Rat Left Ventricle Research Articles

Different LV and RV response to α‐adrenergic stimulation in vivo

Objective: α‐adrenergic agonists (α‐Adr) exert positive cardiac inotropic effects. Recent data suggest that rat left ventricle (LV) and right ventricle (RV) respond differently to α‐Adr in vitro. The purpose of this study is to determine whether α‐Adr exerts different effects on the LV and RV in vivo.Methods: Pigs (n=8) were instrumented with micromanometers and sonomicrometry crystals in the LV and RV. Contractility was determined by preload‐adjusted regional external work, a loading condition‐independent index, and regional blood flow by microspheres. Measurements were obtained at baseline, after CaCl2 15 mg‐kg−1 bolus, and during phenylephrine (PHE) 3–6 μg‐kg−1‐min−1 IV for 60 min.Results: LV and RV contractility both increased similarly after CaCl2 (LV: 143±7% of baseline; RV: 145±10% of baseline); in contrast, PHE increased LV but decreased RV contractility (LV: 147±10% of baseline; RV: 90±19% of baseline; p<0.05 for paired difference) despite increased LV and RV pressure. LV and RV regional blood flow both increased during PHE (LV: 0.8±0.2 and RV 0.6±0.1 ml‐gm−1‐min−1 at baseline; LV: 1.5±0.2 and RV: 1.0±0.2 ml‐gm−1‐min−1 during PHE).Conclusions: The α‐Adr agent phenylephrine exerts positive inotropic effects on the LV but negative inotropic effects on the RV in vivo. This difference is not due to differences in loading conditions or regional myocardial blood flow. Supported by NHLBI.

ANESTHETIC PRECONDITIONING CONFERS ACUTE CARDIOPROTECTION VIA UP-REGULATION OF MANGANESE SUPEROXIDE DISMUTASE AND PRESERVATION OF MITOCHONDRIAL RESPIRATORY ENZYME ACTIVITY

The cellular and molecular mechanisms that underlie cardioprotection against I/R by anesthetic-induced preconditioning (APC) require further elucidation. Using isoflurane as a representative anesthetic, we evaluated the hypothesis that APC induces myocardial protection against I/R by attenuation of excessive reactive oxygen species and restoration of mitochondrial bioenergetics through postischemic up-regulation of manganese superoxide dismutase (MnSOD) expression and preservation of respiratory enzyme activity. Pentobarbital anesthetized open-chest Sprague-Dawley rats were subject to 30-min left coronary artery occlusion, followed by 120-min reperfusion. Before ischemia, rats were randomly assigned to receive 0.9% saline, two cycles of brief coronary artery occlusion and reperfusion, or a 30-min exposure to 1.0 minimum alveolar concentration isoflurane in the absence or presence of a specific mitochondrial adenosine triphosphate-sensitive potassium (KATP) channel blocker, 5-hydroxydecanoate; a membrane-permeable superoxide scavenger, 4-hydroxy-2,2,6,6-tetramethyl piperidinoxyl; or a NOS inhibitor, N(G)-nitro-L-arginine methyl ester. Isoflurane exposure induced an initial increase in myocardial superoxide (O2-), but not NO level. It also significantly decreased infarct size and restored mitochondrial respiratory enzyme activity or ATP production in I/R rat hearts, along with suppression of the O2- surge at reperfusion and increase in MnSOD expression or enzyme activity. These protective effects were abrogated by 5-hydroxydecanoate or 4-hydroxy-2,2,6,6-tetramethyl piperidinoxyl, but not by N(G)-nitro-L-arginine methyl ester pretreatment. These results suggest that opening of mitochondrial KATP channel, followed by O2- signaling, induces postischemic augmentation of MnSOD and preservation of mitochondrial respiratory enzyme activities, leading to attenuated cardiac O2- surge and restored ATP production during reperfusion, and underlie APC-induced cardioprotection.

