u-0126 and Cardiomegaly

Topics: Acetylation; Anacardic Acids; Animals; Butadienes; Cardiomegaly; Cell Survival; Disease Models, Animal; Female; Histone Acetyltransferases; Histones; Male; MAP Kinase Signaling System; MEF2 Transcription Factors; Mice; Myocytes, Cardiac; Nitriles; p300-CBP Transcription Factors; Phenylephrine; Signal Transduction

Hypertrophic cardiomyopathy occurs along with pathological phenomena such as cardiac hypertrophy, myocardial fibrosis and cardiomyocyte activity. However, few of the specific molecular mechanisms underlying this pathological condition have been mentioned.. All target proteins and markers expression in the study was verified by PCR and western bloting. H9c2 cell morphology and behavior were analyzed using immunofluorescent and proliferation assays, respectively. And, the CTGF protein secreted in cell culture medium was detected by ELISA.. We found that high expression of CTGF and low expression of EGFR were regulated by ERK1/2 signaling pathway during the cardiac hypertrophy induced by Ang-II stimulation. CTGF interacted with EGFR, and the interaction is reduced with the stimulation of Ang-II. ERK1/2 serves as the center of signal control during the cardiac hypertrophy.. The ERK1/2 cooperates with GPCR and EGFR signaling, and promotes the occurrence and development of cardiac hypertrophy by regulating the expression and binding states of CTGF and EGFR. The study revealed a regulation model based on ERK1/2, suggesting that ERK1/2 signaling pathway may be an important control link for mitigation of hypertrophic cardiomyopathy treatment.

Topics: Angiotensin II; Animals; Butadienes; Cardiomegaly; Cardiomyopathy, Hypertrophic; Cell Enlargement; Cell Line; Connective Tissue Growth Factor; Disease Models, Animal; ErbB Receptors; Heart Ventricles; MAP Kinase Signaling System; Myocytes, Cardiac; Nitriles; Phosphorylation; Rats; Receptors, G-Protein-Coupled; Signal Transduction

In cardiac wound healing following myocardial infarction (MI), relatively inactive resident cardiac fibroblasts phenoconvert to hypersynthetic/secretory myofibroblasts that produce large quantities of extracellular matrix (ECM) and fibrillar collagen proteins. Our laboratory and others have identified TGFβ1 as being a persistent stimulus in the chronic and inappropriate wound healing phase that is marked by hypertrophic scarring and eventual stiffening of the entire myocardium, ultimately leading to the pathogenesis of heart failure following MI. Ski is a potent negative regulator of TGFβ/Smad signaling with known antifibrotic effects. Conversely, Scleraxis is a potent profibrotic basic helix-loop-helix transcription factor that stimulates fibrillar collagen expression. We hypothesize that TGFβ1 induces Scleraxis expression by a novel Smad-independent pathway. Our data support the hypothesis that Scleraxis expression is induced by TGFβ1 through a Smad-independent pathway in the cardiac myofibroblast. Specifically, we demonstrate that TGFβ1 stimulates p42/44 (Erk1/2) kinases, which leads to increased Scleraxis expression. Inhibition of MEK1/2 using U0126 led to a sequential temporal reduction of phospho-p42/44 and subsequent Scleraxis expression. We also found that adenoviral Ski expression in primary myofibroblasts caused a significant repression of endogenous Scleraxis expression at both the mRNA and protein levels. Thus we have identified a novel TGFβ1-driven, Smad-independent, signaling cascade that may play an important role in regulating the fibrotic response in activated cardiac myofibroblasts following cardiac injury.

