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Logo of nihpaAbout Author manuscriptsSubmit a manuscriptNIH Public Access; Author Manuscript; Accepted for publication in peer reviewed journal;
Circ Cardiovasc Imaging. Author manuscript; available in PMC Nov 25, 2009.
Published in final edited form as:
PMCID: PMC2782621
NIHMSID: NIHMS143290

Molecular MRI Detects Low levels of Cardiomyocyte Apoptosis in a Transgenic Model of Chronic Heart Failure

Abstract

Background

The ability to image cardiomyocyte (CM) apoptosis in heart failure could facilitate more accurate diagnostics and optimize targeted therapeutics. We thus aimed to develop a platform to image CM apoptosis quantitatively and specifically in heart failure in-vivo. The myocardium in heart failure, however, is characterized by very low levels of CM apoptosis and normal vascular permeability, factors thought to preclude the use of molecular MRI.

Methods and Results

Female mice with over-expression of Gaq were studied. Two weeks postpartum these mice develop a cardiomyopathy characterized by low levels of CM apoptosis, and minimal myocardial necrosis or inflammation. The mice were injected with the annexin-labeled nanoparticle (AnxCLIO-Cy5.5) or a control probe (CLIO-Cy5.5) and imaged in-vivo at 9.4 Tesla. Uptake of AnxCLIO-Cy5.5 occurred in isolated clusters, frequently in the subendocardium. Myocardial T2* was significantly lower (7.6 +/− 1.5 versus 16.8 +/− 2.7 ms, p < 0.05) in the mice injected with AnxCLIO-Cy5.5 versus CLIO-Cy5.5, consistent with the uptake of AnxCLIO-Cy5.5 by apoptotic CMs. A strong correlation (r2 = 0.86, p < 0.05) was seen between in-vivo T2* (AnxCLIO-Cy5.5 uptake) and myocardial caspase-3 activity.

Conclusions

The ability of molecular MRI to image sparsely expressed targets in the myocardium is demonstrated in this study. Moreover, a novel platform for high-resolution and specific imaging of CM apoptosis in heart failure is established. In addition to providing novel insights into the pathogenesis of CM apoptosis, the developed platform could facilitate the development of novel anti-apoptotic therapies in heart failure.

Keywords: Apoptosis, Heart Failure, MRI, Molecular Imaging, Cardiomyocyte

Introduction

Cardiomyocyte (CM) apoptosis plays an important role in the development and progression of heart failure,1, 2 and molecular imaging of this process could thus facilitate the development of novel cardioprotective therapies. Molecular imaging of apoptosis is most frequently performed with annexin-labeled imaging agents, which detect phosphatidylserine on the apoptotic cell membrane.3, 4 In a series of breakthrough cardiovascular studies technetium-labeled annexin was used to image cell death in-vivo in acute ischemia and transplant rejection.5, 6 More recently, a magnetofluorescent annexin construct, AnxCLIO-Cy5.5, has been developed and used to image CM apoptosis in-vivo in a mouse model of ischemia reperfusion.7

The level of CM apoptosis in chronic heart failure, however, is substantially lower than that seen in acute conditions such as ischemia and transplant rejection.1, 2, 8, 9 In addition, unlike acutely injured or inflamed tissues, the capillary membrane in chronic heart failure does not become hyperpermeable, potentially reducing the amount of the imaging agent that can be delivered to the interstitial space and the apoptotic CMs. These challenges are particularly relevant to molecular MRI, which involves the use of larger agents than nuclear imaging,10 and has a significantly lower sensitivity. The use of molecular MRI to image CM apoptosis, however, is particularly compelling given the unparalleled ability of MRI to image myocardial structure, function and viability.10

The primary aim of this study was to determine whether molecular MRI could be used to image low levels of CM apoptosis in a mouse model of chronic heart failure. Postpartum mice with 5-fold overexpression of the Gaq transgene were imaged with the apoptosis-sensing nanoparticle AnxCLIO-Cy5.5. These Gaq overexpressing mice develop a well-described postpartum cardiomyopathy characterized by low levels of CM apoptosis (1-2%) in its chronic phase, minimal myocardial inflammation and necrosis, and normal capillary permeability.11, 12 We demonstrate in the study that in-vivo molecular MRI of low levels of CM apoptosis in heart failure is feasible. We show, moreover, that in-vivo uptake of AnxCLIO-Cy5.5 correlates strongly with myocardial caspase-3 activity, demonstrating the sensitivity and specificity of AnxCLIO-Cy5.5 for a sparse population of purely apoptotic CMs. A new platform and readout for basic and translational research of CM apoptosis in heart failure is thus established.

