Ronald J Beyers

Postinfarction Myocardial Scarring in Mice: Molecular MR Imaging with Use of a Collagen-targeting Contrast Agent

PURPOSE To prospectively evaluate a gadolinium-based collagen-targeting contrast agent, EP-3533, for in vivo magnetic resonance (MR) imaging of myocardial fibrosis in a mouse model of healed myocardial infarction (MI). MATERIALS AND METHODS All procedures were performed in accordance with protocols approved by the animal care and use committee. MI was induced in eight mice by means of occlusion of the left anterior descending coronary artery followed by reperfusion. Four MR examinations were performed in each animal: one examination before, one examination 1 day after, and two examinations 6 weeks after the MI. For the latter two examinations, electrocardiographically gated inversion-recovery gradient-echo MR images were acquired before and serially (every 5 minutes) after the intravenous injection of either gadopentetate dimeglumine or EP-3533. The image enhancement kinetic properties of the postinfarction scar, normal myocardium, and blood were compared. RESULTS Dynamic T1-weighted MR imaging revealed the washout time constants for EP-3533 to be significantly longer than those for gadopentetate dimeglumine in regions of postinfarction scarring (mean, 194.8 minutes +/-116.8 [standard deviation] vs 25.5 minutes +/- 4.2; P < .05) and in normal myocardium (mean, 45.4 minutes +/- 16.7 vs 25.1 minutes +/- 9.7; P < .05). Findings on postmortem histologic sections stained for collagen correlated well with EP-3533-enhanced areas seen on inversion-recovery MR images. Fifty minutes after EP-3533 injection, the postinfarction scar tissue samples, as compared with the normal myocardium, had a twofold higher concentration of gadolinium. CONCLUSION Use of the gadolinium-based collagen-targeting contrast agent, EP-3533, enabled in vivo molecular MR imaging of fibrosis in a mouse model of healed postinfarction myocardial scarring.

T2‐weighted MRI of post‐infarct myocardial edema in mice

T2‐weighted, cardiac magnetic resonance imaging (T2w CMR) can be used to noninvasively detect and quantify the edematous region that corresponds to the area at risk (AAR) following myocardial infarction (MI). Previously, CMR has been used to examine structure and function in mice, expediting the study of genetic manipulations. To date, CMR has not been applied to imaging of post‐MI AAR in mice. We developed a whole‐heart, T2w CMR sequence to quantify the AAR in mouse models of ischemia and infarction. The ΔB0 and ΔB1 environment around the mouse heart at 7 T were measured, and a T2‐preparation sequence suitable for these conditions was developed. Both in vivo T2w and late gadolinium enhanced CMR were performed in mice after 20‐min coronary occlusions, resulting in measurements of AAR size of 32.5 ± 3.1 (mean ± SEM)% left ventricular mass, and MI size of 50.1 ± 6.4% AAR size. Excellent interobserver agreement and agreement with histology were also found. This T2w imaging method for mice may allow for future investigations of genetic manipulations and novel therapies affecting the AAR and salvaged myocardium following reperfused MI. Magn Reson Med, 2011.

Cardiac-Selective Expression of Extracellular Superoxide Dismutase After Systemic Injection of Adeno-Associated Virus 9 Protects the Heart Against Post–Myocardial Infarction Left Ventricular Remodeling

Background— Cardiac magnetic resonance imaging has not been used previously to document the attenuation of left ventricular (LV) remodeling after systemic gene delivery. We hypothesized that targeted expression of extracellular superoxide dismutase (EcSOD) via the cardiac troponin-T promoter would protect the mouse heart against both myocardial infarction (MI) and subsequent LV remodeling. Methods and Results— Using reporter genes, we first compared the specificity, time course, magnitude, and distribution of gene expression from adeno-associated virus (AAV) 1, 2, 6, 8, and 9 after intravenous injection. The troponin-T promoter restricted gene expression largely to the heart for all AAV serotypes tested. AAV1, 6, 8, and 9 provided early-onset gene expression that approached steady-state levels within 2 weeks. Gene expression was highest with AAV9, which required only 3.15×1011 viral genomes per mouse to achieve an 84% transduction rate. AAV9-mediated, cardiac-selective gene expression elevated EcSOD enzyme activity in heart by 5.6-fold (P=0.015), which helped protect the heart against both acute MI and subsequent LV remodeling. In acute MI, infarct size in EcSOD-treated mice was reduced by 40% compared with controls (P=0.035). In addition, we found that cardiac-selective expression of EcSOD increased myocardial capillary fractional area and decreased neutrophil infiltration after MI. In a separate study of LV remodeling, after a 60-minute coronary occlusion, cardiac magnetic resonance imaging revealed that LV volumes at days 7 and 28 post-MI were significantly lower in the EcSOD group compared with controls. Conclusions— Cardiac-selective expression of EcSOD from the cardiac troponin-T promoter after systemic administration of AAV9 provides significant protection against both acute MI and LV remodeling.

