Remote Non-infarcted Myocardium Research Articles

Mitochondrial cyclophilin-D as a potential therapeutic target for post-myocardial infarction heart failure

The pharmacological inhibition or genetic ablation of cyclophilin-D (CypD), a critical regulator of the mitochondrial permeability transition pore (mPTP), confers myocardial resistance to acute ischemia-reperfusion injury, but its role in post-myocardial infarction (MI) heart failure is unknown. The aim of this study was to determine whether mitochondrial CypD is also a therapeutic target for the treatment of post-MI heart failure. Wild-type (WT) and CypD–/– mice were subjected to either sham surgery or permanent ligation of the left main coronary artery to induce MI, and were assessed at either 2 or 28 days to determine the long-term effects of CypD ablation. After 2 days, myocardial infarct size was smaller and left ventricular (LV) function was better preserved in CypD–/– mice compared to WT mice. After 28 days, when compared to WT mice, in the CypD–/– mice, mortality was halved, myocardial infarct size was reduced, LV systolic function was better preserved, LV dilatation was attenuated and in the remote non-infarcted myocardium, there was less cardiomyocyte hypertrophy and interstitial fibrosis. Finally, ex vivo fibroblast proliferation was found to be reduced in CypD–/– cardiac fibroblasts, and in WT cardiac fibroblasts treated with the known CypD inhibitors, cyclosporin-A and sanglifehrin-A. Following an MI, mice lacking CypD have less mortality, smaller infarct size, better preserved LV systolic function and undergo less adverse LV remodelling. These findings suggest that the inhibition of mitochondrial CypD may be a novel therapeutic treatment strategy for post-MI heart failure.

Prostanoids suppress the coronary vasoconstrictor influence of endothelin after myocardial infarction

Myocardial infarction (MI) is associated with endothelial dysfunction resulting in an imbalance in endothelium-derived vasodilators and vasoconstrictors. We have previously shown that despite increased endothelin (ET) plasma levels, the coronary vasoconstrictor effect of endogenous ET is abolished after MI. In normal swine, nitric oxide (NO) and prostanoids modulate the vasoconstrictor effect of ET. In light of the interaction among NO, prostanoids, and ET combined with endothelial dysfunction present after MI, we investigated this interaction in control of coronary vasomotor tone in the remote noninfarcted myocardium after MI. Studies were performed in chronically instrumented swine (18 normal swine; 13 swine with MI) at rest and during treadmill exercise. Furthermore, endothelial nitric oxide synthase (eNOS) and cyclooxygenase protein levels were measured in the anterior (noninfarcted) wall of six normal and six swine with MI. eNOS inhibition with N(ω)-nitro-L-arginine (L-NNA) and cyclooxygenase inhibition with indomethacin each resulted in coronary vasoconstriction at rest and during exercise, as evidenced by a decrease in coronary venous oxygen levels. The effect of l-NNA was slightly decreased in swine with MI, although eNOS expression was not altered. Conversely, in accordance with the unaltered expression of cyclooxygenase-1 after MI, the effect of indomethacin was similar in normal and MI swine. L-NNA enhanced the vasodilator effect of the ET(A/B) receptor blocker tezosentan but exclusively during exercise in both normal and MI swine. Interestingly, this effect of L-NNA was blunted in MI compared with normal swine. In contrast, whereas indomethacin increased the vasodilator effect of tezosentan only during exercise in normal swine, indomethacin unmasked a coronary vasodilator effect of tezosentan in MI swine both at rest and during exercise. In conclusion, the present study shows that endothelial control of the coronary vasculature is altered in post-MI remodeled myocardium. Thus the overall vasodilator influences of NO as well as its inhibition of the vasoconstrictor influence of ET on the coronary resistance vessels were reduced after MI. In contrast, while the overall prostanoid vasodilator influence was maintained, its inhibition of ET vasoconstrictor influences was enhanced in post-MI remote myocardium.

