Ischemia/reperfusion injury is increased and cardioprotection by a postconditioning protocol is lost as cardiac hypertrophy develops in nandrolone treated rats

We hypothesized that nandrolone (ND)-abuse induces cardiac hypertrophy, increases myocardial susceptibility to ischemia/reperfusion (I/R) injury, and reduces responsiveness to postconditioning (PostC) cardioprotection. Wistar-rats were ND treated for 2 weeks (short_ND) or 10 weeks (long_ND). Vehicle-treated rats served as controls. Hearts were retrogradely perfused and left ventricular pressure (LVP) was measured before and after 30-min global ischemia. In subgroups of hearts, to induce cardioprotection a PostC protocol (five cycles of 10-s reperfusion and 10-s ischemia) was performed. β-adrenoreceptors, kinases (Akt and GSK-3β) and phosphatases (PP2A sub A and PP2A sub B) were examined by Western blot before and after ischemia. After 120-min reperfusion, infarct size was measured. Short_ND slightly increased cardiac/body weight ratio, but did not affect cardiac baseline nor post-ischemic contractile function or infarct size when compared to vehicle hearts. However, PostC limited cardiac dysfunction much more in short_ND hearts than the other groups. Although cardiac/body weight ratio markedly increased after long_ND, baseline LVP was not affected. Yet, post-ischemic contracture and infarct size were exacerbated and PostC was unable to reduce infarct size and ventricular dysfunction. While short_ND increased phosphatases, non-phosphorylated and phosphorylated Akt, long_ND reduced phosphatase-expression and Akt phosphorylation. Both short_ND and long_ND had no effect on the GSK-3β-phosphorylation but increased the expression of β₂-adrenoreceptors. In reperfusion, PostC increased Akt phosphorylation regardless of protective effects, but reduced phosphatase-expression in protected hearts only. In conclusion, short_ND improves post-ischemic myocardial performance in postconditioned hearts. However, long_ND increases myocardial susceptibility to I/R injury and abolishes cardioprotection by PostC. This increased susceptibility might be related to steroid-induced hypertrophy and/or to altered enzyme expression/phosphorylation.     

Postconditioning and intermittent bradykinin induced cardioprotection require cyclooxygenase activation and prostacyclin release during reperfusion

Postconditioning (PostC), obtained with brief intermittent cycles of ischemia alternating with reperfusion applied after the ischemic event, has been shown to reduce infarct size. Recently, we have shown that PostC triggering includes B(2) receptor activation and its downstream pathway. Moreover, we showed that BK intermittent infusion induces a cardioprotection similar to PostC. The aim of this study was to investigate the involvement of cyclooxygenase-(COX)-derivated prostaglandins, such as prostacyclin (PGI(2)) pathway in the cardioprotective action mediated by intermittent BK infusion.Isolated rat hearts underwent 30 min ischemia and 120 min reperfusion. Myocardial damage was evaluated using nitro-blue-tetrazolium staining. The production of metabolite of PGI(2), 6-keto-PGF1alpha, was evaluated with EIA assay on the samples collected during reperfusion. The perfusion pressure and the left ventricular pressure were monitored. In Control hearts, the infarct size was 64% +/- 4% of risk area. PostC reduced significantly the infarct size (28% +/- 4% P < 0.001 Vs. Control). BK intermittent protocol to mimic PostC, attenuated infarct size (40% +/- 2% P < 0.01 Vs. Control). The BK-intermittent and PostC protections were abolished with COX-inhibition. Intermittent BK and PostC enhanced the release of prostacyclin metabolite, 6-keto-PGF1alpha, in the late phase of reperfusion (i.e., 6-keto-PGF1alpha peaked 30 min after protective maneuvers). Also the stable PGI(2) analogue, Iloprost, given in the early reperfusion reduced infarct size and improved post-ischemic heart function. In conclusion, protection by PostC and intermittent BK requires COX activation and PGI(2) release during late reperfusion. These data suggest that COX must not be inhibited to have PostC protection. This finding should be kept present by future clinical studies on PostC.     

Effects of chronic hypertension and left ventricular hypertrophy on the extent of infarct expansion in rats

