Pregnancies in women with childhood vesicoureteral reflux.

OBJECTIVE: The frequency of pregnancy complications together with renal scarring and voiding dysfunction-related risk factors were investigated in a cohort of women with a history of childhood vesicoureteral reflux (VUR). DESIGN: A retrospective cross-sectional cohort study. POPULATION: Eighty-seven primi- or multiparous middle-aged women diagnosed with primary non-obstructive VUR in childhood. METHODS: Pregnancy outcome was assessed from case records and from patient interviews. Urine flow tests for voiding patterns, renal ultrasound for detecting scars, and blood samples for renal function were investigated. The median follow-up time was 38 years. MAIN OUTCOME MEASURES: Prevalence of pregnancy complications in women with childhood VUR in relation to renal scars and voiding abnormalities. RESULTS: Maternal complications were seen in 64% of the women and fetal complications in 13%. The women with renal scars (48/87) more often had hypertension (33%), proteinuria (40%) and urinary tract infections (42%) during pregnancy than women without scarring. The frequency of fetal complications was not increased by renal scarring or proteinuria during pregnancy. Urinary tract infections during pregnancy (33% of the women) and voiding abnormalities (18%) did not increase the frequency of fetal or maternal complications. The women with fetal complications were more predisposed to frequent urinary tract infections during adult life (55%) than were those without fetal complications (24%) (p = 0.04). CONCLUSIONS: The maternal complication rates in women with childhood VUR were increased only by renal scarring. Frequent urinary tract infections during adulthood seemed to predict an elevated risk of fetal complications.

The association of adult kidney size with childhood vesicoureteral reflux.

BACKGROUND: We used ultrasound to measure kidney volumes in adults with a history of childhood vesicoureteral reflux (VUR) and assessed whether total renal volume, small kidney size or the thickness of the upper pole correlated with renal function or hypertension. METHODS: The kidneys of 123 adults were studied by ultrasound, calculating their volumes using an ellipsoid formula normalised to body surface area (Vol(N)). The thickness of the upper pole parenchyma and the number of small kidneys (<80% of normal volume) were recorded. Blood pressure measurements and laboratory tests were also performed. RESULTS: Kidneys with a history of VUR were 12% smaller than those without known VUR (p < 0.05), and those with prior dilating VUR were 16% smaller than those with non-dilating VUR (p < 0.05). There was a moderate correlation (r = 0.42, p < 0.05) between total Vol(N) and GFR values in the total patient series. Thirteen percent of the patients had a moderate decrease in kidney function. The occurrence of hypertension and proteinuria was not affected by either kidney size or a thin upper pole. CONCLUSIONS: Total Vol(N) in ultrasound in early adulthood could probably predict possible renal deterioration in later life. The occurrence of one small kidney was a common finding and seemed not to affect the prevalence of proteinuria or hypertension.

Collagen XV is necessary for modeling of the extracellular matrix and its deficiency predisposes to cardiomyopathy.

RATIONALE: The extracellular matrix (ECM) is a major determinant of the structural integrity and functional properties of the myocardium in common pathological conditions, and changes in vasculature contribute to cardiac dysfunction. Collagen (Col) XV is preferentially expressed in the ECM of cardiac muscle and microvessels. OBJECTIVE: We aimed to characterize the ECM, cardiovascular function and responses to elevated cardiovascular load in mice lacking Col XV (Col15a1(-/-)) to define its functional role in the vasculature and in age- and hypertension-associated myocardial remodeling. METHODS AND RESULTS: Cardiac structure and vasculature were analyzed by light and electron microscopy. Cardiac function, intraarterial blood pressure, microhemodynamics, and gene expression profiles were studied using echocardiography, telemetry, intravital microscopy, and PCR, respectively. Experimental hypertension was induced with angiotensin II or with a nitric oxide synthesis inhibitor. Under basal conditions, lack of Col XV resulted in increased permeability and impaired microvascular hemodynamics, distinct early-onset and age-dependent defects in heart structure and function, a poorly organized fibrillar collagen matrix with marked interstitial deposition of nonfibrillar protein aggregates, increased tissue stiffness, and irregularly organized cardiomyocytes. In response to experimental hypertension, Col15a1 gene expression was increased in the left ventricle of wild-type mice, and mRNA expression of natriuretic peptides (ANP and BNP) and ECM modeling were abnormal in Col15a1(-/-) mice. CONCLUSIONS: Col XV is necessary for ECM organization in the heart, and for the structure and functions of microvessels. Col XV deficiency leads to a complex cardiac phenotype and predisposes the subject to pathological responses under cardiac stress.