Acute Myocardial Infarction in Rats

With heart failure leading the cause of death in the USA (Hunt), biomedical research is fundamental to advance medical treatments for cardiovascular diseases. Animal models that mimic human cardiac disease, such as myocardial infarction (MI) and ischemia-reperfusion (IR) that induces heart failure as well as pressure-overload (transverse aortic constriction) that induces cardiac hypertrophy and heart failure (Goldman and Tarnavski), are useful models to study cardiovascular disease. In particular, myocardial ischemia (MI) is a leading cause for cardiovascular morbidity and mortality despite controlling certain risk factors such as arteriosclerosis and treatments via surgical intervention (Thygesen). Furthermore, an acute loss of the myocardium following myocardial ischemia (MI) results in increased loading conditions that induces ventricular remodeling of the infarcted border zone and the remote non-infarcted myocardium. Myocyte apoptosis, necrosis and the resultant increased hemodynamic load activate multiple biochemical intracellular signaling that initiates LV dilatation, hypertrophy, ventricular shape distortion, and collagen scar formation. This pathological remodeling and failure to normalize the increased wall stresses results in progressive dilatation, recruitment of the border zone myocardium into the scar, and eventually deterioration in myocardial contractile function (i.e. heart failure). The progression of LV dysfunction and heart failure in rats is similar to that observed in patients who sustain a large myocardial infarction, survive and subsequently develops heart failure (Goldman). The acute myocardial infarction (AMI) model in rats has been used to mimic human cardiovascular disease; specifically used to study cardiac signaling mechanisms associated with heart failure as well as to assess the contribution of therapeutic strategies for the treatment of heart failure. The method described in this report is the rat model of acute myocardial infarction (AMI). This model is also referred to as an acute ischemic cardiomyopathy or ischemia followed by reperfusion (IR); which is induced by an acute 30-minute period of ischemia by ligation of the left anterior descending artery (LAD) followed by reperfusion of the tissue by releasing the LAD ligation (Vasilyev and McConnell). This protocol will focus on assessment of the infarct size and the area-at-risk (AAR) by Evan's blue dye and triphenyl tetrazolium chloride (TTC) following 4-hours of reperfusion; additional comments toward the evaluation of cardiac function and remodeling by modifying the duration of reperfusion, is also presented. Overall, this AMI rat animal model is useful for studying the consequence of a myocardial infarction on cardiac pathophysiological and physiological function.

Effect of Cyclosporine on Left Ventricular Remodeling After Reperfused Myocardial Infarction

This study examined the effect of a single dose of cyclosporine administered at the time of reperfusion on left ventricular (LV) remodeling and function by cardiac magnetic resonance 5 days and 6 months after myocardial infarction. In a human study, administration of cyclosporine at the time of acute reperfusion was associated with a smaller infarct size. Twenty-eight patients of the original cyclosporine study had an acute (at 5 days) and a follow-up (at 6 months) cardiac magnetic resonance study to determine LV volumes, mass, ejection fraction, myocardial wall thickness in infarcted and remote noninfarcted myocardium, and infarct size. There was a persistent reduction in infarct size at 6 months in the cyclosporine group compared with the control group of patients (29 +/- 15 g vs. 38 +/- 14 g; p = 0.04). There was a significant reduction of LV end-systolic volume (and a trend for LV end-diastolic volume; p = 0.07) in the cyclosporine group compared with the control group, both at 5 days and 6 months after infarction. There was no significant difference between the 2 groups in either global LV mass or regional wall thickness of the remote noninfarcted myocardium at 5 days or 6 months. Attenuation of LV dilation and improvement of LV ejection fraction by cyclosporine at 6 months were correlated with infarct size reduction. Cyclosporine used at the moment of acute myocardial infarction reperfusion persistently reduces infarct size and does not have a detrimental effect on LV remodeling. These results are preliminary and must be supported by further studies. (Ciclosporin A and Acute Myocardial Infarction; NCT00403728).

058 Effect of Cyclosporine on Left Ventricle Remodeling after Reperfused Myocardial Infarction

This study examined the effect of cyclosporine A used at the time of reperfusion, on LV remodeling and function by cardiac magnetic resonance (CMR) in the early days and several months after AMI. In a human study, administration of cyclosporine A at the time of acute myocardial infarction (AMI) reperfusion was associated with a smaller infarct size. However, experimental data suggest that cyclosporine A has a detrimental effect on left ventricular remodeling. 28 patients of the original cyclosporine A study had an acute (day 5) and a follow-up (6 months) CMR study. Cine imaging was used to determine LV volumes, mass, ejection fraction and myocardial wall thickness in infarcted and remote non-infarcted myocardium, and late gadolinium imaging was used to determine infarct size. There was a persistent reduction of the absolute infarct size at 6 months in the cyclosporine A group compared with the control group of patients (29±15 grams VS 38±14 grams; P=0.04). There was a significant reduction of LV end-systolic volume (and a trend for LV end-diastolic volume; P=0.07) in the cyclosporine A group, compared with the control group, both at day 5 and at 6 months after infarction. There was no significant difference between the two groups in either global LV mass or regional wall thickness of the remote non-infarcted myocardium at day 5 or at 6 months. Attenuation of LV dilatation and improvement of LV ejection fraction by cyclosporine A at 6 months were correlated with infarct size reduction. Cyclosporine A used at the moment of AMI reperfusion persistently reduces infarct size and does not have a detrimental effect on LV remodeling