Adeno‐associated virus vector‐mediated systemic interleukin‐10 expression ameliorates hypertensive organ damage in Dahl salt‐sensitive rats

Inflammation plays an important role in the pathogenesis of hypertension and hypertensive organ damage. Interleukin (IL)-10, a pleiotropic anti-inflammatory cytokine, exerts vasculoprotective effects in many animal models. In the present study, we examined the preventive effects of adeno-associated virus (AAV) vector-mediated sustained IL-10 expression against hypertensive heart disease and renal dysfunction in Dahl salt-sensitive rats. We injected the rats intramuscularly with an AAV type 1-based vector encoding rat IL-10 or enhanced green fluorescent protein (EGFP) at 5 weeks of age; subsequently, the rats were fed a high-sodium diet from 6 weeks of age. Sustained IL-10 expression significantly improved survival rate of Dahl salt-sensitive rats compared with EGFP expression (62.5% versus 0%, p < 0.001); it also caused 26.0% reduction in systolic blood pressure at 15 weeks (p < 0.0001). Echocardiography exhibited a 22.0% reduction in hypertrophy (p < 0.0001) and a 26.3% improvement in fractional shortening (p < 0.0001) of the rat left ventricle in the IL-10 group compared to the EGFP group. IL-10 expression also caused a 21.7% decrease in the heart weight/body weight index and cardiac atrial natriuretic peptide levels. Histopathological studies revealed that IL-10 decreased inflammatory cell infiltration, fibrosis, and transforming growth factor-beta(1) levels in the failing heart. Furthermore, IL-10 expression significantly reduced urine protein excretion with increased glomerular filtration rates. This is the first study to demonstrate that the anti-inflammatory cytokine IL-10 has a significant anti-hypertensive effect. AAV vector-mediated IL-10 expression potentially prevents the progression of refractory hypertension and hypertensive organ damage in humans.

Effect of β2-agonist, clenbuterol on the mRNA expression of β adrenoceptors in skeletal muscles and left ventricle of rats

Effect of β2-agonist, clenbuterol on the mRNA expression of β adrenoceptors in skeletal muscles and left ventricle of rats

Effects of Castration and Testosterone Replacement on the Antioxidant Defense System in Rat Left Ventricle

There is strong evidence that oxidative stress plays a key role in the pathophysiology of several cardiovascular diseases. On the other hand, the presence of specific receptors for androgens and estrogens in the myocardium implies that sex hormones play a physiological role in cardiac function, myocardial injury, and the regulation of the redox state in the heart. The present study was designed to determine whether castration and androgen replacement result in changes in the capacity of the antioxidant defense system in the left ventricle (LV) of adult male rats. To assess this, the activities of antioxidant enzymes (superoxide dismutase [SOD], glutathione peroxidase [GPX], catalase [CAT], and glutathione reductase [GR]), concentrations of nonenzymatic antioxidants (reduced glutathione [GSH] and alpha- and gamma-tocopherols), and oxidative stress biomarkers (tissue sulfhydryl groups, protein nitrotyrosine levels, and lipid peroxidation) were measured in castrated animals (CAS), castrates replaced with testosterone (CAS+T), and sham-operated controls (Sham). Testosterone was not detectable in serum from gonadectomized rats. The results indicate that castration significantly and negatively affected the antioxidant status of rat LV, as evidenced by a significant decline in activities of all antioxidant enzymes, by a tendency toward lower levels of GSH and protein thiol groups, and by enhanced lipid peroxidation and higher nitrotyrosine concentrations in left ventricular tissue. Increases in LV tissue concentrations of alpha- and gamma-tocopherols seem to be a compensatory response to enhanced oxidative stress induced by gonadectomy. The reestablishment of physiological serum testosterone level by androgen replacement resulted in a tendency toward a further decrease in the antioxidant defense status in the LV tissue.