Left ventricular hypertrophy is an adaptive response to long standing hypertension. However, the influence of left ventricular hypertrophy with hypertension on extent of infarct expansion has not been studied. We compared the effects of left ventricular hypertrophy with hypertension on infarct expansion in spontaneously hypertensive rats (SHR, n = 76), Wistar-Kyoto rats (WKY; n = 46) and spontaneously hypertensive rats treated with delapril, an angiotensin converting enzyme (ACE) inhibitor (SHRD; n = 39). The survival rates at 7 days after myocardial infarction were 41%, 24%, and 46% for WKY, SHR, and SHRD. The survival rate of SHR was significantly lower than those of both SHRD and WKY (P < .05). In the surviving rats (18 SHR, 19 WKY, 18 SHRD), both left ventricular cavity area (LCVA) and the infarct segment length per the noninfarct segment length (FW/IVS), measured as indices of left ventricular dilation, were significantly less in SHR and SHRD than in WKY, and the thickness of the left ventricular free wall (Wth), used as an index of left ventricular thinning, was significantly higher in both SHR and SHRD than in WKY (P < .01). However, there was no significant difference in FW/IVS, LCVA, and Wth between SHR and SHRD. Hemodynamic findings 1 week after coronary occlusion demonstrated that all rats were in heart failure, and there were no significant differences in hemodynamics among the three groups. In conclusion, our findings showed that hypertrophy with hypertension reduced infarct expansion, but that reduction of blood pressure by ACE inhibitor did not reduce infarct expansion more than hypertrophy did. However, this finding suggests that an ACE inhibitor may improve the rate of survival of patients with left ventricular hypertrophy with hypertension.     

Intermittent activation of bradykinin B2 receptors and mitochondrial KATP channels trigger cardiac postconditioning through redox signaling

OBJECTIVE: Postconditioning (PostC) maneuvers allow post-ischemic accumulation of autacoids, which trigger protection. We tested if PostC-triggering includes bradykinin (BK) B2 receptor activation and its downstream pathway. METHODS AND RESULTS: Isolated rat hearts underwent 30 min ischemia and 120 min reperfusion. Infarct size was evaluated using nitro-blue tetrazolium staining. In Control hearts infarct size was 61+/-5% of risk area. PostC (5 cycles of 10 s reperfusion/ischemia) reduced infarct size to 22+/-4% (p<0.01). PostC protection was abolished by B2 BK receptor-antagonists (HOE140 or WIN64338), nitric oxide synthase-inhibitor (L-nitro-arginine-methylester), protein kinase G (PKG)-blocker (8-bromoguanosine-3',5'-cyclic-monophosphorothioate), and mitochondrial K(ATP) (mK(ATP))-blocker (5-hydroxydecanoate) each given for 3 min only. Since 3 min of BK-infusion (100 nM) did not reproduce PostC protection, protocols with Intermittent-BK infusion were used to mimic PostC: a) 5 cycles of 10 s oxygenated-no-BK/oxygenated+BK buffer; b) 5 cycles of 10 s oxygenated-no-BK/hypoxic+BK buffer. Both protocols with Intermittent-BK attenuated infarct size (36+/-5% and 38+/-4%, respectively; p<0.05 vs Control and NS vs PostC for both; NS vs each other). Intermittent-BK protection was abolished by the same antagonists used to prevent PostC protection. Intermittence of re-oxygenation only (5 cycles of 10 s oxygenated/hypoxic buffer) did not reproduce PostC. Yet, cardioprotection was triggered by intermittent mK(ATP) activation with diazoxide, but not by intermittent reactive oxygen species (ROS) generation with purine/xanthine oxidase. ROS scavengers (N-acetyl-L-cysteine or 2-mercaptopropionylglycine), given for 3 min only, abolished PostC-, Intermittent BK-and diazoxide-induced protection. CONCLUSIONS: Intermittent targeting of specific cellular sites (i.e. BK B2 receptors and mK(ATP) channels) during early reperfusion triggers PostC protection via ROS signaling. Since neither intermittent oxygenation nor exogenous ROS generators can trigger protection, it is likely that intermittent autacoid accumulation and ROS compartmentalization may play a pivotal role in PostC-triggering.     

Postconditioning with glucagon like peptide-2 reduces ischemia/reperfusion injury in isolated rat hearts: role of survival kinases and mitochondrial KATP channels

We recently reported that heart expresses functional receptors for the anorexigenic glucagon-like peptide (GLP)-2. Activation of these cardiac receptors affected basal heart performance through extracellular regulated kinase (ERK1/2) activation. Since ERK1/2 is considered one of the prosurvival kinases of postconditioning cardioprotective pathways, we hypothesized that GLP-2 directly protects the heart against ischemia/reperfusion (I/R) injury via prosurvival kinases. Wistar rat hearts were retrogradely perfused on a Langendorff perfusion apparatus. After 40-min stabilization, hearts underwent 30-min global ischemia and 120-min reperfusion (I/R group). In GLP-2 group, the hearts received 20-min GLP-2 (10(-7)?M) infusion at the beginning of the 120-min reperfusion. Perfusion pressure and left ventricular pressure (LVP) were monitored. Infarct size was evaluated by nitroblue-tetrazolium staining. Compared with the I/R group, GLP-2-treated hearts showed a significant reduction of infarct size and of postischemic diastolic LVP (index of contracture), together with a sharp improvement of developed LVP recovery (index of contractility). The protective effects were abolished by co-infusion with phosphatidylinositol 3-kinase inhibitor, Wortmannin (WT), the ERK1/2 inhibitor, PD98059, or the mitochondrial K(ATP) channel blocker, 5-hydroxydecanoate. GLP-2 effects were accompanied by increased phosphorylation of protein kinase B (PKB/Akt), ERK1/2 and glycogen synthase kinase (GSK3β). After 7-min reperfusion, WT blocked Akt and GSK3β phosphorylation. After 30-min reperfusion, WT inhibited phosphorylation of all kinases. In conclusion, the data suggest that GLP-2, given in early reperfusion, as postconditioning, protects against myocardial I/R injury, limiting infarct size, and improving post-ischemic mechanical recovery. It seems that the GLP-2-protection of rat heart involves multiple prosurvival kinases and mitochondrial K(ATP) channels.     