Abstract 4760: Effect of Cyclosporine on Left Ventricle Remodeling After Reperfused Myocardial Infarction

Introduction: In a human study, administration of cyclosporine A at the time of acute myocardial infarction (AMI) reperfusion was associated with a smaller infarct size. However, experimental data suggest that cyclosporine A has a detrimental effect on left ventricular remodeling. This study examined the effect of cyclosporine A used at the time of reperfusion, on LV remodeling and function by cardiac magnetic resonance (CMR) in the early days and several months after AMI. Material and Methods: 28 patients of the original cyclosporine A study had an acute (day 5) and a follow-up (6 months) CMR study. Cine imaging was used to determine LV volumes, mass, ejection fraction and myocardial wall thickness in infarcted and remote non-infarcted myocardium, and late gadolinium imaging was used to determine infarct size. Results: There was a persistent reduction of the absolute infarct size at 6 months in the cyclosporine A group compared with the control group of patients (29±15 grams VS 38±14 grams; P=0.04). There was a significant reduction of LV end-systolic volume (and a trend for LV end-diastolic volume; P=0.07) in the cyclosporine A group, compared with the control group, both at day 5 and at 6 months after infarction. There was no significant difference between the two groups in either global LV mass or regional wall thickness of the remote non-infarcted myocardium at day 5 or at 6 months. Attenuation of LV dilatation and improvement of LV ejection fraction by cyclosporine A at 6 months were correlated with infarct size reduction. Conclusion: Cyclosporine A used at the moment of AMI reperfusion persistently reduces infarct size and does not have a detrimental effect on LV remodeling.

Cell therapy enhances function of remote non-infarcted myocardium

Cell transplantation improves cardiac function after myocardial infarction; however, the underlying mechanisms are not well-understood. Therefore, the goals of this study were to determine if neonatal rat cardiomyocytes transplanted into adult rat hearts 1 week after infarction would, after 8–10 weeks: 1) improve global myocardial function, 2) contract in a Ca2+ dependent manner, 3) influence mechanical properties of remote uninjured myocardium and 4) alter passive mechanical properties of infarct regions. The cardiomyocytes formed small grafts of ultrastructurally maturing myocardium that enhanced fractional shortening compared to non-treated infarcted hearts. Chemically demembranated tissue strips of cardiomyocyte grafts produced force when activated by Ca2+, whereas scar tissue did not. Furthermore, the Ca2+ sensitivity of force was greater in cardiomyocyte grafts compared to control myocardium. Surprisingly, cardiomyocytes grafts isolated in the infarct zone increased Ca2+ sensitivity of remote uninjured myocardium to levels greater than either remote myocardium from non-treated infarcted hearts or sham-operated controls. Enhanced calcium sensitivity was associated with decreased phosphorylation of cTnT, tropomyosin and MLC2, but not changes in myosin or troponin isoforms. Passive compliance of grafts resembled normal myocardium, while infarct tissue distant from grafts had compliance typical of scar. Thus, cardiomyocyte grafts are contractile, improve local tissue compliance and enhance calcium sensitivity of remote myocardium. Because the volume of remote myocardium greatly exceeds that of the grafts, this enhanced calcium sensitivity may be a major contributor to global improvements in ventricular function after cell transplantation.

Delivering Stem Cells to the Heart in a Collagen Matrix Reduces Relocation of Cells to other Organs as Assessed by Nanoparticle Technology

