Search tips
Search criteria 


Logo of nihpaAbout Author manuscriptsSubmit a manuscriptHHS Public Access; Author Manuscript; Accepted for publication in peer reviewed journal;
Circ Cardiovasc Imaging. Author manuscript; available in PMC 2012 September 1.
Published in final edited form as:
PMCID: PMC3178667

Dual Manganese-Enhanced and Delayed Gadolinium-Enhanced MRI Detects Myocardial Border Zone Injury in a Pig Ischemia-Reperfusion Model



Delayed gadolinium (Gd) enhancement MRI (DEMRI) identifies non-viable myocardium, but is non-specific and may overestimate nonviable territory. Manganese (Mn2+)-enhanced MRI (MEMRI) denotes specific Mn2+ uptake into viable cardiomyocytes. We performed a dual-contrast myocardial assessment in a porcine ischemia-reperfusion (IR) model to test the hypothesis that combined DEMRI and MEMRI will identify viable infarct border zone (BZ) myocardium in vivo.

Methods and Results

Sixty-minute LAD ischemia-reperfusion injury (IR) was induced in 13 adult swine. Twenty-one days post-IR, 3T cardiac MRI was performed. MEMRI was obtained after injection (0.7 cc/kg) of Mn2+ contrast agent (EVP1001-1, Eagle Vision Pharmaceutical Corp.). DEMRI was then acquired after 0.2mmol/kg Gd injection. Left ventricular (LV) mass, infarct, and function were analyzed. Subtraction of MEMRI defect from DEMRI signal identified injured border zone myocardium. Explanted hearts were analyzed by 2,3,5-triphenyltetrazolium chloride (TTC) stain and tissue electron microscopy (TEM) to compare infarct, BZ, and remote myocardium. Average LV ejection fraction was reduced (30±7%). MEMRI and DEMRI infarct volumes correlated with TTC (MEMRI: r=0.78; DEMRI: r=0.75; p<0.004). MEMRI infarct volume percentage was significantly lower than DEMRI (14±4%* vs. 23±4%; *p<0.05). BZ MEMRI SNR was intermediate to remote and core infarct SNR (7.5±2.8* vs. 13.2±3.4 and 2.9±1.6; *p<0.0001), and DEMRI BZ SNR tended to be intermediate to remote and core infarct (8.4±5.4 vs. 3.3±0.6 and 14.3±6.6; p>0.05). TEM analysis exhibited preserved cell structure in BZ cardiomyocytes despite transmural DEMRI enhancement.


Dual-contrast MEMRI-DEMRI detects BZ viability within DEMRI infarct zones. This approach may identify injured, at-risk myocardium in ischemic cardiomyopathy.

Keywords: ischemia-reperfusion, magnetic resonance imaging, delayed enhancement MRI, manganese-enhanced MRI, viability imaging

Coronary artery disease and ischemic cardiomyopathy are leading causes of morbidity and mortality. The improved survival of ischemic cardiomyopathy patients with coronary revascularization has been well-documented14; however, the assessment of viable myocardium is critical in determining which patients will benefit from revascularization. DEMRI is considered a gold standard for myocardial viability. This technique exploits the MRI T1-shortening effect of gadolinium (Gd), which distributes primarily within the extracellular space5. Gd accumulates in acutely or chronically infarcted myocardium, and transmural late enhancement is traditionally thought to indicate irreversible myocardial injury. However, Gd-based DEMRI does not provide direct cell viability information due to its nonspecific distribution properties.

Clinically, coronary bypass and percutaneous coronary intervention studies have used DEMRI to predict regional functional recovery after revascularization68. Despite the predictive potential of DEMRI, several groups have reported that DEMRI volume may decrease significantly over time913. Indeed, DEMRI using Gd may be positive in regions of myocardial edema and inflammation, which may cause transient, reversible cardiac injury patterns1417. However, DEMRI may also overestimate infarct areas due to the nonspecific distribution of gadolinium and the kinetics of its T1 signal in vivo913. Recent studies have demonstrated that DEMRI may consistently overestimate the amount of nonviable myocardium by nearly 15%, predominantly within the heterogeneous border zone. There are limited data as to whether viable, and potentially salvageable, cardiomyocytes exist within these ‘non-viable’ DEMRI territories. Similarly, there is no established imaging strategy to identify these border zone areas, which could have a meaningful survival impact for patients who sustain acute myocardial infarction and develop subsequent ischemic cardiomyopathy. An alternative approach utilizing additional contrast agents may complement Gd-based techniques in this capacity.

Manganese (Mn2+) is an essential metal divalent cation that can enter cells via voltage-gated calcium channels. Manganese-enhanced MRI (MEMRI) exploits the T1 shortening effect of Mn2+ and its uptake is dependent upon viable, functioning cells1820. Used widely in neuronal imaging, the uptake of Mn2+ into viable myocardial cells has also been well-documented18, 2123. Recent studies have attempted to characterize infarcted myocardium using MEMRI alone (MEMRI defect area) and found correlation with histopathological infarct volumes16, 18, 24. In this study, we combined DEMRI and MEMRI techniques, and we identified overlapping MEMRI-positive (viable) and DEMRI positive (non-viable) border zone regions from infarcted myocardium in a porcine ischemia-reperfusion (IR) model. This MRI-detectable, mismatched border zone contained live cardiomyocytes with ultrastructural preservation.