Postconditioning cardioprotection against infarct size and post-ischemic systolic dysfunction is influenced by gender

Whether cardioprotection by postconditioning (PostC) is gender dependent is not clear. We studied the effect of PostC in terms of both infarct size (IS) and post-ischemic systolic dysfunction (PSD) reduction. Isolated male and female rat hearts were subjected to 10- or 30-min of global ischemia and 120-min of reperfusion, with or without PostC (i.e., 5 cycles of 10-s reperfusion/ischemia immediately after the ischemia). Surprisingly, after 10-min ischemia, IS and PSD were greater in female than male hearts (IS: 21 ± 2% Vs. 11 ± 2%; P < 0.01), while PostC attenuated IS and PSD in female hearts only. After 30-min ischemia IS was smaller in female than male hearts (52 ± 2% Vs. 61 ± 3%; P < 0.05), whereas PSD was similar in these two groups. PostC reduced IS in both genders, though the effect was smaller (P < 0.05) in females. Yet, PostC reduced PSD in female, but not in male hearts. Contracture development paralleled IS in all groups. To check the effects of buffer perfusion over heart function, additional hearts underwent 150-min buffer perfusion only. Contractile function of these hearts was not significantly affected over time. In conclusion IS, contracture and PSD are differently affected by gender, depending on ischemia duration. Yet, reduction of IS induced by PostC depends on the extension of IS induced by index-ischemia. While in female hearts reduction of PSD paralleled IS reduction, in male it does not occur. Results suggest that improvement of systolic function is mainly due to the anti-necrotic rather than to the anti-stunning effect exerted by PostC. Returned for 1. Revision: 20 August 2008 1. Revision received: 6 October 2008 Returned for 2. Revision: 20 October 2008 2. Revision received: 24 October 2008     

Microvascular responses to ischemia/reperfusion in normotensive and hypertensive rats

The objective of the present study was to determine whether long-term arterial hypertension renders the microvasculature more vulnerable to the deleterious inflammatory responses elicited by ischemia and reperfusion (I/R). Intravital fluorescence microscopy was used to monitor leukocyte adherence and emigration, platelet-leukocyte aggregation, and albumin extravasation in mesenteric postcapillary venules of spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto rats (WKY) after 10 minutes of ischemia and subsequent reperfusion. Significant and comparable increases in leukocyte adherence/emigration and the formation of platelet aggregates were elicited by I/R in both WKY and SHR. Albumin extravasation was enhanced after I/R in SHR, but not in WKY. Monoclonal antibodies directed against the adhesion glycoproteins CD18, P-selectin, or ICAM-1 showed similar patterns of protection against the I/R-induced inflammatory responses in WKY and SHR. The enhanced albumin extravasation noted in postischemic venules of SHR was prevented by immunoneutralization of either CD18 on leukocytes or ICAM-1 on endothelial cells. These results suggest that, whereas long-term arterial hypertension does not significantly modify the leukocyte and platelet recruitment normally elicited in venules by I/R, it does result in an exaggerated albumin leakage response, which is mediated by an interaction between beta(2) (CD18) integrins on leukocytes and ICAM-1 on endothelial cells.     

Postconditioning Protects Against Reperfusion Injury in Hypertensive Dilated Cardiomyopathy by Activating MEK/ERK1/2 Signaling

BackgroundPostconditioning (PostC) cardioprotection has been related to up-regulation of survival kinases; however, the efficacy of PostC and the role of ERK1/2 (extracellular signal-regulated kinase 1/2) remain to be substantiated in hypertension states that may produce “pathologic remodeling.” Therefore, in this work we compared PostC effect and assessed the role of ERK1/2 activation in a model of hypertensive dilated cardiomyopathy (DCM), versus normal (Sham) and compensated hypertrophy (CH) models.Methods and ResultsRats were subjected to angiotensin II administration until development of cardiovascular diseases. Then, isolated hearts underwent ischemia followed by PostC and reperfusion. PostC maintained the double product in all groups. PostC reduced infarct size from 36.16 ± 3% to 9.8% ± 2.2 in Sham, from 37.5 ± 2.4% to 12 ± 3% in CH, and from 40 ± 2.4% to 11.55 ± 3% in DCM. Inhibition of the mitogen-activated protein kinase kinase (MEK)/ERK1/2 pathway had different effects on PostC-conferred cardioprotection in the evaluated groups. Interestingly, although phosphatidylinositol-3-kinase activation was negligible in PostC DCM hearts, we observed Akt activation.ConclusionsPostC confers cardioprotection through alternative survival pathways in normal and CH hearts, and cardiac function recovery in DCM relies mainly on MEK/ERK1/2 cascade. Down-regulation of phosphatidylinositide 3-kinase does not affect the cardioprotective response in DCM, because MEK/ERK1/2 cascade may convey direct Akt activation, strengthening downstream signaling.     