E-NTPDases and ecto-5′-nucleotidase expression profile in rat heart left ventricle and the extracellular nucleotide hydrolysis by their nerve terminal endings

In this study, we have identified the E-NTPDase family members and ecto-5′-nucleotidase/CD73 in rat heart left ventricle. Moreover, we characterize the biochemical properties and enzyme activities from synaptosomes of the nerve terminal endings of heart left ventricle. We observe divalent cation-dependent enzymes that presented optimum pH of 8.0 for ATP and ADP hydrolysis, and 9.5 for AMP hydrolysis. The apparent K M values are 40 μM, 90 μM and 39 μM and apparent V max values are 537, 219 and 111 nmol Pi released/min/mg of protein for ATP, ADP and AMP hydrolysis, respectively. Ouabain, orthovanadate, NEM, lanthanum and levamisole do not affect ATP and ADP hydrolysis in rat cardiac synaptosomes. Oligomycin (2 μg/mL) and sodium azide (0.1 mM), both mitochondrial ATPase inhibitors, inhibit only the ATP hydrolysis. High concentrations of sodium azide and gadolinium chloride show an inhibition on both, ATP and ADP hydrolysis. Suramin inhibit more strongly ATP hydrolysis than ADP hydrolysis whereas Evans blue almost abolish both hydrolysis. AMP hydrolysis is not affected by levamisole and tetramisole, whereas 0.1 mM ammonium molybdate practically abolish the ecto-5′-nucleotidase activity. RT-PCR analysis from left ventricle tissue demonstrate different levels of expression of Entpd1 ( Cd39), Entpd2 ( Cd39L1), Entpd3 ( Cd39L3), Entpd5 ( Cd39L4) Entpd6, ( Cd39L2) and 5′-NT/CD73. By quantitative real-time PCR we identify the Entpd2 as the enzyme with the highest expression in rat left ventricle. Our results contribute to the understanding about the control of the extracellular nucleotide levels in and cardiac system.

Abstract 3294: Superoxide-Dependent Cathepsin Activation System is Associated with Hypertensive Myocardial Remodeling and Represents a Target for Angiotesin II Type 1 Receptor Blocker Therapy

Objective: Cathepsin (Cat) activation contributes to tissue remodeling, and increased Cat activity and superoxide production has been observed in the left ventricle (LV) during heart failure (HF). In the present study we tested the hypothesis that superoxide-dependnet Cat activation inhibition prevents LV remodeling and dysfunction in the development of HF associated with hypertension. Methods and Results: We treated 8% salt-loaded Dahl salt-sensitive hypertensive rats (n=10 for each group) with vehicle hydralazine (5 mg/kg/d) E64d (a synthesis Cat inhibitor, 10 mg/kg/d), or olmesartan (OLM, 5 mg/kg/d) for 8 weeks. The rats fed 0.3% salt served as age-matched controls. The abundance of Cat mRNAs and proteins localized in cardiac myocytes (CMCs) and Cat-dependent activities were increased in the LV of HF rats and/or humans, and were reduced by OLM treatment. OLM suppressed the elastic lamina degradation by 56% (n=6, p&lt;0.01) concomitant with decreased local Cat S expression in intracoronary smooth muscle cells (SMCs) and restored the balance of elastin to collagen in the LV tissue of HF rats (HF 4.6 ± 0.9% vs. OLM 15.5 ± 2.1% elastin content/collagen content (%), n=6, P=0.031; control 22 ± 2.1%). Furthermore, OLM suppressed not only angiotensin converting enzyme and angiotensin II type 1 receptor (AT 1 R) and macrophage infiltration but also levels of NADPH oxidase components (p22 phox , gp91 phox , and p47 phox) concomitant with decreased NADPH activity and O 2 − production in LV tissues of HF rats and humans. These changes were accompanied by improved cardiac fibrosis, stiffness, and dysfunction. Interestingly, LV remodeling and dysfunction were partially improved by treatment with E64d. In vitro, H 2 O 2 stimulated Cat S mRNA and protein expression and activity, and these increases were abolished by pretreatment with MnTmPyp (50 μ mol/L) and N-acetylcysteine (5 mmol/L) as well as apocynin (100 μ mol/L) in culture CMCs (n=6, p&lt;0.01). Conclusions: This is the first report showing that Cats are likely to trigger and promote LV remodeling, and that AT 1 R inhibition exerts inhibitory effect on Cat activation system by the inhibition of NADPH oxidase-dependent superoxide anion production, leading to the prevention of cardiac remodeling and dysfunction.