Ischemia Reperfusion

All animal studies were approved by Stanford University Administrative Panel on Laboratory Animal Care. Thirteen Yorkshire swine were subjected to left anterior descending (LAD) ischemia-reperfusion for 60 minutes, as previously described25, 26. Yorkshire swine (30–45 kg) were anesthetized by inhaled isoflurane (1–2%). A bolus of 300IU/kg heparin was administered intravenously. A 10mm over-the-wire angioplasty balloon was placed in the left anterior descending artery (LAD) proximal to the first diagonal branch and inflated for 60 minutes.

To prevent arrhythmias, 150 mg of amiodarone was administered intravenously (IV) prior to the induction of ischemia. If indicated, non-synchronized direct current defibrillation was attempted at 360 J.

After the 60 minute occlusion, the balloon was deflated and reperfusion of the LAD was documented by coronary angiography (CAG). The vessel sheath was removed and the wound was closed. The animals were weaned from mechanical ventilation and transferred to the animal care facility.


Twenty-one days post-IR, 3T cardiac MRI was performed (Signa 3T-HDx, GE, USA) using an 8-channel chest coil (GE) and cardiac vector gating. Swine were anesthetized by inhaled isoflurane (1–2%). Following localizer images, FIESTA (SSFP: TR3.8ms, TEmin-full, FA45, ST10mm, matrix224×224, FOV35) cine images were obtained in standard long- and short-axis image planes. MEMRI (FGRE-IR: TR4.7 ms, TE1.3 ms, TI 200 ms, FA 10, ST 10 mm, matrix 224×192) was obtained 25–40 min after a 0.7cc/kg IV bolus of EVP1001-1 (Eagle Vision Pharmaceutical Corp., Downington, PA). Following MEMRI imaging, a 30minute washout period preceded infusion of Gd for DEMRI imaging. 3D DEMRI (3D MDE: 3D-FGRE-IR: TR 4.6, TE min, FA 15, ST 1.4mm, matrix 256×256, FOV 35) was acquired 10–20 min post-Gd injection (0.2 mmol/kg Magnevist, Bayer, Germany). Images were analyzed using Osirix (Pixmeo Inc. Geneva, Switzerland) with manual LV mass and infarct volume tracing. MEMRI defect areas and DEMRI enhanced areas were designated infarct areas. These areas were traced in short-axis slices and integrated to determine infarct volumes by MEMRI and DEMRI in matched swine hearts. Percent infarct volume: (infarct volumex100)/total LV mass volume. MEMRI defect volumes were not reliably traced using semiautomatic methods, therefore, both MEMRI and DEMRI infarct volumes were visually traced for consistency. To account for potential partial volume effects with DEMRI tracing, visually traced DEMRI infarct volumes were compared to a subset of semi-automatic full width half maximum (FWHM) tracings (Osirix plugin, courtesy of David Murday), and the two methods were not statistically different (mean infarct volume %: visual tracing, 22.7±4.4; FWHM, 25±6.3, n=6 hearts), exhibiting consistent agreement between individual measurements as well (Bland-Altman Bias of visual vs. FWHM method: −3.1±3.2%). Therefore, visual tracing was employed for both MEMRI and DEMRI infarct volume assessment.

In swine with microvascular obstruction on DEMRI, these hypointense areas were included as infarct area. The core infarct was defined as the central portion of DEMRI signal, which displayed >50% of the maximal Signal-to-Noise Ratio (SNR) on DEMRI images27. The border zone was defined as the regions of overlap between positive MEMRI and positive DEMRI signal. The remote zone was defined as any region that was not DEMRI positive. MEMRI and DEMRI images were analyzed for SNR variation between the remote, border, and infarct zones. Average SNR was computed from each zone by averaging 3 ROIs per zone for each swine heart, and averaging these values across 13 swine hearts. SNR was calculated as: Average SI of tissue/SD of air (SI: signal intensity; SD: standard deviation).

Double Staining to Measure Infarct Size and Areas at Risk for Ischemia

Following the MRI, in situ double-staining with 1% Evans blue dye and 1% solution TTC was performed to delineate areas at risk for ischemia versus infarction as described26. Evans blue dye binds plasma albumin and stains the vascular distribution of injected vessels. In viable myocardium, TTC is converted by mitochondrial dehydrogenase enzymes28 to a red formazan pigment that stains the myocardium red, leaving necrotic myocardium white due to lost dehydrogenase activity. Under anesthesia, the left and right coronary arteries (LCA and RCA) were simultaneously cannulated, and a 0.014 inch guide wire was placed in the LAD, and then the LAD was occluded with an over- the-wire balloon catheter at the site of the previous occlusion, guided by cine and still angiographic images from the original occlusion. In the pig coronary circulation, there is minimal natural collateral growth, providing a reliable and consistent staining method for the evaluation of the myocardial infarction. After confirmation of complete occlusion of LAD by CAG, 1% Evans blue dye was injected into LCA and RCA (60 mL into LCA and 30 mL into RCA) through the guiding catheters. In addition, 20–30 mL of a 1% TTC solution was injected distal to the occluded LAD territory using the guide-wire lumen of the over-the-wire balloon catheter. During this double-staining, animals were anesthetized with 5% isoflurane and then sacrificed with an IV potassium chloride injection. The myocardium supplied by the previously occluded LAD, defined as the area-at-risk for ischemia (AAR), was Evans blue dye-negative. The swine hearts were then excised and left ventricles were sectioned into 10mm cross-sectional myocardial slices parallel to the atrioventricular groove from apex to base. All slices were weighed, photographed, and fixed in 10% formaldehyde. TTC-positive areas (white) were defined as core infarct, TTC-negative areas (red) that were also Evans blue-negative were defined as the AAR. Infarcted sizes were manually measured using Adobe Photoshop Elements 8 (Adobe Inc., San Jose, CA).