Heat shock protein expression in hearts hypertrophied by genetic and nongenetic hypertension

Summary Genetically hypertensive animals are characterized by greater thermosensitivity and overexpression of heat shock proteins (HSP) upon thermal stimulation. We examined HSP72 expression under conditions of brief coronary occlusion or thermal stimulation, and the effects of the severity of these stimuli and of myocardial hypertrophy on the expression in hearts of spontaneously hypertensive rat (SHR) and Wistar Kyoto rat (WKY) groups, A snare was created around the left coronary artery in the SHR (n=16) and WKY (n=19) groups. In 7 WKY rats, the ascending aorta was banded and a snare was created simultaneously (WKY-AoB). By tying the snare, 4 weeks later, we applied 5- or 10-min coronary occlusion without opening the chest. For thermal stimulation, the SHR (n=13) and WKY (n=11) rats were placed in a 42°C chamber for 15 or 40 min. The mRNA or protein level was estimated 1 or 24h after stimulation. In the SHR vs WKY groups, the mRNA and protein levels were higher after 5-min occlusion or 15-min thermal stimulation. After 10-min occlusion or 40-min thermal stimulation the difference was no longer observed. The overexpression was not observed in the WKY-AoB group despite the presence of hypertrophy similar to that seen in the SHR group (3.11±0.11 vs 3.20±0.06 mg/g in left ventricular weight/body weight). The HSP72 was overexpressed in hearts of genetically hypertensive animals after brief ischemia. Differential expression between the two groups was observed after mild stimuli, but not after more severe stimuli. Cardiac hypertrophy was not a major factor for determining the overexpression of HSP72.     

Ischemic preconditioning maintains cardioprotection in aging normotensive and spontaneously hypertensive rats

We determined whether ischemic preconditioning could reduce infarct size and improve cardiac function in both aging normotensive Wistar–Kyoto rats (WKY) and spontaneously hypertensive rats (SHR). The left anterior descending coronary artery was occluded for 1 h followed by 3 h reperfusion in aging (∼16 months old) SHR rats and age-matched WKY rats. Hearts were either preconditioned or not (control group) prior to 1 h of coronary artery occlusion. The preconditioning regimen consisted of three cycles of 3 min occlusion followed by 5 min reperfusion applied prior to the subsequent 1 h occlusion. In WKY (n = 12 each group), the risk zone was similar in the control (51 ± 2%) and preconditioned group (46 ± 2%; p = 0.1). Preconditioning significantly reduced infarct size (as a percentage of the ischemic risk zone) (24 ± 6%) compared to controls (51 ± 5%; p = 0.0026). In SHR rats (n = 9 each group), the risk zone was smaller in the preconditioning group (41 ± 3%) than in the control group (51 ± 3%; p = 0.035). Infarct size (as % of ischemic risk zone) was also significantly reduced in the preconditioned group (13 ± 4%) compared to controls (62 ± 5%; p < 0.0001). For both WKY and SHR rats, for any sized risk zone the infarct size was smaller in preconditioned hearts compared with the control hearts. Preconditioning improved aspects of LV function during ischemia and reperfusion phase in SHR rats, but these benefits were not observed in the WKY rats. Preconditioning maintains powerful cardioprotection in aging normotensive hearts as well as aging hypertrophied hearts.     

Ischemic preconditioning is lost in aging hypertensive rat heart: independent effects of aging and longstanding hypertension