Role of reactive oxygen species in the regulation of cardiac contractility.

Increased production of reactive oxygen species (ROS) has been linked to the pathogenesis of contractile dysfunction in heart failure. However, it is unclear whether ROS can regulate physiological cellular processes in the myocardium. Here, we characterized the role of endogenous ROS production in the acute regulation of cardiac contractility in the intact rat heart. In isolated perfused rat hearts, endothelin-1 (ET-1, 1nmol/L) stimulated ROS formation in the left ventricle, which was prevented by the antioxidant N-acetylcysteine and the NAD(P)H oxidase inhibitor apocynin. N-acetylcysteine, the superoxide dismutase mimetic MnTMPyP, and apocynin significantly attenuated ET-1-mediated inotropic effect, which was accompanied by inhibition of extracellular signal regulated kinase 1/2 (ERK1/2) phosphorylation. Moreover, the mitochondrial K(ATP) channel blocker 5-HD, and the mitochondrial large conductance calcium activated potassium channel blocker paxilline, but not the sarcolemmal K(ATP) channel blocker HMR 1098 attenuated the inotropic response to ET-1. However, ET-1-induced ROS generation was not abolished by inhibiting mitochondrial K(ATP) channel opening. In contrast to ET-1 stimulation, the positive inotropic effect of ß(1)-adrenergic receptor agonist dobutamine (250nmol/L) was significantly augmented by N-acetylcysteine and apocynin. Moreover, dobutamine-induced phospholamban phosphorylation was markedly enhanced by apocynin. In conclusion, NAD(P)H oxidase-derived ROS play a physiological role in the acute regulation of cardiac contractility in the intact rat heart. Our results reveal that ET-1-induced increase in cardiac contractility is partially dependent on enhanced ROS generation, which in turn, activates the ERK1/2 pathway. On the other hand, ß-adrenergic receptor-induced positive inotropic effect and phospholamban phosphorylation is enhanced by NAD(P)H oxidase inhibition.

Parthenolide inhibits STAT3 signaling and attenuates angiotensin II-induced left ventricular hypertrophy via modulation of fibroblast activity.

Parthenolide has shown promise in treatment of various cancers via inhibition of the transcription factor signal transducer and activator of transcription 3 (STAT3). Activation of STAT3 has been observed in left ventricular hypertrophy (LVH); however, its exact role is not known. The aim of the study was to examine the effects of parthenolide on pressure overload-induced LVH in rats. Pressure overload was induced by angiotensin II (Ang II) infusion (33 µg/kg/h) for 1 week in the presence or absence of parthenolide (0.5mg/kg/day, i.p.). Ang II infusion resulted in LVH associated with increased phosphorylation of STAT3 at Tyr705 and Ser727. Parthenolide treatment had no effect on ejection fraction, but abolished the activation of STAT3 and reduced the Ang II-induced LVH (LV posterior wall thickness in end-diastole: 2.28 ± 0.12 mm vs. 1.80 ± 0.06 mm, P<0.001). Importantly, parthenolide treatment had no effect on heart rate or blood pressure. Parthenolide treatment almost completely abolished the Ang II-induced increase in the number of cells positive for prolyl-4-hydroxylase, a marker for collagen-synthesizing cells, as well as Ang II-induced interstitial fibrosis in the left ventricles. This was associated with significant attenuation of Ang II-induced increase in mRNA levels of type 1 collagen and fibronectin. Moreover, parthenolide attenuated the Ang II-induced expression of interleukin-6, a potent pro-hypertrophic fibroblast-derived factor. We conclude that pharmacological inhibition of STAT3 signaling by parthenolide has favorable effects on pressure overload-induced LVH through attenuation of fibroblast activation. Therefore parthenolide may prove as a useful therapy for certain cardiovascular disease.

