Peter Krenek

Isoproterenol-induced heart failure in the rat is associated with nitric oxide-dependent functional alterations of cardiac function

The role of nitric oxide (NO) in heart failure (HF) is complex and remains controversial. We tested the hypothesis that the role of NO in isolated atria and cardiomyocytes is altered in isoproterenol‐induced HF.

Lacidipine prevents endothelial dysfunction in salt-loaded stroke-prone hypertensive rats.

Endothelium-dependent vasorelaxation is defective in hypertensive rats, especially in conduit arteries. In the stroke-prone spontaneously hypertensive rat, impaired endothelium-dependent vasorelaxation appears to contribute to the pathogenesis of stroke independent of blood pressure. Because treatment with lacidipine, a long-acting calcium channel blocker, protects against stroke and cardiovascular remodeling in this model, we investigated the effect of this treatment on endothelium-dependent vasorelaxation in the aorta. Stroke-prone rats were exposed to a salt-rich diet (1% NaCl in drinking water) with or without lacidipine (1 mg · kg−1 · d−1) for 6 weeks. A high-sodium diet (1) increased systolic blood pressure, aortic weight, and wall thickness and plasma renin activity (P <0.05); (2) markedly reduced nitric oxide (NO)-mediated, endothelium-dependent relaxation of aortic rings to acetylcholine and the sensitivity to the relaxing effect of S-nitroso-N-acetylpenicillamine, an NO donor (P <0.001); and (3) induced an elevation of preproendothelin-1 mRNA levels in aortic tissue (P <0.01) without affecting endothelial NO synthase mRNA levels. Lacidipine treatment prevented the salt-dependent functional and structural alterations of the aorta, including the overexpression of the preproendothelin-1 gene, and increased endothelial NO synthase mRNA levels in aortic tissue (P <0.01). In conclusion, lacidipine protects stroke-prone hypertensive rats against the impairment of endothelium-dependent vasorelaxation evoked by a salt-rich diet, and this effect may contribute to its beneficial effect against end-organ damage and stroke.

Rosmarinic acid administration attenuates diabetes-induced vascular dysfunction of the rat aorta

Oxidative stress as well as inflammation processes are engaged in diabetic vascular complications. Rosmarinic acid, a natural phenol antioxidant carboxylic acid, was found to have multiple biological activity, including anti‐inflammatory and antitumour effects, which are a consequence of its inhibition of the inflammatory processes and of reactive oxygen species scavenging. The aim of this work was to study effects of rosmarinic acid administration on vascular impairment induced by experimental diabetes in rats.

Effects of amlodipine and lacidipine on cardiac remodelling and renin production in salt-loaded stroke-prone hypertensive rats

Calcium channel blockers (CCBs) are anti‐hypertensive drugs that are usually considered to act mainly as vasodilators. We investigated the relation between the reduction of blood pressure evoked by two long‐acting CCBs and their protective effect against cardiac and renal damage in salt‐loaded stroke‐prone spontaneously hypertensive rats (SHRSP). SHRSP were exposed to high dietary salt intake (1% NaCl in drinking solution) from 8 to 14 weeks of age, with or without amlodipine or lacidipine at three dosage regimens producing similar effects on blood pressure. The lowest dosages of both drugs had non‐significant effects on blood pressure but inhibited the paradoxical increases in plasma renin activity (PRA) and in renin mRNA in kidney that were found in salt‐loaded SHRSP. The lowest dosage of lacidipine (but not of amlodipine) restored the physiological downregulation of renin production by high salt and reduced left ventricular hypertrophy and mRNA levels of atrial natriuretic factor and transforming growth factor‐β1. The intermediate dosages reduced blood pressure and PRA in a comparable manner, but cardiac hypertrophy was more reduced by lacidipine than by amlodipine. Although the highest doses exhibited a further action on blood pressure, they had no additional effect on cardiac hypertrophy, and they increased PRA and kidney levels of renin mRNA even more than in the absence of drug treatment. We conclude that reduction of blood pressure is not the sole mechanism involved in the prevention of cardiac remodelling by CCBs, and that protection against kidney damage and excessive renin production by low and intermediate dosages of these drugs contributes to their beneficial cardiovascular effects.