Adeno-associated virus serotype 9 administered systemically after reperfusion preferentially targets cardiomyocytes in the infarct border zone with pharmacodynamics suitable for the attenuation of left ventricular remodeling

Adeno‐associated virus serotype 9 (AAV9) vectors provide efficient and uniform gene expression to normal myocardium following systemic administration, with kinetics that approach steady‐state within 2–3 weeks. However, as a result of the delayed onset of gene expression, AAV vectors have not previously been administered intravenously after reperfusion for post‐infarct gene therapy applications. The present study evaluated the therapeutic potential of post‐myocardial infarction gene delivery using intravenous AAV9.

Heart Rate Reduction With Ivabradine Protects Against Left Ventricular Remodeling by Attenuating Infarct Expansion and Preserving Remote-Zone Contractile Function and Synchrony in a Mouse Model of Reperfused Myocardial Infarction.

Background Ivabradine selectively inhibits the pacemaker current of the sinoatrial node, slowing heart rate. Few studies have examined the effects of ivabradine on the mechanical properties of the heart after reperfused myocardial infarction (MI). Advances in ultrasound speckle‐tracking allow strain analyses to be performed in small‐animal models, enabling the assessment of regional mechanical function. Methods and Results After 1 hour of coronary occlusion followed by reperfusion, mice received 10 mg/kg per day of ivabradine dissolved in drinking water (n=10), or were treated as infarcted controls (n=9). Three‐dimensional high‐frequency echocardiography was performed at baseline and at days 2, 7, 14, and 28 post‐MI. Speckle‐tracking software was used to calculate intramural longitudinal myocardial strain (Ell) and strain rate. Standard deviation time to peak radial strain (SD Tpeak Err) and temporal uniformity of strain were calculated from short‐axis cines acquired in the left ventricular remote zone. Ivabradine reduced heart rate by 8% to 16% over the course of 28 days compared to controls (P<0.001). On day 28 post–MI, the ivabradine group was found to have significantly smaller end‐systolic volumes, greater ejection fraction, reduced wall thinning, and greater peak Ell and Ell rate in the remote zone, as well as globally. Temporal uniformity of strain and SD Tpeak Err were significantly smaller in the ivabradine‐treated group by day 28 (P<0.05). Conclusions High‐frequency ultrasound speckle‐tracking demonstrated decreased left ventricular remodeling and dyssynchrony, as well as improved mechanical performance in remote myocardium after heart rate reduction with ivabradine.

Whole-heart T2-weighted (T2w) sequence for imaging post-infarct edematous area at risk (AR) in mice

Introduction T2w cardiac magnetic resonance (CMR) defines the area at risk (AR) in large mammals by imaging post myocardial infarct (MI) edema. Here, we expand the T2w CMR to cover the entire left ventricle (LV) in mice and jointly applied late gadolinium enhanced (LGE) CMR to noninvasively define MI size as percent AR. Fast murine heart rate, flow and motion present challenges to T2w CMR success. The typical 60 ms echo time for edema contrast occupies a large portion of the murine cardiac cycle which prohibits traditional T2w methods.

Arterial spin labeling CMR quantifies increased perfusion in hearts of mice treated with cardioprotective, AAV9-mediated EcSOD gene therapy prior to myocardial infarction

Introduction Experimental myocardial infarction (MI), direct gene transfer with adeno-associated viral (AAV) vectors and CMR in mice are powerful tools for studying the roles of individual genes in MI and post-MI left ventricular (LV) remodeling. Arterial spin labeling (ASL) enables the quantification of myocardial perfusion (MP) by CMR, but is sensitive to variable heart rates and irregular respiration, prohibiting accurate measurement early after MI. We developed a cardio-respiratory gated (CRG) ASL method that is insensitive to these factors to measure MP in mice.

2093 CMR reveals that cardiac-specific overexpression of the inducible form of nitric oxide synthase induces Left Ventricular Hypertrophy in wild-type mice after AAV-Mediated direct gene transfer

Introduction CMR has previously shown that left ventricular remodeling (LVR) resulting from reperfused myocardial infarction (MI) is dramatically reduced in iNOS knock-out mice [1]. However, previous studies using transgenic mice with cardiac-specific overexpression of iNOS have yielded disparate results. One study reported a benign phenotype while a second reported that iNOS overexpression resulted in cardiac fibrosis, hypertrophy and dilatation. One explanation is that transgenic mice may develop compensatory mechanisms to down-regulate iNOS overexpression. Another possiblity is that the phenotype of iNOS overexpression may be variable in its penetrance due to limited availability of a necessary co-factor (tetrahydrobiopterin).