Tissue Electron Microscopy

Myocardial tissue was excised from DEMRI negative (remote) and DEMRI positive (infarct) regions by visually correlating the mid-ventricular, cross-sectional gross specimen with mid-ventricular in vivo short axis DEMRI images. Samples were also excised from myocardial regions with overlapping positive MEMRI and DEMRI signal. The samples were cut into 2mm3 pieces and fixed with 2% glutaraldehyde and 4% paraformaldehyde in sodium cacodylate buffer. Post- xation was performed with 1% osmiumtetroxyde 1h at 4°C. After dehydration and embedding, sections were analysed by a JEOL 1230 tissue electron microscope (JEOL Ltd., Tokyo, Japan) at 80keV. Photos were taken using a Gatan Multiscan 791 digital camera (Gatan Inc., Pleasanton, CA).

Two to three nuclei were identified per cardiomyocyte, with 10–15 myocytes analyzed per zone (remote, infarct, and border). Cell ultrastructural analysis was performed by blinded observers, assessing 16 ‘healthy’ and ‘unhealthy’ features of cell integrity and sarcomeric organization29. A scoring system assessed the relative presence or absence of each of the 16 features. For ‘healthy’ features, each identified cardiomyocyte was graded on whether it exhibited: ‘5’–high abundance; ‘4’–moderate abundance; ‘3’–low abundance; ‘2’–rare; or ‘1’–complete absence of that feature (Table: black). Conversely, for an ‘unhealthy’ TEM feature (Table: bold italics): ‘5’–indicated that the nucleus displayed complete absence; ‘4’–rare; ‘3’–low abundance; ‘2’–moderate abundance; and ‘1’–high abundance of the ‘unhealthy’ feature. For example, the complete absence of a ‘healthy’ feature or the high abundance of an ‘unhealthy’ feature yielded a score of ‘1’. Individual scores from 10–15 myocytes per zone were averaged to generate an overall zone score for that feature (from 1–5). The 16 scores were averaged to generate a composite score for each zone that reflected the overall structural integrity of the cells within each zone, with ‘5’ being the best, and ‘1’ being the worst score. Agreement on TEM scoring between two independent observers was high (Kappa = 0.79). Composite scores were compared between remote zone, border zone, and core infarct zone TEM.

Cardiomyocyte Ultrastructure Score by Tissue Electron Microscopy

Statistical Analyses

Results in TEM score (range between 1 and 5) are shown as mean +/− standard deviation. Significant differences (p<0.05) were tested using the Kruskal-Wallis test for the composite scores of the three myocardial zones, and a Bonferoni post-test ANOVA test for SNR comparison. DEMRI, MEMRI and TTC methods were compared using a Wilcoxon signed rank test (two-tailed) on the related measures of infarct volumes between the groups. Spearman correlation tested for association between MRI measurements of infarct volume (dependent variable) and TTC infarct volume (independent variable).


Left ventricular systolic function/morphology

At three weeks post-IR, left ventricular ejection fraction was markedly reduced (30±7%*, n=13), compared to age-matched control swine (EF = 66±3%, n=3). Left ventricular volumes were dilated (end-diastolic volume: 141±23ml; end-systolic volume: 100±23ml) and all swine displayed significant thinning and hypokinesis/akinesis of the anterolateral and septal walls.

Quantitative correlation of MEMRI-DEMRI infarct volume and histopathology

MEMRI from the IR swine showed reproducible, homogeneous myocardial Mn2+ uptake in remote (normal) areas of myocardium (Figure 1). Within the anteroseptal, anterior, and anterolateral walls, MEMRI signal defect (infarct) was consistently observed and corresponded to the core infarct zone of DEMRI images. Due to higher signal intensity, DEMRI-positive myocardium was easily demarcated against any residual MEMRI signal. MEMRI infarct volume correlated highly with TTC infarct volume (r=0.78, p=0.002) and DEMRI infarct volume also correlated closely with TTC infarct volume (r=0.75, p=0.003).

Figure 1

MEMRI versus DEMRI infarct volume

Quantitative comparison revealed that the MEMRI infarct volume percentage (14±4%*, n = 13) was significantly (*p<0.001) lower than both DEMRI infarct volume percentage (23±4%, n=13) and TTC infarct volume percentage (19±3%). In addition, an overlapping border zone (BZ) that was both MEMRI and DEMRI positive was consistently observed (Figure 1). To determine whether the high correlation between MEMRI/DEMRI and TTC measures also exhibited agreement, Bland-Altman plots30 of infarct volumes were created to show the differences agreement between TTC - DEMRI or TTC - MEMRI (Figure 1B). The measurement differences from TTC to DEMRI were all negative, and the limits of agreement did not include 0, indicating that DEMRI consistently overestimates the infarct volumes. Conversely, MEMRI measures are consistently lower than TTC, showing positive measurement differences between TTC and MEMRI. Moreover, the differences were proportional to the mean (larger differences for larger infarct volume). However, the Bland-Altman plots of infarct volume differences between TTC–MEMRI (all positive values) and TTC - DEMRI (all negative values) were within two standard deviations of the mean differences for each contrast agent (Figure 1B), indicating that the reduced infarct volume % of MEMRI compared to DEMRI was a consistent finding within the group.

Tissue Characterization of the Border Zone (BZ)

In order to analyze the tissue characteristics of the BZ, signal-to-noise ratios (SNR) from the border, core infarct, and remote zones were obtained for both MEMRI and DEMRI. By MEMRI, the border zone SNR (7.5±2.8*,#, n=13) was significantly lower when compared to remote zone SNR (13.2±3.4), yet significantly higher than MEMRI SNR from the core infarct zone (2.9±1.6*, Figure 2; *p<0.05 vs remote; #p<0.05 vs core infarct MEMRI SNR). Similarly, by DEMRI, the SNR of the border zone tended to be lower than core infarct SNR, but higher than remote zone SNR. These intermediate SNR levels further distinguished the border zone from both the remote and core infarct zones, indicating potentially heterogeneous cell populations within the border zone.