Overexpression of vascular endothelial growth factor-B in mouse heart alters cardiac lipid metabolism and induces myocardial hypertrophy.

Vascular endothelial growth factor (VEGF)-B is poorly angiogenic but prominently expressed in metabolically highly active tissues, including the heart. We produced mice expressing a cardiac-specific VEGF-B transgene via the alpha-myosin heavy chain promoter. Surprisingly, the hearts of the VEGF-B transgenic mice showed concentric cardiac hypertrophy without significant changes in heart function. The cardiac hypertrophy was attributable to an increased size of the cardiomyocytes. Blood capillary size was increased, whereas the number of blood vessels per cell nucleus remained unchanged. Despite the cardiac hypertrophy, the transgenic mice had lower heart rate and blood pressure than their littermates, and they responded similarly to angiotensin II-induced hypertension, confirming that the hypertrophy does not compromise heart function. Interestingly, the isolated transgenic hearts had less cardiomyocyte damage after ischemia. Significantly increased ceramide and decreased triglyceride levels were found in the transgenic hearts. This was associated with structural changes and eventual lysis of mitochondria, resulting in accumulation of intracellular vacuoles in cardiomyocytes and increased death of the transgenic mice, apparently because of mitochondrial lipotoxicity in the heart. These results suggest that VEGF-B regulates lipid metabolism, an unexpected function for an angiogenic growth factor.

A novel p38 MAPK target dyxin is rapidly induced by mechanical load in the heart.

Dyxin is a novel LIM domain protein acting as a transcriptional cofactor with GATA transcription factors. Here, we characterized dyxin as a p38 mitogen-activated protein kinase (MAPK) regulated gene, since combined upstream MAPK kinase 3b and wild-type p38 alpha MAPK gene transfer increased left ventricular dyxin mRNA and protein levels in vivo. We also studied cardiac dyxin expression in experimental models of pressure overload and myocardial infarction (MI) in vivo. Angiotensin II infusion increased left ventricular dyxin mRNA levels (9.4-fold, p<0.001) rapidly at 6 h followed by induction of protein levels. Furthermore, simultaneous administration of p38 MAPK inhibitor SB203580 abolished angiotensin II-induced activation of dyxin gene expression. During the post-infarction remodeling process, increased dyxin mRNA levels (7.7-fold, p<0.01) were noted at day 1 followed by the increase in proteins levels at 2 weeks after MI (1.5-fold, p<0.05). Moreover, direct wall stretch by using isolated rat heart preparation as well as direct mechanical stretch of cardiomyocytes in vitro activated dyxin gene expression within 1 h. Our results indicate that dyxin expression is rapidly upregulated in response to mechanical load, this increase being at least partly mediated by p38 MAPK. These results suggest that dyxin may play an important role in regulating hypertrophic process.

Increase in tissue endothelin-1 and ETA receptor levels in human aortic valve stenosis.