Biochemical characterization of ecto-nucleotide pyrophosphatase/phosphodiesterase (E-NPP, E.C. 3.1.4.1) from rat heart left ventricle

In the present study we investigate the biochemical properties of the members of NPP family in synaptosomes prepared from rat heart left ventricles. Using p-nitrophenyl-5'-thymidine monophosphate (p-Nph-5'-TMP) as substrate for E-NPPs in rat cardiac synaptosomes, we observed an alkaline pH dependence, divalent cation dependence and the K ( M ) value corresponded to 91.42 +/- 13.97 microM and the maximal velocity (V ( max )) value calculated was 63.79 +/- 3.59 nmol p-nitrophenol released/min/mg of protein (mean +/- SD, n = 4). Levamisole (1 mM), was ineffective as inhibitor of p-Nph-5'-TMP hydrolysis in pH 8.9 (optimum pH for the enzyme characterized). Suramin (0.25 mM) strongly reduced the hydrolysis of p-Nph-5'-TMP by about 46%. Sodium azide (10 and 20 mM) and gadolinium chloride (0.3 and 0.5 mM), E-NTPases inhibitors, had no effects on p-Nph-5'-TMP hydrolysis. RT-PCR analysis of left ventricle demonstrated the expression of NPP2 and NPP3 enzymes, but excluded the presence of NPP1 member. By quantitative real-time PCR we identified the NPP3 as the enzyme with the highest expression in rat left ventricle. The demonstration of the presence of the E-NPP family in cardiac system, suggest that these enzymes could contribute with the fine-tuning control of the nucleotide levels at the nerve terminal endings of left ventricles that are involved in several cardiac pathologies.

Specific adaptations of estrogen receptor α and β transcripts in liver and heart after endurance training in rats

Estrogens exert their biological roles mainly through estrogen receptors (ER) that function as ligand-activated transcription factors. ER content in a cell is regulated by many factors and is decisive for estrogen action. The purpose of the present study was to investigate the influence of an 8-wk endurance training program on ER expression in the liver, right atrium (RA), and left ventricle (LV) of intact and ovariectomized (Ovx) rats. We measured ERalpha and ERbeta mRNA content by reverse transcription-polymerase chain reaction (RT-PCR). We found an important increase in ERalpha mRNA levels in the liver (300%; P < 0.01) and in ERbeta mRNA levels in the RA (200%; P < 0.05), and a marked decrease in ERalpha (80%; P < 0.01) and ERbeta (40%; P < 0.05) transcripts content in the LV of intact rats after endurance training. On the other hand, ERalpha mRNA levels were depressed by 50% (P < 0.01) in the liver, and increased by 60% (P < 0.01) in LV of Ovx rats after exercise training. These results first indicate that endurance training is associated with modifications of ER transcripts levels in the liver, LV, and RA of female rats. More specifically, these effects are tissue and isoform-specific and the direction of the response (increase or decrease) is different in intact and Ovx rats. It is suggested that some of the adaptations to endurance training in liver and heart may be mediated by an ER-dependent mechanism.

Effects of acute mechanical stretch on the expression of mechanosensitive potassium channel TREK-1 in rat left ventricle

To explore the role of mechanosensitive potassium channel TREK-1, Western blot analysis was used to investigate the expression changes of TREK-1 in left ventricle in acute mechanically stretched heart. Forty Wistar rats were randomly divided into 8 groups (n=5 in each group), subject to single Langendorff perfusion for 0, 30, 60, 120 min and acute mechanical stretch for 0, 30, 60, 120 min respectively. With Langendorff apparatus, an acute mechanically stretched heart model was established. There was no significant difference in the expression of TREK-1 among single Langendorff perfusion groups (P>0.05). As compared to non-stretched Langendorff-perfused heart, only the expression of TREK-1 in acute mechanically stretched heart (120 min) was greatly increased (P<0.05). This result suggested that some course of mechanical stretch could up-regulate the expression of TREK-1 in left ventricle. TREK-1 might play an important role in mechanoelectric feedback, so it could reduce the occurrence of arrhythmia that was induced by extra mechanical stretch.

