles; Hypertrophy; LIM Domain Proteins; Membrane Proteins; Mice; Myocardium; Myocytes, Cardiac; Phenylephrine; Protein Serine-Threonine Kinases; Proteins; Rats; Rats, Sprague-Dawley; RNA, Messenger

This study investigates the effect of hypertrophy, using one kidney and one kidney/one clip rats, and development, comparing 3- and 12-week-old rats, on the expression of the 28-amino acid atrial natriuretic peptide (ANP(1-28)) binding sites in rat kidney. Here we report an increased B(max) value of glomerular binding sites for ANP(1-28) and C-type natriuretic peptide 1-22 (CNP(1-22)) in hypertrophied and developing kidney, without modifying their affinity, an effect that was prevented in the presence of the synthetic des[Gln(18), Ser(19), Gly(20), Leu(21), Gly(22)]ANP(4-23)-amide (C-ANF), suggesting that natriuretic peptide receptor (NPR)-C binding sites might be enhanced. The enhanced B(max) was only detected in the high affinity binding site for CNP(1-22), which has been identified as the 67 kDa NPR-C-like protein. A similar effect was observed in renal glomeruli from 3-week-old rats compared with 12-week-old rats. Our results indicate that ANP(1-28), CNP(1-22) and C-ANF inhibited cAMP synthesis stimulated by the physiological agonists histamine and 5-hydroxytryptamine or directly by forskolin. The inhibitory effect was found to be significantly greater in 1-kidney and 1-kidney/1-clip rats than in controls, and in 3-week-old rats compared with 12-week-old rats. Our observations suggest that this effect must be attributed to the 67 kDa NPR-C-like protein due to the enhanced B(max) values and the reported inhibitory role for this receptor on adenylyl cyclase activity. The enhanced inhibitory role of natriuretic peptides on cAMP synthesis in hypertrophied and developing kidney may influence glomerular function in the rat kidney and suggests a role for the 67 kDa NPR-C-like protein in growth.

Topics: Animals; Atrial Natriuretic Factor; Binding Sites; Binding, Competitive; Colforsin; Cyclic AMP; Histamine; Hypertrophy; Kidney; Kidney Glomerulus; Kinetics; Male; Natriuretic Peptide, C-Type; Nephrectomy; Peptide Fragments; Rats; Rats, Inbred WKY; Receptors, Atrial Natriuretic Factor; Serotonin

Glycogen synthase kinase 3beta (GSK-3beta) negatively regulates cardiac hypertrophy. A potential target mediating the antihypertrophic effect of GSK-3beta is eukaryotic translation initiation factor 2Bepsilon (eIF2Bepsilon). Overexpression of GSK-3beta increased the cellular kinase activity toward GST-eIF2Bepsilon in neonatal rat cardiac myocytes, whereas LiCl (10 mmol/L) or isoproterenol (ISO) (10 micromol/L), a treatment known to inhibit GSK-3beta, decreased it. Immunoblot analyses using anti-S535 phosphospecific eIF2Bepsilon antibody showed that S535 phosphorylation of endogenous eIF2Bepsilon was decreased by LiCl or ISO, suggesting that GSK-3beta is the predominant kinase regulating phosphorylation of eIF2Bepsilon-S535 in cardiac myocytes. Decreases in eIF2Bepsilon-S535 phosphorylation were also observed in a rat model of cardiac hypertrophy in vivo. Overexpression of wild-type eIF2Bepsilon alone moderately increased cell size (+31+/-11%; P<0.05 versus control), whereas treatment of eIF2Bepsilon-transduced myocytes with LiCl (+73+/-22% versus eIF2Bepsilon only; P<0.05) or ISO (+84+/-33% versus eIF2Bepsilon only; P<0.05) enhanced the effect of eIF2Bepsilon. Overexpression of eIF2Bepsilon-S535A, which is not phosphorylated by GSK-3beta, increased cell size (+107+/-35%) as strongly as ISO (+95+/-25%), and abolished antihypertrophic effects of GSK-3beta, indicating that S535 phosphorylation of eIF2Bepsilon critically mediates antihypertrophic effects of GSK-3beta. Furthermore, expression of eIF2Bepsilon-F259L, a dominant-negative mutant, inhibited ISO-induced hypertrophy, indicating that eIF2Bepsilon is required for beta-adrenergic hypertrophy. Interestingly, expression of eIF2Bepsilon-S535A partially increased cytoskeletal reorganization, whereas it did not increase expression of atrial natriuretic factor gene. These results suggest that GSK-3beta is the predominant kinase mediating phosphorylation of eIF2Bepsilon-S535 in cardiac myocytes, which in turn plays an important role in regulating cardiac hypertrophy primarily through protein synthesis.

Topics: Adenoviridae; Adrenergic beta-Agonists; Adrenergic beta-Antagonists; Animals; Animals, Newborn; Atrial Natriuretic Factor; Cardiomegaly; Cells, Cultured; Eukaryotic Initiation Factor-2B; Gene Expression Regulation; Genetic Vectors; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; Heart Ventricles; Hypertrophy; Isoproterenol; Lithium Chloride; Male; Myocardial Infarction; Myocytes, Cardiac; Myosin Heavy Chains; Peptide Chain Initiation, Translational; Phosphoserine; Propranolol; Protein Processing, Post-Translational; Rats; Rats, Wistar; Recombinant Fusion Proteins; Transduction, Genetic; Ventricular Remodeling

In vivo, left ventricular remodeling after myocardial infarction involves hypertrophy generally attributed to increased cardiac workload. We hypothesized that hypoxia/reoxygenation directly induces cardiomyocyte hypertrophy and studied several participating kinases and transcription factors in isolated cardiomyocytes. Hypoxia for 6 h followed by 42 h reoxygenation induced cardiomyocyte hypertrophy assessed by 3H leucine incorporation and immunohistochemistry. Inhibition of reactive oxygen species (ROS), serine/threonine kinase AKT, and ERK abolished reoxygenation-induced hypertrophy. In addition, a beta2-adrenergic receptor (beta2-AR) antagonist, as well as Gi inhibitor pertussis toxin, blocked reoxygenation-induced hypertrophy. Hypoxia for 6 h increased transcription factors CREB, NF-kappaB, and GATA DNA binding activities. However, only CREB DNA-binding was sustained during reoxygenation. Inhibition of PI3-kinase, ERK, and PKA abrogated reoxygenation-induced CREB DNA-binding without affecting CREB serine-133 phosphorylation. These same pathways were found to regulate hypoxia/reoxygenation-induced GSK3beta kinase activity and CREB serine-129 de-phosphorylation. GSK3beta mutants resistant to phosphorylation blocked the stimulation of CRE-dependent transcription induced by hypoxia/reoxygenation. Transfection of cardiomyocytes with a dominant-negative mutant of CREB abrogated hypoxia/reoxygenation-induced hypertrophy. We suggest that hypoxia/reoxygenation induces cardiomyocyte hypertrophy through CREB activation. Inactivation of GSK3beta by hypoxia/reoxygenation, possibly integrating PI3-kinase and ERK pathways downstream of beta2-AR and ROS, is a prerequisite for CRE-dependent transcription. Transient hypoxia may contribute to cardiac hypertrophy in ischemic heart disease independent of cardiac workload.

Topics: Adrenergic beta-Antagonists; Animals; Atrial Natriuretic Factor; Cell Hypoxia; Cyclic AMP Response Element-Binding Protein; DNA-Binding Proteins; Enzyme Inhibitors; GATA4 Transcription Factor; Genes, Reporter; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; Hypertrophy; Hypoxia-Inducible Factor 1, alpha Subunit; I-kappa B Proteins; MAP Kinase Signaling System; Myocytes, Cardiac; NF-kappa B; NF-KappaB Inhibitor alpha; Oxygen; PC12 Cells; Pertussis Toxin; Phosphatidylinositol 3-Kinases; Phosphorylation; Protein Kinases; Protein Processing, Post-Translational; Rats; Rats, Wistar; Reactive Oxygen Species; Receptors, Adrenergic, beta; Recombinant Fusion Proteins; Signal Transduction; Transcription Factors; Transfection

Transforming growth factor-beta (TGF-beta) has been associated with the onset of cardiac cell hypertrophy, but the mechanisms underlying this dissociation are not completely understood. By a previous study, we investigated the involvement of a MAP3K, ZAK, which in cultured H9c2 cardiac cells is a positive mediator of cell hypertrophy. Our results showed that expression of a dominant-negative form of ZAK inhibited the characteristic TGF-beta-induced features of cardiac hypertrophy, including increased cell size, elevated expression of atrial natriuretic factor (ANF), and increased organization of actin fibers. Furthermore, dominant-negative MKK7 effectively blocked both TGF-beta-and ZAK-induced ANF expression. In contrast, a JNK/SAPK specific inhibitor, sp600125, had little effect on TGF-beta- or ZAK-induced ANF expression. Our findings suggest that a ZAK mediates TGF-beta-induced cardiac hypertrophic growth via a novel TGF-beta signaling pathway that can be summarized as TGF-beta>ZAK>MKK7>ANF.