Topics: 3T3 Cells; Animals; Basic Helix-Loop-Helix Transcription Factors; Butadienes; Cardiomegaly; Chlorocebus aethiops; COS Cells; Fibrosis; Male; MAP Kinase Signaling System; Mice; Mitogen-Activated Protein Kinase 1; Myocytes, Cardiac; Nitriles; Primary Cell Culture; Proto-Oncogene Proteins; Rats; Rats, Sprague-Dawley; Signal Transduction; Smad Proteins; Transforming Growth Factor beta1

Evidence suggests that upregulation of soluble epoxide hydrolase (sEH) is associated with the development of myocardial infarction, dilated cardiomyopathy, cardiac hypertrophy, and heart failure. However, the upregulation mechanism is still unknown. In this study, we treated H9C2 cells with buthionine sulfoximine (BSO) to explore whether oxidative stress upregulates sEH gene expression and to identify the molecular and cellular mechanisms behind this upregulatory response. Real-time PCR and Western blot analyses were used to measure mRNA and protein expression, respectively. We demonstrated that BSO significantly upregulated sEH at mRNA levels in a concentration- and time-dependent manner, leading to a significant increase in the cellular hypertrophic markers, atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP). Furthermore, BSO significantly increased the cytosolic phosphorylated IκB-α and translocation of NF-κB p50 subunits, as measured by Western blot analysis. This level of translocation was paralleled by an increase in the DNA-binding activity of NF-κB P50 subunits. Moreover, our results demonstrated that pretreatment with the NF-κB inhibitor PDTC significantly inhibited BSO-mediated induction of sEH and cellular hypertrophic marker gene expression in a dose-dependent manner. Additionally, mitogen-activated protein kinases (MAPKs) were transiently phosphorylated by BSO treatment. To understand further the role of MAPKs pathway in BSO-mediated induction of sEH mRNA, we examined the role of extracellular signal-regulated kinase (ERK), c-JunN-terminal kinase (JNK), and p38 MAPK. Indeed, treatment with the MEK/ERK signal transduction inhibitor, PD98059, partially blocked the activation of IκB-α and translocation of NF-κB p50 subunits induced by BSO. Moreover, pretreatment with MEK/ERK signal transduction inhibitors, PD98059 and U0126, significantly inhibited BSO-mediated induction of sEH and cellular hypertrophic marker gene expression. These results clearly demonstrated that the NF-κB signaling pathway is involved in BSO-mediated induction of sEH gene expression, and appears to be associated with the activation of the MAPK pathway. Furthermore, our findings provide a strong link between sEH-induced cardiac dysfunction and involvement of NF-κB in the development of cellular hypertrophy.

Topics: Animals; Antioxidants; Atrial Natriuretic Factor; Butadienes; Buthionine Sulfoximine; Cardiomegaly; Cell Line; Cell Survival; Enzyme Activation; Epoxide Hydrolases; Extracellular Signal-Regulated MAP Kinases; Flavonoids; Gene Expression Regulation; Glutathione; Heart Failure; I-kappa B Proteins; JNK Mitogen-Activated Protein Kinases; MAP Kinase Signaling System; Myoblasts, Cardiac; Natriuretic Peptide, Brain; NF-kappa B p50 Subunit; NF-KappaB Inhibitor alpha; Nitriles; Oxidative Stress; p38 Mitogen-Activated Protein Kinases; Phosphorylation; Proline; Rats; RNA, Messenger; Thiocarbamates; Transcription Factor RelA; Up-Regulation

