The Dual-Specificity Phosphatase MKP-1 Limits the Cardiac Hypertrophic Response In Vitro and In Vivo
Abstract—Mitogen-activated protein kinase (MAPK) signaling pathways are important regulators of cell growth, proliferation, and stress responsiveness. A family of dual-specificity MAP kinase phosphatases (MKPs) act as critical counteracting factors that directly regulate the magnitude and duration of p38, c-Jun N-terminal kinase (JNK), and extracellular signal-regulated kinase (ERK) activation. Here we show that constitutive expression of MKP-1 in cultured primary cardiomyocytes using adenovirus-mediated gene transfer blocked the activation of p38, JNK1/2, and ERK1/2 and prevented agonist-induced hypertrophy. Transgenic mice expressing physiological levels of MKP-1 in the heart showed (1) no activation of p38, JNK1/2, or ERK1/2; (2) diminished developmental myocardial growth; and (3) attenuated hypertrophy in response to aortic banding and catecholamine infusion. These results provide further evidence implicating MAPK signaling factors as obligate regulators of cardiac growth and hypertrophy and demonstrate the importance of dual-specificity phosphatases as counterbalancing regulatory factors in the heart.
Cardiac hypertrophy is associated with the activation of numerous signal transduction factors, including G protein–coupled receptors, receptor tyrosine kinases, gp130, Ras, Rac-1, protein kinase C (PKC), calcineurin, and members of the mitogen-activated protein kinase (MAPK) signaling cascade.1 2 3 The MAPK signaling cascade consists of a hierarchy of kinases that ultimately phosphorylate and activate 1 of 3 main terminal effector kinases: p38s, c-Jun N-terminal kinases (JNKs), or extracellular signal-regulated kinases (ERKs).4 These terminal effector kinases are each regulated by reversible dual phosphorylation of adjacent Tyr and Thr residues within the motifs TGY, TPY, and TEY (p38, JNK, and ERK, respectively).4
In response to MAPK activation, a family of dual-specificity phosphatases become transcriptionally induced, leading to the specific dephosphorylation and inactivation of p38, JNK, and ERK signaling factors within 30 to 60 minutes.5 There are ≈9 dual-specificity phosphatase family members, each of which has a slightly different substrate specificity (p38 versus ERKs versus JNKs), tissue distribution, subcellular localization, or inducible expression profile.5 MKP-1 is an important member of the dual-specificity phosphatase family that is expressed in the heart, where it regulates inactivation of nuclear p38, JNK1/2, and ERK1/2.6 7 8 Transient transfection of an MKP-1–encoding expression vector into cardiomyocytes was shown to downregulate multiple hypertrophy-induced promoter constructs in vitro, suggesting that MAPK signaling cascades are important regulators of hypertrophic transcriptional responses.9 10
Although MAPK signaling factors are well established transducers of growth and stress responses in many cell types, considerable controversy exists concerning their importance as mediators of cardiomyocyte hypertrophy and developmental growth of the heart. A number of culture-based studies have demonstrated necessary roles for ERKs, p38, or JNKs as hypertrophic mediators.1 11 However, an almost equal number of culture-based studies have disputed the involvement of 1 or more of the MAPK signaling branches as necessary mediators of hypertrophy.1 11 In addition, very few studies have evaluated the role that MAPK signaling pathways might play in vivo, and almost nothing is known about the role that counteracting dual-specificity phosphatases play in regulating reactive MAPK signaling and the effects on cardiomyocyte hypertrophy. The results of the present study indicate that MAPK signaling factors are important regulators of both developmental cardiac hypertrophy and adult-onset hypertrophy in response to pathophysiological stimuli.