Methods

Generation of the Model

Heterozygous FVB/N mice with 5-fold overexpression of the Gaq transgene were kindly provided by Dr. Gerald Dorn.11, 12 Genotypic characterization of the female pups was performed with a real time quantitative PCR system (QPCR), after purifying genomic DNA from the tail. Male mice not needed to maintain the line were euthanized at birth. Heterozygous female pups were housed until 3 months of age, at which time they were mated with wildtype males. Postpartum females were identified on the day of delivery and imaged 10-14 days after delivery. While higher levels of CM apoptosis have been documented in the early postpartum period (days 1-4),13, 14 by 10-14 days postpartum apoptosis is seen in only 1-2% of the CMs in this model.11, 12 16 postpartum mice were imaged in two phases: In the initial phase, ex-vivo fluorescence reflectance imaging was performed in 6 postpartum Gaq mice to demonstrate feasibility and proof-of-principle. In the second phase in-vivo molecular MRI, ex-vivo MRI and FRI were performed in 10 postpartum Gaq mice, and the imaging data were correlated with myocardial caspase-3 activity and levels of cleaved PARP-1.

Phase 1: Ex-Vivo Fluorescence Reflectance Imaging

Postpartum Gaq mice were injected (tail vein) with 3mg Fe/kg of AnxCLIO-Cy5.5 (n = 3) or the unlabeled control probe CLIO-Cy5.5 (n = 3). The properties of AnxCLIO-Cy5 have been previously described,15 although it should be noted that the transverse relaxivity of the current agent is > 80 mM−1s−1. AnxCLIO-Cy5.5 is < 50 nm in size and has a biological activity similar to that of unmodified annexin.15 The superparamagnetic cross linked iron-oxide (CLIO) moiety on the probe provides an MRI readout, while the near infrared fluorochrome Cy5.5 allows fluorescence imaging and microscopy of the agent to be performed.

Fluorescence reflectance imaging (FRI) was performed 48 hours after injection of the imaging agent, providing a low-resolution but cheap and rapid (5 minutes per heart) screen of the pharmacokinetics and target organ uptake of AnxCLIO-Cy5.5. The blood half life of AnxCLIO-Cy5.5 in mice is approximately 3 hours, while that of CLIO-Cy5.5 is approximately 10 hours.16 An imaging time point 48 hours after injection was chosen to ensure that both agents, particularly CLIO-Cy5.5, had been completely cleared from the blood pool. Selection of this time point ensured that any differences in myocardial signal between the two probes would reflect the ability of AnxCLIO-Cy5.5 to cross the capillary membrane, enter the interstitial space and bind to apoptotic CMs.

Prior to FRI, the excised hearts were bisected in the short axis at the midventricular level. FRI was performed using a high performance digital CCD camera (Sensicam, Cooke Corporation, Auburn Hills MI) in reflection mode. The fluorescence images were obtained using a 672 nm diode laser, an excitation strength of 550 mA, a 60 second exposure time and a Cy5.5 filter. The fluorescence signal in both portions of each heart was averaged and mapped linearly. FRI data from phase-1 of the study were pooled with the FRI data from phase-2 (see below). The fluorescence intensity in the mice injected with AnxCLIO-Cy5.5 and CLIO-Cy5.5 was then compared using a Mann-Whitney test (Prism, Graphpad, La Jolla CA).

Phase 2: In-Vivo MRI

The 10 mice in the second phase of the study were injected with 10 mg Fe/kg of AnxCLIO-Cy5.5 (n=5) or CLIO-Cy5.5 (n=5). In-vivo MRI was performed 48 hours following probe injection on a horizontal bore 9.4 Tesla small animal MRI scanner (Biospec, Bruker, Billerica MA) with cardiorespiratory gating (SA instruments, Stonybrook, NY) and a cardiac-tailored surface coil. Gradient echo cines were acquired from the base to the apex of the heart to quantify left ventricular volumes and ejection fraction (EF). The following imaging parameters were used: FOV 30×30 mm, slice 1mm, matrix 200 × 200 (150 um resolution), flip angle 30 degrees, TE 2ms, 16 frames/RR interval, Nex 4.