Activation of Ca2+/Calmodulin protein kinase II (CaMKII) is an important step in signaling of cardiac hypertrophy. The molecular mechanisms by which CaMKII integrates with other pathways in the heart are incompletely understood. We hypothesize that CaMKII association with extracellular regulated kinase (ERK), promotes cardiac hypertrophy through ERK nuclear localization.. In H9C2 cardiomyoblasts, the selective CaMKII peptide inhibitor AntCaNtide, its penetratin conjugated minimal inhibitory sequence analog tat-CN17β, and the MEK/ERK inhibitor UO126 all reduce phenylephrine (PE)-mediated ERK and CaMKII activation and their interaction. Moreover, AntCaNtide or tat-CN17β pretreatment prevented PE induced CaMKII and ERK nuclear accumulation in H9C2s and reduced the hypertrophy responses. To determine the role of CaMKII in cardiac hypertrophy in vivo, spontaneously hypertensive rats were subjected to intramyocardial injections of AntCaNtide or tat-CN17β. Left ventricular hypertrophy was evaluated weekly for 3 weeks by cardiac ultrasounds. We observed that the treatment with CaMKII inhibitors induced similar but significant reduction of cardiac size, left ventricular mass, and thickness of cardiac wall. The treatment with CaMKII inhibitors caused a significant reduction of CaMKII and ERK phosphorylation levels and their nuclear localization in the heart.. These results indicate that CaMKII and ERK interact to promote activation in hypertrophy; the inhibition of CaMKII-ERK interaction offers a novel therapeutic approach to limit cardiac hypertrophy.

Topics: Animals; Butadienes; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Cardiomegaly; Gene Expression Regulation; Heart; Humans; Mitogen-Activated Protein Kinase 3; Myoblasts, Cardiac; Nitriles; Phenylephrine; Phosphorylation; Rats

IL-6 has been shown to play a major role in collagen up-regulation process during cardiac hypertrophy, although the precise mechanism is still not known. In this study we have analyzed the mechanism by which IL-6 modulates cardiac hypertrophy. For the in vitro model, IL-6-treated cultured cardiac fibroblasts were used, whereas the in vivo cardiac hypertrophy model was generated by renal artery ligation in adult male Wistar rats (Rattus norvegicus). During induction of hypertrophy, increased phosphorylation of STAT1, STAT3, MAPK, and ERK proteins was observed both in vitro and in vivo. Treatment of fibroblasts with specific inhibitors for STAT1 (fludarabine, 50 μM), STAT3 (S31-201, 10 μM), p38 MAPK (SB203580, 10 μM), and ERK1/2 (U0126, 10 μM) resulted in down-regulation of IL-6-induced phosphorylation of specific proteins; however, only S31-201 and SB203580 inhibited collagen biosynthesis. In ligated rats in vivo, only STAT3 inhibitors resulted in significant decrease in collagen synthesis and hypertrophy markers such as atrial natriuretic factor and β-myosin heavy chain. In addition, decreased heart weight to body weight ratio and improved cardiac function as measured by echocardiography was evident in animals treated with STAT3 inhibitor or siRNA. Compared with IL-6 neutralization, more pronounced down-regulation of collagen synthesis and regression of hypertrophy was observed with STAT3 inhibition, suggesting that STAT3 is the major downstream signaling molecule and a potential therapeutic target for cardiac hypertrophy.

Topics: Animals; Antineoplastic Agents; Butadienes; Cardiomegaly; Cells, Cultured; Collagen; Disease Models, Animal; Down-Regulation; Enzyme Inhibitors; Extracellular Signal-Regulated MAP Kinases; Fibroblasts; Humans; Imidazoles; Interleukin-6; Male; MAP Kinase Signaling System; Nitriles; p38 Mitogen-Activated Protein Kinases; Phosphorylation; Pyridines; Rats; Rats, Wistar; STAT1 Transcription Factor; STAT3 Transcription Factor; Vidarabine

Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) and extracellular signal-regulated kinase (ERK) have pivotal roles in endothelin-1 (ET-1)-induced cardiomyocyte hypertrophy. We here tested whether a novel CaM antagonist, DY-9760e inhibits ET-1-induced hypertrophy through inhibition of CaMKII and ERK activities. We first confirmed that Ca(2+) oscillation induced by ET-1 treatment elicits transient activation of CaMKII and ERK in cultured cardiomyocytes. DY-9760e treatment with 3 microM totally and partially inhibited the ET-1-induced CaMKII and ERK activation, respectively. The ET-1-induced ERK activation was also partially blocked by a CaMKII inhibitor, KN93. To confirm involvement of CaMKII activity in the ERK activation by ET-1 and A23187, cultured cardiomyocytes were transfected with a constitutively active CaMKII. The transfection with the active CaMKII elicited ERK activation in cultured cardiomyocytes and cotransfection with dominant negative CaMKII eliminated its ERK activation. Consistent with inhibitory actions of DY-9760e on the ET-1-induced CaMKII and ERK activation, induction of hypertrophy-related genes including atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) was significantly inhibited by DY-9760e treatment. Combination treatment with DY-9760e and U0126, a MEK inhibitor, totally blocked the ET-1-induced ANP and BNP expression. DY-9760e treatment (3 microM) significantly inhibited the ET-1-induced hypertrophy and combination treatment with DY-9760e and U0126 totally blocked the ET-1-induced hypertrophy in cultured cardiomyocytes. These results suggest that DY-9760e elicits antihypertrophic action on ET-1-induced cardiac hypertrophy through inhibition of CaMKII and ERK activation and that CaMKII activity in part mediates ET-1-induced ERK activation.

Topics: Animals; Animals, Newborn; Atrial Natriuretic Factor; Benzylamines; Butadienes; Calcimycin; Calcium Signaling; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Cardiomegaly; Cell Proliferation; Cell Size; Cells, Cultured; DNA Replication; Endothelin-1; Extracellular Signal-Regulated MAP Kinases; Indazoles; Ionophores; Myocytes, Cardiac; Natriuretic Peptide, Brain; Nitriles; Phosphorylation; Protein Kinase Inhibitors; Rats; Rats, Wistar; RNA, Messenger; Sulfonamides; Time Factors; Transfection

The purpose of this study was to investigate whether prostaglandin E2 (PGE2) induces Signal transducer and activator of transcription 3 (Stat3) activation in neonatal rat ventricular cardiomyocytes and if so to determine the possible role of this activation in PGE2-induced hypertrophic responses.. Stat3 activation and its nuclear phosphorylation were determined by electrophoretic mobility shift assay (EMSA) and by Western blots, respectively. Protein synthesis was assessed by [3H]-leucine incorporation into total protein and cell surface was quantified by microscopic analysis.. We found that PGE2 induces a concentration- (1-100 nM) and time-dependent increase in Stat3 activation, reaching maximal values after 90 min of stimulation. Experiments with agonists and antagonists of the PGE2 receptor subtypes EP1-EP4 indicate that PGE2 activates Stat3 mainly through the EP4 receptor. We further observed that the extracellular signal-regulated kinase 1/2 (ERK1/2) inhibitor U0126 abolishes PGE2-induced Stat3 activation whereas the p38 MAP kinase blocker SB203580 has no effect. Nuclear Stat3 phosphorylation induced by PGE2 is also suppressed by the translation and transcription inhibitors, cycloheximide and actinomycin D, respectively. Transfecting ventricular cardiomyocytes with a small interfering RNA (siRNA) targeting rat Stat3, we obtained an approximately 70% reduction in Stat3 expression, 24 and 48 h after electroporation. In these Stat3-silenced cells, the PGE2-induced increase in protein synthesis and cell surface is strongly inhibited.. In ventricular cardiomyocytes, PGE2 induces the activation of Stat3 which plays an essential role in PGE2-induced increase in cell size and protein synthesis. The activation of Stat3 occurs mainly through EP4 and involves ERK1/2 as well as newly synthesized protein(s).