Figure 2
SNR Heterogeneity within Border Zone Regions

One potential limitation of this study was that Mn2+ administration prior to Gd might affect the T1-shortening properties of Gd images. To explore this possibility, we examined several swine with a reverse order protocol (DEMRI-before-MEMRI, or DEMRI-alone), and we did not observe any noticeable affect on DEMRI infarct % (21±9% with EF 29±7%; n=8). Intermediate BZ DEMRI SNR was similarly observed in DEMRI-before-MEMRI hearts (BZ SNR 6.9±2.2; n=8; p>0.05) compared to DEMRI-after-MEMRI BZ SNR). Notably, MEMRI defect was not interpretable on images with the DEMRI-before-MEMRI reverse protocol, due to the high intensity T1 signal of DEMRI in the infarct zone. In summary, digital subtraction of MEMRI-negative infarct regions from DEMRI-positive infarct regions revealed overlapping positive signal designated as border zone areas. This overlap suggested a significant number of viable myocytes in these transmural DEMRI-positive regions, which may contribute to the intermediate MEMRI and DEMRI SNR.

TEM of infarct, border, and remote zone myocardium

To further characterize the ultrastructure of the contractile apparatus within these border zone regions, TEM was performed from the core infarct, border zone, and remote zone regions, using established characteristics of normal nuclear, chromatin, mitochondrial and sarcomeric structure and organization29 (Figure 3). As noted above, these outer edges of TTC-positive and DEMRI-positive (non-viable) signal, when superimposed upon MEMRI images, overlapped significantly with the outer regions of positive MEMRI signal (viable). The core infarct, border, and remote zones were quantitatively analyzed. Border zone cells displayed significant ultrastructural preservation (overall TEM score: 4.4±0.4, n=10), similar to remote zone TEM (4.9±0.1, n=10), and unlike infarct zone cells (1.3±0.3*, n=10, *p<0.05 versus remote zone score). These results are displayed in the Table. Together, the intact cell structure and observed Mn2+ uptake of border zone cells suggest viable cardiomyocyte populations within the areas of transmural DEMRI.

Figure 3
TEM of Infarct, Border, and Remote Myocardium


In this study, we applied a novel, dual-contrast MEMRI-DEMRI strategy to determine if viable myocardium was detectable within regions of positive, transmural DEMRI. MEMRI infarct volume was found to be significantly (39%) lower than DEMRI infarct volume in this 21-day swine IR model. This signal mismatch identified a border zone that was positive for both MEMRI signal (viable) and transmural DEMRI signal (non-viable), and which also displayed an intermediate SNR by MEMRI, compared to core infarct and remote zones. TEM analysis also revealed preservation of cell architecture and contractile elements within this border zone region. Overall, these results suggest that dual contrast MEMRI-DEMRI may allow accurate detection of viable myocardium within the infarct border zone that appears nonviable by DEMRI alone.

Although most patients who suffer an MI will survive the acute event31, they often develop heart failure32 and a progressive decline in ventricular function that is believed to involve apoptosis and collagen deposition, particularly in border zone areas33,34. Revascularization decisions for these patients are frequently made at intermediate and chronic timepoints post-MI and are often based upon viability assessment. In this study, a 21-day timepoint post-IR was chosen to explore this intermediate timepoint, and DEMRI infarct volume measurements were significantly higher than either MEMRI or TTC infarct volumes. These data suggest that DEMRI may overestimate infarct volume at intermediate post-IR timepoints, and that combination with MEMRI may accurately delineate these injured, but live populations of cardiomyocytes.

Intermediate MEMRI SNR measurements also reflected tissue heterogeneity within this mismatched border zone, which has not been previously reported using MEMRI in a myocardial IR model. While prior studies have reported heterogenous DEMRI signal in infarct border zones14, Gd accumulation is non-specific and provides limited information on actual cell viability. Because of specific Mn2+ uptake into viable, functioning cells, the observed MEMRI SNR heterogeneity points to significant populations of cardiomyocytes that are alive with intact Ca2+-channel function (MEMRI positive) in this region despite surrounding necrotic tissue (DEMRI positive). The intermediate degree of Mn2+ uptake within the border zone demonstrates that this region is not only visually and quantitatively distinct from the core infarct zone by TEM analysis, but biologically distinct as well. Prior MEMRI studies have imaged myocardial infarct areas16, 18, 24; however, these studies focused either on the acute ischemia phase or have looked at chronic timepoints with ex vivo analysis. Intermediate MEMRI and DEMRI SNR, with intact cell structure, suggests that the DEMRI signal may still be evolving at 21 days post-IR and not yet reflective of true infarct size. Future studies may examine earlier and later time points to further characterize this mismatch region.