AIMS: Aortic valve stenosis (AS) is an actively regulated process like atherosclerosis, which is accompanied by changes e.g. in endothelin-related genes. However, the role of endothelin peptides in AS is unknown. METHODS AND RESULTS: We characterized the expression of the endothelin system in aortic valves of patients with normal valves (n = 12), regurgitation, and fibrosis (n = 6) and AS (n = 18) by reverse-transcriptase-polymerase chain reaction and immunohistochemistry. The number of endothelin-1 (ET-1) positive cells was higher in AS than in control valves, while levels of ET-1 mRNA did not differ between groups. Endothelin receptor-A (ET(A)) mRNA levels were upregulated in stenotic valves (4.3-fold, P = 0.032) associated with a remarkable increase in number of ET(A)-immunopositive cells. ET(B)-receptor mRNA levels did not change during disease progression. Endothelin-converting enzyme-1 (ECE-1) mRNA levels were 42% lower (P = 0.007) in stenotic valves. Finally, because ET-1 and ECE-1 have binding site for activator protein-1 (AP-1), we measured AP-1 DNA binding by gel shift assays, which showed significantly lower (76%, P = 0.003) activity in AS. CONCLUSION: AS is characterized by distinct upregulation of ET-1 and its target receptor ET(A), promoting growth, inflammation, and fibrosis. These findings suggest therapeutic potential for ET(A)-receptor antagonists in aortic valve calcification.

Hypoxia-induced expression of bradykinin type-2 receptors in endothelial cells triggers NO production, cell migration, and angiogenesis.

Bradykinin receptors are differentially expressed in the coronary vascular endothelium of rat and human hearts during the pathogenesis of heart failure, but the mechanisms responsible for this regulation have remained vague. Here we show by quantitative real-time PCR, Western blot analysis, and immunohistochemistry, that hypoxia triggers the expression of bradykinin type-2 receptors (BK-2Rs) in cultured human coronary artery endothelial cells (HCAECs), in isolated rat cardiac microvascular endothelial cells (RCMECs), and in rat hearts subjected to ligation of the left anterior descending coronary artery. Mild hypoxia (5% O(2)) induced a fourfold temporal increase in BK-2R mRNA expression in HCAECs, which was also observed at the protein level, whereas severe hypoxia (1% O(2)) slightly inhibited the mRNA expression of BK-2Rs. In addition, HOE-140, a BK-2R antagonist, inhibited mRNA and protein expression of BK-2Rs. The BK-2Rs induced by mild hypoxia were biologically active, that is, capable of inducing intracellular production of nitric oxide (NO) upon activation of HCAECs with bradykinin (BK), a response attenuated by HOE-140. In rat hearts recovering from myocardial infarction, BK-2Rs were upregulated in the endothelium of vessels forming at the border zone between fibrotic scar tissue and healthy myocardium. Furthermore, in an in vitro wound-healing assay, RCMEC migration was increased under mild hypoxic culture conditions in the presence of BK and was attenuated with HOE-140. Our present results show that mild hypoxia triggers a temporal expression of functional BK-2Rs in human and rat endothelial cells and support a role for BK-2Rs in hypoxia-induced angiogenesis.

Urodynamical findings on women with voiding problems and earlier vesico-ureteral reflux.

AIMS: Examine bladder function behind an abnormal urine flow curve pattern or lower urinary tract symptoms (LUTS) in women with earlier vesicoureteral reflux (VUR). METHODS: Seventeen earlier female VUR patients with weak, fractionated or tower-shaped flow pattern, 10 patients with normal voiding and 2 patients with a big bladder were urodynamically studied. RESULTS: A third of the patients had a large (>800 ml) cystometric bladder capacity. Sensitivity of the bladder was decreased especially in weak urine flow group. Half of these patients felt the "first sensation" when at least 40% of the bladder capacity was filled. Neither the earlier treatment modality nor grade of childhood VUR had an influence on the bladder sensitivity. Urinary tract infections were reported significantly more often (P = 0.028) in patients with decreased bladder sensitivity than in the other patients. Overactive detrusor and decreased bladder compliance were uncommon findings. The detrusor pressure was good exceeding 15 cm H(2)O during the maximum flow rate in almost all patients. Abnormal sphincteric EMG activity during voiding was found in 70% of all patients and especially in fractionated flow group (89%). The EMG activity was seen twice as often in patients with earlier bilateral dilating VUR as in those who had suffered of non-dilating VUR (P = 0.005). CONCLUSIONS: Despite of high frequency of symptoms in female patients with earlier VUR, detrusor overactivity was a rarity, but decreased sensitivity and large capacity of the bladder were found frequently. The patients with weak or fractionated urine flow seemed to suffer from an overactive urethral sphincter.