Topics: Animals; Animals, Newborn; Blotting, Western; Butadienes; Cardiomegaly; Cells, Cultured; Cycloheximide; Dactinomycin; Dinoprostone; Electrophoretic Mobility Shift Assay; Enzyme Inhibitors; Extracellular Signal-Regulated MAP Kinases; Gene Silencing; Genistein; Imidazoles; Janus Kinases; Myocytes, Cardiac; Nitriles; Phosphorylation; Protein Synthesis Inhibitors; Pyridines; Rats; Rats, Wistar; Receptors, Prostaglandin E; Receptors, Prostaglandin E, EP4 Subtype; RNA Interference; Signal Transduction; STAT3 Transcription Factor; Stress, Mechanical

Alpha2-macroglobulin (alpha2M) is an acute phase protein released to the serum upon challenges such as cardiac hypertrophy and infarction. Here we report on the role of alpha2M in the induction of hypertrophic cell growth, contractile responsiveness of rat ventricular cardiomyocytes, and on the underlying extracellular regulated kinase 1,2 (ERK1,2) and phosphoinositide 3-kinase (PI3-kinase)/Akt pathways.. Cell volume and cross-sectional areas were assessed as parameters of hypertrophic growth, and real time RT-PCR for the analysis of hypertrophy-related genes was performed. Protein synthesis was analyzed by 14C-phenylalanine incorporation. Activation of ERK1,2, PI3-kinase and Akt was assessed by immunohistochemical analysis of phosphorylated proteins. Contractile responsiveness was investigated by determination of cell shortening following electrical field stimulation. Intracellular calcium concentration [Ca2+]i was determined by fluo-3 microfluorometry.. Treatment of ventricular cardiomyocytes for 24 h with alpha2M significantly increased cell volume and protein synthesis as well as expression of hypertrophy-associated genes [brain natriuretic protein (BNP), beta-myosin heavy chain (beta-MHC), myosin light chain-2 (MLC-2), atrial natriuretic factor (ANF), and skeletal alpha-actin]. Comparable effects were achieved by treatment of cells with an antibody directed against the alpha2M-receptor LDL receptor-related protein-1 (LRP-1) and counteracted upon coincubation with receptor-associated protein (RAP), suggesting an involvement of alpha2M-LRP-1 signalling. Furthermore, alpha2M treatment increased sarcoplasmic reticulum Ca2+-ATPase (SERCA-2a) expression, diastolic and systolic [Ca2+]i, and contractile responsiveness after electrical stimulation. Shortly after alpha2M stimulation, activation of ERK1,2, Akt, and PI3-kinase pathways was observed. Consequently, alpha2M-induced protein synthesis was inhibited upon treatment with the ERK1,2 inhibitor UO126 as well as by LY294002 and wortmannin, which inhibit PI3-kinase, and by rapamycin, which inhibits mammalian target of rapamycin (mTOR) downstream of Akt.. Our data show that alpha2M induces hypertrophic cell growth in rat ventricular cardiomyocytes via ERK1,2 and PI3-kinase/Akt and improves cardiac cell function.

Topics: alpha-Macroglobulins; Androstadienes; Animals; Butadienes; Calcium; Cardiomegaly; Cells, Cultured; Chromones; Enzyme Activation; Immunohistochemistry; Male; Morpholines; Nitriles; Phenylalanine; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; Rats; Rats, Wistar; Reverse Transcriptase Polymerase Chain Reaction; Sarcoplasmic Reticulum Calcium-Transporting ATPases; Signal Transduction; Sirolimus; Wortmannin

To assess the contribution of signal transducer and activator of transcription 3 (JAK-STAT3) pathway, extracellular signal-regulated kinases1/2 (ERK1/2) pathway, and phosphatidylinositol 3-kinase (PI3-K) pathway to cardiomyocytes hypertrophy induced by cardiotrophin-1 (CT-1), a new member of interleukin-6 (IL-6) family of cytokines.. STAT3, ERK1/2, and PI3-K were assessed by Western blot analysis. Activity of ERK1/2 was also confirmed by in-gel kinase assay. Hypertrophy of cardiomyocyte was evaluated by [3H]leucine incorporation and cellular protein-to-DNA ratio.. CT-1 simultaneously activated phosphorylation of STAT3, ERK1/2, and PI3-K in rat cardiomyocytes. Parthenolide, an inhibitor of STAT, suppressed CT-1-induced [3H]leucine incorporation by 88.3 % and protein-to-DNA ratio by 75.0 %. U0126, an MEK1/2 inhibitor, increased CT-1-induced the phosphorylation of STAT3 in a dose-dependent manner and, consistently, augmented CT-1-induced increase in [3H]leucine incorporation and cellular protein-to-DNA ratio by 17.6 % and 16.3 %, respectively. Wortmannin, a PI3-K inhibitor, did not influence CT-1-induced [3H]leucine incorporation and cellular protein-to-DNA ratio.. The hypertrophic effect of CT-1 was essentially mediated by STAT3, independent of PI3-K, and negatively regulated by ERK1/2 via inhibiting the phosphorylation of STAT3. The interaction between STAT3 and ERK1/2 in CT-1-induced signaling contributes to development of cardiac hypertrophy.

