Hanna Leskinen

Circulating Natriuretic Peptide Concentrations in Patients With End-Stage Renal Disease: Role of Brain Natriuretic Peptide as a Biomarker for Ventricular Remodeling

OBJECTIVES To determine levels of natriuretic peptides (NPs) in patients with end-stage renal disease (ESRD) and to examine the relationship of these cardiovascular peptides to left ventricular hypertrophy (LVH) and to cardiac mortality. PATIENTS AND METHODS One hundred twelve dialysis patients without clinical evidence of congestive heart failure underwent plasma measurement of NP concentrations and echocardiographic investigation for left ventricular mass index (LVMI). RESULTS Plasma atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) concentrations correlated positively with LVMI and inversely with left ventricular ejection fraction, whereas C-type NP and Dendroaspis NP levels did not correlate with LVMI. In dialysis patients with LVH (LVMI >125 g/m2), plasma ANP and BNP concentrations were increased compared with those in dialysis patients without LVH (both P<001). In a subset of 15 dialysis patients without LVH or other concomitant diseases, plasma BNP concentrations were not significantly increased compared with those in 35 controls (mean +/- SD, 20.1+/-13.4 vs 13.5+/-9.6 pg/mL; P=.06), demonstrating that the BNP concentration was not increased by renal dysfunction alone. Furthermore, the BNP level was significantly higher in the 16 patients who died from cardiovascular causes compared with survivors (mean +/- SD, 129+/-13 vs 57+/-7 pg/mL; P<.003) and was significantly associated with greater risk of cardiovascular death in Cox regression analysis (P<.001), as was the ANP level (P=.002). CONCLUSIONS Elevation of the plasma BNP concentration is more specifically related to LVH compared with the other NP levels in patients with ESRD independent of congestive heart failure. Thus, BNP serves as an important plasma biomarker for ventricular hypertrophy in dialysis patients with ESRD.

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 constitu‐tively active form of HIF‐1α 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.—Ronkainen V.‐P., Ronkainen, J. J., Hanninen, S. L., Leskinen, H., Ruas, J. L., Pereira, T., Poellinger, L., Vuolteenaho, O., Tavi P. Hypoxia inducible factor regulates the cardiac expression and secretion of apelin. FASEB J. 21, 1821–1830 (2007)

Distinct Upregulation of Extracellular Matrix Genes in Transition From Hypertrophy to Hypertensive Heart Failure

Cardiac hypertrophy in response to pressure overload is initially beneficial but eventually leads to heart failure, a major cause of morbidity and mortality in the Western countries. Although abnormalities in left ventricular (LV) diastolic filling are early features associated with pressure overload-induced LV hypertrophy, the molecular mechanisms regulating transition to diastolic heart failure are poorly understood. We analyzed global changes in gene expression in 12-, 16-, and 20-month-old spontaneously hypertensive rats (SHR) and their age-matched controls, Wistar Kyoto rats, using DNA microarrays. In SHR, a progressive LV hypertrophy was associated with increased expression of hypertrophy-associated genes including contractile protein and natriuretic peptide genes. Echocardiography indicated that 16-month-old SHR had features of diastolic dysfunction leading to diastolic failure at age 20 months without significant changes in LV systolic function. Comparison analysis revealed that the extracellular matrix genes strikingly dominated the list of altered genes after transition to the heart failure, whereas there was no major shift in gene expression patterns involved in calcium homeostasis and neurohumoral activation, as well as myofilament contractile and cytoskeletal proteins. The microarray analysis also revealed differential gene expression of several novel factors, such as thrombospondin-4 and matrix Gla protein, as well as unknown expressed sequence tags. Our data show that transition from LV hypertrophy to diastolic hypertensive heart failure is almost exclusively associated with progressive remodeling of the extracellular matrix and provide new insights into the pathogenesis of hypertrophy by suggesting existence of novel regulators of LV remodeling.

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 α-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.

Mechanisms of mechanical load-induced atrial natriuretic peptide secretion: role of endothelin, nitric oxide, and angiotensin II.