Upregulation of cardiac matrix Gla protein expression in response to hypertrophic stimuli.

Matrix Gla protein (MGP) expression is increased in cardiac hypertrophy, but the precise mechanisms regulating its expression are unknown. Here we characterized the effect of pressure overload and myocardial infarction in vivo as well as mechanical stretch and hypertrophic agonists in vitro on MGP expression. When angiotensin II (Ang II) was administered by osmotic minipumps, left ventricular (LV) MGP mRNA levels increased significantly from 6 h to 2 weeks, whereas intravenous arginine(8)-vasopressin increased LV MGP mRNA levels within 4 h. During post-infarction remodeling process, MGP mRNA levels were elevated at 24 h (1.3-fold, p<0.05) and the maximal increase was observed at 4 weeks (2.8-fold, p<0.01). Ang II increased MGP mRNA levels 20% (p<0.05) in neonatal rat cardiac myocytes and 40% (p<0.05) in cardiac fibroblasts, whereas endothelin-1 decreased MGP mRNA levels 30% (p<0.01) in myocytes and had no effect in fibroblasts. Cyclic mechanical stretch resulted in reduction of MGP gene expression in both cardiac myocytes and fibroblasts. These results demonstrate that MGP is rapidly upregulated in response to cardiac overload well before the development of LV hypertrophy and post-infarction remodeling process. Our results also suggest that Ang II may be involved in mediating load-induced activation of MGP expression.

Distinct downregulation of C-type natriuretic peptide system in human aortic valve stenosis.

BACKGROUND: Aortic valve calcification is an actively regulated process that displays hallmarks of atherosclerosis. Natriuretic peptides (A-, B-, and C-type natriuretic peptides [ANP, BNP, and CNP]) have been reported to have a role in the pathogenesis of vascular atherosclerosis, but their expression in aortic valves is not known. Here, we characterized and compared expression of natriuretic peptide system in aortic valves of patients with normal valves (n=4), aortic regurgitation (n=11), regurgitation and fibrosis (n=6), and aortic valve stenosis (n=21). METHODS AND RESULTS: By reverse-transcription polymerase chain reaction, all 3 natriuretic peptides were found to be expressed in aortic valves. CNP mRNA levels were 92% lower (P<0.001) in stenotic valves, whereas no significant changes in the expression of ANP and BNP genes were found compared with valves obtained from patients with aortic regurgitation. CNP was localized by immunohistochemistry with specific CNP (32-53) antibody to valvular endothelial cells and myofibroblasts. Gene expression of furin, which proteolytically cleaves proCNP into active CNP, was 54% lower in aortic valve stenosis (P=0.04). Moreover, natriuretic peptide receptor-A and natriuretic peptide receptor-B mRNA levels were 78% and 76% lower, respectively, in stenotic valves. In contrast, gene expression of corin, a proANP- and proBNP-converting enzyme, and natriuretic peptide receptor-C did not differ between groups. CONCLUSIONS: We show that natriuretic peptides, their processing enzymes, and their receptors are expressed in human aortic valves. Aortic valve stenosis is characterized by distinct downregulation of gene expression of CNP, its processing enzyme furin, and the target receptors natriuretic peptide receptor-B and natriuretic peptide receptor-A, which suggests that CNP acts as a paracrine regulator of the aortic valve calcification process.

Adaptive or maladaptive response to adenoviral adrenomedullin gene transfer is context-dependent in the heart.