Topics: Androstadienes; Animals; Animals, Newborn; Butadienes; Cardiomegaly; Cell Size; Cells, Cultured; Cytokines; DNA-Binding Proteins; Heart Ventricles; Leucine; MAP Kinase Kinase 1; Mitogen-Activated Protein Kinase 3; Myocytes, Cardiac; Nitriles; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Phosphorylation; Rats; Rats, Sprague-Dawley; Sesquiterpenes; Signal Transduction; STAT3 Transcription Factor; Trans-Activators; Wortmannin

Extracellular-signal-regulated protein kinases (ERKs) are activated rapidly and transiently in response to phenylephrine (PE) and endothelin-1 (ET-1) in cardiac myocytes, but whether this is linked to the subsequent development of the hypertrophic phenotype remains equivocal. To investigate this, we examined the dependence of the hypertrophic response on the length of exposure to PE in neonatal myocyte cultures. In addition to the initial transient activation of ERKs (maximum at 5-10 min), PE (10 microM) induced a second, more prolonged peak of activity several hours later. The activity of a transfected atrial natriuretic factor-luciferase reporter gene was increased 10- to 24-fold by PE. This response was inhibited by the alpha(1)-antagonist prazosin (100 nM) and by U0126 (10 microM) and PD184352 (1 microM), inhibitors of ERK activation, irrespective of whether these were added before or up to 24 h after the addition of PE. Prazosin had no effect on ET-1 (50 nM)-stimulated atrial natriuretic factor-luciferase activity. Protein synthesis was enhanced by 35+/-6% by PE, and this was blocked by prazosin added 1 h after the addition of PE, but decreased only by half when added 8 h after PE. Similarly, PE (48 h) increased myocyte area by 49% and this was prevented by prazosin added 1 h after PE, but decreased only by half when added at 24 h. These results demonstrate that prolonged exposure to PE is required to elicit alterations in gene expression, protein synthesis and cell size, characteristic of hypertrophied myocytes, and they confirm that the initial peak of ERK activity is insufficient to trigger hypertrophic responses.

Topics: Adrenergic alpha-Antagonists; Animals; Atrial Natriuretic Factor; Benzamides; Butadienes; Cardiomegaly; Cell Size; Cells, Cultured; Endothelin-1; Enzyme Activation; Enzyme Inhibitors; Gene Expression Regulation; Luciferases; Mitogen-Activated Protein Kinases; Myocytes, Cardiac; Nitriles; Phenylephrine; Prazosin; Protein Biosynthesis; Proteins; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Time Factors