Although in experimental hypertension the cardioprotective effects of ischemic preconditioning (PC) appear to be maintained, most studies have examined the short-term hypertension in juvenile animals. However, aging may be an additional factor that influences the effectiveness of PC. The aim of this study was to characterise the effects on PC of LVH and aging simultaneously. Hearts from spontaneously hypertensive rats (SHR) and age-matched normotensive Wistar-Kyoto rats (WKY) were studied. Excised hearts were Langendorff-perfused to give equivalent coronary flow per gram heart weight. The left main coronary artery was occluded for 35 min followed by 120 min reperfusion. Infarct size was determined by tetrazolium staining. Heart size was assessed as left ventricle/body weight ratio (LV/BW). PC was effected with 2 x 5 min periods of global ischemia prior to coronary occlusion. Hearts were studied at 3-4 months (juvenile), 7-8 months (mature) or 12-13 months (aging). LV/BW ratio in SHR increased relative to WKY controls by 20%, 32% and 40% in juvenile, mature and aging hearts, respectively, but ischemic risk zone size was similar in all groups (52-59% of LV). PC was equally effective at limiting infarct size in juvenile and mature SHR and WKY hearts but was ineffective in aging hearts from both WKY and SHR. Since angiotensin-converting enzyme inhibitors enhance sub-threshold PC in normal myocardium, we also examined the action of captopril (Cap) in aging hearts. Additional aging hearts received treatment with Cap 200 microM as an adjunct to PC. Although Cap+PC was able to induce modest protection in aging WKY hearts, this was not seen in aging SHR hearts. We conclude that PC is lost in longstanding hypertension through independent contributions of both hypertension and aging. These findings may have implications for the clinical development of preconditioning-based therapies since elderly patients with longstanding hypertension are at high risk of developing ischemic heart disease.     

Comparative effects of candesartan and enalapril on augmented vasoconstrictive responses to endothelin-1 in coronary vessels of spontaneously hypertensive rats

BACKGROUND: The aim of this study was to compare the effect of angiotensin type-1 receptor blockade (ARB) on augmented vasoconstrictive response to endothelin-1 (ET-1) in coronary vessels of hypertensive hearts with angiotensin converting enzyme (ACE) inhibitor, candesartan cilexetil (CAN) or enalapril was administered for 3 weeks in spontaneously hypertensive rats (SHR). METHODS: We used SHR (9 to 12 weeks old, n = 18) and Wistar-Kyoto (WKY) rats (n = 6). Systolic blood pressure was measured once a week. Spontaneously hypertensive rats were divided into three groups. Enalapril malate (10 mg/day) or CAN (10 mg/day) was administered orally in each of six SHR in each group receiving treatment for 3 weeks. The control group (n = 6) received no treatment. At the end of this experiment, the hearts were isolated. Isolated hearts mounted on a Langendorff apparatus after weighing were then perfused with modified Krebs-Henseleit buffer at constant pressure (75 mm Hg). The coronary perfusion pressure and coronary flow were measured during perfusion of isolated hearts. Coronary vascular resistance (CVR; mm Hg/mL/min/100 g) was calculated. RESULTS: The ET-1 elicited increases in CVR dose-dependently in both normotensive and hypertensive rat hearts. However, the responses were significantly greater in SHR than in WKY rat. Chronic treatment with enalapril or candesartan inhibited the development of hypertension and cardiac hypertrophy equally in SHR. Augmented vasoconstrictive responses to ET-1 were significantly reduced in treated SHR. There was no difference in these effects between enalapril and candesartan. CONCLUSION: These findings suggest that both ACE inhibitors and ARB can equally inhibit augmented coronary vascular response to ET-1 in hypertensive hearts.     

Hypertension impairs postnatal vasculogenesis: role of antihypertensive agents

We analyzed the effect of hypertension on postischemic vasculogenesis. Ischemia was induced by right femoral artery ligature in Wistar Kyoto rats (WKY) or spontaneously hypertensive rats (SHR) treated with or without angiotensin-converting enzyme inhibitor (Perindopril, 0.76 mg/kg/d) and angiotensin type 1 receptor blocker (losartan, 30 mg/kg/d). Basal postischemic neovascularization was reduced in SHR compared to WKY (P<0.05, n=8). Treatment with ACE inhibitor or angiotensin type 1 receptor blocker decreased blood pressure levels by 1.4- and 1.3-fold (P<0.001), respectively and restored vessel growth in SHR to WKY levels. Interestingly, 14 days after bone-marrow mononuclear cell (BM-MNC) transfusion, angiographic scores, capillary density, and foot perfusion were decreased by 1.4-, 1.5-, and 1.2-fold, respectively in SHR transfused with BM-MNCs isolated from SHR compared to those receiving BM-MNCs of WKY (P<0.05, n=6). Alteration in BM-MNCs proangiogenic potential was likely related to the reduction in their ability to mobilize into peripheral circulation, as revealed by the 2.9-fold decrease in number of circulating CD34+/CD117+ cells (P<0.001) and to differentiate into cells with endothelial phenotype, as revealed by the 2.1-fold reduction in percentages of DilLDL/BS-1 lectin positive cells (P<0.001). In addition, reactive oxygen species (ROS) levels were increased by 2.2-fold in SHR BM-MNCs compared to WKY BM-MNCs (P<0.01), as assessed by L-012 luminescence. Cotreatment with ACE inhibitor, angiotensin type 1 receptor blocker, or antioxidants (NAC 3 mmol/L, Apocynin 200 micromol/L) reduced ROS levels, improved the number of DilLDL/BS-1 lectin-positive cells by around 1.5-fold, and restored BM-MNCs proangiogenic effects in ischemic hindlimb. In conclusion, alteration in progenitor cell proangiogenic function may participate to the hypertension-induced impairment in postischemic revascularization.     