Viable myocardium that falls outside the core infarct zone has been imaged previously using a combination of agents, including radio-labeled and fluorescent microparticles8, 25, 3538. However, in this study, MEMRI infarct volume was compared directly to DEMRI infarct volume and was found to be significantly smaller, pointing to viable cardiomyocytes within areas of transmural DEMRI. A technical limitation of this study, and of DEMRI in general, is the potential for partial volume effects during infarct volume quantification. The use of 3D DEMRI images mitigated the partial volume effects by employing a thinner slice thickness (1.4mm) than standard 2D DEMRI images (10mm). As described above, a subset of DEMRI infarct volumes were analyzed using a semiautomatic FWHM method, and infarct sizes were not statistically different from visual tracing results. Our results are consistent with two recent studies, in which manual tracing of DEMRI volumes was equivalent to both FWHM- and Standard Deviation-based semiautomatic methods39, 40. While some partial volume effect may contribute to border zone signal, the TEM analysis shows relative preservation of cytoarchitecture in this region, lending support to the overlap of the positive MEMRI and DEMRI signals in the border zone territories. These findings are consistent with previous publications that document DEMRI overestimation of infarct volume913. Absolute and relative degrees of DEMRI infarct volume have also been observed to decrease over time following a myocardial infarction, which has been attributed to cell debris removal and resolution of myocardial edema11. This diminishing zone of positive DEMRI underscores the need for improved imaging approaches to detect viable myocardial territories at early and intermediate time points post-MI with high accuracy to assist decisions on revascularization. Specifically, patients in the peri-infarct period may benefit from an imaging strategy that better delineates the amount of injured, yet viable myocardium that would be jeopardized if not revascularized.

The use of manganese as a MRI contrast agent has been limited by its potential for adverse cardiovascular effects, which is mainly attributed to its competition for the L-type Ca2+-channel on the sarcolemmal membrane41. To mitigate these effects, the EVP-1011 used for MEMRI contains calcium as well, and no toxicity was observed in swine dosed at 0.7cc/kg. Indeed, clinical manganese MRI imaging agents are already in use, namely mangafodopir trisodium (MnDPDP), which is FDA-approved for liver tumor imaging42. Although no approval for cardiac imaging with manganese exists at present, EVP-1011 is currently being evaluated for FDA approval.

T2-weighted imaging (T2WI) has also been validated for characterizing peri-infarct zone biology, including ‘area-at-risk’ (AAR), in both the acute and subacute settings43, 44. The presence of T2WI edema is particularly useful in the acute coronary syndrome (ACS) setting, as it was recently demonstrated to independently predict the presence of obstructive coronary disease and even 6-month survival in ACS patients45. However, by 14–28 days post-infarct, myocardial edema may have dissipated completely46,4749, which lessens the utility of T2WI at intermediate and chronic timepoints. MEMRI-DEMRI dual contrast imaging affords unique information about infarct zone biology that may be particularly effective at intermediate and chronic timepoints post-infarct. Moreover, in contrast to T2WI, which assesses AAR lying primarily outside DEMRI territory50, the MEMRI-DEMRI strategy presented herein detects live cardiomyocyte populations within DEMRI-positive zones.

An important limitation of this study is that no revascularization data is available to confirm the functional viability of these overlapping DEMRI- and MEMRI-positive regions. Consequently, there is no evidence that these live cells are capable of contributing to overall ventricular function if they were to be salvaged with revascularization. Indeed, these populations of live cells may represent arrhythmogenic foci, as recent work has linked the border or ‘gray’ zone areas of DEMRI, which displays SNR heterogeneity, with increased propensity for inducible ventricular tachycardia51 and cardiovascular events27. Future revascularization studies would be required to understand how these pockets of viable myocardium may affect long-term outcome, and whether aggressive restoration of coronary blood flow to these incompletely scarred regions is beneficial.

In summary, the infarct border zone represents a heterogeneous region comprised of complex post-infarction biology. In order to better characterize this dynamic process, contrast agents that enable both anatomical (DEMRI) and biological (MEMRI) information may be advantageous. This study demonstrates a non-invasive, dual-contrast MEMRI-DEMRI detection method for viable myocardium within the infarct border zone. Cells in the border zone display relatively intact cytoarchitecture, which suggests salvageable myocardium and/or arrhythmogenic foci. Future studies are needed to determine how these pockets of viable tissue may impact revascularization strategy in ischemic cardiomyopathy.

Clinical Summary

Improved survival of ischemic cardiomyopathy patients with coronary revascularization has been well documented. Clinically, the assessment of viable myocardium is crucial for determining which patients will benefit from revascularization. Delayed gadolinium-enhanced MRI (DEMRI) is considered a gold standard for myocardial viability. Transmural DEMRI signal is thought to indicate irreversible myocardial injury; however, DEMRI signal is not specific for cell viability and may overestimate true myocardial infarct volumes by nearly 15% in some studies.

In this study, we demonstrate a novel, dual-contrast MRI strategy to non-invasively characterize the injured border zone of the myocardium post-infarction. Manganese (Mn2+) is an essential metal cation that is avidly taken up by live myocardial cells. Manganese-enhanced MRI (MEMRI) signal is, therefore, a specific marker for viable cardiomyocytes, producing a bright T1 signal in live myocardium, and a signal defect in infarcted myocardium. Adult farm pigs underwent one-hour ischemia-reperfusion injury and were imaged by cardiac MRI 21days later with MEMRI and DEMRI sequences. Both contrast agents correlated well with histopathologic scar volume; however, MEMRI infarct volume was significantly (39%) smaller than DEMRI infarct volume. There was significant overlap in positive signal from MEMRI and DEMRI, and these overlap regions displayed intermediate SNR and preserved cytoarchitecture by electron microscopy, indicating that live cells may reside within regions of transmural DEMRI-positive myocardium. The potential clinical impact of this novel imaging strategy is significant, as MEMRI-DEMRI dual contrast may provide complementary information on complex, heterogeneous border zone biology that could help guide therapy for ischemic heart disease patients.


We would like to thank the following people/entities for their assistance and support:

Alfredo Green and John Perrino for technical assistance.