End diastolic and end systolic volumes were measured offline in each slice using a freeware Dicom reader (Osirix, University of Geneva, Switzerland) and summed to calculate left ventricular end diastolic volume, end systolic volume and EF. Midsystolic T2* maps were created in the short axis of the left ventricle, at the midventricular level, from gradient echo images acquired with the parameters above and at echo times (TE) of 3.5, 5.0, 6.5 and 8 ms. T2* values were calculated using a mono-exponential decay model, as previously described.7 Susceptibility artifacts from the lungs precluded accurate interpretation of myocardial T2* values over the inferior portion of the left ventricle and these areas were thus excluded from the analysis. In-vivo T2* values in the mice injected with AnxCLIO-Cy5.5 and CLIO-Cy5.5 were compared using a Mann-Whitney test (Prism, Graphpad, La Jolla CA).

Measurement of Myocardial Caspase-3 Activity and Cleaved PARP-1

Following MRI, the mice were euthanized and the excised hearts were bisected at the midventricular level for further analysis. The basal half of the left ventricle was flash frozen and used to assess myocardial caspase-3 activity and levels of cleaved PARP-1, while the apical half was embedded for microscopy. Caspase-3 activity was measured with an assay based on the cleavage of a fluorogenic DEVD-AMC substrate. A western blot assay was performed to measure levels of cleaved PARP-1, which reflects the downstream activity of caspase-3. Recent reports, however, suggest that PARP-1 can be cleaved during autophagy as well.17 Left ventricular EF and the in-vivo uptake of AnxCLIO-Cy5.5 (T2* values) were correlated with caspase-3 activity and PARP-1 using both Pearson and Spearman correlation (Prism, Graphpad, La Jolla CA).

The Western blot assay for cleaved PARP-1 was performed with a commercially available antibody for cleaved PARP-1 (Santa Cruz Biotech, Santa Cruz, CA). The gels were then stripped with Restore buffer (Pierce, Rockford IL) and, if possible, stained again with an anti-GAPDH (glyceraldehyde-3-phosphate) antibody (Rockland Immuno-chemicals, Gilbertsville PA). If not possible then a separate gel was run for the loading controls. Blots were developed with Western Lightning chemiluminescence reagent (PerkinElmer, Waltham MA) and molecular weights were compared to bands for Precision Plus Protein WesternC standards (BioRad, Hercules CA). Densitometry was preformed with a custom macro executed in ImageJ software to treat all samples identically. The fluorogenic readout of caspase-3 activity involved diluting homogenized myocardial samples in a pH 7.4 buffer containing 100 M 7-amino-4-methylcoumarin-derived caspase-3 substrate (DEVD-AMC). An AMC standard curve was used to convert arbitrary fluorescent units into moles of product AMC generated. Fluorescence measurements (excitation 342 nm, emission 441 nm) were made with a fluorescence plate reader both before and after a 6-hour incubation with substrate. Bicinchoninic acid protein assays (Pierce, Rockford, IL) were performed to determine the total protein concentration in each heart sample and used to normalize the fluorescence signal per mg of tissue.

Ex-Vivo MRI, FRI and Histology

Ten histological sections, each 5 um thick with 10 um gaps, were obtained from the midventricular portion of each embedded heart. The remaining portion of the myocardium was allowed to thaw, rinsed in PBS, and then placed in batches of two in a hydrocarbon matching medium for ex-vivo MR microscopy. MR microscopy was performed at 9.4 Tesla using a tailored solenoid radiofrequency coil and a 3D gradient echo sequence with an isotropic spatial resolution of 65 um and a flip angle of 30 degrees. Two data sets, each taking 55 minutes, were obtained with echo times of 2.9 and 8 ms respectively. FRI of the myocardium was then performed to corroborate the MRI and prior (phase-1) FRI findings.

CM apoptosis was identified on the histological sections with a terminal uridine nick-end labeling (TUNEL) assay (Integreen, New York, NY). The sections were counterstained with blue hematoxylin and commercially available human lymph node sections provided positive controls. Fluorescence microscopy of AnxCLIO-Cy5.5 uptake was performed using an upright epifluorescence microscope (Eclipse 80i, Nikon Instruments, Melville, New York) with the following filters: excitation 650+/−22.5 nm, emission 680 nm longpass and 710+/−25 nm bandpass.

Averaged results throughout the paper are reported as mean +/− standard error of measurement (SEM). A p-value of < 0.05 was regarded as significant in the Mann-Whitney and correlation tests. All experiments were performed in accordance with the guidelines for the humane care of research animals at our institution. The authors had full access to the data and take responsibility for its integrity. All authors have read and agree to the manuscript as written.