Time course of changes in the expression of DHPR, RyR2, and SERCA2 after myocardial infarction in the rat left ventricle

Postinfarction left ventricular remodeling leads to the functional decline of the left ventricle (LV). Since dihydropyridine receptor (DHPR), ryanodine receptor (RyR(2)), and sarco-endoplasmic reticulum (SR) Ca(2+)-ATPase2 (SERCA2a) play a major role in the contractility of the heart, the aim of our study was to evaluate the time course of changes in the expression of these proteins 1 day, 2 weeks and 4 weeks after myocardial infarction (MI). Myocardial infarction was produced by ligation of left anterior descending coronary artery of the rat. Transthoracic echocardiography was performed to characterize structural and functional changes after MI. To evaluate protein mRNA levels and the relative amount of proteins, real-time quantitative RT-PCR and Western blotting were used. LV ejection fraction and fractional shortening decreased significantly during the 4-week follow-up period (P < 0.001). Typical features of LV remodeling after MI were seen, with a decrease in anterior wall thickness (P < 0.001) and dilatation of the LV (P < 0.001). Expression of DHPR and RyR(2) mRNAs decreased and Serca2a mRNA tended to decrease 1 day after MI (P < 0.001, P < 0.01 and P = 0.06, respectively), followed by recovery of the expression during the next 4 weeks. In the infarcted hearts the quantities of SERCA2 proteins in the LV were significantly decreased at the time of 4 weeks. In conclusion, MI was associated with transient decrease in the expression of the DHPR and RyR(2) mRNAs and a reduced quantity of SERCA2 proteins in the LV. Since they have a key role in the contraction of the heart, changes in the expression of these proteins may be important regulators of LV systolic function after MI.

TNF-α blockade decreases oxidative stress in the paraventricular nucleus and attenuates sympathoexcitation in heart failure rats

Oxidative stress plays an important role in the pathophysiology of cardiovascular disease. Recent evidence suggests that cytokines induce oxidative stress and contribute to cardiac dysfunction. In this study, we investigated whether increased circulating and tissue levels of tumor necrosis factor (TNF)-alpha in congestive heart failure (CHF) modulate the expression of NAD(P)H oxidase subunits, Nox2 and its isoforms, in the paraventricular nucleus (PVN) of the hypothalamus and contribute to exaggerated sympathetic drive in CHF. Heart failure was induced in Sprague-Dawly rats by coronary artery ligation and was confirmed using echocardiography. Pentoxifylline (PTX) was used to block the production of cytokines for a period of 5 wk. CHF induced a significant increase in the production of reactive oxygen species (ROS) in the left ventricle (LV) and in the PVN. The mRNA and protein expression of TNF-alpha, Nox1, Nox2, and Nox4 was significantly increased in the LV and PVN of CHF rats. CHF also decreased ejection fraction, increased Tei index, and increased circulating catecholamines (epinephrine and norepinephrine) and renal sympathetic activity (RSNA). In contrast, treatment with PTX in CHF rats completely blocked oxidative stress and decreased the production of TNF-alpha and Nox2 isoforms both in the LV and PVN. PTX treatment also decreased catecholamines and RSNA and prevented further decrease in cardiac function. In summary, TNF-alpha blockade attenuates ROS and sympathoexcitation in CHF. This study unveils new mechanisms by which cytokines play a role in the pathogenesis of CHF, thus underscoring the importance of targeting cytokines in heart failure.