A treatment with rosuvastatin induced a reduction of arterial pressure and a decrease of oxidative stress in spontaneously hypertensive rats

The aim of this study was to appreciate consequences of rosuvastatin administration on hemodynamic function, vascular oxidative stress and ischemia/reperfusion disorders in normotensive and hypertensive rats. At 10 weeks of age, spontaneously hypertensive rats (SHR, n=20) and normotensive Wistar Kyoto male rats (WKY, n=20) were divided into four groups and given, either vehicle or 10 mg/kg/day of rosuvastatin by gavage for 3 weeks. Systolic blood pressure was assessed every week. At the end of these treatments, vascular NADPH oxidase activity was evaluated by chemiluminescence (lucigenin 0.5 microM). Hearts were isolated and perfused according to the Langendorff method and were subjected to 30 min of global ischemia. Reactive oxygen species (ROS) produced during reperfusion were quantified by electron spin resonance (ESR) spectroscopy using a spin probe (CP-H, 1 mM). After one week of treatment, rosuvastatin reduced the arterial pressure in SHR rats (180.3 +/- 2.1, SHR vs 169.7 +/- 2.3 mmHg, SHR+rosuvastatin; p < 0.01), without lowering plasma cholesterol levels; these effects were not observed in WKY. NADPH activity was 25% higher in control SHR rat aortas compared to control WKY, and was reduced by rosuvastatin in SHR rats. In isolated rat hearts subjected to ischemia/reperfusion sequences, there was a deterioration in functional parameters in control SHR compared to control WKY hearts. Rosuvastatin decreased post-ischemic contracture in WKY hearts by 50% (41.5 +/- 7.5, WKY control vs 18.4 +/- 4.6 mmHg, WKY+rosuvastatin; p < 0.01) and increased left ventricular developed pressure. This beneficial effect was accompanied by a decrease in ROS detected by ESR during reperfusion (312.5 +/- 45.3, WKY control; vs 219.3 +/- 22.9 AUC/mL, WKY+rosuvastatin; p < 0.05). In conclusion, these results are in accordance with the hypothesis that oxidative stress plays a crucial role in the pathogenesis of cardiovascular diseases including hypertension, and demonstrate the beneficial effects of rosuvastatin.     

Cardiac Mechanical Dysfunction Induced by Ischemia‐reperfusion in Perfused Heart Isolated from Stroke‐prone Spontaneously Hypertensive Rats

We investigated the difference in mechanical function after ischemia and reperfusion between Wistar‐Kyoto rats (WKY) and spontaneously hypertensive rats (SHR) or stroke‐prone SHR (SHRSP) using the isolated working heart model, in order to examine postischemic mechanical dysfunction in the severely hypertrophied heart. Systolic blood pressure of SHRSP was higher than that of SHR and WKY, and the left ventricular wall in SHRSP was thicker than in WKY. Mechanical dysfunction of the heart during reperfusion following ischemia (11 min) in SHRSP was severer than that in SHR and WKY, and recovery of the cardiac energy charge potential (ECP) level in SHRSP was lower than that in SHR and WKY. Twenty‐five, 12 and 11 min‐ischemia in WKY, SHR and SHRSP, respectively, caused a similar level of cardiac mechanical damage. Also, the ECP levels were almost equivalent among them at the end of 20 min reperfusion following each time of ischemia. Under each ischemic condition, a Ca^{2 +}‐channel blocker, diltiazem, and an adenosine potentiator, dilazep, produced a beneficial effect on the post‐ischemic dysfunction in SHR and WKY. However, neither cardioprotective drug led to recovery of the mechanical dysfunction of the heart during reperfusion following ischemia in SHRSP. Thus, the severely hypertrophied heart such as that in SHRSP was more susceptible to cardiac reperfusion dysfunction, than the moderately hypertrophied heart such as that in SHR. These results suggest that the cardioprotective effects of drugs may be deteriorated in severe hypertrophied hearts.     