Topics: Actins; Angiotensin II; Animals; Anthracenes; Atrial Natriuretic Factor; Cell Size; Cells, Cultured; Humans; Hypertrophy; Interleukins; MAP Kinase Kinase 7; MAP Kinase Kinase Kinases; Myocytes, Cardiac; Phenylephrine; Protein Kinase Inhibitors; Protein Kinases; Rats; Recombinant Fusion Proteins; Signal Transduction; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha; Vasoconstrictor Agents

Yeast silent information regulator 2 (Sir2), a nicotinamide adenine dinucleotide-dependent histone deacetylase (HDAC) and founding member of the HDAC class III family, functions in a wide array of cellular processes, including gene silencing, longevity, and DNA damage repair. We examined whether or not the mammalian ortholog Sir2 affects growth and death of cardiac myocytes. Cardiac myocytes express Sir2alpha predominantly in the nucleus. Neonatal rat cardiac myocytes were treated with 20 mmol/L nicotinamide (NAM), a Sir2 inhibitor, or 50 nmol/L Trichostatin A (TSA), a class I and II HDAC inhibitor. NAM induced a significant increase in nuclear fragmentation (2.2-fold) and cleaved caspase-3, as did sirtinol, a specific Sir2 inhibitor, and expression of dominant-negative Sir2alpha. TSA also modestly increased cell death (1.5-fold) but without accompanying caspase-3 activation. Although TSA induced a 1.5-fold increase in cardiac myocyte size and protein content, NAM reduced both. In addition, NAM caused acetylation and increases in the transcriptional activity of p53, whereas TSA did not. NAM-induced cardiac myocyte apoptosis was inhibited in the presence of dominant-negative p53, suggesting that Sir2alpha inhibition causes apoptosis through p53. Overexpression of Sir2alpha protected cardiac myocytes from apoptosis in response to serum starvation and significantly increased the size of cardiac myocytes. Furthermore, Sir2 expression was increased significantly in hearts from dogs with heart failure induced by rapid pacing superimposed on stable, severe hypertrophy. These results suggest that endogenous Sir2alpha plays an essential role in mediating cell survival, whereas Sir2alpha overexpression protects myocytes from apoptosis and causes modest hypertrophy. In contrast, inhibition of endogenous class I and II HDACs primarily causes cardiac myocyte hypertrophy and also induces modest cell death. An increase in Sir2 expression during heart failure suggests that Sir2 may play a cardioprotective role in pathologic hearts in vivo.

Topics: Acetylation; Alkaloids; Animals; Apoptosis; Atrial Natriuretic Factor; Benzamides; Benzophenanthridines; Cell Nucleus; Cell Size; Cell Survival; Cells, Cultured; Culture Media, Serum-Free; Cysteine Proteinase Inhibitors; Dogs; Gene Silencing; Genes, Dominant; Genes, p53; Heart Failure; Heart Ventricles; Hydroxamic Acids; Hypertrophy; Hypertrophy, Left Ventricular; Longevity; Mice; Myocytes, Cardiac; Naphthols; Niacinamide; Phenanthridines; Protein Processing, Post-Translational; Rats; Rats, Wistar; Recombinant Fusion Proteins; Sirtuin 1; Sirtuins; Transcription, Genetic; Tumor Suppressor Protein p53

Because cell shape and alignment, cell-matrix adhesion, and cell-cell contact can all affect growth, and because mechanical strains in vivo are multiaxial and anisotropic, we developed an in vitro system for engineering aligned, rod-shaped, neonatal cardiac myocyte cultures. Photolithographic and microfluidic techniques were used to micropattern extracellular matrices in parallel lines on deformable silicone elastomers. Confluent, elongated, aligned myocytes were produced by varying the micropattern line width and collagen density. An elliptical cell stretcher applied 2:1 anisotropic strain statically to the elastic substrate, with the axis of greatest stretch (10%) either parallel or transverse to the myofibrils. After 24 h, the principal strain parallel to myocytes did not significantly alter myofibril accumulation or expression of atrial natriuretic factor (ANF), connexin-43 (Cx-43), or N-cadherin (by indirect immunofluorescent antibody labeling and immunoblotting) compared with unstretched controls. In contrast, 10% transverse principal strain resulted in continuous staining of actin filaments (rhodamine phalloidin); increased immunofluorescent labeling of ANF, Cx-43, and N-cadherin; and upregulation of protein signal intensity by western blotting. By using microfabrication and microfluidics to control cell shape and alignment on an elastic substrate, we found greater effects for transverse than for longitudinal stretch in regulating sarcomere organization, hypertrophy, and cell-to-cell junctions.

Topics: Actins; Animals; Animals, Newborn; Anisotropy; Atrial Natriuretic Factor; Cadherins; Cell Adhesion; Cell Polarity; Cell Size; Cells, Cultured; Collagen; Connexin 43; Elasticity; Heart Ventricles; Hypertrophy; Membranes, Artificial; Micromanipulation; Myocytes, Cardiac; Photography; Rats; Rats, Sprague-Dawley; Rheology; Silicone Elastomers; Stress, Mechanical

Natriuretic peptide signaling is important in the regulation of blood pressure as well as in the growth of multiple cell types. To examine the role of natriuretic peptide signaling in atherosclerosis, we crossbred mice that lack natriuretic peptide receptor A (NPRA; Npr1-/-) with atherosclerosis-prone mice that lack apolipoprotein E (apoE; Apoe-/-).. Doubly deficient Npr1-/-Apoe-/- mice have increased blood pressure relative to Npr1+/+Apoe-/- mice (118+/-4 mm Hg compared with 108+/-2 mm Hg, P<0.05) that is coincident with a 64% greater atherosclerotic lesion size (P<0.005) and more advanced plaque morphology. Additionally, aortic medial thickness is increased by 52% in Npr1-/-Apoe-/- mice relative to Npr1+/+Apoe-/- mice (P<0.0001). Npr1-/-Apoe-/- mice also have significantly greater cardiac mass (9.0+/-0.3 mg/g body weight) than either Npr1+/+Apoe-/- mice (5.8+/-0.2 mg/g) or Npr1-/-Apoe+/+ mice (7.1+/-0.2 mg/g), suggesting that the lack of both NPRA and apoE synergistically enhances cardiac hypertrophy.. These data provide evidence that NPR1 is an atherosclerosis susceptibility locus and represents a potential link between atherosclerosis and cardiac hypertrophy. Our results also suggest roles for Npr1 as well as Apoe in regulation of hypertrophic cell growth.

Topics: Animals; Aorta; Apolipoproteins E; Arteriosclerosis; Atrial Natriuretic Factor; Blood Pressure; Cholesterol; Creatinine; Female; Genetic Predisposition to Disease; Genotype; Guanylate Cyclase; Hypertrophy; Longevity; Male; Mice; Mice, Knockout; Mice, Mutant Strains; Muscle, Smooth, Vascular; Receptors, Atrial Natriuretic Factor; Triglycerides

In overloaded heart the cardiomyocytes adapt to increased mechanical and neurohumoral stress by activation of hypertrophic program, resulting in morphological changes of individual cells and specific changes in gene expression. Accumulating evidence suggests an important role for the zinc finger transcription factor GATA-4 in hypertrophic agonist-induced cardiac hypertrophy. However, its role in stretch-induced cardiomyocyte hypertrophy is not known. We employed an in vitro mechanical stretch model of cultured cardiomyocytes and used rat B-type natriuretic peptide promoter as stretch-sensitive reporter gene. Stretch transiently increased GATA-4 DNA binding activity and transcript levels, which was followed by increases in the expression of B-type natriuretic peptide as well as atrial natriuretic peptide and skeletal alpha-actin genes. The stretch inducibility mapped primarily to the proximal 520 bp of the B-type natriuretic peptide promoter. Mutational studies showed that the tandem GATA consensus sites of the proximal promoter in combination with an Nkx-2.5 binding element are critical for stretch-activated B-type natriuretic peptide transcription. Inhibition of GATA-4 protein production by adenovirus-mediated transfer of GATA-4 antisense cDNA blocked stretch-induced increases in B-type natriuretic peptide transcript levels and the sarcomere reorganization. The proportion of myocytes with assembled sarcomeres in control adenovirus-infected cultures increased from 14 to 59% in response to stretch, whereas the values for GATA-4 antisense-treated cells were 6 and 13%, respectively. These results show that activation of GATA-4, in cooperation with a factor binding on Nkx-2.5 binding element, is essential for mechanical stretch-induced cardiomyocyte hypertrophy.

Topics: Animals; Atrial Natriuretic Factor; DNA-Binding Proteins; GATA4 Transcription Factor; Hypertrophy; Mechanotransduction, Cellular; Myocytes, Cardiac; Natriuretic Peptide, Brain; Nuclear Proteins; Promoter Regions, Genetic; Rats; Rats, Sprague-Dawley; RNA, Messenger; Transcription Factor AP-1; Transcription Factors; Transcriptional Activation

The effects of low-dose oral spironolactone (SPIRO) in a rat model of hypertensive heart failure (spontaneously hypertensive heart failure rat) were compared with its effects when combined with captopril (CAP). Twenty-six spontaneously rats with hypertensive heart failure were treated with either placebo (CON), SPIRO (20 mg/kg/d by mouth), CAP (100 mg/kg/d by mouth), or both SPIRO and CAP for 12 weeks. This dose of oral SPIRO did not affect blood pressure, left ventricular end-diastolic diameter, left ventricular ejection fraction, plasma atrial natriuretic peptide concentration, or cardiac fibrosis; however, in combination with CAP, it exerted a significant depressor effect after 12 weeks of treatment that was accompanied by increased urine output and decreased urinary protein excretion. These effects were significantly greater than those with CAP treatment alone. A significant increase in plasma aldosterone level was observed only in CON (174 +/- 21%). These data suggest that the addition of low-dose SPIRO to angiotensin I-converting enzyme inhibitor treatment may prevent progression into end-stage congestive heart failure through synergistic effects on diuresis and renoprotection.

Topics: Administration, Oral; Aldosterone; Angiotensin-Converting Enzyme Inhibitors; Animals; Atrial Natriuretic Factor; Blood Pressure; Captopril; Diuresis; Drug Therapy, Combination; Fibrosis; Heart Failure; Heart Ventricles; Hypertension; Hypertrophy; Male; Mineralocorticoid Receptor Antagonists; Myocardium; Organ Size; Rats; Rats, Inbred SHR; Spironolactone; Time Factors; Ultrasonography

The aryl hydrocarbon receptor (AhR), a ligand activated transcription factor, is the receptor for the polycyclic aromatic hydrocarbons found in tobacco smoke, polychlorinated biphenyls, and the environmental pollutant, dioxin. To better understand the role of the AhR in the heart, echocardiography, invasive measurements of aortic and left ventricular pressures, isolated working heart preparations, as well as morphological and molecular analysis were used to investigate the impact of AhR inactivation on the mouse heart using the AhR knockout as a model. Cardiac hypertrophy is an early phenotypic manifestation of the AhR knockout. Although the knockout animals were not hypertensive at the ages examined, cardiomyopathy accompanied by diminished cardiac output developed. Despite the structural left ventricular remodeling, the hearts of these animals exhibit minimal fibrosis and do not have the expected increases in surrogate molecular markers of cardiac hypertrophy. The anatomic remodeling without typical features of molecular remodeling is not consistent with hypertrophic growth secondary to pressure or volume overload, suggesting that increased cardiomyocyte size may be a direct consequence of the absence of the AhR in this cell type.