BACKGROUND: Adrenomedullin (AM) is a potent vasodilator and natriuretic peptide produced in the heart, but controversy persists regarding its cardiac effects. We explored the potential role of AM on cardiac function and remodeling by direct recombinant adenoviral AM gene delivery into the anterior wall of the left ventricle (LV). METHODS: AM was overexpressed in healthy rat hearts and in hearts during the remodeling process in response to pressure overload and myocardial infarction. The AM effects were analysed with echocardiography and in an isolated perfused rat heart preparation. The expression of AM and the activation of underlying signaling pathways were also investigated. RESULTS: AM mRNA increased by 20.9-fold (p < 0.001) in healthy rat heart and improved fractional shortening by 14% (p < 0.05) and ejection fraction by 8% (p < 0.05). In isolated perfused hearts, an increase (p < 0.05) in the first derivative of isovolumic LV pressure rise (dP/dt(max)) without alteration in diastolic properties was noted. The overexpression of AM activated protein kinase Cepsilon and Cdelta isoforms in the LV, whereas p38 mitogen-activated protein kinase activity decreased. Angiotensin II-induced LV hypertrophy was significantly attenuated by AM (p < 0.01) without compromising cardiac contractility. By contrast, AM enhanced LV dilatation (p < 0.01) and anterior wall thinning (p < 0.001) and augmented the deterioration of LV function (p < 0.05) post-infarction. CONCLUSIONS: The results obtained in the present study show that AM overexpression improves LV systolic function without altering cardiac diastolic properties in the normal heart. Moreover, AM is a potent context-dependent modulator of LV remodeling because it promotes an adaptive response in pressure overload-induced LV hypertrophy and triggers a maladaptive process in post-infarction remodeling.

Thrombospondin-4 expression is rapidly upregulated by cardiac overload.

The precise mechanisms regulating gene expression of thrombospondins (TSPs) in the heart remain incompletely understood. Here we characterized cardiac TSP-4 expression in response to pressure overload and myocardial infarction in vivo. Arginine(8)-vasopressin (AVP) infusion increased left ventricular (LV) TSP-4 mRNA levels within 30 min. Also angiotensin II infusion rapidly activated LV TSP-4 expression, TSP-4 mRNA levels being highest at 6h and protein at 72 h and 2 weeks. During remodeling process following myocardial infarction, LV TSP-4 mRNA levels increased at day one, as studied by quantitative RT-PCR. TSP-4 immunostaining was localized to endothelial cells in hypertrophied hearts of spontaneously hypertensive rats. AVP-infusion increased LV TSP-1 mRNA levels similarly to TSP-4 within 30 min showing that rapid induction of gene expression, well before the development of cardiac hypertrophy, is typical for the thrombospondin family. These results further suggest that TSP-4 may be an endothelial specific marker of cardiac overload.

Hypoxia inducible factor regulates the cardiac expression and secretion of apelin.

Apelin and its G-protein-coupled receptor APJ have various beneficial effects on cardiac function and blood pressure. The mechanisms that regulate apelin gene expression are not known. Because apelin gene expression has been shown to increase in cardiac ischemia, we investigated if apelin (Apln) gene expression was sensitive to hypoxia. Here we show that hypoxia increases the apelin expression in rat myocardium and in cultured cardiomyocytes. Pharmacological activation of hypoxia inducible factor by desferrioxamine (DFO) or expression of a constitutively active form of HIF-1alpha increased apelin expression in cardiomyocyte cultures. The induction of apelin by hypoxia was abolished on transient expression of the HIF inhibitory PAS protein in cardiomyocytes. Increased apelin expression induced by hypoxia or DFO was accompanied by the processing of the cellular storage form proapelin into smaller apelin peptides and increased secretion of these biologically active forms of apelin. In a rat in vivo model, acute myocardial infarction (24 h) led to a transient increase in ventricular apelin mRNA levels. Our results indicate that apelin gene is regulated by hypoxia in cardiac myocytes via the HIF pathway, suggesting a role for apelin as a potential marker for acute cardiac hypoxia with a possible compensatory role in myocardial tissue suffering from oxygen deprivation.

Identification of cell cycle regulatory and inflammatory genes as predominant targets of p38 mitogen-activated protein kinase in the heart.