A limitation of cell therapy for heart disease is the fact that stem cells injected directly into the myocardium are capable of entering the vasculature and migrating to remote organs. We determined whether retention of mesenchymal stem cells (MSCs) in the infarcted myocardium could be improved by implanting the cells in a collagen matrix. A myocardial infarction was induced by ligation of the left anterior descending coronary artery in Fischer rats. A total of 7 days after myocardial infarction, saline (n = 12), saline plus 2 million bone marrow-derived rat MSCs labeled with isotopic colloidal nanoparticles containing europium (n = 13), collagen (n = 13) or collagen plus 2 million labeled MSCs (n = 13) were directly injected into the infarcted myocardium. Tissues from the infarcted myocardium, noninfarcted myocardium, lung, liver, spleen and kidney were sampled 4 weeks later. Distribution of grafted MSCs was quantitatively analyzed by measuring the nanoparticle radioactivity in these tissues. Cardiac function was assessed by left ventriculography. There were zero nanoparticles detected in the tissues that received saline or collagen alone into the heart. Nanoparticles were detected in the heart and remote organs in the saline plus MSC group. Labeled cells (expressed as cell number/g tissue weight) were present in three out of 13 lungs (mean of 12,724 +/- 7060 cells/g), four out of 13 livers (12,301 +/- 5924 cells/g), 11 out of 13 spleens (57,228 +/- 11,483 cells/g), zero out of 13 kidneys, 13 out of 13 infarcted myocardium (8,006,835 +/- 1,846,462 cells/g) and nine out of 13 noninfarcted myocardium (167,331 +/- 47,007 cells/g). However, compared with the saline plus MSC group, nanoparticles were detected to a lesser extent in remote organs in collagen plus MSC group. Nanoparticles were detected in two out of 13 lungs (4631 +/- 3176 cells/g; p = NS), zero out of 13 livers (0 cells/g; p <0.05 vs saline plus MSC), four out of 13 spleens (24,060 +/- 17,373 cells/g; p <0.05), zero out of 13 kidneys (p = NS) and five out of 13 noninfarcted myocardium (51,522 +/- 21,548 cells/g; p <0.05). In the collagen plus MSC group, nanoparticles were detected in 12 out of 13 infarcted myocardium (4,830,050 +/- 592,215 cells/g), which did not significantly differ from that in the saline plus MSC group (p = NS). Both saline plus MSCs and collagen alone improved left ventricular ejection fraction compared with saline treatment. However, collagen plus MSCs failed to improve cardiac function. Collagen matrix as a delivery vehicle significantly reduced the relocation of transplanted MSCs to remote organs and noninfarcted myocardium.

Prevention of Myofilament Dysfunction by β-Blocker Therapy in Postinfarct Remodeling

Myofilament contractility of individual cardiomyocytes is depressed in remote noninfarcted myocardium and contributes to global left ventricular pump dysfunction after myocardial infarction (MI). Here, we investigated whether beta-blocker therapy could restore myofilament contractility. In pigs with a MI induced by ligation of the left circumflex coronary artery, beta-blocker therapy (bisoprolol, MI+beta) was initiated on the first day after MI. Remote left ventricular subendocardial biopsies were taken 3 weeks after sham or MI surgery. Isometric force was measured in single permeabilized cardiomyocytes. Maximal force (F(max)) was lower, whereas Ca(2+) sensitivity was higher in untreated MI compared with sham (both P<0.05). The difference in Ca(2+) sensitivity was abolished by treatment of cells with the beta-adrenergic kinase, protein kinase A. beta-blocker therapy partially reversed F(max) and Ca(2+) sensitivity to sham values and significantly reduced passive force. Despite the lower myofilament Ca(2+) sensitivity in MI+beta compared with untreated myocardium, the protein kinase A induced reduction in Ca(2+) sensitivity was largest in cardiomyocytes from myocardium treated with beta-blockers. Phosphorylation of beta-adrenergic target proteins (myosin binding protein C and troponin I) did not differ among groups, whereas myosin light chain 2 phosphorylation was reduced in MI, which coincided with increased expression of protein phosphatase 1. beta-blockade fully restored the latter alterations and significantly reduced expression of protein phosphatase 2a. beta-blockade reversed myofilament dysfunction and enhanced myofilament responsiveness to protein kinase A in remote myocardium after MI. These effects likely contribute to the beneficial effects of beta-blockade on global left ventricular function after MI.

Intravenous mesenchymal stem cell therapy early after reperfused acute myocardial infarction improves left ventricular function and alters electrophysiologic properties

Direct intramyocardial injection of mesenchymal stem cells (MSCs) improves left ventricular ejection fraction (LVEF) and may increase ventricular arrhythmia in hearts with myocardial infarction (MI). We hypothesized that intravenous MSCs given early after acute MI would engraft in injured myocardium, improve LV function, and result in pro-arrhythmic electrical remodeling. We created an apical infarction in swine by balloon occlusion/reperfusion, administered diI-labeled allogeneic bone marrow derived MSCs intravenously 30 min post-reperfusion and measured LVEF and wall thickness at baseline, 1 month, and 3 months. Epicardial effective refractory periods (ERPs) were determined before sacrifice. At 3 months, treated pigs [ n = 7] had significantly higher LVEF than controls [ n = 8] (49 ± 2% vs. 44 ± 3%, P = 0.015)and significantly less wall thickening of non-infarcted myocardium. ERPs were significantly shorter than controls at all pacing cycle lengths ( P ≤ 0.002), suggesting a pro-arrhythmic potential. DiI was found in the lungs, in infarct, and peri-infarct myocardium. Conclusion: IV infusion of MSCs soon after acute MI in swine improves LVEF and limits wall thickening in the remote non-infarcted myocardium, consistent with a beneficial effect on post-MI ventricular remodeling. Since there is no need for immune suppression or clinical expertise, IV infusion of MSCs may expand the potential clinical application of stem cell therapy.