Topics: Actins; Animals; Aorta; Atrial Natriuretic Factor; Biomarkers; Blood Pressure; Cardiomegaly; Cardiovascular Physiological Phenomena; Echocardiography; Hypertrophy; In Vitro Techniques; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Myocardium; Myocytes, Cardiac; Organ Size; Receptors, Aryl Hydrocarbon; RNA, Messenger; Ventricular Function, Left; Ventricular Remodeling

During hypertrophy the heart increases its utilization of glucose and decreases that of fatty acids, resuming a fetal pattern of substrate metabolism. As demonstrated here, GLUT1 protein expression is increased in association with in vivo pressure-overload-induced hypertrophy. The relationship of changes in GLUT1 to enhanced glucose uptake and to cardiomyocyte hypertrophy and survival is not known. To explore this question we first examined the effect of prostaglandin F2alpha (PGF2alpha), an established hypertrophic agonist, on GLUT1 expression and glucose uptake in neonatal rat ventricular myocytes (NRVMs). PGF2alpha treatment for 24 h led to a fivefold increase in GLUT1 expression and a sixfold increase in glucose uptake. However, NRVMs cultured in the absence of glucose or with 3-O-methyl glucose, a competitive inhibitor of glucose uptake, still exhibited PGF2alpha-induced hypertrophic growth. In addition, we determined that overexpression of GLUT1 using adenovirus was insufficient to cause an increase in cell size, myofibrillar organization, or atrial natriuretic factor (ANF) expression. On the other hand, adenoviral overexpression of antisense GLUT1 (which blocked PGF2alpha-induced increases in GLUT1 protein) prevented PGF2alpha-stimulated cell enlargement and increases in ANF transcription. Overexpression of GLUT1 or addition of PGF2alpha also protected cells against serum deprivation-induced apoptosis; this effect was blocked by antisense GLUT1 but, surprisingly, was not dependent on glucose. Together, these data suggest that upregulation of GLUT1 serves a role in agonist-induced hypertrophy and survival which can be dissociated from its role in glucose transport.

Topics: Adenoviridae; Animals; Aorta; Apoptosis; Atrial Natriuretic Factor; Biological Transport; Blotting, Western; Cell Survival; Cells, Cultured; Dinoprost; Enzyme-Linked Immunosorbent Assay; Glucose; Glucose Transporter Type 1; Hypertrophy; Immunohistochemistry; Male; Mice; Mice, Inbred C57BL; Monosaccharide Transport Proteins; Muscle Cells; Myocytes, Cardiac; Oligonucleotides, Antisense; Pressure; Rats; Rats, Sprague-Dawley; Subcellular Fractions; Time Factors; Up-Regulation

Activation of G(q)-coupled alpha(1)-adrenergic receptors leads to hypertrophic growth of neonatal rat ventricular cardiomyocytes that is associated with increased expression of hypertrophy-related genes, including atrial natriuretic peptide (ANP) and myosin light chain-2 (MLC), as well as increased ribosome synthesis. The role of inositol phosphates in signaling pathways involved in these changes in gene expression was examined by overexpressing inositol phosphate-metabolizing enzymes and determining effects on ANP, MLC, and 45 S ribosomal gene expression following co-transfection of appropriate reporter gene constructs. Overexpression of enzymes that metabolize inositol 1,4,5-trisphosphate did not reduce ANP or MLC responses, but overexpression of the enzyme primarily responsible for metabolism of inositol 4,5-bisphosphate (Ins(1,4)P(2)), inositol polyphosphate 1-phosphatase (INPP), reduced ANP and MLC responses associated with alpha(1)-adrenergic receptor-mediated hypertrophy. Similarly overexpressed INPP reduced ANP and MLC responses associated with contraction-induced hypertrophy. In addition, overexpression of INPP reduced the increase in ribosomal DNA transcription associated with both hypertrophic models. Hypertrophied cells from both cell models as well as ventricular tissue from mouse hearts hypertrophied by pressure overload in vivo contained heightened levels of Ins(1,4)P(2), suggesting reduced INPP activity in three different models of hypertrophy. These studies provide evidence for an involvement of Ins(1,4)P(2) in hypertrophic signaling pathways in ventricular myocytes.

Topics: Animals; Atrial Natriuretic Factor; Blotting, Western; Cardiac Myosins; Cells, Cultured; CHO Cells; Cloning, Molecular; Cricetinae; DNA, Complementary; DNA, Ribosomal; Electrophoresis, Polyacrylamide Gel; Gene Library; Genes, Reporter; Humans; Hypertrophy; Inositol Phosphates; Mice; Mice, Inbred C57BL; Models, Chemical; Myocardium; Myosin Light Chains; Phosphoric Monoester Hydrolases; Promoter Regions, Genetic; Protein Binding; Protein Kinase C; Rats; Signal Transduction; Time Factors; Transcription, Genetic; Transfection

The contractile force of the cardiomyocyte is transmitted through the adherens junction, a component of the intercalated disc, enabling the myocardium to function as a syncytium. The cadherin family of cell adhesion receptors, located in the adherens junction, interact homophilically to mediate strong cell-cell adhesion. Ectopic expression of cadherins is associated with changes in tumor cell behavior and pathology. To examine the effect of cadherin specificity on cardiac structure and function, we expressed either the epithelial cadherin, E-cadherin, or N-cadherin in the heart of transgenic mice. E-cadherin was localized to the intercalated disc structure in these animals similar to endogenous N-cadherin. Both N- and E-cadherin transgenic animals developed dilated cardiomyopathy. However, misexpression of E-cadherin led to earlier onset and increased mortality compared with N-cadherin mice. A dramatic decrease in connexin 43 was associated with the hypertrophic response in E-cadherin transgenic mice. Myofibril organization appeared normal although, vinculin, which normally localizes to the intercalated disc, was redistributed to the cytoplasm in the E-cadherin transgenic mice. Furthermore, E-cadherin induced cyclin D1, nuclear reduplication, and karyokinesis in the absence of cytokinesis, resulting in myocytes with two closely opposed nuclei. By contrast, N-cadherin overexpressing transgenic mice did not exhibit an increase in cyclin D1, suggesting that E-cadherin may provide a specific growth signal to the myocyte. This study demonstrates that modulation of cadherin-mediated adhesion can lead to dilated cardiomyopathy and that E-cadherin can stimulate DNA replication in myocytes normally withdrawn from the cell cycle.

Topics: Animals; Atrial Natriuretic Factor; Cadherins; Cardiomyopathies; Cell Adhesion; Connexin 43; Female; Humans; Hypertrophy; Intercellular Junctions; Mice; Mice, Transgenic; Myocardium; Myosin Heavy Chains; Vinculin

Norepinephrine (NE) causes hypertrophic growth of cardiac myocytes via stimulation of alpha1-adrenergic receptors (alpha1-AR). Reactive oxygen species (ROS) can act as signaling molecules for cell growth. Accordingly, we tested the hypothesis that ROS mediate alpha1-AR-stimulated hypertrophic growth in adult rat ventricular myocytes (ARVM). NE increased the level of intracellular ROS as assessed by lucigenin chemiluminescence or cytochrome c reduction, and this effect was prevented by the superoxide dismutase (SOD)-mimetic MnTMPyP. NE also caused the induction of MnSOD mRNA. alpha1-AR stimulation with NE (1 microM) in the presence of propranolol (2 microM) for 48-96 h caused a hypertrophic growth phenotype characterized by a 36+/-3% increase in 3H-leucine incorporation, a 49+/-14% increase in protein accumulation, a six-fold induction of atrial natriuretic peptide mRNA, actin filament reorganization, and the induction of MnSOD mRNA. These responses were all prevented by pretreatment with the alpha1-AR-selective antagonist prazosin (100 n M) or the SOD-mimetics MnTMPyP (50 microM) and Euk-8 (100 microM). MnTMPyP had no effect on alpha1-AR-stimulated 3H-inositol phosphate turnover or the hypertrophic phenotype caused by the protein kinase C activator phorbol-12-myristate-13-acetate. Thus, ROS play a critical role in mediating the hypertrophic growth response to alpha1-AR-stimulation in ARVM.

Topics: Actin Cytoskeleton; Actins; Animals; Atrial Natriuretic Factor; Cell Division; Enzyme Induction; Ethylenediamines; Gene Expression Regulation; Heart; Heart Ventricles; Hypertrophy; Inositol Phosphates; Myocardium; Norepinephrine; Organometallic Compounds; Porphyrins; Prazosin; Rats; Reactive Oxygen Species; Receptors, Adrenergic, alpha-1; RNA, Messenger; Signal Transduction; Superoxide Dismutase

We have demonstrated that two hypertrophic agents, interleukin-1 beta (IL-1 beta) and leukemic inhibitory factor (LIF), altered cardiac myocyte morphology with striking similarity and prompted us to investigate the common actions of these cytokines. We compared the phosphorylation/activation of signal tranducer and activator of transcription 3 (STAT3), extracellular signal-regulated kinase (ERK), p38(MAPK), and c-Jun N-terminal kinase mitogen-activated protein kinases (MAPKs). The phosphorylation of STAT3 by IL-1 beta was delayed (>60 min), whereas the response to LIF was rapid (<10 min) and transient. We confirmed that IL-1 beta potently stimulated all three MAPK subfamilies. In contrast, LIF promoted strong activation of ERKs, marginal activation of p38(MAPK), and no c-Jun N-terminal kinase activation. To test the roles of ERKs and p38(MAPK), myocytes were pretreated with PD98059 and SB203580. Either inhibitor alone prevented STAT3 phosphorylation, implicating ERKs and p38(MAPK) in the delayed STAT3 response to IL-1 beta. The interplay of MAPKs and STAT3 phosphorylation in regulating IL-1 beta-stimulated hypertrophy was investigated by evaluating the effect of MAPK inhibitors on atrial natriuretic factor (ANF) expression and myocyte morphology. The specific inhibition of either ERK or p38(MAPK) attenuated the IL-1 beta- or LIF-stimulated ANF expression by up to 70%. Inhibition was not further increased in the presence of both inhibitors. Furthermore, although individual inhibition of ERK or p38(MAPK) did not affect morphology, co-treatment with both inhibitors abrogated the hypertrophic morphology stimulated by IL-1 beta but not by LIF. Taken together, our data indicate that the activation of ERK and p38(MAPK) is essential in regulating a delayed STAT3 phosphorylation as well as changes in ANF expression and morphology that follow IL-1 beta treatment. Thus, the role of MAPKs in the hypertrophic response can be dictated at least partly by the nature of the hypertrophic agent employed.