David Murday for Osirix FWHM plugin.

National Institute of Health/NHLBI

General Electric Healthcare

Sources of Funding

National Institute of Health/NHBLI (RD, PCY)

General Electric Healthcare (MM, PCY)



General Electric Healthcare (MM, PCY)

Boehringer-Ingelheim, Research (PCY)


1. Cameron A, Davis KB, Green G, Schaff HV. Coronary bypass surgery with internal-thoracic-artery grafts--effects on survival over a 15-year period. N Engl J Med. 1996;334:216–219. [PubMed]
2. Cameron AA, Green GE, Brogno DA, Thornton J. Internal thoracic artery grafts: 20-year clinical follow-up. J Am Coll Cardiol. 1995;25:188–192. [PubMed]
3. Fitzgibbon GM, Kafka HP, Leach AJ, Keon WJ, Hooper GD, Burton JR. Coronary bypass graft fate and patient outcome: angiographic follow-up of 5,065 grafts related to survival and reoperation in 1,388 patients during 25 years. J Am Coll Cardiol. 1996;28:616–626. [PubMed]
4. Lytle BW, Cosgrove DM, Loop FD, Borsh J, Goormastic M, Taylor PC. Perioperative risk of bilateral internal mammary artery grafting: analysis of 500 cases from 1971 to 1984. Circulation. 1986;74:III37–41. [PubMed]
5. Kanderian AS, Renapurkar R, Flamm SD. Myocardial viability and revascularization. Heart Fail Clin. 2009;5:333–348. vi. [PubMed]
6. Hillenbrand HB, Kim RJ, Parker MA, Fieno DS, Judd RM. Early assessment of myocardial salvage by contrast-enhanced magnetic resonance imaging. Circulation. 2000;102:1678–1683. [PubMed]
7. Rogers WJ, Jr, Kramer CM, Geskin G, Hu YL, Theobald TM, Vido DA, Petruolo S, Reichek N. Early contrast-enhanced MRI predicts late functional recovery after reperfused myocardial infarction. Circulation. 1999;99:744–750. [PubMed]
8. Fieno DS, Kim RJ, Chen EL, Lomasney JW, Klocke FJ, Judd RM. Contrast-enhanced magnetic resonance imaging of myocardium at risk: distinction between reversible and irreversible injury throughout infarct healing. J Am Coll Cardiol. 2000;36:1985–1991. [PubMed]
9. Suranyi P, Kiss P, Brott BC, Simor T, Elgavish A, Ruzsics B, Saab-Ismail NH, Elgavish GA. Percent infarct mapping: an R1-map-based CE-MRI method for determining myocardial viability distribution. Magn Reson Med. 2006;56:535–545. [PubMed]
10. Judd RM, Lugo-Olivieri CH, Arai M, Kondo T, Croisille P, Lima JA, Mohan V, Becker LC, Zerhouni EA. Physiological basis of myocardial contrast enhancement in fast magnetic resonance images of 2-day-old reperfused canine infarcts. Circulation. 1995;92:1902–1910. [PubMed]
11. Saeed M, Bremerich J, Wendland MF, Wyttenbach R, Weinmann HJ, Higgins CB. Reperfused myocardial infarction as seen with use of necrosis-specific versus standard extracellular MR contrast media in rats. Radiology. 1999;213:247–257. [PubMed]
12. Saeed M, Lund G, Wendland MF, Bremerich J, Weinmann H, Higgins CB. Magnetic resonance characterization of the peri-infarction zone of reperfused myocardial infarction with necrosis-specific and extracellular nonspecific contrast media. Circulation. 2001;103:871–876. [PubMed]
13. Amado LC, Gerber BL, Gupta SN, Rettmann DW, Szarf G, Schock R, Nasir K, Kraitchman DL, Lima JA. Accurate and objective infarct sizing by contrast-enhanced magnetic resonance imaging in a canine myocardial infarction model. J Am Coll Cardiol. 2004;44:2383–2389. [PubMed]
14. Fenster BE, Chan FP, Valentine HA, Yang E, McConnell MV, Berry GJ, Yang PC. Images in cardiovascular medicine. Cardiac magnetic resonance imaging for myocarditis: effective use in medical decision making. Circulation. 2006;113:e842–843. [PubMed]
15. Eitel I, Behrendt F, Schindler K, Kivelitz D, Gutberlet M, Schuler G, Thiele H. Differential diagnosis of suspected apical ballooning syndrome using contrast-enhanced magnetic resonance imaging. Eur Heart J. 2008;29:2651–2659. [PubMed]
16. Haghi D, Fluechter S, Suselbeck T, Borggrefe M, Papavassiliu T. Delayed hyperenhancement in a case of Takotsubo cardiomyopathy. J Cardiovasc Magn Reson. 2005;7:845–847. [PubMed]
17. Syed IS, Martinez MW, Feng DL, Glockner JF. Cardiac magnetic resonance imaging of eosinophilic endomyocardial disease. Int J Cardiol. 2008;126:e50–52. [PubMed]
18. Yamada M, Gurney PT, Chung J, Kundu P, Drukker M, Smith AK, Weissman IL, Nishimura D, Robbins RC, Yang PC. Manganese-guided cellular MRI of human embryonic stem cell and human bone marrow stromal cell viability. Magn Reson Med. 2009;62:1047–1054. [PubMed]
19. Bruvold M, Nordhoy W, Anthonsen HW, Brurok H, Jynge P. Manganese-calcium interactions with contrast media for cardiac magnetic resonance imaging: a study of manganese chloride supplemented with calcium gluconate in isolated Guinea pig hearts. Invest Radiol. 2005;40:117–125. [PubMed]