Recent studies indicate that cardiac T-type Ca2+ current (ICaT) reappears in hypertrophied ventricular cells. The aim of this study was to investigate the role of angiotensin II (Ang II), a major inducer of cardiac hypertrophy, in the reexpression of T-type channel in left ventricular hypertrophied myocytes. We induced cardiac hypertrophy in rats by abdominal aorta stenosis for 12 weeks and thereafter animals were treated for 2 weeks with losartan (12 mg/kg per day), an antagonist of type 1 Ang II receptors (AT1). In hypertrophied myocytes, we showed that the reexpressed ICaT is generated by the CaV3.1 and CaV3.2 subunits. After losartan treatment, ICaT density decreased from 0.40+/-0.05 pA/pF (n=26) to 0.20+/-0.03 pA/pF (n=27, P<0.01), affecting CaV3.1- and CaV3.2-related currents. The amount of CaV3.1 mRNA increased during hypertrophy and retrieved its nonhypertrophic level after losartan treatment, whereas the amount of CaV3.2 mRNA was unaffected by stenosis. In cultured newborn ventricular cells, chronic Ang II application (0.1 micromol/L) also increased ICaT density and CaV3.1 mRNA amount. UO126, a mitogen-activated protein kinase kinase-1/2 (MEK1/2) inhibitor, reduced Ang II-increased ICaT density and CaV3.1 mRNA amount. Bosentan, an endothelin (ET) receptor antagonist, reduced Ang II-increased ICaT density without affecting the amount of CaV3.1 mRNA. Finally, cotreatment with bosentan and UO126 abolished the Ang II-increased ICaT density. Our results show that AT1-activated MEK pathway and autocrine ET-activated independent MEK pathway upregulate T-type channel expression. Ang II-increased of ICaT density observed in hypertrophied myocytes may play a role in the pathogenesis of Ca2+ overload and arrhythmias seen in cardiac pathology.

Topics: Angiotensin II; Angiotensin Receptor Antagonists; Animals; Animals, Newborn; Bosentan; Butadienes; Calcium Channels, T-Type; Calcium-Calmodulin-Dependent Protein Kinases; Cardiomegaly; Constriction, Pathologic; Dose-Response Relationship, Drug; Endothelin Receptor Antagonists; Endothelin-1; Enzyme Inhibitors; Flavonoids; Gene Expression; Losartan; Male; Membrane Potentials; Mitogen-Activated Protein Kinase Kinases; Myocytes, Cardiac; Nickel; Nitriles; Oligopeptides; Peptides, Cyclic; Piperidines; Rats; Rats, Wistar; Receptors, Angiotensin; Receptors, Endothelin; RNA, Messenger; Signal Transduction; Sulfonamides

The extracellular signal-regulated kinase (ERK) pathway is activated by hypertrophic stimuli in cardiomyocytes. However, whether ERK plays an essential role or is implicated in all major components of cardiac hypertrophy remains controversial. Using a selective MEK inhibitor, U0126, and a selective Raf inhibitor, SB-386023, to block the ERK signaling pathway at two different levels and adenovirus-mediated transfection of dominant-negative Raf, we studied the role of ERK signaling in response of cultured rat cardiomyocytes to hypertrophic agonists, endothelin-1 (ET-1), and phenylephrine (PE). U0126 and SB-386023 blocked ET-1 and PE-induced ERK but not p38 and JNK activation in cardiomyocytes. Both compounds inhibited ET-1 and PE-induced protein synthesis and increased cell size, sarcomeric reorganization, and expression of beta-myosin heavy chain in myocytes with IC(50) values of 1-2 microm. Furthermore, both inhibitors significantly reduced ET-1- and PE-induced expression of atrial natriuretic factor. In cardiomyocytes transfected with a dominant-negative Raf, ET-1- and PE-induced increase in cell size, sarcomeric reorganization, and atrial natriuretic factor production were remarkably attenuated compared with the cells infected with an adenovirus-expressing green fluorescence protein. Taken together, our data strongly support the notion that the ERK signal pathway plays an essential role in ET-1- and PE-induced cardiomyocyte hypertrophy.

Topics: Animals; Base Sequence; Butadienes; Cardiomegaly; DNA Primers; Endothelin-1; Enzyme Inhibitors; Enzyme-Linked Immunosorbent Assay; MAP Kinase Signaling System; Mitogen-Activated Protein Kinases; Nitriles; Phenylephrine; Rats; Rats, Sprague-Dawley