Materials and Methods
Primary Cardiomyocyte Cultures and Immunocytochemistry
Cardiomyocyte cultures were prepared from 1- to 2-day-old rat pups and processed for immunofluorescence as described previously.12 13 For quantification of atrial natriuretic factor (ANF) expression, cardiomyocytes showing intense perinuclear staining were counted from 15 randomly selective fields in 3 independent experiments. Cell surface areas from α-actinin–labeled cardiomyocytes were calculated as described previously.12 13 At least 100 cardiomyocytes in 15 to 25 fields were examined in 3 separate experiments.
Replication-Deficient Adenovirus and Transgenic Mouse Production
MKP-1 mouse cDNA (EST ai006592) was sequenced and cloned into pACCMVpLpA14 to permit adenovirus production. The construction, characterization, and procedures for cardiomyocyte infections with replication-deficient adenovirus were performed as previously described.12 13 14 The MKP-1 cDNA was also cloned into the 5.5-kb murine α-myosin heavy chain (MHC) promoter construct to permit transgenic mouse production (a gift of J. Robbins, University of Cincinnati, Cincinnati, Ohio).
SDS-PAGE and Immunoblot Analysis
The generation of protein extracts from cultured cardiomyocytes or heart tissue and their subsequent Western blotting were performed as described previously.12 13 Antibodies that were used included phospho-MEK1/2, phospho-MKK3/6, phospho-MKK4, phospho-ERK1/2, ERK1/2, phospho-p38, p38, phospho-JNK, JNK, and Akt (New England Biolabs) or MKP-1 (M18 and V15) (Santa Cruz).
Dot-Blot Analysis of Hypertrophic Markers
Cardiac gene expression of hypertrophic molecular markers was assessed by RNA dot-blot analysis as previously described.15 Hybridization signals were quantified with a Storm 860 PhosphorImager and ImageQuant software (Molecular Dynamics). Cardiac gene expression was normalized to GAPDH signal.
Murine Abdominal Aorta Constriction and Isoproterenol Infusion
Eight- to 10-week-old male mice (FVBN strain) from either genotype were anesthetized with isoflurane, and a laparotomy was performed. The abdominal aorta was isolated from annexed tissue, and the artery was partially ligated immediately below the celiac trunk with 6-0 silk around a 25-gauge blunted needle, which was subsequently removed to generate a calibrated constriction. Sham-operated mice went through the same procedure, except that the abdominal aorta was not stenosed.
Alzet osmotic minipumps (model 2002; Alza Corporation) that contained either isoproterenol (60 mg · kg–1 · d–1 for 2 weeks) or PBS were implanted dorsally and subcutaneously in 2-month-old mice (FVBN strain) of either genotype under isoflurane anesthesia without adverse effects or any deaths.
MKP-1 low-expressing transgenic mice or littermate wild-type mice at 8 weeks of age were anesthetized with 2% isoflurane, and echocardiography was performed with a Hewlett Packard Sonos 5500 instrument with a 15-MHz transducer. Cardiac ventricular dimensions and wall thicknesses were measured on 2-dimensional M-mode images at least 3 times for each animal.
Statistical analyses were performed between the experimental groups with a Student’s t test or 1-way ANOVA. Data are reported as mean±SEM. P<0.05 was considered significant.