Identification of PDE1 and PDE5 in adult rat left ventricular cardiomyocytes

The cardiac tissue heterogeneity has raised question on the existence of cyclic nucleotide phosphodiesterase (PDE) isozymes, namely PDE1 and PDE5 in cardiomyocytes. PDE isozyme activities were characterized in adult rat left ventricle (LV) whole homogenate by using specific PDE inhibitors. Cardiomyocytes derived from adult rat LV were cultured; PDE activities of cardiomyocyte soluble fraction were resolved by MonoQ HPLC. PDE expression w as investigated by Western blot on cardiomyocyte whole homogenate.

Transmural variations in gene expression of stretch-modulated proteins in the rat left ventricle

The properties of left ventricular cardiac myocytes vary transmurally. This may be related to the gradients of stress and strain experienced in vivo across the ventricular wall. We tested the hypothesis that within the rat left ventricle there are transmural differences in the expression of genes for proteins that are involved in mechanosensitive pathways and in associated physiological responses. Real time reverse transcription polymerase chain reaction was used to measure messenger RNA (mRNA) levels of selected targets in sub-epicardial (EPI) and sub-endocardial (ENDO) myocardium. Carbon fibres were attached to single myocytes to stretch them and to record contractility. We observed that the slow positive inotropic response to stretch was not different between EPI and ENDO myocytes and consistent with this, that the mRNA expression of two proteins implicated in the slow response, non-specific cationic mechanosensitive channels (TRPC-1) and Na/H exchanger, were not different. However, mRNA levels of other targets, e.g. the mechanosensitive K+ channel TREK-1, Brain Natriuretic Peptide and Endothelin-1 receptor B, were significantly greater in ENDO than EPI. No targets had significantly greater mRNA levels in EPI than ENDO. On the basis of these findings, we suggest that the response of the ventricle to stretch will depend upon both the regional differences in stimuli and the relative expression of the mechanosensitive targets and that generally, stretch sensitivity is predicted to be greater in ENDO.

Exercise training maintains cardiac output and stroke volume in hypertensive TG (mREN‐2)27 rats with impaired diastolic function

The role of the renin-angiotensin system (RAS) on in vivo cardiac function, blood pressure, and markers of maladaptive cardiac hypertrophy were determined after low-intensity exercise training (ET) in Ren-2 rats and age-matched Sprague Dawley (SD) controls (3–4 weeks). After 5 weeks of treadmill ET, echocardiography demonstrated that both cardiac output (ml / min) and stroke volume (ml) per body weight (g) were increased (p<0.05) by ET in both SD and Ren-2. The slope of mitral deceleration time (m/sec2), a non-invasive measure of diastolic function, was lower in the Ren-2 rats, but not changed by ET in either rat strain (p>0.05). Systolic, diastolic, and mean (MAP) blood pressures via tail-cuff method in Ren-2 rats were higher compared to the SD controls, while ET had no effect. LV collagen deposition, as assessed by hydroxyproline assay, was not affected by rat strain or ET. B-type natriuretic peptide (BNP) mRNA levels measured in the left ventricle (LV) by real-time PCR were higher in the Ren-2 rats (100%), but not affected by ET (p=0.81). Both α (~14.5 fold) and β (~2.5 fold) myosin heavy chain mRNA were higher in the LV of Ren-2 rats compared to SD (p < 0.05), but were not changed by ET. These data suggest that ET restores cardiac index and stroke volume in the Ren-2 rat.

Formation of leptofibrils is associated with remodelling of muscle cells and myofibrillogenesis in the border zone of myocardial infarction