Topics: Acute-Phase Proteins; Animals; Animals, Newborn; Atrial Natriuretic Factor; Cell Nucleus; Cells, Cultured; Culture Media, Serum-Free; Cycloheximide; DNA-Binding Proteins; Enzyme Inhibitors; Flavonoids; Growth Inhibitors; Heart; Heart Ventricles; Hypertrophy; Imidazoles; Interleukin-1; Interleukin-6; JNK Mitogen-Activated Protein Kinases; Kinetics; Leukemia Inhibitory Factor; Lymphokines; MAP Kinase Signaling System; Mitogen-Activated Protein Kinases; Myocardium; p38 Mitogen-Activated Protein Kinases; Phenylephrine; Phosphorylation; Protein Transport; Pyridines; Rats; Rats, Sprague-Dawley; STAT3 Transcription Factor; Trans-Activators

ANG II arrests LLC-PK1 cells in the G1 phase of the cell cycle and induces hypertrophy, an effect mediated by induction of p27Kip1. We studied whether atrial natriuretic peptide (ANP) may modulate ANG II-induced hypertrophy and p27Kip1 expression in tubular LLC-PK1 cells. ANP, through its fragments 3---28 and 4---27, prevented ANG II-induced cell cycle arrest. ANP inhibited >80% of ANG II-induced p27Kip1 protein expression (Western blots). ANP stimulated expression of MKP-1, a phosphatase involved in dephosphorylation of p44/42 mitogen-activated protein (MAP) kinase, up to 12 h. ANP prevented the ANG II-mediated phosphorylation peak of MAP kinase after 12 h of stimulation. 8-Bromo-cGMP mimicked all the effects of ANP. Transfection with MKP-1 antisense, but not sense, oligonucleotides abolished the modifying role of ANP on ANG II-mediated cell cycle arrest. The effect of ANP on ANG II-mediated hypertrophy of LLC-PK1 cells is regulated on the level of MAP kinase phosphorylation, a key step in the induction of p27Kip1. Although ANP and ANG II both stimulate generation of reactive oxygen species, ANP additionally induces expression of MKP-1, leading to interference with ANG II-mediated MAP kinase phosphorylation.

Topics: Angiotensin II; Animals; Atrial Natriuretic Factor; Cell Line; Hypertrophy; Kidney Tubules; Mitogen-Activated Protein Kinases; Oligonucleotides, Antisense; Phosphorylation; Protein Tyrosine Phosphatases; Superoxides; Swine; Transfection

Several studies have demonstrated that static stretch of cardiomyocytes induces cardiomyocyte hypertrophy. We investigated the effects of cyclic stretch, a more physiological stimulus, on protein synthesis and DNA synthesis of rat ventricular cardiomyocytes and cardiofibroblasts. Further-more, we investigated whether these effects are caused by autocrine mechanisms. In addition, we studied the paracrine influences of stretched cardiofibroblasts on cardiomyocyte growth. Short-term cyclic stretch (0-24 h) of cardiomyocytes induced a growth response indicative of cardiomyocyte hypertrophy, given the fact that increased rates of protein synthesis and DNA synthesis were accompanied by an elevated release of atrial natriuretic peptide into the culture medium. In cardiofibroblasts, short-term cyclic stretch also induced a growth response as indicated by an increased rate of protein synthesis and DNA synthesis. Furthermore, incubation of stationary cardiofibroblasts with conditioned medium derived from stretched cardiofibroblasts revealed an autocrine effect of stretch as illustrated by an increased rate of protein synthesis in stationary cardiofibroblasts. In analogy, there was an autocrine effect of stretch on stationary cardiomyocytes incubated with conditioned medium derived from stretched cardiomyocytes. Moreover, we observed a paracrine effect of the conditioned medium derived from stretched cardiofibroblasts on stationary cardiomyocytes. Thus, short-term cyclic stretch of cardiomyocytes and cardiofibroblasts induces growth responses that are the result of direct, autocrine, and paracrine effects. These autocrine/paracrine effects of stretch are most probably due to release of factors from stretched cells.

Topics: Animals; Animals, Newborn; Atrial Natriuretic Factor; Autocrine Communication; Cell Division; Cells, Cultured; DNA; Fibroblasts; Hyperplasia; Hypertrophy; Muscle Contraction; Myocardium; Paracrine Communication; Rats; Rats, Wistar; Time Factors

Oxidative stress stimulates both growth and apoptosis in cardiac myocytes in vitro. We investigated whether oxidative stress mediates hypertrophy and apoptosis in cyclically stretched ventricular myocytes. Neonatal rat ventricular myocytes cultured on laminin-coated silastic membranes were stretched cyclically (1 Hz) at low (nominal 5%) and high (nominal 25%) amplitudes for 24 hours. Stretch caused a graded increase in superoxide anion production as assessed by superoxide dismutase (SOD)-inhibitable cytochrome c reduction or electron paramagnetic resonance spectroscopy. The role of reactive oxygen species (ROS) was assessed using the cell-permeable SOD/catalase mimetics Mn(II/III)tetrakis(1-methyl-4-peridyl) (MnTMPyP) and EUK-8. Stretch-induced increases in protein synthesis ((3)H-leucine incorporation) and cellular protein content were completely inhibited by MnTMPyP (0.05 mmol/L) at both low and high amplitudes of stretch. In contrast, while MnTMPyP inhibited basal atrial natriuretic factor (ANF) mRNA expression, the stretch-induced increase in ANF mRNA expression was not inhibited by MnTMPyP. In contrast to hypertrophy, only high-amplitude stretch increased myocyte apoptosis, as reflected by increased DNA fragmentation on gel electrophoresis and an approximately 3-fold increase in the number of TUNEL-positive myocytes. Similarly, only high-amplitude stretch increased the expression of bax mRNA. Myocyte apoptosis and bax expression stimulated by high-amplitude stretch were inhibited by MnTMPyP. Both low- and high-amplitude stretch caused rapid phosphorylation of ERK1/2, while high-, but not low-, amplitude stretch caused phosphorylation of JNKs. Activation of both ERK1/2 and JNKs was ROS-dependent. Thus, cyclic strain causes an amplitude-related increase in ROS, associated with differential activation of kinases and induction of hypertrophic and apoptotic phenotypes.

Topics: Animals; Animals, Newborn; Apoptosis; Atrial Natriuretic Factor; bcl-2-Associated X Protein; Cells, Cultured; Ethylenediamines; Free Radical Scavengers; Gene Expression Regulation; Heart Ventricles; Hypertrophy; Leucine; Organometallic Compounds; Porphyrins; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-bcl-2; Rats; Reactive Oxygen Species; RNA, Messenger; Signal Transduction; Stress, Mechanical; Superoxides; Tritium

1. Calcium channel blockers (CCBs) are anti-hypertensive drugs that are usually considered to act mainly as vasodilators. We investigated the relation between the reduction of blood pressure evoked by two long-acting CCBs and their protective effect against cardiac and renal damage in salt-loaded stroke-prone spontaneously hypertensive rats (SHRSP). 2. SHRSP were exposed to high dietary salt intake (1% NaCl in drinking solution) from 8 to 14 weeks of age, with or without amlodipine or lacidipine at three dosage regimens producing similar effects on blood pressure. 3. The lowest dosages of both drugs had non-significant effects on blood pressure but inhibited the paradoxical increases in plasma renin activity (PRA) and in renin mRNA in kidney that were found in salt-loaded SHRSP. The lowest dosage of lacidipine (but not of amlodipine) restored the physiological downregulation of renin production by high salt and reduced left ventricular hypertrophy and mRNA levels of atrial natriuretic factor and transforming growth factor-beta1. 4. The intermediate dosages reduced blood pressure and PRA in a comparable manner, but cardiac hypertrophy was more reduced by lacidipine than by amlodipine. 5. Although the highest doses exhibited a further action on blood pressure, they had no additional effect on cardiac hypertrophy, and they increased PRA and kidney levels of renin mRNA even more than in the absence of drug treatment. 6. We conclude that reduction of blood pressure is not the sole mechanism involved in the prevention of cardiac remodelling by CCBs, and that protection against kidney damage and excessive renin production by low and intermediate dosages of these drugs contributes to their beneficial cardiovascular effects.

Topics: Actins; Amlodipine; Animals; Atrial Natriuretic Factor; Blood Pressure; Calcium Channel Blockers; Collagen Type I; Dihydropyridines; Dose-Response Relationship, Drug; Fibrosis; Gene Expression Regulation; Heart Ventricles; Hypertension; Hypertrophy; Kidney; Male; Muscle, Skeletal; Rats; Rats, Inbred SHR; Renin; RNA, Messenger; Sodium Chloride, Dietary; Transforming Growth Factor beta; Transforming Growth Factor beta1

Several investigations have demonstrated the regional heterogeneity of myocardial phenotype, and hypertrophy may also induce regionally disparate changes. We have utilized the direct DNA injection technique to study regional variations in overload-induced ANF expression. Pressure overload was induced by stenosis of the ascending aorta in canines. ANF promoter reporters were injected into the left ventricle; in different regions including the base, the midwall region, and the apex. Injections were made at different depths to include the epicardial and endocardial layers. The animals were sacrificed 7 days following surgery and the left ventricle harvested for tissue analysis. Under normotensive conditions, ANF reporter expression was similar throughout the heart. PO increased ANF expression and the increases were greater in the endocardium than in the epicardium. PO also significantly increased expression in the midwall and base regions, but not in the apex. It is unknown from these experiments, whether the greater increases in midwall expression are a function of greater wall stress, metabolic demand, or phenotypic differences in the midwall myocytes. These findings do indicate that regional differences in overload-induced changes in gene expression are evident and may be functionally important in determining myocardial response to increased functional demand.