AdMKP-1 Prevents Cardiomyocyte Hypertrophy In Vitro
To evaluate the ability of dual-specificity phosphatases to negatively regulate MAPK activation in cultured cardiomyocytes, an MKP-1–expressing adenovirus (AdMKP-1) was generated. Adenoviral infection routinely resulted in 98% to 100% infection of cultured neonatal cardiomyocytes, consistent with previous descriptions.12 13 AdMKP-1–infected cardiomyocytes demonstrated primarily nuclear MKP-1 localization, although some MKP-1 was also detected in the cytoplasm (data not shown). At 24 hours after infection, cardiomyocyte cultures were stimulated with the hypertrophic agonists phenylephrine (PE) or endothelin-1 (endo-1), and protein extracts were generated for MAPK Western blot analyses. Phosphorylation-specific antibodies demonstrated that p38, JNK1/2, and ERK1/2 are each activated by PE or endo-1 stimulation in control adenoviral β-galactosidase (Adβgal)-infected cultures at 30 minutes, 1 hour, and 4 hours (Figure 1A⇓ and data not shown). In contrast, AdMKP-1–infected cardiomyocytes demonstrated a loss of p38, JNK, and ERK1/2 phosphorylation at all time points investigated, whereas absolute protein levels of each MAPK factor were invariant (Figure 1A⇓). To verify the specificity of MKP-1, the phosphorylation status of MEK1, MKK4, and MKK3/6 was also examined by Western blotting. AdMKP-1 infection did not negatively affect the upstream MAPKK factors MEK1, MKK4, and MKK3/6 or affect Akt phosphorylation in PE-stimulated cardiomyocytes (Figure 1B⇓). These data suggest that MKP-1 is specific for the 3 MAPK terminal effectors and is not acting upstream in the MAPK cascade or nonspecifically on other signaling proteins (see Discussion).
The loss of agonist-induced MAPK activation was also associated with a complete inhibition of cardiomyocyte hypertrophy. Cardiomyocyte morphology, sarcomeric organization, and hypertrophy were assessed in α-actinin– and ANF-immunostained cultures. Adβgal-infected control cultures treated with PE or endo-1 (48 hours) demonstrated increased cell surface area, pronounced sarcomeric organization, and significant ANF expression (Figures 2C⇓ through 2F) compared with unstimulated cells (Figures 2A⇓ and 2B⇓). In contrast, AdMKP-1–infected cultures were refractory to PE- or endo-1–induced hypertrophy and ANF expression (Figures 2I⇓ through 2L). Quantification of these effects demonstrated that both PE and endo-1 significantly increased the 2-dimensional area of cardiomyocytes infected with Adβgal to 2827±79 and 2792±69 μm2, respectively (Figure 2M⇓). AdMKP-1–infected cardiomyocytes failed to increase in 2-dimensional area after either PE or endo-1 treatment (1288±41 and 1275±34 μm2, respectively) (P<0.05) (Figure 2M⇓).
We also observed that unstimulated AdMKP-1–infected cardiomyocytes were slightly smaller in appearance compared with Adβgal-infected cells (Figures 2G⇑ and 2H⇑ compared with Figures 2A⇑ and 2B⇑), suggesting a potential negative influence of the AdMKP-1 adenovirus. However, stimulation of AdMKP-1–infected cardiomyocytes with 10% FBS (multifactorial agonist) still promoted a significant increase in cell surface area and ANF expression compared with untreated AdMKP-1–infected cultures, suggesting that MKP-1 did not simply promote a “sick” phenotype (P<0.05) (Figures 2M⇑ and 2N⇑). Consistent with these data, AdMKP-1 infection did not induce TUNEL labeling or otherwise affect myocardial cell viability in agonist-stimulated or unstimulated cultures (data not shown). Last, both PE and endo-1 significantly increased ANF immunoreactivity in cardiomyocytes infected with Adβgal (34±2% and 57±3%, respectively), which was prevented by AdMKP-1 infection (6±1% and 8±2%, respectively) (P<0.05) (Figure 2N⇑). Taken together, these results indicate that constitutive MKP-1 expression prevents agonist-induced cardiomyocyte hypertrophy by blocking the activity of the MAPK terminal effectors.
MKP-1 Transgenic Mice Have Reduced MAPK Activation in the Heart
Previous in vivo studies have shown MAPK activation in response to pressure overload, ischemia, and agonist treatment.16 17 18 19 However, the global necessity of MAPK signaling factors as mediators of the hypertrophic response remains an area of ongoing investigation. To this end, we generated transgenic mice expressing MKP-1 under the control of the α-MHC cardiac-specific promoter. Three lines were generated, of which 2 passed the transgene into the germ-line. Western blotting of heart extracts from lines 1 and 2 demonstrated MKP-1 protein levels that were ≈3.7- and ≈1.8-fold above endogenous levels, respectively (Figure 3A⇓). To control for MKP-1 antibody specificity, protein extracts from AdMKP-1– and Adβgal-infected cardiomyocytes were electrophoresed together with the indicated heart extracts (Figure 3A⇓).