Results

Pooled analysis of the FRI data from both phases of the study (Figure 1) showed a significantly greater fluorescence signal (7.6 +/− 0.3 versus 6.1 +/− 0.3, p < 0.05) in the mice injected with AnxCLIO-Cy5.5 (n=8) than in those injected with CLIO-Cy5.5 (n=8). Representative in-vivo MR images from phase 2 of the study are shown in Figure 2. Myocardial T2* values were significantly lower in the mice injected with AnxCLIO-Cy5.5 than in those injected with CLIO-Cy5.5 (7.6 +/− 1.5 ms versus 16.8 +/− 2.7 ms, p < 0.05). T2* weighted images at an echo time of 8 ms are also shown in Figure 2. Multiple discrete foci of signal hypointensity, consistent with probe uptake, could be seen in the mice injected with AnxCLIO-Cy5.5 but not in those injected with CLIO-Cy5.5. Uptake of the probe was irregular, patchy and frequently most prominent in the subendocardium (Figure 2).

Figure 1
FRI of mice injected with 3mg Fe/kg of (A) AnxCLIO-Cy5.5 (n=8) and (B) the control probe CLIO-Cy5.5 (n=8). A-C: Fluorescence activity was significantly higher in the mice injected with AnxCLIO-Cy5.5. FRI provided a low resolution but rapid assessment ...
Figure 2
Molecular MRI of CM apoptosis in-vivo. Panel A, In-vivo T2* map in a Gaq mouse injected with 10 mg Fe/kg of AnxCLIO-Cy5.5. Susceptibility artifacts from the lungs and the iron-oxide laden liver precluded interpretation of the T2* maps over the inferior ...

Only those mice with an EF < 60% were included in the comparison of AnxCLIO-Cy5.5 and CLIO-Cy5.5 accumulation. This ensured that the analysis of probe uptake was performed between two groups of mice with similar phenotypes. 9/10 of the Gaq mice had an EF < 60% (mean 49 +/− 2%), and were thus included in the analysis. The EF of one of the mice injected with AnxCLIO-Cy5.5, however, was fully preserved at 68% and this mouse was thus excluded from the uptake comparison of AnxCLIO-Cy5.5 and CLIO-Cy5.5 (Figure 2).

Ex-vivo MRI of the excised hearts confirmed the in-vivo findings. Multiple discrete areas of signal hypointensity, consistent with probe uptake, were seen in the mice injected with AnxCLIO-Cy5.5 but not in those injected with CLIO-Cy5.5 (Figure 3). Fluorescence microscopy of the mice injected with AnxCLIO-Cy5.5 revealed that the probe was bound to the cell membrane of apoptotic CMs, many showing gross morphological features of apoptosis such as blebbing (Figure 3). Uptake of AnxCLIO-Cy5.5 occurred in discrete isolated foci, each consisting of 1-5 apoptotic CMs. No evidence of CLIO-Cy5.5 uptake was seen by fluorescence microscopy.

Figure 3
Fluorescence microscopy and ex-vivo MRI of the apical portion of the left ventricle. A, B: T2* weighted ex-vivo MRI (spatial resolution 65 × 65 × 65 um, scale bar = 1.5 mm) in a mouse injected with AnxCLIO-Cy5.5 at echo times (TE) of (A) ...

Levels of GAPDH were similar in all mice (Figure 4). Likewise, no significant difference in myocardial caspase-3 activity was seen between the two groups of mice. However, presumably due to random variation in the model, levels of cleaved PARP-1 were significantly lower (0.49 +/− 0.08 versus 0.84 +/− 0.04, p < 0.005) in the mice injected with AnxCLIO-Cy5.5, and spanned a large range (Figure 4). In the mouse with the fully preserved EF (68%) the level of cleaved PARP-1 was barely higher than that seen in healthy wildtype mice. Caspase-3 activity in this mouse was also significantly lower than any of the other 9 mice in phase 2 of the study. TUNEL staining in all mice revealed only occasional apoptotic CMs (Figure 4), even in those mice with the highest levels of myocardial caspase-3 activity. While higher levels of apoptosis are seen in this model in the first 4 days postpartum,13, 14 similarly low levels (1-2%) of apoptosis have been previously reported in this model two weeks postpartum.11, 12

Figure 4
Molecular assays of CM apoptosis in the Gaq mice that underwent in-vivo imaging. Wt = wildtype mouse. A, Western blot for cleaved PARP-1 and (B) for the control protein GAPDH. 9/10 of the Gaq mice had an EF < 60%. The level of cleaved PARP-1 in ...