Discrepancy between increased left ventricular mass and “normal” QRS voltage is associated with decreased connexin 43 expression in early stage of left ventricular hypertrophy in spontaneously hypertensive rats

BACKGROUND AND PURPOSE On the basis of our previous results of animal and human studies, we assume that the discrepancies between increased left ventricular mass (LVM) and electrocardiographic (ECG) findings not exceeding the upper normal limits in left ventricular hypertrophy (LVH) are conditioned by the electrical remodeling of hypertrophied myocardium. We assumed that these discrepancies observed in the early stage of LVH in spontaneously hypertensive rats (SHR) are associated with a decreased expression of connexin 43. METHODS Standard 12-lead ECG was recorded in 20-week-old male SHR and age-matched and sex-matched normotensive Wistar rats (Institute of Experimental Pharmacology SAV, Dobra Voda, Slovakia). The approximated maximum QRS spatial vector magnitude (QRSmax) was calculated from leads V(2), aVF, and V(5). Left ventricular mass was weighed, and the specific potential (SP) of myocardium was calculated as the QRSmax-to-LVM ratio. Left ventricular protein levels of connexin 43 were analyzed with sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting. RESULTS The LVM values were significantly higher in SHR than in normotensive controls (0.96 +/- 0.03 g and 0.680 +/- 0.07 g, respectively; P < .001). The QRSmax values in SHR did not follow the increase either in systolic blood pressure or in LVM. The SP values in SHR were significantly lower than those in control rats (0.92 +/- 0.11 mV/g and 1.358 +/- 0.06 mV/g, respectively; P < .01). A 37% decrease in connexin 43 level was observed in SHR. CONCLUSIONS The QRS voltage did not follow the increase in the LVM in 20-week-old SHR, and the values of connexin 43 were lower in SHR than in normotensive controls. We believe that the discrepant findings between ECG voltage and LVM can be caused by the electrical remodeling in the early stages of LVH.

Effect of chronic nNOS inhibition on blood pressure, vasoactivity, and arterial wall structure in Wistar rats.

While the unequivocal pattern of endothelial nitric oxide (NO) synthase (eNOS) inhibition in cardiovascular control has been recognised, the role of NO produced by neuronal NOS (nNOS) remains unclear. The purpose of the present study was to describe the cardiovascular effects of NO production interference by inhibition of nNOS with 7-nitroindazole (7-NI). Wistar rats (10 weeks old) were used: control and experimental rats were administered 7-NI 10 mg/kg b.w./day in drinking water for 6 weeks. Systolic blood pressure (BP) was measured by the tail-cuff plethysmographic method. Isolated thoracic aortas (TAs) were used to study vasomotor activity of the conduit artery in vitro. The BP response of anaesthetised animals was used to follow the cardiovascular-integrated response in vivo. Geometry of the TA was measured after perfusion fixation (120 mm Hg) by light microscopy. Expression of eNOS was measured in the TA by immunoblot analysis. Although 6 weeks of nNOS inhibition did not alter systolic BP, the heart/body weight ratio was decreased. Relaxation of the TA in response to acetylcholine (10(-9)-10(-5)mol/L) was moderately inhibited. However, no difference in the BP hypotensive response after acetylcholine (0.1, 1, 10 microg) was observed. The contraction of TA in response to noradrenaline (10(-10)-10(-5)mol/L), and the BP pressor response to noradrenaline (0.1, 1 microg) was attenuated. The inner diameter of the TA was increased, and the wall thickness, wall cross-sectional area, and wall thickness/inner diameter ratio were decreased. The expression of eNOS in the TA was increased. In summary, cardiac and TA wall hypotrophy, underlined by decreased contractile efficiency, were observed. The results suggested that two constitutive forms of NOS (nNOS, eNOS) likely participate in regulation of cardiovascular tone by different mechanisms.

Enalaprilat increases PPARβ/δ expression, without influence on PPARα and PPARγ, and modulate cardiac function in sub-acute model of daunorubicin-induced cardiomyopathy

Anthracycline therapy is limited by a cardiotoxicity that may eventually lead to chronic heart failure which is thought to be prevented by ACE inhibitors (ACEi). However, the protective effect of ACEi in early stages of this specific injury remains elusive. Activated nuclear transcription factors peroxisome proliferator-activated receptors (PPAR) regulate cellular metabolism, but their involvement in anthracycline cardiomyopathy has not been investigated yet. For this purpose, Wistar rats were administered with daunorubicin (i.p., 3 mg/kg, in 48 h intervals) or co-administered with daunorubicine and enalaprilat (i.p., 5 mg/kg in 12 h intervals). Control animals received vehicle. Left ventricular function was measured invasively under anesthesia. Cell-shortening was measured by videomicroscopy in isolated cardiomyocytes. Expression of PPARs mRNA in cardiac tissue was measured by Real-Time PCR. Although the hemodynamic parameters of daunorubicin-treated rats remained altered upon ACEi co-administration, ACEi normalized daunorubicin-induced QT prolongation. On cellular level, ACEi normalized altered basal and isoproterenol-stimulated cardiac cell shortening in daunorubicine-treated group. Moreover, anthracycline administration significantly up-regulated heart PPARα mRNA and its expression remained increased after ACEi co-administration. On the other hand, the expression of cardiac PPARβ/δ was not altered in anthracycline-treated animals, whereas co-administration of ACEi increased its expression. Conclusively, effect of ACEi can be already detected in sub-acute phase of anthracycline-induced cardiotoxicity. Altered expression of heart PPARs may suggest these nuclear receptors as a novel target in anthracycline cardiomyopathy.