Hearts from high expressing MKP-1 transgenic mice (line 1) at postnatal day 10 were assayed for MAPK activation in response to acute systemic PE delivery via subcutaneous injection (10 mg/kg). Thirty minutes after injection, the hearts were removed and protein extracts were generated for Western blot analysis. The results demonstrate that acute PE injection was associated with p38, JNK1/2, and ERK1/2 phosphorylation in the nontransgenic heart (Figure 3B⇑). Remarkably, PE-induced p38, JNK1/2, and ERK1/2 phosphorylations were each blocked in the hearts of high expressing MKP-1 transgenic mice (Figure 3B⇑). Identical results were obtained in 3 independent experiments from separate cohorts, assayed 30 or 60 minutes after PE injection (data not shown). p38, JNK1/2, and ERK1/2 activation was also examined in lower expressing MKP-1 transgenic mice (line 2) after acute PE injection. Low expressing MKP-1 transgenic mice also demonstrated a lack of p38 and JNK activation similar to high expressing transgenic mice, although ERK1/2 dephosphorylation was less affected in the low expressing line, which is consistent with the known specificity of MKP-1 (see Discussion) (Figure 3C⇑). These data indicate that constitutive MKP-1 expression in the mouse heart inactivates reactive MAPK signaling.
Constitutive MKP-1 Expression Attenuates Developmental Hypertrophy
High expressing MKP-1 transgenic mice displayed compromised developmental hypertrophy that resulted in lethality at 7 to 15 postnatal days (founder mouse was viable due to mosaicism). Histological analysis of hearts from high expressing MKP-1 mice demonstrated severe and uniform ventricular dilation, without significant terminal deoxynucleotidyl transferase biotin-dUTP nick end labeling (TUNEL) (Figures 4A⇓ and 4B⇓ and data not shown). The observation of ventricular dilation in high expressing MKP-1 transgenic mice in the absence of appreciable apoptosis suggests that the heart is compromised in its ability to undergo developmental hypertrophy (see Discussion).
Microscopic histological analysis of H&E- and trichrome-stained heart sections from high expressing MKP-1 mice revealed disorganized and smaller myofibrils, without fibrosis (Figures 4C⇑ through 4F). To characterize myofiber cross-sectional areas in more detail, histological sections were stained with wheat germ agglutinin–tetrarhodamine isothiocyanate (TRITC) (50 μg/mL). Histological analysis showed significantly smaller myofibrils in MKP-1 transgenic hearts, further suggesting a defect in neonatal developmental hypertrophy (Figures 4G⇑ and 4H⇑).
Constitutive MKP-1 Expression Inhibits Pressure-Overload Hypertrophy In Vivo
Lower levels of constitutive MKP-1 expression, as characterized in line 2 mice, did not result in neonatal or adult lethality (up to 15 months at present). However, low expressing MKP-1 transgenic mice demonstrated mild left ventricular dilation on echocardiography at 8 weeks of age (Table⇓), which was not readily apparent through histological methods (Figures 5A⇓ and 5B⇓). Furthermore, careful histological analyses revealed no associated pathology or interstitial cell fibrosis in low expressing MKP-1 hearts (Figures 5C⇓ through 5F). However, wheat germ agglutinin–TRITC–stained histological sections revealed a 13% decrease in myofiber cross-sectional area in MKP-1 line 2 transgenic hearts at 8 weeks of age (nontransgenic 219±5.7 μm, transgenic 192±3.9 μm) (P<0.05) (Figures 5G⇓ through 5I). These data are consistent with the observed decrease in myofiber cross-sectional area characteristic of high expressing MKP-1 transgenic mice and support the notion that MAPK signaling pathways regulate, in part, developmental growth of the myocardium. MKP-1 transgenic mice did not demonstrate increased TUNEL labeling in histological sections at 2 and 8 weeks of age (data not shown).