Topics: Animals; Aortic Valve Stenosis; Atrial Natriuretic Factor; DNA; Dogs; Gene Expression Regulation; Genes, Reporter; Hypertrophy; Luciferases; Myocardium; Phenotype; Plasmids; Pressure; Promoter Regions, Genetic

Cardiac hypertrophy is a major predictor of future morbidity and mortality. Recent investigation has centered around identifying the molecular signaling pathways that regulate cardiac myocyte reactivity with the goal of modulating pathologic hypertrophic programs. One potential regulator of cardiomyocyte hypertrophy is the calcium-sensitive phosphatase calcineurin. We show here that calcineurin enzymatic activity, mRNA, and protein levels are increased in cultured neonatal rat cardiomyocytes by hypertrophic agonists such as angiotensin II, phenylephrine, and 1% fetal bovine serum. This induction of calcineurin activity was associated with an increase in calcineurin Abeta (CnAbeta) mRNA and protein, but not in CnAalpha or CnAgamma. Agonist-dependent increases in calcineurin enzymatic activity were specifically inhibited with an adenovirus expressing a noncompetitive peptide inhibitor of calcineurin known as cain [Lai, M. M., Burnett, P. E., Wolosker, H., Blackshaw, S. & Snyder, S. H. (1998) J. Biol. Chem. 273, 18325-18331]. Targeted inhibition of calcineurin with cain or an adenovirus expressing only the calcineurin inhibitory domain of AKAP79 attenuated cardiomyocyte hypertrophy and atrial natriuretic factor expression in response to angiotensin II, phenylephrine, and 1% fetal bovine serum. These data demonstrate that calcineurin is an important regulator of cardiomyocyte hypertrophy in response to certain agonists and suggest that cyclosporin A and FK506 function to attenuate cardiac hypertrophy by specifically inhibiting calcineurin.

Topics: Adenoviridae; Angiotensin II; Animals; Apoptosis Regulatory Proteins; Atrial Natriuretic Factor; Calcineurin; Calcineurin Inhibitors; Cardiomegaly; Carrier Proteins; Cattle; Cells, Cultured; DNA-Binding Proteins; Fetal Blood; Gene Expression Regulation; Genes; Genetic Therapy; Genetic Vectors; Hypertrophy; Myocardium; NFATC Transcription Factors; Nuclear Proteins; Phenylephrine; Rats; Reverse Transcriptase Polymerase Chain Reaction; Transcription Factors; Transfection

The transcriptional regulation of an isolated rat phospholamban (PL) promoter fragment in rat cardiomyocytes was analyzed by applying a new method to reach substantially higher transfection efficiencies: gene gun biolistics. The gene gun transfection method was optimized for application to primary cultures of rat neonatal cardiomyocytes. Cells, cultured at different densities (0.75-1.50x10(5)cells/cm(2)) in serum-free medium, were transfected with DNA coated gold particles. A transfection efficiency of up to 10% could be achieved (compared to <1% with other methods) by the gene gun as checked using a RSV- beta-Gal construct. Cardiomyocytes were stimulated by endothelin-1 (ET-1) (10(-8)M) to induce hypertrophy, thereby yielding the characteristic changes in gene expression (upregulation of Atrial Natriuretic Factor (ANF) and downregulation of PL). The basal activity of an ANF promoter fragment (increasing from the lowest to highest density 2.6-fold) and its ET-1 inducibility (only significant upregulation of 2.6-fold, at lowest density) appeared to be dependent on the plating density of the cardiomyocytes. A PL promoter fragment was isolated, sequenced and 1.4 kb was subcloned in a luciferase reporter vector. The basal activity of the PL promoter fragment was not dependent on the plating density. ET-1 did not downregulate the PL promoter, rather a significant upregulation (1.4-fold) was found at the highest plating density. In conclusion, plating density of the cardiomyocytes can influence promoter activity as shown with an ANF promoter fragment. A newly isolated and sequenced rat PL promoter fragment did not direct gene expression as expected on basis of downregulation of the PL gene by ET-1 observed in this model.

Topics: Animals; Animals, Newborn; Atrial Natriuretic Factor; Base Sequence; Biolistics; Calcium-Binding Proteins; Cells, Cultured; Culture Media, Serum-Free; Endothelin-1; Gene Expression Regulation; Genes, Reporter; Heart; Hypertrophy; Luciferases; Molecular Sequence Data; Muscle Proteins; Myocardium; Promoter Regions, Genetic; Rats; Recombinant Fusion Proteins; Transcription, Genetic; Transfection

The signal transduction mechanisms mediating hypertrophic responses in myocardial cells (MCs) remain uncertain. We investigated the role of the extracellular signal-regulated kinase (ERK) cascade in myocardial cell hypertrophy by the strategy of using the adenovirus-mediated overexpression of mitogen-activated protein kinase (MAPK)/ERK kinase (MEK), which is the upstream activator of ERK. We generated recombinant adenoviruses expressing constitutively active MEK1 (MEK1 EE) and dominant negative MEK1 (MEK1 DN). Overexpression of MEK1 EE in MCs activated ERK1/2 and subsequently induced atrial natriuretic peptide (ANP) mRNA expression. In addition, MEK1 EE overexpression resulted in an increase in cell size and sarcomeric reorganization. In contrast, overexpression of MEK1 DN in MCs inhibited endothelin-1 (ET-1)-, phenylephrine (PE)-, leukemia inhibitory factor (LIF)-, isoproterenol (ISP)-, and mechanical stretch-induced ERK activation and ANP mRNA expression. MEK1 DN overexpression inhibited ET-1-, PE-, LIF-, and ISP-induced increases in cell size and sarcomeric reorganization. Consistent with the observed effects on cellular morphology, overexpression of MEK1 EE resulted in an increase in amino acid incorporation, while overexpression of MEK1 DN inhibited ET-1-, PE-, LIF-, ISP-, and mechanical stretch-induced increases in amino acid incorporation. These results indicate that the ERK cascade plays an important role in the signaling pathway leading to the development of myocardial cell hypertrophy.

Topics: Adenoviridae; Animals; Atrial Natriuretic Factor; Cardiomegaly; Cells, Cultured; Enzyme Activation; Genetic Vectors; Hypertrophy; MAP Kinase Kinase 1; MAP Kinase Signaling System; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Mitogen-Activated Protein Kinase Kinases; Mitogen-Activated Protein Kinases; Myocardium; Protein Serine-Threonine Kinases; Rats; Rats, Sprague-Dawley; RNA, Messenger; Sarcomeres

Cultured neonatal rat cardiac myocytes have been used extensively to study cellular and molecular mechanisms of cardiac hypertrophy. However, there are only a few studies in cultured mouse myocytes despite the increasing use of genetically engineered mouse models of cardiac hypertrophy. Therefore, we characterized hypertrophic responses in low-density, serum-free cultures of neonatal mouse cardiac myocytes and compared them with rat myocytes. In mouse myocyte cultures, triiodothyronine (T3), norepinephrine (NE) through a beta-adrenergic receptor, and leukemia inhibitory factor induced hypertrophy by a 20% to 30% increase in [(3)H]phenylalanine-labeled protein content. T3 and NE also increased alpha-myosin heavy chain (MyHC) mRNA and reduced beta-MyHC. In contrast, hypertrophic stimuli in rat myocytes, including alpha(1)-adrenergic agonists, endothelin-1, prostaglandin F(2alpha), interleukin 1beta, and phorbol 12-myristate 13-acetate (PMA), had no effect on mouse myocyte protein content. In further contrast with the rat, none of these agents increased atrial natriuretic factor or beta-MyHC mRNAs. Acute PMA signaling was intact by extracellular signal-regulated kinase (ERK1/2) and immediate-early gene (fos/jun) activation. Remarkably, mouse but not rat myocytes had hypertrophy in the absence of added growth factors, with increases in cell area, protein content, and the mRNAs for atrial natriuretic factor and beta-MyHC. We conclude that mouse myocytes have a unique autonomous hypertrophy. On this background, T3, NE, and leukemia inhibitory factor activate hypertrophy with different mRNA phenotypes, but certain Gq- and protein kinase C-coupled agonists do not.

Topics: Adrenergic alpha-Agonists; Animals; Animals, Newborn; Atrial Natriuretic Factor; Cells, Cultured; Dinoprost; Endothelin-1; Fetus; Growth Inhibitors; Growth Substances; Heart; Hypertrophy; Interleukin-1; Interleukin-6; Leukemia Inhibitory Factor; Lymphokines; Mice; Microscopy, Phase-Contrast; Models, Animal; Myocardium; Myosin Heavy Chains; Myosins; Norepinephrine; Phenotype; Phenylalanine; Protein Biosynthesis; Proteins; Rats; RNA, Messenger; Tetradecanoylphorbol Acetate; Triiodothyronine

Topics: Animals; Atrial Appendage; Atrial Natriuretic Factor; Disease Models, Animal; Dogs; Goats; Guinea Pigs; Heart Diseases; Hemodynamics; Humans; Hypertrophy; Sheep; Stroke; Thirst; Thrombosis

Accumulating evidence suggests that the local synthesis of endothelin-1 (ET-1) plays a role in the development of heart failure in vivo. We investigated the role of endothelin-converting enzyme-1 (ECE-1), which mediates the conversion of big ET-1 to mature ET-1, in the development of alpha1-adrenergic-stimulated hypertrophy in cultured neonatal rat cardiac myocytes.. Phenylephrine (PE) induced the expression of ET-1 in rat cardiac myocytes and accelerated the conversion of big ET-1 to ET-1. The ECE-1 mRNA levels were markedly increased 3 hours after PE stimulation (3.6-fold compared with saline stimulation, P<0.005). A specific ECE-1 antagonist, FR901533, inhibited the PE-stimulated increase in protein synthesis rate by 45% (P<0.05). As genetic markers for the hypertrophic response, FR901533 inhibited the PE-stimulated transcriptional activities of the 3.5-kb beta-myosin heavy chain promoter by 79% (P<0.01) but did not affect that of the 3.4-kb atrial natriuretic factor (ANF) promoter. In Bio14.6 Syrian cardiomyopathic hamsters, ventricular ET-1 and ANF mRNA levels did not correlate at 2 different stages.. ET-1-independent pathways may mediate activation of the ANF gene program in ventricular myocytes both in vitro and in vivo. These results also indicate that the conversion of big ET-1 to ET-1 in rat cardiac myocytes is required for the development of alpha1-adrenergic-stimulated hypertrophy and beta-myosin heavy chain gene transcription.