Leptofibrils, or leptomeres, remain the least studied cytoskeletal structures in muscle cells, and their function and mechanism of assembly are still poorly understood. Our ultrastructural study of the surviving cardiac myocytes located in the perinecrotic border zone of the infarcted left ventricle in rats revealed intense formation of leptofibrils and leptofibrillar clusters during 4–15 days following experimental myocardial infarction. In the perinecrotic myocytes, leptofibrils developed predominantly in the subsarcolemmal areas, near disassembled intercalated discs and at the sites of intense myofibrillogenesis in the peripheral zones of the sarcoplasm. We found that the development of these structures occurred before or at the time of assembly of myofibrils. In our material, leptofibrils consisted of longitudinally oriented filamentous bundles inserted in electron dense Z-band-like material and periodically crossed by 3–8 bands of this material with the period of cross-striation of 120–210 nm. The presence of leptofibrils in growing cytoplasmic processes and ruffles developing in the border zone in the areas of lost intercellular contacts indicates their formation de novo during post-infarction period. We observed four major morphological types of localization of these structures: (1) direct contact of one end of leptofibrils with Z bands of nascent, mature or disassembling myofibrils; (2) direct contact with the sarcolemma: (a) multifocal attachment of leptofibrils to the sarcolemma through the lateral surfaces of their minute Z band-like structures; (b) attachment of one or both ends of leptofibrils to the sarcolemma without contacts or in contact with myofibrils; (3) attachment of leptofibrils to subsarcolemmal accumulations of electron dense Z-band material in newly formed fasciae adherentes of the remodeled intercalated disks; (4) clustering and contacts of leptofibrils with one another predominantly at the level of their Z bands. Interestingly, most leptofibrils of all four types were topographically associated with the system of T-tubules, the sarcoplasmic reticulum and subsarcolemmal vesicles. Serial sections through the areas containing leptofibrils indicate their spindle-like or nearly cylindrical shape. Thus, we found that leptofibrils assemble in terminally differentiated cardiac myocytes following destabilization of their differentiated state and partial dedifferentiation induced by myocardial infarction. The results of this study demonstrate that formation of leptofibrils, earlier described mainly in the developing and malignant muscle, is temporally associated with adaptive structural remodelling and the activation of myofibrillogenesis in functionally overloaded cardiac myocytes of adult animals. Our findings suggest that re-expression of some structural characteristics of the embryonic muscle appear to represent one of the mechanisms that underlie adaptive plasticity of the myocardium following injury and under conditions of hyperfunction.

Soluble Guanylate Cyclase Stimulation on Cardiovascular Remodeling in Angiotensin II–Induced Hypertensive Rats

It is unknown whether long-term pharmacological stimulation of soluble guanylate cyclase (sGC), elevating intracellular cGMP levels, has a beneficial effect on hypertension. The purpose of this study is to investigate the effects of BAY41-2272, an orally available sGC stimulator, on cardiovascular remodeling in hypertensive rats. Eight-week-old male Wistar rats with hypertension induced by angiotensin II infused subcutaneously at 250 ng/kg per minute were treated orally with a low ([L] 2 mg/kg per day) or high ([H] 10 mg/kg per day) dose of BAY41-2272 for 14 days. BAY41-2272-H partially suppressed the rise in blood pressure and reduced the heart weight (4.20+/-0.34 versus 3.68+/-0.20 mg/g; P<0.01), whereas BAY41-2272-L had no effect. However, both doses decreased the angiotensin II-induced left ventricular accumulation of collagen in the perivascular area (L, -20%, P<0.05; H, -30%, P<0.01) and myocardial interstitium (L, -21%, P<0.05; H, -38%, P<0.01), reducing the number of activated fibroblasts surrounding coronary arteries (L, -74%; H, -79%; P<0.05). BAY41-2272 downregulated the angiotensin II-induced left ventricular gene expression of type 1 collagen (L, -41%, P<0.05; H, -49%, P<0.01) and transforming growth factor-beta1 (L, -49%, P<0.05; H, -65%, P<0.01). cGMP levels were elevated by BAY41-2272 not only in the left ventricle, but also in cultured cardiac fibroblasts, resulting in reduced thymidine incorporation into the cells. Thus, stimulation of sGC by BAY41-2272 attenuates fibrosis of the left ventricle in rats with angiotensin II-induced hypertension partly in a pressure-independent manner, suggesting an important role for sGC generating cGMP in inhibiting cardiovascular remodeling.