20. Mendonca-Dias MH, Gaggelli E, Lauterbur PC. Paramagnetic contrast agents in nuclear magnetic resonance medical imaging. Semin Nucl Med. 1983;13:364–376. [PubMed]
21. Wendland MF. Applications of manganese-enhanced magnetic resonance imaging (MEMRI) to imaging of the heart. NMR Biomed. 2004;17:581–594. [PubMed]
22. Storey P, Danias PG, Post M, Li W, Seoane PR, Harnish PP, Edelman RR, Prasad PV. Preliminary evaluation of EVP 1001-1: a new cardiac-specific magnetic resonance contrast agent with kinetics suitable for steady-state imaging of the ischemic heart. Invest Radiol. 2003;38:642–652. [PubMed]
23. Storey P, Chen Q, Li W, Seoane PR, Harnish PP, Fogelson L, Harris KR, Prasad PV. Magnetic resonance imaging of myocardial infarction using a manganese-based contrast agent (EVP 1001-1): preliminary results in a dog model. J Magn Reson Imaging. 2006;23:228–234. [PubMed]
24. Delattre BM, Braunersreuther V, Hyacinthe JN, Crowe LA, Mach F, Vallee JP. Myocardial infarction quantification with Manganese-Enhanced MRI (MEMRI) in mice using a 3T clinical scanner. NMR Biomed. 2010;23:503–513. [PubMed]
25. Kaneda H, Ikeno F, Inagaki K, Mochly-Rosen D. Preserved coronary endothelial function by inhibition of delta protein kinase C in a porcine acute myocardial infarction model. Int J Cardiol. 2009;133:256–259. [PMC free article] [PubMed]
26. Suzuki Y, Lyons JK, Yeung AC, Ikeno F. The porcine restenosis model using thermal balloon injury: comparison with the model by coronary stenting. J Invasive Cardiol. 2008;20:142–146. [PubMed]
27. Heidary S, Patel H, Chung J, Yokota H, Gupta SN, Bennett MV, Katikireddy C, Nguyen P, Pauly JM, Terashima M, McConnell MV, Yang PC. Quantitative tissue characterization of infarct core and border zone in patients with ischemic cardiomyopathy by magnetic resonance is associated with future cardiovascular events. J Am Coll Cardiol. 2010;55:2762–2768. [PubMed]
28. Klein HHSP, Schaper J, Schaper W. The Mechanism of the Tetrazolium Reaction in Identifying Experimental Myocardial Infarction. Virchows Archiv. A, Pathological anatomy and histopathology. 1981;393:287–297.
29. Misfeld M, Szabo K, Kraatz EG, Grossherr M, Schmidtke C, Pilgrim M, Kuhnel W, Sievers HH. Electron-microscopic findings after transmyocardial laser revascularization in an acute ischemic pig model. Eur J Cardiothorac Surg. 1998;13:398–403. [PubMed]
30. Bland JM, Altman DG. Measuring agreement in method comparison studies. Stat Methods Med Res. 1999;8:135–160. [PubMed]
31. Lloyd-Jones D, Adams R, Carnethon M, De Simone G, Ferguson TB, Flegal K, Ford E, Furie K, Go A, Greenlund K, Haase N, Hailpern S, Ho M, Howard V, Kissela B, Kittner S, Lackland D, Lisabeth L, Marelli A, McDermott M, Meigs J, Mozaffarian D, Nichol G, O’Donnell C, Roger V, Rosamond W, Sacco R, Sorlie P, Stafford R, Steinberger J, Thom T, Wasserthiel-Smoller S, Wong N, Wylie-Rosett J, Hong Y. Heart disease and stroke statistics--2009 update: a report from the American Heart Association Statistics Committee and Stroke Statistics Subcommittee. Circulation. 2009;119:480–486. [PubMed]
32. Mielniczuk LM, Lamas GA, Flaker GC, Mitchell G, Smith SC, Gersh BJ, Solomon SD, Moye LA, Rouleau JL, Rutherford JD, Pfeffer MA. Left ventricular end-diastolic pressure and risk of subsequent heart failure in patients following an acute myocardial infarction. Congest Heart Fail. 2007;13:209–214. [PubMed]
33. Olivetti G, Quaini F, Sala R, Lagrasta C, Corradi D, Bonacina E, Gambert SR, Cigola E, Anversa P. Acute myocardial infarction in humans is associated with activation of programmed myocyte cell death in the surviving portion of the heart. J Mol Cell Cardiol. 1996;28:2005–2016. [PubMed]
34. van den Borne SW, Isobe S, Zandbergen HR, Li P, Petrov A, Wong ND, Fujimoto S, Fujimoto A, Lovhaug D, Smits JF, Daemen MJ, Blankesteijn WM, Reutelingsperger C, Zannad F, Narula N, Vannan MA, Pitt B, Hofstra L, Narula J. Molecular imaging for efficacy of pharmacologic intervention in myocardial remodeling. JACC Cardiovasc Imaging. 2009;2:187–198. [PubMed]
35. Misra P, Lebeche D, Ly H, Schwarzkopf M, Diaz G, Hajjar RJ, Schecter AD, Frangioni JV. Quantitation of CXCR4 expression in myocardial infarction using 99mTc-labeled SDF-1alpha. J Nucl Med. 2008;49:963–969. [PMC free article] [PubMed]
36. Zhao M, Zhu X, Ji S, Zhou J, Ozker KS, Fang W, Molthen RC, Hellman RS. 99mTc-labeled C2A domain of synaptotagmin I as a target-specific molecular probe for noninvasive imaging of acute myocardial infarction. J Nucl Med. 2006;47:1367–1374. [PubMed]