Effect of ubiquinone on contractile function and antioxidant status of the myocardium in spontaneously hypertensive rats

During the period of aging of spontaneously hypertensive rats (SHR) between 6 and 13 weeks the systolic arterial pressure increased from 131+/-2 up to 176+/-3 mm Hg while in the control group of WKY rats it reached 122+/-2 mmHg. The hypertension was combined with myocardial hypertrophy -- the relative weight of SHR heart was 24% higher. The contractile myocardial function of the isolated isovolumic heart of SHR group did not differ from WKY group in a wide range of coronary perfusion rates. During oxidative stress induced by 40-min intracoronary introduction of H(2)O(2) function of hypertrophied SHR hearts fell significantly deeper. This coincided with decreased myocardial activity of superoxide dismutase and glutathione peroxidase by 29-30%, and increased catalase activity by 18%. The rate of generation of active forms of oxygen (hydroxyl radicals HO(.-)) in mitochondria from SHR hearts was higher as compared with WKY. Thus, the development of hypertension was combined with decreased antioxidant protection of the myocardium. The addition of ubiquinone to drinking water (approximately 10 mg/kg/day) for 6 weeks did not affect arterial pressure level, but was associated with two times lesser degree of myocardial hypertrophy. The hearts of SHR that received ubiquinone differed from those not treated with ubiquinone by increased maximal level of myocardial contractile function, and by improved myocardial relaxability and distensibility. After administration of H(2)O(2), myocardial function of SHR was kept on higher level. That was combined with less myocardial oedema, better preservation of antioxidant enzymes and reduced rate of succinate-dependent generation of superoxide radicals in mitochondria from hearts of ubiquinone treated SHR. The results have shown, that administration of ubiquinone to rats with hereditary hypertension reduces degree of myocardial hypertrophy, improves functional properties of the myocardium, promotes effective protection of antioxidant enzymes and increases the resistance of the cardiac muscle to oxidative stress.     

Cardiac mechanical dysfunction induced by ischemia-reperfusion in perfused heart isolated from stroke-prone spontaneously hypertensive rats

We investigated the difference in mechanical function after ischemia and reperfusion between Wistar-Kyoto rats (WKY) and spontaneously hypertensive rats (SHR) or stroke-prone SHR (SHRSP) using the isolated working heart model, in order to examine postischemic mechanical dysfunction in the severely hypertrophied heart. Systolic blood pressure of SHRSP was higher than that of SHR and WKY, and the left ventricular wall in SHRSP was thicker than in WKY. Mechanical dysfunction of the heart during reperfusion following ischemia (11 min) in SHRSP was severer than that in SHR and WKY, and recovery of the cardiac energy charge potential (ECP) level in SHRSP was lower than that in SHR and WKY. Twenty-five, 12 and 11 min-ischemia in WKY, SHR and SHRSP, respectively, caused a similar level of cardiac mechanical damage. Also, the ECP levels were almost equivalent among them at the end of 20 min reperfusion following each time of ischemia. Under each ischemic condition, a Ca2+-channel blocker, diltiazem, and an adenosine potentiator, dilazep, produced a beneficial effect on the post-ischemic dysfunction in SHR and WKY. However, neither cardioprotective drug led to recovery of the mechanical dysfunction of the heart during reperfusion following ischemia in SHRSP. Thus, the severely hypertrophied heart such as that in SHRSP was more susceptible to cardiac reperfusion dysfunction, than the moderately hypertrophied heart such as that in SHR. These results suggest that the cardioprotective effects of drugs may be deteriorated in severe hypertrophied hearts.     

Protection from postconditioning depends on the number of short ischemic insults in anesthetized pigs

Abstract Postconditioning in the early reperfusion period confers protection to the heart after a potentially lethal episode of prolonged ischemia. Protection from this novel intervention has been documented in rat, rabbit and canine hearts, but one group has reported that it is ineffective in pigs, a large-animal species that should be most relevant to humans. We hypothesized that this negative result was related to an inappropriate postconditioning protocol rather than the species. The present study, therefore, tested whether an effective postconditioning protocol could be identified that limits infarct size in anesthetized pigs. Domestic Landrace pigs weighing 25–29 kg were anesthetized, and after a mid-sternal thoracotomy and pericardiotomy the left anterior descending coronary artery was ligated for 60 min followed by 3 h of reperfusion. Three groups were studied: control group (n = 5) with no other intervention, 4–30 PostC group (n = 5) with 4 cycles of 30-s reperfusion/30-s ischemia, and 8–30 PostC group (n = 6) with 8 cycles of 30-s reperfusion/30-s ischemia. The two postconditioning protocols started immediately after termination of the 60-min coronary occlusion. Region at risk and infarct size were delineated with the aid of pre-mortem monastral blue injection and postmortem staining with triphenyltetrazolium chloride, respectively. In control hearts 33.5 ± 7.6% of the risk zone infarcted and 36.7 ± 3.7% in the 4–30 PostC group (P = NS). But there was only 10.5 ± 0.5% infarction in the 8–30 PostC group (P < 0.01 vs. the other two groups). Postconditioning confers protection in pigs but requires more than 4 ischemia/reperfusion cycles. Postconditioning may protect by inhibiting mitochondrial permeability transition pore formation by keeping the heart acidotic as it is reoxygenated. If true, then it would be difficult to employ too many occlusion cycles.     