Effect of diastolic pressure on MLC2v phosphorylation in the rat left ventricle

The effect of passive muscle stretch on the extent of MLC2v phosphorylation was investigated. We used an isolated rat heart preparation and controlled the passive pressure of the left ventricle (LV) at 0 or 15 mmHg. The hearts were flash frozen and the LV free wall was split into epicardial and the endocardial halves. The samples were solubilized using a novel method that minimizes changes in the phosphate content of MLC2v under non-denaturing conditions. The proteins were separated by urea glycerol PAGE and identified by mass spectrometry and Western blots. At 0 mmHg passive pressure, the extent of MLC2v phosphorylation of the epicardium (34.1 ± 1.7%) was the same as that of the endocardium (35.3 ± 3.4%). At 15 mmHg passive pressure, we found a significant increase in MLC2v phosphorylation in the epicardium (to 41.5 ± 2.0%) and a significant reduction in the endocardium (to 24.2 ± 1.2%), giving rise to a gradient in the extent of MLC2v phosphorylation from epicardium (high) to endocardium (low). These changes in MLC2v phosphorylation that take place in response to increased diastolic pressure are likely to impact on the calcium sensitivity of actomyosin interaction (with an increased sensitivity towards the epicardium) and may play a role in the Frank–Starling mechanism of the heart.

Twelve Weeks of Exercise Training Upregulates Mn-SOD while Reducing iNOS and Oxidative Stress in the Rat Left Ventricle

Twelve Weeks of Exercise Training Upregulates Mn-SOD while Reducing iNOS and Oxidative Stress in the Rat Left Ventricle

Effect of Exercise on the Expression of HSP20 in the Hypertensive Rat Heart

Previous observations show that levels of Hsp20, a small heat shock protein, are elevated in the rat left ventricle by exercise training. PURPOSE: To determine whether exercise training would induce an increase in the levels of Hsp20 in hearts of spontaneously hypertensive rats (SHR). METHODS: SHR were randomly assigned to a sedentary (SHR-SED) or trained (SHR-TRN) group. The trained group was exercised on a treadmill at 0% grade for eight weeks, using a high intensity interval training protocol. After 3 weeks of introductory running, 2.5 min of sprint running at 29–45 m/min was interspersed with steady running at 24 m/min for five minutes for a total duration of 37 minutes. The number of sprints was increased progressively so that the duration was approximately 60 minutes after eight weeks. During the last week of the study, sedentary rats were habituated to the treadmill for 5 days for 10 min each at 15 m/min. On the last day of the study, sedentary and exercise-trained rats were run for 10 minutes at 15 or 25 m/min respectively, and hearts were harvested at various times (t=0,10,30,60,180 min) after the last bout of exercise. From groups of SHR-SED and SHR-TRN rats, heart left (LV) and right ventricle (RV) free walls and septum (SEP) were used to measure the levels of Hsp20. Western blotting was performed using 12% gels, PVDF membranes, and an enhanced chemiluminescent detection method. A rabbit polyclonal anti- HSP20 antibody was used to detect the protein. Levels of Hsp20 were normalized to those of glyceraldehyde-3-phosphate dyhydrogenase (GAPDH). RESULTS: The level of HSP20 expression was significantly increased in response to exercise training in the soluble fraction (+61%) and whole homogenate (+70%) of the RV. In the SEP, the increase in HSP20 expression level was observed only in the soluble fraction (+54%) while the expression level was not statistically different in the whole homogenate. Exercise training did not trigger increased expression of HSP20 in the hypertensive LV free wall. In the RV and LV free walls, the level of HSP20 was higher in the soluble fraction than in the whole homogenate, and the ratio of the two measurements was unchanged by exercise-training. On the other hand, the level of HSP20 in the SEP was greater in the whole homogenate, and the soluble-to-whole homogenate ratio was increased as a result of exercise training (p < 0.05). CONCLUSION: Exercise training triggers increased expression of HSP20 in the unaffected RV free wall of hypertensive rats, but not in the pressure-overloaded LV free wall. Supported by the American Heart Association Midwest Affiliate and the Undergraduate Research Opportunity Program at the University of Michigan