Topics: Adrenergic alpha-Agonists; Animals; Animals, Newborn; Aspartic Acid Endopeptidases; Atrial Natriuretic Factor; Cells, Cultured; Cricetinae; Endothelin-1; Endothelin-Converting Enzymes; Hypertrophy; Mesocricetus; Metalloendopeptidases; Myocardium; Myosin Heavy Chains; Phenylephrine; Polymerase Chain Reaction; Rats; Rats, Sprague-Dawley; RNA, Messenger; Tetracyclines; Transcription, Genetic

To evaluate the role of angiotensin II (AII) on diastolic function during post-myocardial infarction (MI) ventricular remodeling, coronary ligation or sham operation was performed in male Sprague-Dawley rats. Experimental animals were maintained on either irbesartan, a selective AT1-receptor antagonist, or no treatment. Measurement of cardiac hypertrophy, diastolic function, and sarcoendoplasmic reticulum adenosine triphosphatase (ATPase; SERCA) and phospholamban (PLB) gene expression was assessed at 6 weeks after MI. Myocardial infarction caused a significant increase in myocardial mass and left ventricular (LV) filling pressure, whereas LV systolic pressure and +dP/dt were reduced. The time constant of isovolumic relaxation (tau) was markedly prolonged after MI. Post-MI hypertrophy was associated with substantial increases in the messenger RNA (mRNA) expression of atrial natriuretic peptide (ANP), but no significant changes in SERCA or PLB levels. Although irbesartan treatment did not significantly alter post-MI LV systolic or filling pressures, it nevertheless effectively decreased ventricular hypertrophy, improved tau, and normalized ANP expression. These results demonstrate that AT1-receptor antagonism has important effects on myocardial hypertrophy and ANP gene expression, which are independent of ventricular loading conditions. In addition, the improvement in diastolic function was not related to changes in SERCA and PLB gene expression, suggesting that enhanced myocardial relaxation was related to the blockade of AII effects on myocyte function or through a reduction of ventricular hypertrophy itself or both.

Topics: Angiotensin Receptor Antagonists; Animals; Antihypertensive Agents; Atrial Natriuretic Factor; Biphenyl Compounds; Body Weight; Calcium-Binding Proteins; Calcium-Transporting ATPases; Cardiomegaly; Diastole; Gene Expression; Heart Ventricles; Hemodynamics; Hypertrophy; Irbesartan; Male; Myocardial Infarction; Organ Size; Rats; Rats, Sprague-Dawley; Receptor, Angiotensin, Type 1; Receptor, Angiotensin, Type 2; RNA, Messenger; Tetrazoles

Agonist-induced hypertrophy of cultured neonatal rat ventricular myocytes (NRVM) has been attributed to biochemical signals generated during receptor activation. However, NRVM hypertrophy can also be induced by spontaneous or electrically stimulated contractile activity in the absence of exogenous neurohormonal stimuli. Using single-cell imaging of fura 2-loaded myocytes, we found that low-density, noncontracting NRVM begin to generate intracellular Ca2+ concentration ([Ca2+]i) transients and contractile activity within minutes of exposure to the alpha 1-adrenergic agonist phenylephrine (PE; 50 microM). However, NRVM pretreated with verapamil and then stimulated with PE failed to elicit [Ca2+]i transients and beating. We therefore examined whether PE-induced [Ca2+]i transients and contractile activity were required to elicit specific aspects of the hypertrophic phenotype. PE treatment (48-72 h) increased cell size, total protein content, total protein-to-DNA ratio, and myosin heavy chain (MHC) isoenzyme content. PE also stimulated sarcomeric protein assembly and prolonged MHC half-life. However, blockade of voltage-gated L-type Ca2+ channels with verapamil, diltiazem, or nifedipine (10 microM) blocked PE-induced total protein and MHC accumulation and prevented the time-dependent assembly of myofibrillar proteins into sarcomeres. Inhibition of actin-myosin cross-bridge cycling with 2,3-butanedione monoxime (7.5 mM) also prevented PE-induced total protein and MHC accumulation, indicating that mechanical activity, rather than [Ca2+]i transients per se, was required. In contrast, blockade of [Ca2+]i transients and contractile activity did not prevent the PE-induced increase in cell surface area, activation of the mitogen-activated protein kinases ERK1 and ERK2, or upregulation of atrial natriuretic factor gene expression. Thus contractile activity is required to elicit some but not all aspects of the the hypertrophic phenotype induced by alpha 1-adrenergic receptor activation.

Topics: Adrenergic alpha-Agonists; Animals; Animals, Newborn; Atrial Natriuretic Factor; Calcium; Calcium-Calmodulin-Dependent Protein Kinases; Enzyme Activation; Gene Expression; Heart; Hypertrophy; Intracellular Membranes; Myocardial Contraction; Myocardium; Osmolar Concentration; Phenylephrine; Rats; Rats, Sprague-Dawley; Sarcomeres

Sodium and water retention are common in acromegaly and upon GH administration. The underlying mechanisms, however, have not been clearly characterized as yet. Therefore, the aim of this study was to examine possible alterations of atrial natriuretic peptide (ANP), an endogenous regulator of volume homeostasis, in response to chronic elevated GH. We used GH-transgenic mice (GH-TM) as a model for chronic hypersomatotropinemia and moreover investigated 7 and 27 week old animals, respectively, in order to discriminate between short and long term effects of GH overexpression. Hematocrit values were reduced in GH-TM compared to control animals and it is known that plasma volume is increased in these animals. Structural lesions of the kidney were found in the GH-TM, however, in the animals studied there were no signs of renal insufficiency as evidenced by serum creatinine and urea levels. The serum concentration of immunoreactive ANP (IR-ANP) determined by RIA was significantly (P < 0.005) elevated in the young GH-TM as compared to control littermates (81.7+/-13.3 vs. 50.9+/-10.8 fmol/ml). The increase in serum IR-ANP of 27 week old GH-TM, however did not reach the level of significance (57.13+/-16.3 vs. 50.25+/-16.4 fmol/ml). Serum samples of control mice as well as of the 7 week old animals mainly contained ANP 99-126, known to be the circulating form of ANP. In contrast, serum of 27 week old GH-TM predominantly showed the cardiac storage form of ANP, ANP 1-126. Cardiac expression of ANP was quantified by Northern blot analysis. mRNA coding for ANP was found 1.2- and 2-fold increased in the atria of 7 and 27 week old GH-TM, respectively. In parallel, a 2.2-fold (7 week) and 2-fold (27 week) increase of IR-ANP was observed in transgenic atria compared to tissue of control animals. In contrast, no significant difference of ANP mRNA expression or of content of IR-ANP was observed in the ventricles of both groups of animals. In conclusion, GH-TM show various alterations in their ANP status suggesting an influence of the peptide on the effect of GH in fluid retention.

Topics: Aging; Animals; Atrial Natriuretic Factor; Blotting, Northern; Body Weight; Chromatography, High Pressure Liquid; Growth Hormone; Hematocrit; Hypertrophy; Kidney; Liver; Mice; Mice, Transgenic; Myocardium; Organ Size; Peptide Fragments; Plasma Volume; Protein Precursors; Radioimmunoassay

Left ventricular remodeling after myocardial infarction (MI) involves the hypertrophic growth of cardiomyocytes and the accumulation of fibrillar collagen in the interstitial space. We evaluated the role of kinins in postinfarction ventricular remodeling and their potential contribution to the antiremodeling effects of ACE inhibition and angiotensin II type 1 (AT1) receptor blockade.. Rats underwent coronary artery ligation followed by chronic B2 kinin receptor blockade with icatibant. Additional groups of infarcted rats were treated with the ACE inhibitor lisinopril or the AT1 receptor antagonist ZD7155, each separately and in combination with icatibant. B2 kinin receptor blockade enhanced the interstitial deposition of collagen after MI, whereas morphological and molecular markers of cardiomyocyte hypertrophy (cardiac weight, myocyte cross-sectional area, prepro-atrial natriuretic factor mRNA expression) were not affected. Chronic ACE inhibition and AT1 receptor blockade reduced collagen deposition and cardiomyocyte hypertrophy after MI. The inhibitory action of ACE inhibition and AT1 receptor blockade on interstitial collagen was partially reversed by B2 kinin receptor blockade. However, B2 kinin receptor blockade did not attenuate the effects of ACE inhibition and AT1 receptor blockade on cardiomyocyte hypertrophy.. (1) Kinins inhibit the interstitial accumulation of collagen but do not modulate cardiomyocyte hypertrophy after MI. (2) Kinins contribute to the reduction of myocardial collagen accumulation by ACE inhibition and AT1 receptor blockade. (3) The effects of ACE inhibition and AT1 receptor blockade on cardiomyocyte hypertrophy are related to a reduced generation/receptor blockade of angiotensin II.