Because MKP-1 transgenic mice have attenuated MAPK activation in the heart, it was of interest to determine their ability to hypertrophy in response to pressure overload. Adult wild-type mice (n=7) subjected to abdominal aortic banding for 14 days developed a 19±1% increase in heart weight–to–body weight ratio compared with sham-operated mice (n=4) (Figures 6A⇓ and 6B⇓). However, low expressing MKP-1 transgenic mice (n=5) showed only a 2±1% increase in heart weight–to–body weight ratio compared with sham-operated transgenic mice (n=5) (P<0.05) (Figures 6A⇓ and 6B⇓). Hearts from aortic-banded low expressing MKP-1 transgenic mice did not transition into greater dilation (Figure 6A⇓). These results demonstrate a significant attenuation of load-induced hypertrophy in MKP-1 transgenic mice.
MKP-1 Expression Attenuates Catecholamine-Induced Hypertrophy In Vivo
As an additional model of cardiac hypertrophy, MKP-1 transgenic mice were infused with isoproterenol with the use of osmotic minipumps as previously described.20 At baseline, hearts from MKP-1 transgenic mice demonstrated a subtle reduction in ERK1/2 phosphorylation and a pronounced reduction in p38 phosphorylation compared with littermate control hearts (Figure 6C⇑). Isoinfusion over 14 days was associated with irregular ERK1/2 activation in nontransgenic hearts, which was blocked in MKP-1 transgenic hearts (Figure 6C⇑). p38 phosphorylation was reduced more dramatically after 14 days of isoproterenol infusion in MKP-1 hearts (Figure 6C⇑). A similar decrease in ERK1/2 and p38 phosphorylation was also observed in MKP-1 transgenic hearts after aortic banding (data not shown). We were unable to detect JNK1/2 phosphorylation in adult hearts of wild-type MKP-1 transgenic mice after 14 days of chronic isoinfusion (data not shown). We also observed that chronic isoinfusion actually promoted a downregulation of p38 activation in nontransgenic mice (Figure 6C⇑), which is consistent with the report of Hines et al21 in which the induction of cardiomyocyte hypertrophy by chronic pacing promoted a gradual downregulation of p38 MAPK. Isoinfusion induced a 21±2% increase in heart weight–to–body weight ratio in wild-type mice (n=9) compared with saline-infused controls (n=4). In contrast, isoinfusion induced only a 8±1% increase in MKP-1 transgenic mice (n=10) compared with saline-infused MKP-1 controls (n=4) (P<0.05) (Figure 6B⇑). These results show a 62% inhibition of isoproterenol-induced cardiac hypertrophy in MKP-1 transgenic mice.
To examine the hypertrophic response of MKP-1 mice in greater detail, we measured the expression of hypertrophy-associated genes such as ANF, b-type natriuretic peptide (BNP), skeletal α-actin, and β-MHC. At baseline, low expressing MKP-1 transgenic hearts analyzed at 1, 3, 8, or 14 weeks of age did not show altered mRNA expression of BNP, skeletal α-actin, or β-MHC compared with wild-type littermate controls (data not shown). However, MKP-1 transgenic hearts analyzed at 8 weeks of age (4 individual hearts) showed mild expression of ANF mRNA, whereas hearts harvested at 1, 3, or 14 weeks of age showed no ANF induction (Figure 6D⇑ and data not shown). These results suggest that ANF is transiently activated at baseline in low expressing MKP-1 hearts at 8 weeks of age. Despite this transient developmental effect, isoinfusion induced an even greater comparative increase in ANF mRNA expression in wild-type hearts compared with low expressing MKP-1 transgenic hearts at 8 weeks (Figure 6D⇑). In addition, the induction of BNP mRNA expression was blocked in low expressing MKP-1 transgenic hearts (Figure 6D⇑). Collectively, these data indicate that MKP-1 expression partially suppresses the molecular program for cardiac hypertrophy in response to elevated catecholamines.