Pycnogenol® improves left ventricular function in streptozotocin-induced diabetic cardiomyopathy in rats

We studied whether Pycnogenol® (PYC) may attenuate the development of experimental streptozotocin‐induced diabetic cardiomyopathy in rat. In addition, we aimed to study whether PYC affects cardiac oxidative stress and the protein expression of reactive oxygen species (ROS)‐producing molecules (gp91phox‐containing NADPH oxidase and NO‐signalling proteins). Experimental diabetes mellitus was manifested by hyperglycaemia and impaired cardiac function estimated using left ventricular catheterisation in vivo. PYC lowered fasting plasma glucose and normalized basal cardiac function. Excessive oxidative stress in streptozotocin (STZ) hearts, evidenced by 40% increase (P < 0.05) of thiobarbituric acid reactive substances (TBARS) concentration, was associated with increased expression of gp91phox (by 75%, P < 0.05), iNOS (by 40%, P < 0.05) and alpha‐tubulin (by 49%, P < 0.05), but unchanged expression of eNOS and its alosteric regulators, as compared to CON. PYC failed to affect these expression abnormalities. Our study shows that PYC corrects diabetic cardiac dysfunction, probably by its metabolic and direct radical scavenging activity without affecting the molecular maladaptations of ROS‐producing enzymes and cytoskeletal components. Copyright

Perinatally administered losartan augments renal ACE2 expression but not cardiac or renal Mas receptor in spontaneously hypertensive rats

Since the identification of the alternative angiotensin converting enzyme (ACE)2/Ang‐(1‐7)/Mas receptor axis, renin‐angiotensin system (RAS) is a new complex target for a pharmacological intervention. We investigated the expression of RAS components in the heart and kidney during the development of hypertension and its perinatal treatment with losartan in young spontaneously hypertensive rats (SHR). Expressions of RAS genes were studied by the RT‐PCR in the left ventricle and kidney of rats: normotensive Wistar, untreated SHR, SHR treated with losartan since perinatal period until week 9 of age (20 mg/kg/day) and SHR treated with losartan only until week 4 of age and discontinued until week 9. In the hypertrophied left ventricle of SHR, cardiac expressions of Ace and Mas were decreased while those of AT1 receptor (Agtr1a) and Ace2 were unchanged. Continuous losartan administration reduced LV weight (0.43 ± 0.02; P < 0.05 versus SHR) but did not influence altered cardiac RAS expression. Increased blood pressure in SHR (149 ± 2 in SHR versus 109 ± 2 mmHg in Wistar; P < 0.05) was associated with a lower renal expressions of renin, Agtr1a and Mas and with an increase in ACE2. Continuous losartan administration lowered blood pressure to control levels (105 ± 3 mmHg; P < 0.05 versus SHR), however, only renal renin and ACE2 were significantly up‐regulated (for both P < 0.05 versus SHR). Conclusively, prevention of hypertension and LV hypertrophy development by losartan was unrelated to cardiac or renal expression of Mas. Increased renal Ace2, and its further increase by losartan suggests the influence of locally generated Ang‐(1‐7) in organ response to the developing hypertension in SHRs.

Pioglitazone, a PPARγ agonist, provides comparable protection to angiotensin converting enzyme inhibitor ramipril against adriamycin nephropathy in rat.

Peroxisome proliferator-activated receptor γ (PPARγ) agonists have been shown to ameliorate diabetic nephropathy, but much less are known about their effects in non-diabetic nephropathies. In the present study, metabolic parameters, blood pressure, aortic endothelial function along with molecular and structural markers of glomerular and tubulointerstitial renal damage, were studied in a rat model of normotensive nephropathy induced by adriamycin and treated with PPARγ agonist pioglitazone (12mg/kg, po), angiotensin converting enzyme (ACE) inhibitor ramipril (1mg/kg, po) or their combination. Pioglitazone had no effect on systolic blood pressure, marginally reduced glycemia and improved aortic endothelium-dependent relaxation. In the kidney, pioglitazone prevented the development of proteinuria and focal glomerulosclerosis to the similar extent as blood-pressure lowering ramipril. Renoprotection provided by either treatment was associated with a reduction in the cortical expression of profibrotic plasminogen activator inhibitor-1 and microvascular damage-inducing endothelin-1, and a limitation of interstitial macrophage influx. Treatment with PPARγ agonist, as well as ACE inhibitor comparably affected renal expression of the renin-angiotensin system (RAS) components, normalizing increased renal expression of ACE and enhancing the expression of Mas receptor. Interestingly, combined pioglitazone and ramipril treatment did not provide any additional renoprotection. These results demonstrate that in a nondiabetic renal disease, such as adriamycin-induced nephropathy, PPARγ agonist pioglitazone provides renoprotection to a similar extent as an ACE inhibitor by interfering with the expression of local RAS components and attenuating related profibrotic and inflammatory mechanisms. The combination of the both agents, however, does not lead to any additional renal benefit.