Topics: Adrenergic beta-Antagonists; Angiotensin Receptor Antagonists; Angiotensin-Converting Enzyme Inhibitors; Animals; Atrial Natriuretic Factor; Bradykinin; Bradykinin Receptor Antagonists; Collagen; Enzyme Induction; Extracellular Matrix; Gene Expression Regulation; Heart Ventricles; Hemodynamics; Hypertrophy; Kinins; Male; Myocardial Contraction; Myocardial Infarction; Myocardium; Naphthyridines; Nitric Oxide Synthase; Peptidyl-Dipeptidase A; Polymerase Chain Reaction; Rats; Rats, Sprague-Dawley; Receptor, Bradykinin B2; Single-Blind Method

In the present study, we compare expression, storage and secretion of the atrial natriuretic factor (ANF) in atrial and ventricular adult rat cardiomyocytes (aARC and vARC) in long-term culture. The influence of insulin-like growth factor-I (IGF-I) and of basic fibroblast growth factor (bFGF) on ANF production and secretion, as well as on the expression of a structural component, alpha-smooth muscle actin (alpha-sm actin), was studied in the two cell types. Antibodies against alpha-ANF were used for immunocytochemical localization of ANF. aARC contained more ANF-granules than vARC, and they were distributed throughout the cell bodies. Quantitative determination of ANF storage and secretion was done by radioimmunoassay (RIA; 125I), and it was demonstrated that aARC stored and secreted ANF 18- and 16-times more, respectively, when compared to vARC. Immuno-electron microscopy confirmed that ANF storing secretory granules were present in both types of cardiomyocytes. Expression of ANF and alpha-sm actin in aARC and vARC responded differently to treatment with either IGF-I or bFGF. In aARC, neither IGF-I nor bFGF had an influence on expression of ANF. In vARC, expression of ANF was downregulated by IGF-I and upregulated by bFGF with regard to both immunoreactivity and message. In contrast to vARC, expression of alpha-sm actin was not affected by IGF-I in aARC, whereas bFGF produced a strong upregulation similar to that found in vARC. Mitogen-activated protein kinases (MAPK) 42 and 44, though, were equally activated by bFGF and IGF-I in both aARC and vARC.

Topics: Actins; Animals; Atrial Natriuretic Factor; Calcium-Calmodulin-Dependent Protein Kinases; Cells, Cultured; Enzyme Activation; Female; Fibroblast Growth Factor 2; Gene Expression Regulation; Heart Atria; Heart Ventricles; Hypertrophy; Insulin-Like Growth Factor I; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Mitogen-Activated Protein Kinases; Organ Specificity; Rats; Rats, Sprague-Dawley

In cardiac hypertrophy, both excessive enlargement of cardiac myocytes and progressive interstitial fibrosis are well known to occur simultaneously. In the present study, to investigate the interaction between ventricular myocytes (MCs) and cardiac nonmyocytes (NMCs), mostly fibroblasts, during cardiocytes hypertrophy, we examined the change in cell size and gene expression of atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) in cultured MCs as markers for hypertrophy in the neonatal rat ventricular cardiac cell culture system.. The size of cultured MCs significantly increased in the MC-NMC coculture. Concomitantly, secretions of ANP and BNP into culture media were significantly increased in the MC-NMC coculture compared with in the MC culture (with the possible contamination of NMC <1% of MC). Moreover, in the MC culture, enlargement of MC and an increase in ANP and BNP secretions were induced by treatment with conditioned media of the NMC culture. A considerable amount of endothelin (ET)-1 production was detected in the NMC-conditioned media. BQ-123, an ET-A receptor antagonist, and bosentan, a nonselective ET receptor antagonist, significantly blocked the hypertrophic response of MCs induced by treatment with NMC-conditioned media. Angiotensin II (Ang II) (10(-10) to 10(-6) mol/L) and transforming growth factor-beta1 (TGF-beta1) (10(-13) to 10(-9) mol/L), both of which are known to be cardiac hypertrophic factors, did not induce hypertrophy in MC culture, but both Ang II and TGF-beta1 increased the size of MCs and augmented ANP and BNP productions in the MC-NMC coculture. This hypertrophic activity of Ang II and TGF-beta1 was associated with the potentiation of ET-1 production in the MC-NMC coculture, and the effect of Ang II or TGF-beta1 on the secretions of ANP and BNP in the coculture was significantly suppressed by pretreatment with BQ-123.. These results demonstrate that NMCs regulate MC hypertrophy at least partially via ET-1 secretion and that the interaction between MCs and NMCs plays a critical role during the process of Ang II- or TGF-beta1-induced cardiocyte hypertrophy.

Topics: Angiotensin II; Animals; Atrial Natriuretic Factor; Cardiomegaly; Cattle; Coculture Techniques; Culture Media, Conditioned; Endothelin Receptor Antagonists; Endothelin-1; Hypertrophy; Lipoproteins, LDL; Myocardium; Natriuretic Peptide, Brain; Nerve Tissue Proteins; Paracrine Communication; Rats; Rats, Wistar; Receptors, Angiotensin; Receptors, Endothelin; RNA, Messenger; Transforming Growth Factor beta; Ventricular Function

Topics: Animals; Animals, Newborn; Atrial Natriuretic Factor; Cardiomegaly; Cells, Cultured; Gene Expression Regulation; Genes, ras; Genes, Reporter; Genes, src; Hypertrophy; Luciferases; Myocardium; Phenotype; Promoter Regions, Genetic; Rats

Atrial natriuretic peptide (ANP) inhibits the growth of a variety of cell types in vitro including mesangial cells. The effects of ANP on the growth of the kidney in vivo were evaluated. A 2-h infusion of 0.2 microgram/250 g body wt per minute of ANP suppressed the subsequent uptake of [3H]thymidine into the renal DNA of uninephrectomized but not intact rats. This treatment also depressed the ratio of RNA/DNA in kidneys undergoing compensatory growth. Correlative physiologic studies revealed enhanced GFR in rats with two kidneys infused with ANP, but no increase in the GFR of uninephrectomized rats. It was concluded that ANP may oppose the growth factor(s) mediating compensatory renal growth.

Topics: Animals; Atrial Natriuretic Factor; Body Weight; DNA; Glomerular Filtration Rate; Hypertrophy; Kidney; Male; Mitosis; Nephrectomy; Organ Size; Rats; Rats, Sprague-Dawley

Mechanical loading and alpha-adrenergic receptor stimulation have both been shown to induce hypertrophy in isolated neonatal heart cells. The present study examined the effects of adrenergic hormones and contractile activity on the hypertrophic response in isolated adult feline cardiomyocytes maintained for more than 14 days in insulin- and serum-supplemented medium. Measurements of the hypertrophic response included cell size, total protein content, myosin heavy chain content, and the time course of activation of increased protein synthesis. Reactivation of the "fetal" gene program was evaluated by secretion of atrial natriuretic factor (ANF) into the medium. Significant myocyte hypertrophy was induced in both quiescent myocytes treated with alpha 1-adrenergic agonists and in beating myocytes treated with beta-adrenergic agonists. However, there were both quantitative and qualitative differences in the response to each type of stimulation. alpha-Adrenergic agonists promoted an increase in cell size, protein content, and ANF secretion but not myofibrillar reorganization, which was observed only in beating myocytes. In contrast to results reported for neonatal heart cells, determinants of hypertrophy in beating myocytes exceeded those in nonbeating alpha 1-adrenergic agonist-treated heart cells in every parameter examined. In addition, in the case of both beating and alpha-adrenergic stimulation, there were marked time-dependent variations in rates of protein synthesis over the interval of 4 hours to 7 days of treatment with each type of stimulus. Differences were also encountered in correlations between rates of protein synthesis and protein accumulation over this interval. The effect of beating was particularly important both to the reorganization of myofibrillar structure and the metabolism of myosin heavy chain. In cultures in which beating was inhibited with the calcium channel antagonist nifedipine, the loss of myosin heavy chain was significantly greater than that of total protein.

Topics: Animals; Atrial Natriuretic Factor; Atrophy; Blood Physiological Phenomena; Cats; Cell Division; Cells, Cultured; Culture Media; Hypertrophy; Insulin; Muscle Proteins; Myocardial Contraction; Myocardium; Myosins; Sympathomimetics; Time Factors

Hypertrophy is a fundamental adaptive process employed by postmitotic cardiac and skeletal muscle in response to mechanical load. How muscle cells convert mechanical stimuli into growth signals has been a long-standing question. Using an in vitro model of load (stretch)-induced cardiac hypertrophy, we demonstrate that mechanical stretch causes release of angiotensin II (Ang II) from cardiac myocytes and that Ang II acts as an initial mediator of the stretch-induced hypertrophic response. The results not only provide direct evidence for the autocrine mechanism in load-induced growth of cardiac muscle cells, but also define the pathophysiological role of the local (cardiac) renin-angiotensin system.

Topics: Actins; Angiotensin I; Angiotensin II; Angiotensinogen; Animals; Animals, Newborn; Atrial Natriuretic Factor; Cardiomegaly; Cells, Cultured; Cytoplasmic Granules; Endothelins; Gene Expression Regulation; Genes, fos; Hypertrophy; In Vitro Techniques; Mechanoreceptors; Myocardium; Peptidyl-Dipeptidase A; Rats; Renin; RNA, Messenger; Stress, Mechanical

Calmodulin (CaM) levels are developmentally regulated in the mouse heart. During late gestational and early postnatal stages, CaM levels decline several-fold in close temporal association with the declining population of proliferating cardiomyocytes. This correlation suggests that CaM may influence cardiomyocyte cell cycle activity, particularly since CaM is implicated in cell cycle control in several eukaryotic nonmuscle cells. To test this possibility, nucleotides -500 to 77 of the human atrial natriuretic factor gene were linked to a chicken CaM minigene to establish two pedigrees of transgenic mice that express 3- to 5-fold increased levels of CaM in cardiomyocytes. Developmental overexpression of CaM in mouse cardiomyocytes produced a markedly exaggerated cardiac growth response, characterized by the presence of cardiomyocyte hypertrophy in regions demonstrated to overexpress CaM and by cardiomyocyte hyperplasia, apparent at early developmental stages. Early postnatal suppression of fusion gene expression in the cardiac ventricles correlated with regression of the ventricular growth response in transgenic relative to nontransgenic mice between 3 days and 6-10 weeks of age, but was not apparent in the cardiac atria, where levels of CaM remained constitutively elevated until advanced stages. To test the possibility that increased cytosolic Ca2+ buffering contributes to the growth response induced by CaM over-expression, two additional lines of transgenic mice were generated using the same human atrial natriuretic factor promoter to target expression of a CaM mutant (amino acids 75-82 deleted) in cardiomyocytes. This mutant has previously been shown to bind Ca2+ with kinetic properties similar to those of wild-type CaM, but was unable to activate several CaM-dependent target enzymes in vitro. Despite high level expression of the CaM mutant, no growth response was apparent in the hearts of transgenic relative to those of nontransgenic mice, suggesting that increased Ca2+ buffering is unlikely to contribute to the growth response induced by CaM overexpression. Taken together, these findings reveal that cardiomyocyte growth regulation is specifically influenced by CaM concentrations in transgenic mice.