Heart Failure Following Anterior Myocardial Infarction: An Indication for Ventricular Restoration, a Surgical Method to Reverse Post-Infarction Remodeling

Anterior myocardial infarction produces abrupt left ventricular (LV) dysynergy and global systolic dysfunction. Rapid intense neurohumoral activation, infarct expansion, and early ventricular chamber dilatation all contribute to restoring a normal stroke volume despite a persistently depressed ejection fraction. Continued neurohumoral activation provokes late remodeling of the remote non-infarcted myocardium, characterized by an abnormal progressively increasing LV volume/mass ratio that leads to further LV remodeling. Heart failure is a progressive disorder of LV remodeling. Heart failure from post-infarction remodeling is unique because of the persistent non-functioning scar that self- perpetuates abnormal loading conditions and neurohumoral activation. Medical therapy attenuates remodeling and improves survival but does not change the size of the scar. Surgical ventricular restoration to exclude the non-functioning infarct from the ventricular cavity decreases ventricular volumes, increases global ejection fraction, attenuates neurohumoral activation and yields an excellent 5-year survival. Combined medical and surgical therapy is recommended in this patient population.

Exercise unmasks autonomic dysfunction in swine with a recent myocardial infarction

Severe congestive heart failure is associated with autonomic imbalance consisting of an increased sympathetic and decreased parasympathetic activity. In the present study, we investigated the influence of alterations in autonomic balance on cardiovascular function in 11 swine with left ventricular (LV) dysfunction produced by a 2- to 3-week-old myocardial infarction (MI). Swine underwent permanent occlusion of the left circumflex coronary artery resulting in MI of the lateral LV wall. Autonomic activity was studied 2-3 weeks later using blockers of muscarinic (atropine), alpha-adrenergic (phentolamine) and beta-adrenergic (propranolol) receptors. Under resting conditions, parasympathetic and sympathetic control of the heart and coronary circulation were similar in MI and normal swine. In contrast, during exercise of MI compared to normal swine, (i) there was a more pronounced gradual inhibition of parasympathetic control of heart rate with increasing exercise intensity; (ii) circulating catecholamines increased excessively, resulting in an increased beta-adrenergic influence on heart rate, while (iii) the beta-adrenergic influence on global left ventricular contractility was decreased, reflecting a blunted left ventricular beta-adrenergic responsiveness. Furthermore, (iv) an alpha-adrenergic vasoconstrictor influence was absent in the anterior LV wall of both MI and normal swine, while (v) the beta-adrenergic vasodilator influence in the coronary circulation was not different between normal and MI swine, which, in conjunction with the elevated catecholamine levels during exercise, suggests a diminished beta-adrenergic responsiveness of coronary resistance vessels within remote non-infarcted myocardium in MI swine. Swine with a recent MI display autonomic dysfunction, which is characterized by a more pronounced inhibition of parasympathetic influence and an exaggerated increase in sympathetic drive during exercise, as well as reduced myocardial and coronary vascular beta-adrenergic responsiveness.

Progressive left ventricular remodeling, myocyte apoptosis, and protein signaling cascades after myocardial infarction in rabbits

To determine the temporal changes in oxidative stress, mitogen-activated protein (MAP) kinases and mitochondrial apoptotic proteins, and their relationship to myocyte apoptosis in the remote noninfarcted myocardium after myocardial infarction (MI), rabbits were randomly assigned to either coronary artery ligation to produce MI or sham operation. The animals were sacrificed at 1, 4, 8, or 12 weeks after coronary artery occlusion. Sham rabbits were sacrificed at 12 weeks after surgery. MI rabbits exhibited progressive increases of left ventricular (LV) end-diastolic pressure and end-diastolic dimension, and progressive decreases of LV fractional shortening and dP/dt over 12 weeks. The LV remodeling with LV chamber dilation and LV systolic dysfunction was temporally associated with progressive increases of cardiac oxidative stress as evidenced by decreased myocardial reduced-to-oxidized-glutathione ratio and increased myocardial 8-hydroxydeoxyguanosine and myocyte apoptosis. The ERK and JNK activities were decreased while p38 MAP kinase activity was increased with age of MI. The extent of p38 MAP kinase activation correlated with Bcl-2 phosphorylation. Bcl-2 protein was decreased in both mitochondrial and cytosolic fractions with age of MI. Bax protein was increased in both mitochondrial and cytosolic fractions. Cytochrome c was reduced in mitochondrial fraction and increased in cytosolic fraction in a time-dependent manner after MI. Cleaved caspase 9 and caspase 3 proteins were time-dependently increased after MI. These data suggest that p38 MAP kinase activation is not only time-dependent after MI, but also correlates with oxidative stress, Bcl-2 phosphorylation, and myocyte apoptosis. These changes in the remote noninfarcted myocardium may contribute to LV remodeling and dysfunction after MI.