To establish the function of MAPK signaling cascades as necessary regulators of developmental hypertrophy and adult-onset hypertrophy, we antagonized p38, JNK1/2, and ERK1/2 activation by increasing the expression of the dual-specificity phosphatase MKP-1. Adenovirus-mediated gene transfer of MKP-1 in cultured cardiomyocytes prevented p38, JNK1/2, and ERK1/2 activation, resulting in an abrogation of myocyte growth/hypertrophy in response to agonist or mitogens. Transgenic mice constitutively expressing MKP-1 in the heart showed attenuation in normal developmental hypertrophy and concomitant ventricular dilation. Similarly, adult MKP-1 transgenic mice failed to mount a significant hypertrophic response when stressed by pressure overload or isoproterenol infusion.
Role of MAPK Signaling in the Regulation of Cardiac Hypertrophy
Although it is widely accepted that hypertrophic stimuli result in MAPK activation in cardiac myocytes, the causality between p38, JNK, or ERK activity and the promotion of hypertrophy is an area of controversy. ERK1/2 have been shown to mediate certain aspects of cardiomyocyte hypertrophy in cultured neonatal ventricular myocytes.22 23 However, other studies have disputed a role for ERK signaling in cultured cardiomyocyte hypertrophy,24 25 26 27 28 29 and 1 study even suggested that ERK activation was associated with hypertrophy prevention.30
p38 is activated in response to pressure-overload hypertrophy and in failing human hearts.16 31 Adenovirus-mediated gene transfer of an activated MKK6 factor (activates p38) or transfection of the MKK6 cDNA was reported to induce cardiomyocyte hypertrophy in vitro, demonstrating a sufficient role for p38 in hypertrophy.16 28 However, pacing-induced cardiomyocyte hypertrophy was associated with a downregulation of p38 activity, despite progressive hypertrophy,21 and pharmacological p38 inhibitors were reported to be ineffective in blocking certain aspects of endo-1–induced cardiomyocyte hypertrophy.29
More definitive data have emerged regarding a role for JNK signaling in the hypertrophic response.17 27 29 JNKs have been shown to be activated by pressure-overload hypertrophy and myocardial infarction.17 18 Adenovirus-mediated gene transfer of MKK7 (upstream activator of JNK1/2) was shown to induce cardiac hypertrophy in cultured cardiomyocytes.32 Last, adenovirus-mediated gene transfer of dnSEK-1 attenuated pressure-overload hypertrophy in aortic-banded rats, which stands as the only in vivo assessment of MAPK necessity yet performed in vivo.17 Even though a number of studies have implicated JNKs as hypertrophic mediators, 1 study reported that JNKs actually antagonize hypertrophic signaling.33
The current study was designed to address the controversy surrounding the necessity of MAPK signaling factors as mediators of cardiac hypertrophy using MKP-1 overexpression as a global inhibitory strategy. In vitro, AdMKP-1 infection of cardiomyocytes resulted in relatively high levels of MKP-1 protein expression, which blocked JNK1/2, p38, and ERK1/2 activation. In vivo, high expressing MKP-1 transgenic mice also lacked activation of all 3 MAPK branches, whereas low expressing transgenic mice showed only significant inactivation of p38 and JNK1/2. These observations are consistent with the known mechanism of MKP-1 action, whereby p38 and JNK1/2 are preferred substrates over ERK1/2.7 As such, the inhibition of cardiac hypertrophy seen in low expressing MKP-1 transgenic mice preferentially implicates p38 and JNK1/2 signaling as necessary mediators of cardiac hypertrophy in vivo.