Mitogen-activated protein kinases (MAPKs) regulate cardiomyocyte growth and apoptosis in response to extracellular stimulation, but the downstream effectors that mediate their pathophysiological effects remain poorly understood. We determined the targets and role of p38 MAPK in the heart in vivo by using local adenovirus-mediated gene transfer of constitutively active upstream kinase mitogen-activated protein kinase kinase 3b (MKK3bE) and wild-type p38alpha in rats. DNA microarray analysis of animals with cardiac-specific overexpression of p38 MAPK revealed that 264 genes were upregulated more than 2-fold including multiple genes controlling cell division, cell signaling, inflammation, adhesion, and transcription. A large number of previously unknown p38 target genes were found. Using gel mobility-shift assays we identified several cardiac transcription factors that were directly activated by p38 MAPK. Finally, we determined the functional significance of the altered cardiac gene-expression profile by histological analysis and echocardiographic measurements, which indicated that p38 MAPK overexpression-induced gene expression results in myocardial cell proliferation, inflammation, and fibrosis. In conclusion, we defined the novel target genes and transcription factors as well as the functional effects of p38 MAPK in the heart. Expression profiling of p38 MAPK overexpression identified cell cycle regulatory and inflammatory genes critical for pathological processes in the adult heart.

Long-term postinfarction melatonin administration alters the expression of DHPR, RyR2, SERCA2, and MT2 and elevates the ANP level in the rat left ventricle.

We investigated the effect of 2 wk continuous postinfarction subcutaneous melatonin supply on the expression of the rat left ventricular (LV) dihydropyridine receptor (DHPR), ryanodine receptor (RyR(2)), and sarco-endoplasmic reticulum Ca(2+)-ATPase2 (SERCA2), as they are fundamental proteins in cardiac contractility. The levels of plasma and LV atrial (ANP) and brain natriuretic peptide and melatonin were also measured, as was the expression of LV MT(1) and MT(2) receptors and pineal arylalkylamine N-acetyltransferase. Myocardial infarction (MI) was induced by ligation of the left anterior descending coronary artery and vehicle or melatonin (4.5 mg/kg per day) was administered by subcutaneous osmotic pumps. Echocardiography, real-time quantitative reverse transcription-polymerase chain reaction, and western blotting were used to analyze the samples. Echocardiography revealed that MI induced serious systolic LV dysfunction. The expression of DHPR, RyR(2), and SERCA2 mRNAs was significantly lower in the LVs of melatonin-treated MI rats compared with vehicle-treated rats (P < 0.01 for DHPR and P < 0.05 for RyR(2) and SERCA2). Melatonin also elevated the amount of LV MT(2) receptors to 1.9-fold (P < 0.05) and the concentration of LV ANP to over fivefold (P < 0.05) compared with vehicle rats after MI. Therefore, the results suggest that melatonin may influence the cardiac contractility after MI by regulating the expression of DHPR, RyR(2), and SERCA2, and melatonin receptors, particularly MT(2)s, might contribute to the postinfarction cardioprotective actions of melatonin. Furthermore, the finding of the relationship between melatonin and ANP suggests a novel mechanism for melatonin in protecting the heart after MI.

Nuclear factor-kappaB signaling contributes to severe, but not moderate, angiotensin II-induced left ventricular remodeling.

OBJECTIVE: The transcription factor nuclear factor-kappaB (NF-kappaB) has been implicated in cardiomyocyte hypertrophy in vitro as well as in vivo; however, it is unknown if activation of NF-kappaB plays a mandatory role in the hypertrophic process. Here we characterize the importance of NF-kappaB signaling in moderate and severe left ventricular (LV) hypertrophy in rats with chronic pressure overload induced by angiotensin II (Ang II) infusion. METHODS AND RESULTS: Electrophoretic mobility shift assay analysis revealed that Ang II infusion (2.5 microg/kg per min) for 6 days increased LV NF-kappaB/DNA-binding activity in a biphasic manner in Sprague-Dawley rats. Pyrrolidine dithiocarbamate (PDTC) (100 mg/kg per day), an NF-kappaB inhibitor, abolished Ang II-induced NF-kappaB activation and concomitant increase in tumor necrosis factor-alpha gene expression, while activator protein-1/DNA binding was not affected. Inhibition of NF-kappaB signaling for 6 days significantly attenuated Ang II-induced increases in LV/body weight ratio, LV mean wall thickness and cardiomyocyte cross-sectional area, without compromising LV systolic function. Moreover, PDTC abolished Ang II-induced cardiomyocyte apoptosis and interstitial fibrosis, and attenuated the gene expression of type I collagen. In contrast, a moderate LV hypertrophy induced by Ang II at a lower dose (0.5 microg/kg per min) was not associated with a significant activation of NF-kappaB, and PDTC treatment had no effect on the hypertrophic indices. CONCLUSION: Our in-vivo data indicate a critical role of NF-kappaB signaling in the advanced stage of the remodeling process, whereas development of moderate LV hypertrophy is not dependent on NF-kappaB activation.