Abstract There are three members in the natriuretic peptide hormone family, atrial natriuretic peptide (ANP), B-type natriuretic peptide (BNP, brain natriuretic peptide), and C-type natriuretic peptide (CNP), that are involved in the regulation of blood pressure and fluid homeostasis. CNP is found principally in the central nervous system and vascular endothelial cells while ANP and BNP are cardiac hormones. ANP is synthesized mainly in the atria of the normal adult heart, while BNP is produced by both the atria and ventricles. The mechanisms controlling ANP release have been the subject of intense research, and are now fairly well understood. The major determinant of ANP secretion is myocyte stretch. Although much less is known about the factors regulating BNP release from the heart, myocyte stretch has also been reported to stimulate BNP release from both atria and ventricles. However, whether wall stretch acts directly or via factors such as endothelin-1, nitric oxide, or angiotensin II liberated in response to distension has not been established. Recent studies show that by stimulating endothelin type A receptors endothelin plays an important physiological role as a mediator of acute-volume load-induced ANP secretion from atrial myocytes in conscious animals. In fact, endogenous paracrine/autocrine factors liberated in response to atrial wall stretch rather than direct stretch appears to be responsible for activation of ANP secretion in response to volume load, as evidenced by almost complete blockade of ANP secretion during combined inhibition of endothelin type A/B and angiotensin II receptors. Furthermore, under certain experimental conditions angiotensin II and nitric oxide may also exert a significant modulatory effect on stretch-activated ANP secretion. The molecular mechanisms by which endothelin-1, angiotensin II, and nitric oxide synergistically regulate stretch-activated ANP release are yet unclear.

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 p38α 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.

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/mm2 vs. 1934±86/mm2, 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. —Tenhunen, O., Soini, Y., Ilves, M., Rysä, J., Tuukkanen, J., Serpi, R., Pennanen, H., Ruskoaho, H., Leskinen, H. p38 Kinase rescues failing myocardium after myocardial infarction: evidence for angiogenic and anti‐apoptotic mechanisms. FASEB J. 20, E1276‐E1286 (2006)

Combined Inhibition of Endothelin and Angiotensin II Receptors Blocks Volume Load–Induced Cardiac Hormone Release

Volume expansion has been shown to increase plasma atrial natriuretic peptide (ANP) levels, but the precise role of paracrine and autocrine factors in stretch-induced cardiac hormone release is not clear. In the present study, we report the effects of endothelin (ET) and angiotensin receptor (AT receptor) antagonists on baseline and atrial stretch-induced immunoreactive ANP (IR-ANP) and immunoreactive N-terminal ANP (IR-NT-ANP) release in vivo by using BQ-123 (ETA receptor antagonist), bosentan (ETA and ETB receptor antagonist), and losartan (AT1 receptor antagonist). Intravenous administration of BQ-123 had no significant effect on baseline hemodynamics in conscious rats, whereas bosentan (10 mg/kg) and losartan (10 mg/kg) decreased slightly (4 to 7 mm Hg, P < .05 to .001) the mean arterial pressure. Both the ETA receptor antagonist BQ-123 and ETA/ETB receptor antagonist bosentan decreased plasma ANP and NT-ANP responses to volume load (P < .05 to .001), whereas the AT1 receptor antagonist losartan had no significant effect on this response. The relative increase in plasma IR-ANP corresponding to a 3 mm Hg increase in right atrial pressure was 2.7-fold in the vehicle-treated group. BQ-123 (0.3 and 1.0 mg/kg) decreased this response 2.5- and 2.1-fold (P < .05); bosentan (3 and 10 mg/kg), 1.7-fold (P < .001) and 1.9-fold (P < .05); and bosentan (10 mg/kg)+losartan (10 mg/kg), 1.6-fold (P < .001). The responses in plasma IR-NT-ANP decreased simultaneously. These results indicate that combined inhibition of ETA/B and AT1 receptors almost completely blocks ANP response to acute volume load. Therefore, our study shows that endogenous paracrine and/or autocrine factors liberated in response to atrial wall stretch rather than myocyte stretch itself are responsible for the activation of ANP peptide secretion in response to acute volume load. Our results also show that ETA receptors are more important in the regulation of mechanical stretch-induced changes in cardiac hormone secretion than AT1 receptors.

Effects of levosimendan on cardiac remodeling and cardiomyocyte apoptosis in hypertensive Dahl/Rapp rats

Progression of heart failure in hypertensive Dahl rats is associated with cardiac remodeling and increased cardiomyocyte apoptosis. This study was conducted to study whether treatment with a novel inotropic vasodilator compound, levosimendan, could prevent hypertension‐induced cardiac remodeling and cardiomyocyte apoptosis.

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 &agr;-actin and &bgr;-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.