Topics: Animals; Atrial Natriuretic Factor; Calmodulin; Cell Division; Gene Expression; Heart Atria; Heart Ventricles; Humans; Hypertrophy; Mice; Mice, Inbred ICR; Mice, Transgenic; Microscopy, Electron; Mutagenesis, Site-Directed; Myocardium; Organ Size; Recombinant Fusion Proteins

In the present study we investigated the in vivo and in vitro renal responsiveness to ANF, and the adaptation of ANF receptors in compensatory renal hypertrophy in the rat. One week after left nephrectomy (UNx), plasma levels of immunoreactive ANF, blood pressure (MAP), hematocrit (Hct), and urine flow rate (V) were unaltered compared to control (C) rats. Baseline GFR and potassium excretion (UKV) were significantly higher, and sodium excretion (UNaV) tended to be elevated in UNx rats. Administered ANF led to similar dose-related decreases in MAP and increases in Hct in UNx and C rats. However, at each dose of infused ANF, absolute values and the increase in GFR and UNaV were higher in UNx than in C rats. Hypertrophied (H) kidneys were removed from UNx and perfused in vitro to determine distribution and density of ANF receptors, responsiveness to ANF, and receptor-mediated organ clearance of 125I-ANF1-28. The density of ANF receptors in cortex, outer medulla, and papilla of H kidneys was not significantly different from that in C kidneys. In H isolated kidneys, ANF led to dose-related increases in GFR, V, UNaV, and UKV that were indistinguishable (P greater than 0.05) from those in C kidneys. Receptor-mediated organ clearance of 125I-ANF1-28 in isolated H kidneys was 2.8 +/- .02 ml/min, a value not significantly different (P greater than 0.05) from that in C kidneys.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Atrial Natriuretic Factor; Glomerular Filtration Rate; Hypertrophy; In Vitro Techniques; Kidney; Male; Natriuresis; Nephrectomy; Perfusion; Rats; Rats, Inbred Strains; Receptors, Atrial Natriuretic Factor; Receptors, Cell Surface

Seven-month-old, lean male SHHF/Mcc-cp rats, a model of spontaneous hypertension, progressive renal dysfunction, and congestive heart failure (CHF), were treated with either clonidine (CL) or enalapril (EN) or received no treatment (CON) for 20 weeks. CL significantly decreased systolic blood pressure (SBP), kidney weights, and severity of renal lesions as compared with untreated CON. EN produced a decrease in SBP comparable to that in CL. Kidney weights and severity of renal histologic changes in the EN group were intermediate between those of the CL and CON groups. Despite similar plasma atrial natriuretic peptide (ANP) concentrations, CL treatment resulted in a significant increase in the density of guanylate cyclase-linked glomerular ANP receptors, whereas EN treatment resulted in a significant decrease in the total number of ANP receptors and in the number of nonguanylate cyclase-linked receptors and an increase in overall binding affinity. These findings demonstrate that antihypertensive agents will slow progression of renal injury in SHHF/Mcc-cp rats and that CL is more effective than EN in alleviating progressive kidney damage in this model. Furthermore, different classes of antihypertensive drugs may alter the density or ratio of biologically active and clearance ANP receptor sites in the glomerulus.

Topics: Animals; Atrial Natriuretic Factor; Binding Sites; Clonidine; Cyclic GMP; Enalapril; Hypertension; Hypertrophy; Kidney; Kidney Diseases; Kidney Glomerulus; Male; Rats; Rats, Inbred Strains; Receptors, Atrial Natriuretic Factor; Receptors, Cell Surface

Plasma concentrations of immunoreactive (ir) atrial (ANP) and brain (BNP) natriuretic peptides were measured in the prehypertensive and hypertensive phases in spontaneously hypertensive rats (SHR) and in the malignant phase of hypertension caused by deoxycorticosterone acetate (DOCA)-salt in SHR. The secretory rate of ANP and BNP were examined in the perfusion of isolated beating heart before and after atrial removal. Plasma irANP and irBNP in mature SHR were higher than those of control Wistar-Kyoto (WKY) rats, whereas ANP and BNP values in young SHR did not differ from those of control WKY rats. DOCA-salt treatment for 8 weeks markedly increased blood pressure, ventricular weight, and plasma irANP and irBNP in SHR. ANP and BNP values were positively correlated with ventricular weight in DOCA-salt SHR. The secretory rate of ANP and BNP from the perfused whole heart were much higher in DOCA-salt SHR than other rat groups. A large amount of BNP was secreted from the hypertrophied ventricles in DOCA-salt SHR. In contrast, ANP was mainly secreted from the atrium in all rat groups. High-performance liquid chromatography profiles of extract in plasma showed that a major component of irANP and irBNP corresponded to synthetic rat ANP-(1-28) and rat BNP-45, respectively. Results suggest that both rat ANP-(1-28) and rat BNP-45 are markedly increased in plasma in DOCA-salt-induced malignant hypertension of SHR and that the major source of circulating BNP is the hypertrophied ventricles in this model.

Topics: Aging; Animals; Atrial Natriuretic Factor; Cerebral Ventricles; Chromatography, High Pressure Liquid; Desoxycorticosterone; Hypertension, Malignant; Hypertrophy; In Vitro Techniques; Myocardium; Natriuretic Peptide, Brain; Nerve Tissue Proteins; Osmolar Concentration; Radioimmunoassay; Rats; Rats, Inbred SHR; Rats, Inbred WKY; Sodium Chloride

Vascular remodeling is central to the pathophysiology of hypertension and atherosclerosis. Recent evidence suggests that vasoconstrictive substances, such as angiotensin II (AII), may function as a vascular smooth muscle growth promoting substance. To explore the role of the counterregulatory hormone, atrial natriuretic polypeptide (ANP) in this process, we examined the effect of ANP (alpha-rat ANP [1-28]) on the growth characteristics of cultured rat aortic smooth muscle (RASM) cells. ANP (10(-7) M) significantly suppressed the proliferative effect of 1% and 5% serum as measured by 3H-thymidine incorporation and cell number, confirming ANP as an antimitogenic factor. In quiescent RASM cells, ANP (10(-7), 10(-6) M) significantly suppressed the basal incorporations of 3H-uridine and leucine by 50 and 30%, respectively. ANP (10(-7), 10(-6) M) also suppressed AII-induced RNA and protein syntheses (by 30-40%) with the concomitant reduction of the cell size. Furthermore, ANP also significantly attenuated the increase of 3H-uridine and leucine incorporations caused by transforming growth factor-beta (4 x 10(-11), 4 x 10(-10) M), a potent hypertrophic factor. These results indicate that ANP possesses an antihypertrophic action on vascular smooth muscle cells. Down-regulation of protein kinase C by 24-h treatment with phorbol 12,13-dibutyrate did not inhibit ANP-induced suppression on 3H-uridine incorporation. Based on the observation that ANP was more potent than a ring-deleted analogue of ANP on inhibiting 3H-uridine incorporation, we conclude that the ANP's inhibitory effect is primarily mediated via the activation of a guanylate cyclase-linked ANP receptor(s). Indeed 8-bromo cGMP mimicked the antihypertrophic action of ANP. Accordingly, we speculate that in addition to its vasorelaxant and natriuretic effects, the antihypertrophic action of ANP observed in the present study may serve as an additional compensatory mechanism of ANP in hypertension.

Topics: Angiotensin II; Animals; Aorta; Atrial Natriuretic Factor; Cell Division; Cell Line; Cyclic GMP; Down-Regulation; Hypertrophy; Muscle, Smooth, Vascular; Protein Kinase C; Rats; Receptors, Atrial Natriuretic Factor; Receptors, Cell Surface; Transforming Growth Factor beta

Topics: Angiotensin II; Atrial Natriuretic Factor; Bartter Syndrome; Humans; Hyperaldosteronism; Hypertrophy; Hypokalemia; Juxtaglomerular Apparatus; Kallikreins; Kidney Tubules; Kinins; Prostaglandins

In the renal compensatory adaptation after the definition and the description of the fundamental phenomena of the functional compensation as well as of the structural adaptation is reported on mediator concepts and on modulations of the renal adaptation processes. Issuing from the central position of the sodium balance a mediator concept on natriuretic hormones (Auriculin and Endoxin) is developed which is supplemented by the renotropin mediator concept. The authors deal with the modulation of the renal compensatory adaptation (e.g. influences of age diet and so on). The pharmacotherapeutic modulation of the renal compensatory adaptation is discussed with regard to the stimulation of the tubulosecretory transport of foreign substances with para-amino hippuric acid as principal substance (including own investigations with cyclopenthiazide [Benesal].

Topics: Adaptation, Physiological; Animals; Atrial Natriuretic Factor; Blood Proteins; Cardenolides; Digoxin; Feeding Behavior; Glomerular Filtration Rate; Growth Substances; Humans; Hypertrophy; Intercellular Signaling Peptides and Proteins; Kidney; Kidney Function Tests; Kidney Transplantation; Nephrectomy; Saponins; Sodium-Potassium-Exchanging ATPase; Water-Electrolyte Balance