A strong correlation (Spearman r = 0.8, Pearson r = 0.93 and r2 = 0.86, p < 0.05) was seen in the mice injected with AnxCLIO-Cy5.5 between the in-vivo T2* values, reflecting the degree of probe uptake, and myocardial caspase-3 activity (Figure 5). Likewise, a strong correlation (Spearman r = 0.87, Pearson r = 0.92 and r2 = 0.85, p < 0.0005) was seen between EF and levels of cleaved PARP-1 (Figure 5). The strength of this correlation (EF and PARP-1) is also demonstrated in Figure 6, showing two mice injected with AnxCLIO-Cy5.5 with significantly different EFs and levels of cleaved PARP-1.

Figure 5
Panel A, a strong correlation (r2 = 0.86, p < 0.05) was seen in the mice injected with AnxCLIO-Cy5.5 between in-vivo T2* values (AnxCLIO-Cy5.5 uptake) and normalized myocardial caspase-3 activity. B, The correlation between EF and normalized levels ...
Figure 6
Cine MRI of postpartum Gaq mice. Images at the midventricular level are shown: Panels A, B: End diastolic and systolic images in a mouse with high levels of PARP-1 and an EF of 51%. C, D: End diastolic and systolic images in a mouse with low levels of ...

Discussion

Transgenic mouse models have shown that low but persistent levels of CM apoptosis can overwhelm the limited regenerative capacity of the myocardium and result in heart failure.1, 12 The ability to serially image CM apoptosis in heart failure could thus facilitate more accurate diagnostics and the development of targeted therapeutics.2 We demonstrate in the current study that molecular MRI with the apoptosis sensing nanoparticle, AnxCLIO-Cy5.5, can non-invasively image low levels of CM apoptosis in a transgenic model of heart failure. Moreover, we show that in-vivo quantification of AnxCLIO-Cy5.5 uptake correlates strongly with myocardial caspase-3 activity. A novel platform allowing integrated anatomical, physiological and molecular imaging of CM apoptosis in heart failure is thus established.

Molecular imaging of apoptosis has most frequently exploited the ability of annexin V or the C2 domain of synaptotagmin to bind to phosphatidylserine on the apoptotic cell membrane.3, 4, 18 Pioneering work with a technetium labeled annexin construct showed that cell death could be imaged in patients with acute ischemic syndromes and transplant rejection.5, 6 More recently, the same construct has been used to image cell death in eight patients with dilated cardiomyopathy.19 The etiology of the cardiomyopathy in these patients, however, was not defined and biopsies to exclude myocarditis could not be performed. Nevertheless, the 4 patients who showed evidence of probe uptake had a more rapid decline in their clinical status, while the patients with no probe uptake remained stable.19

Despite the success of these highly pioneering clinical studies, concerns have been raised that the use of annexin-based probes in the complex and multifaceted milieu of cardiovascular injury could reflect binding to both apoptotic and necrotic CMs,20 apoptotic macrophages,21, 22 and even non apoptotic lymphocytes.23, 24 The sensitivity and specificity of technetium labeled annexin for a population of purely apoptotic CMs has thus been difficult to determine. In contrast, the transgenic mouse model used in this study is a controlled model of pure CM apoptosis with minimal CM necrosis or inflammation.11, 12 The results of this study thus show that the binding of annexin to a population of purely apoptotic cells can generate adequate signal to be imaged non-invasively in-vivo. This finding has important implications both for the utility of annexin as an imaging marker of CM apoptosis and also, more generally, for the sensitivity of molecular MRI.

The correlation between AnxCLIO-Cy5.5 uptake and myocardial caspase-3 activity was extremely strong (Figure 5). The correlation between EF, however, was stronger with cleaved PARP-1 than with caspase-3 (Figure 5). This likely reflects a well-described phenomenon in CMs where caspase-3 activation results in the rapid expression of phosphatidylserine on the outer cell membrane,25, 26 but does not necessarily result in PARP-1 cleavage, nuclear fragmentation and cell death.27 Caspase-3 activation in these CMs leads to the translocation of phosphatidylserine on the cell membrane and the cleavage of some cytosolic and myofibrillar proteins,28 but the cell remains viable.29, 30 It has been hypothesized that these myofibrillar CMs may represent an interrupted or forme fruste of CM apoptosis.29, 30

Complete execution of the apoptotic cascade with cleavage of PARP-1 and nuclear fragmentation may represent the response of the CM to a more severe insult, in which its upregulated pro-survival signals are overwhelmed.30 This would account for the strength of the correlation seen between EF and cleaved PARP-1. It should also be noted that while levels of cleaved PARP-1 largely represent its cleavage by caspase-3, recent reports suggest that autophagy can also result in the cleavage of PARP-1.17 Cleaved PARP-1 may thus be a composite marker of both caspase-3 activity (apoptosis) and autophagy, also explaining the strength of its correlation with EF.