p38 Kinase rescues failing myocardium after myocardial infarction: evidence for angiogenic and anti-apoptotic mechanisms.

As a leading cause of heart failure, postinfarction left ventricular remodeling represents an important target for therapeutic interventions. Mitogen-activated protein kinases regulate critical cellular processes including stress response and survival, but their role in left ventricular remodeling is unknown. In the present study, rats were subjected to myocardial infarction by ligating the left anterior descending coronary artery. Western blot and kinase assay analysis revealed an inactivation of p38 kinase after myocardial infarction. Local adenovirus-mediated cotransfection of wild-type (WT) p38 kinase and constitutively active MKK3b reduced infarct size (26+/-3% vs. 47+/-4%, P<0.05 vs. LacZ-treated control) associated with improved ejection fraction (66.9+/-5.5% vs. 44.4+/-4.0%, P<0.001), fractional shortening (30.2+/-2.1% vs. 19.7+/-2.2%, P<0.001), and decreased left ventricular diastolic diameter (8.5+/-0.4 mm vs. 9.5+/-0.2 mm, P<0.01). p38 kinase gene transfer increased capillary density (2423+/-107/mm(2) vs. 1934+/-86/mm(2), P<0.001) and resulted in microvessel enlargement in the ischemic border zone. Apoptosis (35+/-7 vs. 69+/-13 cells, P<0.01) and fibrosis (16+/-3% vs. 34+/-8%, P<0.05) were reduced, while the number of c-kit positive cardiac stem-like cells remained unchanged. These results indicate that reduced p38 signaling predisposes to adverse postinfarction remodeling. The rescue of failing myocardium with p38 kinase may be a potential new therapy for heart failure after myocardial infarction.

Evidence for a functional role of angiotensin II type 2 receptor in the cardiac hypertrophic process in vivo in the rat heart.

BACKGROUND: The precise function of angiotensin II type 2 receptor (AT2-R) in the mammalian heart in vivo is unknown. Here, we investigated the role of AT2-R in cardiac pressure overload. METHODS AND RESULTS: Rats were infused with vehicle, angiotensin II (Ang II), PD123319 (an AT2-R antagonist), or the combination of Ang II and PD123319 via subcutaneously implanted osmotic minipumps for 12 or 72 hours. Ang II-induced increases in mean arterial pressure, left ventricular weight/body weight ratio, and elevation of skeletal alpha-actin and beta-myosin heavy chain mRNA levels were not altered by PD123319. In contrast, AT2-R blockade resulted in a marked increase in the gene expression of c-fos, endothelin-1, and insulin-like growth factor-1 in Ang II-induced hypertension. In parallel, Ang II-stimulated mRNA and protein expression of atrial natriuretic peptide were significantly augmented by AT2-R blockade. Moreover, PD123319 markedly increased the synthesis of B-type natriuretic peptide. Furthermore, the expression of vascular endothelial growth factor and fibroblast growth factor-1 was downregulated by Ang II only in the presence of AT2-R blockade. CONCLUSIONS: Our results provide evidence that AT2-R plays a functional role in the cardiac hypertrophic process in vivo by selectively regulating the expression of growth-promoting and growth-inhibiting factors.