37. Beller GA, Smith TW, Hood WB., Jr Effects of ischemia and coronary reperfusion on myocardial digoxin uptake. Am J Cardiol. 1975;36:902–907. [PubMed]
38. Kenis H, Zandbergen HR, Hofstra L, Petrov AD, Dumont EA, Blankenberg FD, Haider N, Bitsch N, Gijbels M, Verjans JW, Narula N, Narula J, Reutelingsperger CP. Annexin A5 uptake in ischemic myocardium: demonstration of reversible phosphatidylserine externalization and feasibility of radionuclide imaging. J Nucl Med. 2010;51:259–267. [PubMed]
39. Flett AS, Hasleton J, Cook C, Hausenloy D, Quarta G, Ariti C, Muthurangu V, Moon JC. Evaluation of techniques for the quantification of myocardial scar of differing etiology using cardiac magnetic resonance. JACC Cardiovasc Imaging. 2011;4:150–156. [PubMed]
40. Harrigan CJ, Peters DC, Gibson CM, Maron BJ, Manning WJ, Maron MS, Appelbaum E. Hypertrophic cardiomyopathy: quantification of late gadolinium enhancement with contrast-enhanced cardiovascular MR imaging. Radiology. 2011;258:128–133. [PubMed]
41. Silva AC, Lee JH, Aoki I, Koretsky AP. Manganese-enhanced magnetic resonance imaging (MEMRI): methodological and practical considerations. NMR Biomed. 2004;17:532–543. [PubMed]
42. Federle M, Chezmar J, Rubin DL, Weinreb J, Freeny P, Schmiedl UP, Brown JJ, Borrello JA, Lee JK, Semelka RC, Mattrey R, Dachman AH, Saini S, Harms SE, Mitchell DG, Anderson MW, Halford HH, 3rd, Bennett WF, Young SW, Rifkin M, Gay SB, Ballerini R, Sherwin PF, Robison RO. Efficacy and safety of mangafodipir trisodium (MnDPDP) injection for hepatic MRI in adults: results of the U.S. Multicenter phase III clinical trials. Efficacy of early imaging. J Magn Reson Imaging. 2000;12:689–701. [PubMed]
43. Cury RC, Shash K, Nagurney JT, Rosito G, Shapiro MD, Nomura CH, Abbara S, Bamberg F, Ferencik M, Schmidt EJ, Brown DF, Hoffmann U, Brady TJ. Cardiac magnetic resonance with T2-weighted imaging improves detection of patients with acute coronary syndrome in the emergency department. Circulation. 2008;118:837–844. [PubMed]
44. Edwards NC, Routledge H, Steeds RP. T2-weighted magnetic resonance imaging to assess myocardial oedema in ischaemic heart disease. Heart. 2009;95:1357–1361. [PubMed]
45. Raman SV, Simonetti OP, Winner MW, 3rd, Dickerson JA, He X, Mazzaferri EL, Jr, Ambrosio G. Cardiac magnetic resonance with edema imaging identifies myocardium at risk and predicts worse outcome in patients with non-ST-segment elevation acute coronary syndrome. J Am Coll Cardiol. 2010;55:2480–2488. [PubMed]
46. Schulz-Menger J, Gross M, Messroghli D, Uhlich F, Dietz R, Friedrich MG. Cardiovascular magnetic resonance of acute myocardial infarction at a very early stage. J Am Coll Cardiol. 2003;42:513–518. [PubMed]
47. Nilsson JC, Nielsen G, Groenning BA, Fritz-Hansen T, Sondergaard L, Jensen GB, Larsson HB. Sustained postinfarction myocardial oedema in humans visualised by magnetic resonance imaging. Heart. 2001;85:639–642. [PMC free article] [PubMed]
48. Ripa RS, Nilsson JC, Wang Y, Sondergaard L, Jorgensen E, Kastrup J. Short- and long-term changes in myocardial function, morphology, edema, and infarct mass after ST-segment elevation myocardial infarction evaluated by serial magnetic resonance imaging. Am Heart J. 2007;154:929–936. [PubMed]
49. Dall’armellina E, Karia N, Lindsay AC, Karamitsos TD, Ferreira V, Robson MD, Kellman P, Francis JM, Forfar C, Prendergast B, Banning AP, Channon KM, Kharbanda RK, Neubauer S, Choudhury RP. Dynamic Changes of Edema and Late Gadolinium Enhancement after Acute Myocardial Infarction and Their Relationship to Functional Recovery and Salvage Index. Circ Cardiovasc Imaging. 2011;4:228–236. [PMC free article] [PubMed]
50. Payne AR, Casey M, McClure J, McGeoch R, Murphy A, Woodward R, Saul A, Bi X, Zuehlsdorff S, Oldroyd KG, Tzemos N, Berry C. Bright Blood T2 Weighted MRI Has Higher Diagnostic Accuracy Than Dark Blood STIR MRI for Detection of Acute Myocardial Infarction and for Assessment of the Ischemic Area-at-Risk and Myocardial Salvage. Circ Cardiovasc Imaging. 2011;4:210–219. [PubMed]
51. Schmidt A, Azevedo CF, Cheng A, Gupta SN, Bluemke DA, Foo TK, Gerstenblith G, Weiss RG, Marban E, Tomaselli GF, Lima JA, Wu KC. Infarct tissue heterogeneity by magnetic resonance imaging identifies enhanced cardiac arrhythmia susceptibility in patients with left ventricular dysfunction. Circulation. 2007;115:2006–2014. [PMC free article] [PubMed]