Increased apoptosis in remote non-infarcted myocardium in multivessel coronary disease

Background: Multivessel coronary disease after myocardial infarction is a major risk factor for unfavorable cardiac remodeling and death due to pump failure, but underlying pathophysiologic mechanisms are still uncompletely established. Post-infarction myocardial apoptosis has been recently implicated as a cause of ongoing cell loss leading to cardiac failure. Our aim was to assess the role of post-infarction myocardial apoptosis and pro-apoptotic factor expression in the non-infarcted remote myocardium of subjects with multivessel coronary disease. Methods: Twenty-one males dying after recent myocardial infarction with permanent occlusion of the infarct-related artery were selected at autopsy. Apoptosis was assessed at viable myocardial regions remote from infarction by co-staining for in situ end-labeling of DNA fragmentation and cleaved caspase-3. Expression of pro-apoptotic factor bax and hypoxia-induced factor-1alpha was evaluated by immunohistochemistry. Results: Subjects with multivessel disease ( N=11) showed a significantly two-fold higher myocardial apoptosis in comparison to subjects with single vessel disease ( N=10) (0.9% vs. 0.5%, p=0.013). Similarly, myocardial bax expression was increased in patients with multivessel disease (3.0% vs. 1.4%, p=0.029). Stratification for the number of diseased coronary vessels confirmed the association between extent of coronary disease and apoptotic rates ( p=0.022). Even in subjects dying over 30 days after infarction multivessel disease remained predictive of enhanced myocardiocyte apoptosis at remote regions ( p=0.033). Conclusions: Post-infarction myocardial apoptosis and bax expression in remote left ventricular regions are significantly increased in male patients with multivessel coronary disease in comparison to those with isolated infarct-related artery occlusion. These findings suggest that apoptotic cell loss in the viable non-infarcted myocardium, possibly due ongoing ischemia, may play a relevant role in the unfavorable clinical course typical of multivessel disease after myocardial infarction.

The potential of contrast-enhanced magnetic resonance imaging for predicting left ventricular remodeling.

To determine whether the myocardial injury size on day 2 measured after gadolinium (Gd)-mesoporphyrin and Gd-diethylenetriamine-pentaacetic acid (DTPA) administration can be used for predicting left ventricular (LV) remodeling 8 weeks later, and to monitor the structural and functional changes in the infarct, peri-infarct rim, and remote myocardium in reperfused infarction using contrast-enhanced and functional magnetic resonance imaging (MRI) MATERIALS AND METHODS: Myocardial infarction (MI) was induced in 27 rats by 1 hour of coronary occlusion/reperfusion. Rats were imaged 2 days and 8 weeks after MI using MRI to determine LV function and size of myocardial injury. All animals received 0.05 mmol/kg Gd-mesoporphyrin 12 hours before the first MRI. A subgroup of 13 rats received 0.3 mmol/kg Gd-DTPA in addition to Gd-mesoporphyrin, and seven rats received 0.05 mmol/kg Gd-mesoporphyrin 12 hours before the second MRI for detection of healed MI. True infarct size (IS) and LV mass were measured postmortem. LV volumes, mass, function, and wall thickness were determined in both imaging sessions. A close correlation was found between contrast-enhanced MRI and postmortem measurements for IS (r = 0.94, P < 0.001 for Gd-mesoporphyrin; r = 0.91, P < 0.001, N = 13 for Gd-DTPA). IS measured on Gd-mesoporphyrin-enhanced images correlated well with end-systolic LV volumes (r = 0.68, P < 0.001) and ejection fraction (r = -0.75, P < 0.001) 8 weeks after MI. Similar correlation with parameters of LV remodeling were found on Gd-DTPA-enhanced MRI. Healed infarcts showed no enhancement on Gd-mesoporphyrin-enhanced MRI. Contrast-enhanced MRI can be used as a noninvasive method for determining the initial size of myocardial injury and predicting later LV remodeling. MRI demonstrates the structural and functional changes in infarct, peri-infarct rim, and remote non-infarcted myocardium. The complementary use of functional and contrast-enhanced MRI may provide reliable assessment of therapeutic interventions to reduce IS and LV remodeling.