The small size (< 50 nm) and long circulation half-life of AnxCLIO-Cy5.5 allow it to move into the interstitial space via slow transport processes such as diffusion, in a manner analogous to which large immunoglobulins and lymphotrophic nanoparticles reach the interstitial space.31 The ability of AnxCLIO-Cy5.5 to penetrate the interstitial space of the myocardium has been previously demonstrated,7 but only in the setting of acute injury and increased vascular permeability.7 We now show that AnxCLIO-Cy5.5 is able to cross a normal capillary membrane, access the interstitial space of the myocardium and bind to a target expressed on only 1-2% of CMs.12 The high affinity of AnxCLIO-Cy5.5 for apoptotic cells,15 its high magnetic relaxivity and the stability of its signal over time allowed a detectable MR signal to be generated even though only a small fraction of the injected dose likely reached the interstitial space of the myocardium. The excellent sensitivity of molecular MRI shown in this study suggests that it could support the imaging of a variety of sparsely expressed targets in the myocardium, even in the presence of normal vascular permeability.

The uptake of AnxCLIO-Cy5.5 in the ex-vivo images (and in the portions of myocardium visualized in-vivo) was seen in all regions of the left ventricle but was frequently most predominant in the subendocardium and in the anterior and lateral walls (Figures 2, ,3).3). In addition, uptake of the probe was patchy, consistent with the presence of scattered clusters of apoptotic CMs in the myocardium (Figures 2, ,3).3). Similar patterns of CM apoptosis have been documented in humans and in animal models of heart failure: Focal uptake of technetium-labeled annexin was seen in the anterolateral wall of the left ventricle in a patient with dilated cardiomyopathy and heart failure.19 Likewise clusters and scattered groups of apoptotic CMs have been documented histologically in patients with dilated cardiomyopathy and heart failure,8, 9 with the subendocardium most frequently involved.8, 32 A strong correlation has been reported between wall stress, levels of the pro-apoptotic protein Bax and CM apoptosis, all of which were highest in the subendocardium.32 Further study will be needed to elucidate the mechanisms underlying this spatial pattern of CM apoptosis and underscores the value of molecular MRI, which allows CM apoptosis and left ventricular mechanics to be imaged with high spatial resolution in a single integrated dataset.10

The dose of iron-oxide used in this study was substantially lower than the doses used in many previous studies in mice.33, 34 Moreover, the use of magnetic nanoparticles with similar sizes and properties to CLIO but significantly higher relaxivities should allow even lower doses of iron-oxide to be used.35, 36 The design of this study did not allow the tissue elimination of bound AnxCLIO-Cy5.5 to be determined which, based on prior experience with analogous magnetic nanoparticles, could take up to 1-2 weeks.37 Nevertheless, elimination over 1-2 weeks would still allow serial imaging in a chronic condition such as heart failure to be performed at several physiologically meaningful time points.

In conclusion, a novel platform for basic and translational research of anti-apoptotic therapies in heart failure is established in this study. The postpartum Gaq overexpressing mouse recapitulates a variant of heart failure seen in humans and provides a highly pure and specific model of CM apoptosis. In addition to its implications for future research in heart failure, the current study also demonstrates the ability of molecular MRI to image sparsely expressed molecular targets in the myocardium, thus expanding the scope and potential applications of molecular MRI.

Acknowledgements

We thank Sarafima Zaltsman for her assistance in maintaining the Gaq mouse colony.

Funding: This study was supported in part by the following grants from the National Institutes of Health: DES (R01 HL093038 and K08 HL079984), LJ (R01EB004472), RW (CA92782, CA86355), AR (HL073363, HL077543, HL059521), and a Leducq Network of Research Excellence Award (AR).

Footnotes

Disclosures: RW is a consultant and shareholder in Visen Medical, Woburn MA.

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