In contrast, high expressing MKP-1 transgenic mice, which have inactivation of p38, JNK1/2, and ERK1/2, are characterized by reduced developmental hypertrophy, resulting in severe ventricular dilation and neonatal lethality. These data suggest that global inhibition of all 3 MAPK signaling branches compromises the maturation of the heart. Because the lower expressing transgenic mice undergo adequate developmental hypertrophy and survive into adulthood, it is likely that ERK1/2 activation is the critical component that can rescue heart maturation in the absence of p38 and JNK1/2. In any event, future approaches designed to specifically inhibit each of the MAPK signaling pathways separately will likely provide additional relevant information.
Specificity of MKP-1
Dual-specificity phosphatase expression is transcriptionally induced by stress or mitogen stimulation, which results in a delayed and coordinated inactivation of all 3 MAPK signaling pathways simultaneously. Likewise, most stimuli that result in MAPK activation coordinately induce all 3 signaling branches, suggesting that endogenous MAPK activation and subsequent inactivation are always coordinated. Indeed, pressure-overload hypertrophy in vivo is associated with a coordinated activation of all 3 MAPK signaling branches.17 Given that MAPK signaling branches are both activated and inactivated in a coordinated manner, it predicts that the overexpression of MKP-1 models the endogenous mechanism whereby MAPK responses are normally antagonized. The use of MKP-1 described in the present report models the mechanism of “physiological” MAPK regulation, resulting in attenuation of 2 or all 3 signaling branches.
MKP-1 recognizes a complex phosphorylation motif present only within the terminal kinases (TEY, TPY, and TGY), so the mild overexpression of MKP-1 is unlikely to affect targets other than MAPK factors.5 Indeed, forced expression of MKP-1 did not significantly downregulate MEK1, MKK4, MKK3/6, or Akt phosphorylation (Figure 1B⇑). Specificity is also suggested by the observation that overexpression of a different phosphatase, calcineurin (PP2B), actually induces hypertrophy in both cultured cardiomyocytes and transgenic mice,12 34 whereas the phosphatase MKP-1 blocks hypertrophy.
MAPK Signaling Is a Central Component of a Multifactorial Response
Numerous studies have demonstrated a requirement for various intracellular signaling factors in the regulation of cardiac hypertrophy.35 Such observations have suggested a model of reactive signaling in the heart such that certain central pathways are absolutely necessary for the initiation and maintenance of a balanced hypertrophy response. The results presented in this report indicate that MAPK signaling pathways are central regulators of reactive signaling in cardiac myocytes. The ability to directly downregulate intermediate MAPK signaling responses in the myocardium is predicted to effectively blunt cardiac hypertrophy in response to a wide array of receptor-activated stimuli and pathological insults. Because divergent receptor-activated signaling events (eg, Ang II, adrenergic, and endo-1 receptors) all appear to uniformly promote MAPK activation, it suggests that MAPK signaling pathways represent a convergence point whereby heterogeneous stimuli might mediate a common hypertrophic program. This notion suggests that members of the MAPK signaling cascade would be ideal targets for pharmacological intervention to treat maladaptive cardiac hypertrophy of multifactorial etiology. Alternatively, the specific upregulation of endogenous dual-specificity phosphatases in the heart might offer an additional therapeutic strategy to benefit certain forms of heart disease.
This work was supported by NIH grants HL-69562, HL-62927, and HL-52318 and a Pew Charitable Trust Scholar Award (Dr Molkentin). Dr Bueno was supported by NIH grant F32-HL-10336. We would like to thank the veterinary staff of Children’s Hospital for technical support with animal surgery and the Center for Environmental Genetics of the University of Cincinnati for transgenic mouse production.
Original received September 15, 2000; revision received October 30, 2000; accepted October 31, 2000.
- © 2001 American Heart Association, Inc.
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