Expression of Heme Oxygenase-1 in Response to Myocardial Infarction in Rats

P. Lakkisto, E. Palojoki, T. Bäcklund, A. Saraste, I. Tikkanen, L.-M. Voipio-Pulkki and K. Pulkki. Expression of Heme Oxygenase-1 in Response to Myocardial Infarction in Rats. Journal of Molecular and Cellular Cardiology (2002) 34, 1357–1365. Heme oxygenase-1 (HO-1) is a heat shock protein catalysing the degradation of heme to yield biliverdin, carbon monoxide and iron. Several recent studies have proposed the stress-inducible HO-1 to participate in cellular protection also in the heart. We, therefore, examined the expression and localization of HO-1 in a rat experimental myocardial infarction model. Male Wistar rats were subjected to left anterior coronary artery ligation or sham-operation and sacrificed at 1 day, 1 week and 4 weeks after ligation. The expression of HO-1 mRNA was assessed by real-time quantitative RT-PCR and the localization of HO-1 protein by immunoconfocal microscopy. At day 1, HO-1 mRNA was increased 3.9-fold in the peri-infarct border area vs sham-operated hearts (P<0.001) and 2.9-fold vs remote areas of the same hearts (P<0.001). At 1 week, HO-1 mRNA levels remained significantly higher (5-fold) in the peri-infarct border area than in sham-operated hearts (P<0.001). In addition, HO-1 mRNA transiently increased 1.6-fold in the remote non-infarcted myocardium vs sham operated hearts (P<0.05). HO-1 mRNA returned to basal levels by 4 weeks. The increase in HO-1 mRNA was accompanied by increased immunoreactivity of HO-1 protein in the vascular walls throughout the myocardium, and in the cardiomyocytes and fibroblast-like cells of the peri-infarct border areas. Cardiomyocytes showed immunoreactivity at the intercalated disc area, and in the sarcoplasmic reticulum as indicated by the striated pattern of staining. The results suggest that the induction of HO-1 may have an important role in the heart during the first days after myocardial infarction.

Expression of Heme Oxygenase-1 in Response to Myocardial Infarction in Rats

P. Lakkisto, E. Palojoki, T. Bäcklund, A. Saraste, I. Tikkanen, L.-M. Voipio-Pulkki and K. Pulkki. Expression of Heme Oxygenase-1 in Response to Myocardial Infarction in Rats. Journal of Molecular and Cellular Cardiology (2002) 34, 1357–1365. Heme oxygenase-1 (HO-1) is a heat shock protein catalysing the degradation of heme to yield biliverdin, carbon monoxide and iron. Several recent studies have proposed the stress-inducible HO-1 to participate in cellular protection also in the heart. We, therefore, examined the expression and localization of HO-1 in a rat experimental myocardial infarction model. Male Wistar rats were subjected to left anterior coronary artery ligation or sham-operation and sacrificed at 1 day, 1 week and 4 weeks after ligation. The expression of HO-1 mRNA was assessed by real-time quantitative RT-PCR and the localization of HO-1 protein by immunoconfocal microscopy. At day 1, HO-1 mRNA was increased 3.9-fold in the peri-infarct border area vs sham-operated hearts (P<0.001) and 2.9-fold vs remote areas of the same hearts (P<0.001). At 1 week, HO-1 mRNA levels remained significantly higher (5-fold) in the peri-infarct border area than in sham-operated hearts (P<0.001). In addition, HO-1 mRNA transiently increased 1.6-fold in the remote non-infarcted myocardium vs sham operated hearts (P<0.05). HO-1 mRNA returned to basal levels by 4 weeks. The increase in HO-1 mRNA was accompanied by increased immunoreactivity of HO-1 protein in the vascular walls throughout the myocardium, and in the cardiomyocytes and fibroblast-like cells of the peri-infarct border areas. Cardiomyocytes showed immunoreactivity at the intercalated disc area, and in the sarcoplasmic reticulum as indicated by the striated pattern of staining. The results suggest that the induction of HO-1 may have an important role in the heart during the first days after myocardial infarction.

Left ventricular remodeling after infarction: sequential MR imaging with oral nicorandil therapy in rat model.