Remote postconditioning

Objectives A series of brief coronary artery reperfusions and reocclusions applied during the early minutes of coronary artery reflow (“postconditioning”) attenuates reperfusion injury. However, it is not known whether brief ischemia–reperfusion applied to a distant organ at the onset of myocardial reperfusion (i.e. “remote postconditioning”, remote PostC) reduces infarct size in the reperfused myocardium. In an in vivo anesthetized rat model of myocardial infarction induced by coronary artery occlusion and reperfusion, this study tested the hypothesis that remote postC induced by a single 5 minute episode of renal artery (RA) occlusion and reperfusion applied immediately before the onset of coronary artery reperfusion protects the myocardium from reperfusion injury by mechanisms involving endogenous adenosine receptor activation. Methods All rats were subjected to a total of 30 minutes of left coronary artery occlusion (LCAO) and 3 hours of reperfusion. The rats were randomized to one of six groups: 1) Control: LCAO and reperfusion only with no other intervention; 2) Remote PostC: after 24 minutes of LCAO the RA was occluded for 5 minutes and released 1 min before coronary artery reperfusion; 3) Permanent RA occlusion: the RA was permanently occluded after 24 minutes LCAO continuing to the end of reperfusion; 4) Delayed Remote PostC: after 26 minutes LCAO the RA was occluded for 5 minutes, and its release was delayed until 1 min after coronary artery reperfusion; 5) CON + SPT: rats with LCAO and reperfusion received 10 mg/kg IV of the non–selective adenosine receptor antagonist 8–sulfophenyl theophylline [SPT] administered 5 minutes before coronary artery reperfusion; and 6) Remote PostC + SPT: after 24 minutes of LCAO the RA was occluded for 5 minutes and released 1 minute before coronary artery reperfusion in the presence of 10 mg/kg SPT given 5 min before coronary artery reperfusion. Results Myocardial infarct size (percentage necrosis/area at risk, mean ± SEM) was reduced by 50% in Remote PostC (25 ± 4%) compared to Control (49 ± 4%, p = 0.003), consistent with a reduction in plasma CK activity (44 ± 5 vs. 67 ± 6 U/ml, p = 0.023). In contrast, permanent RA occlusion before LCAO and reperfusion failed to reduce myocardial infarct size (47 ± 5%) vs Control. Delaying the release of the RA occlusion (delayed Remote PostC) abrogated the myocardial infarct reduction observed with Remote PostC (48 ± 6%). SPT alone had no effect on infarct size (47 ± 4% in CON + SPT vs. 49 ± 4% in CON); however, Remote PostC+SPT abrogated the myocardial infarct size reduction in Remote PostC (50 ± 3% in Remote PostC + SPT vs. 25 ± 4% in Remote PostC). Conclusions Remote renal postconditioning applied immediately before the onset of coronary artery reperfusion provides potent myocardial infarct size reduction likely exerted during the first minutes of coronary artery reperfusion. This inter–organ remote postconditioning phenomenon is likely mediated in part by release of adenosine by the ischemic–reperfused kidney and subsequent activation of adenosine receptors.     

Myocardial function in normal and spontaneously hypertensive rats during reperfusion after a period of global ischaemia

Isolated working hearts of 16 month old spontaneously hypertensive rats (SHR, n = 8) and age matched Wistar-Kyoto (WKY, n = 8) rats were exposed to 30 min global normothermic ischaemia followed by 60 min reperfusion. The hearts were routinely perfused at an afterload level of 13.3 kPa and a preload level of 1.0 kPa. The control values of left ventricular pressure, its maximal positive first derivative (dP1v/dtmax), coronary flow per gram heart tissue, and release of lactate and enzymes such as lactate dehydrogenase and aspartate aminotransferase were comparable in both groups. WKY rat hearts ejected almost twice as much perfusate per gram heart weight as the SHR hearts. In pressure-flow curves, obtained during the control period in SHR hearts, cardiac output was independent of changes in afterload, varying between 10.7 and 18.7 kPa. In contrast, in WKY rat hearts increases in afterload resulted in a progressive decrease in cardiac output. Reperfusion of the SHR hearts after 30 min of global normothermic ischaemia resulted in a poor recovery of cardiac output (13% of the control values) and dP1v/dtmax (32%) compared with the values in the WKY rat hearts (66% and 91% of the control values respectively). Reactive hyperaemia was prominent in the WKY rat hearts but completely absent in the SHR hearts. During one hour reperfusion, SHR hearts lost 3.5 times more lactate dehydrogenase and 2.5 times more aspartate aminotransferase than the WKY rat hearts. Pressure-flow curves, obtained during the reperfusion period, showed modest recovery of myocardial function of the WKY rat hearts at the lowest afterload level tested but completely depressed myocardial function of the SHR hearts at all afterload levels. Heart tissue contents of adenosine triphosphate and creatine phosphate after one hour of reperfusion were lower in the SHR than in the WKY rats, but compared with native values a comparable percentage decrease was seen in both groups of rats.