To use magnetic resonance (MR) imaging in quantification of the short- and long-term effects of therapy with orally administered nicorandil on left ventricular (LV) geometry and function independent of infarction size. Forty-six rats were subjected to reperfused infarction and randomly divided into two groups. Group 1 rats (n = 21) were treated with nicorandil (3 mg/kg/day in drinking water) for 4 days before infarction and 8 weeks after infarction (hereafter, the nicorandil group). Group 2 rats (n = 25) received tap water for the same period and served as the control group. Mesoporphyrin- (as a necrosis-specific agent) enhanced MR imaging was used to define necrotic myocardium on day 2 after infarction in all 46 animals. Contrast material-enhanced MR images showed large but identical infarction size in 11 control and 11 nicorandil rats. Only these 22 rats underwent repeat MR imaging at 8 weeks after infarction. The following variables were measured: LV volumes, ejection fraction, mass, wall thickness, and infarction size. Student t test and analysis of variance for repeated measurements were used for statistical analysis. The size of the necrotic region on mesoporphyrin-enhanced MR images was 39% +/- 3 of the size of the left ventricle in the control group and 41% +/- 2 in the nicorandil group (difference not significant, unpaired Student t test). Pretreatment with nicorandil for 6 days before imaging did not reduce LV dilation or improve function compared with those in control animals with identical infarction size. Eight weeks after infarction, control animals showed deterioration in LV function, wall thinning, and gradient in regional dysfunction (analysis of variance test). Nicorandil produced significant salutary effects on LV ejection fraction (37% +/- 3 in the nicorandil group vs 24% +/- 3 in the control group), end-diastolic volume (0.53 mL +/- 0.03 vs 0.65 mL +/- 0.04), end-systolic volume (0.36 mL +/- 0.03 vs 0.49 mL +/- 0.05), LV wall thickening in remote noninfarcted myocardium (28% +/- 2 vs 19% +/- 1), and a rim of infarction (16% +/- 2 vs 8% +/- 1) (P <.05 for all parameters). The increase in LV mass was reduced in the nicorandil group (0.73 g +/- 0.03) compared with that in the control group (0.89 g +/- 0.04) (P <.05). In animals studied longitudinally, MR imaging demonstrated the deleterious changes in LV geometry and function in the period after infarction and the salutary effects of medical therapy.

Antioxidant Therapy Attenuates JNK Activation and Apoptosis in the Remote Noninfarcted Myocardium after Large Myocardial Infarction

We hypothesized that the concomitant occurrence of increased oxidative stress, JNK activation, and myocyte apoptosis in the remote myocardium (RM) following a large myocardial infarction (MI) are causally related. Three days following coronary ligation, rats were randomized to treatment with probucol and PDTC (MI-T) or vehicle (MI). Control rats (C) underwent sham operation. At 7 weeks, TBARS assay showed increased level of lipid-peroxidation within the RM in the MI group vs C, which was completely inhibited in the MI-T group. Similarly, Western blot analysis showed a twofold increase in p-JNK in the MI group, vs C, which was attenuated in MI-T, a result confirmed by a JNK-kinase activity. Furthermore, apoptosis was increased within the RM in MI vs C, while this was inhibited in MI-T. We conclude that long-term antioxidant therapy with probucol and PDTC attenuates oxidative stress, JNK activation, and myocyte apoptosis within the RM after large MI.

Myocyte contractile function is intact in the post-infarct remodeled rat heart despite molecular alterations.

To investigate the cellular mechanisms underlying global and regional LV dysfunction in the post-infarct (MI) remodeled rat hearts. LV remodeling and function were quantified by echocardiography, morphometry, in vivo hemodynamics, and isolated perfused heart studies in 6 weeks post-MI and sham-operated rats. LV myocytes from sham and MI hearts were used for morphometric and functional studies. Myocyte contractile function and intracellular calcium kinetics were measured at different stimulation frequencies (0.2-2 Hz), temperatures (30 and 37 degrees C), and external viscous load (1, 15, 200 and 300 centipoise). Myocyte apoptosis was measured by DNA laddering; BCL-2, BAX, Na(+)-Ca(2+) exchanger, and SERCA-2 proteins by western blot; and brain natriuretic peptide (BNP), SERCA-2 mRNA by RT-PCR. MI hearts were remodeled (Echo LV diameter 7.3+/-0.38 vs. 5.9+/-0.16 mm, P<0.03), and showed global (Echo % fractional shortening 30+/-2.4 vs. 58+/-3, P<0.001), and regional contractile dysfunction of non-infarcted myocardium (Echo % systolic posterior wall thickening 36+/-2 vs. 57+/-1.7, P<0.001). In vivo hemodynamic and isolated heart function studies confirmed depressed LV systolic and diastolic function and increased volumes. Whereas, myocytes isolated from infarcted hearts were remodeled (40% longer and 10% wider), their contractile function and calcium kinetics under basal conditions and at high stimulation frequency, temperature and viscous load were similar to sham myocytes. The mRNA for BNP was increased whereas that for SERCA-2 decreased, but the SERCA-2 protein was normal. Despite myocyte hypertrophy, ventricular septal thickness was reduced in infarcted hearts (2.2+/-0.1 vs. 2. 6+/-0.07 mm, P<0.01), and showed increased apoptosis. Myocytes from remote non-infarcted myocardium of the remodeled hearts have normal contractile function, despite structural remodeling and altered gene expression. Non-myocyte factors may be more important in genesis of contractile dysfunction in the remodeled heart, for up to 6 weeks after MI.