Desiree Abdurrachim

Good and bad consequences of altered fatty acid metabolism in heart failure: Evidence from mouse models

The shift in substrate preference away from fatty acid oxidation (FAO) towards increased glucose utilization in heart failure has long been interpreted as an oxygen-sparing mechanism. Inhibition of FAO has therefore evolved as an accepted approach to treat heart failure. However, recent data indicate that increased reliance on glucose might be detrimental rather than beneficial for the failing heart. This review discusses new insights into metabolic adaptations in heart failure. A particular focus lies on data obtained from mouse models with modulations of cardiac FA metabolism at different levels of the FA metabolic pathway and how these differently affect cardiac function. Based on studies in which these mouse models were exposed to ischaemic and non-ischaemic heart failure, we discuss whether and when modulations in FA metabolism are protective against heart failure.

Cardiac diastolic dysfunction in high-fat diet fed mice is associated with lipotoxicity without impairment of cardiac energetics in vivo

Obesity is often associated with abnormalities in cardiac morphology and function. This study tested the hypothesis that obesity-related cardiomyopathy is caused by impaired cardiac energetics. In a mouse model of high-fat diet (HFD)-induced obesity, we applied in vivo cardiac (31)P magnetic resonance spectroscopy (MRS) and magnetic resonance imaging (MRI) to investigate cardiac energy status and function, respectively. The measurements were complemented by ex vivo determination of oxygen consumption in isolated cardiac mitochondria, the expression of proteins involved in energy metabolism, and markers of oxidative stress and calcium homeostasis. We also assessed whether HFD induced myocardial lipid accumulation using in vivo (1)H MRS, and if this was associated with apoptosis and fibrosis. Twenty weeks of HFD feeding resulted in early stage cardiomyopathy, as indicated by diastolic dysfunction and increased left ventricular mass, without any effects on systolic function. In vivo cardiac phosphocreatine-to-ATP ratio and ex vivo oxygen consumption in isolated cardiac mitochondria were not reduced after HFD feeding, suggesting that the diastolic dysfunction was not caused by impaired cardiac energetics. HFD feeding promoted mitochondrial adaptations for increased utilization of fatty acids, which was however not sufficient to prevent the accumulation of myocardial lipids and lipid intermediates. Myocardial lipid accumulation was associated with oxidative stress and fibrosis, but not apoptosis. Furthermore, HFD feeding strongly reduced the phosphorylation of phospholamban, a prominent regulator of cardiac calcium homeostasis and contractility. In conclusion, HFD-induced early stage cardiomyopathy in mice is associated with lipotoxicity-associated oxidative stress, fibrosis, and disturbed calcium homeostasis, rather than impaired cardiac energetics.

Quantitative 1H MRI, 19F MRI, and 19F MRS of cell-internalized perfluorocarbon paramagnetic nanoparticles

In vivo molecular imaging with targeted MRI contrast agents will require sensitive methods to quantify local concentrations of contrast agent, enabling not only imaging-based recognition of pathological biomarkers but also detection of changes in expression levels as a consequence of disease development, therapeutic interventions or recurrence of disease. In recent years, targeted paramagnetic perfluorocarbon emulsions have been frequently applied in this context, permitting high-resolution (1)H MRI combined with quantitative (19)F MR imaging or spectroscopy, under the assumption that the fluorine signal is not altered by the local tissue and cellular environment. In this in vitro study we have investigated the (19)F MR-based quantification potential of a paramagnetic perfluorocarbon emulsion conjugated with RGD-peptide to target the cell-internalizing α(ν)β(3)-integrin expressed on endothelial cells, using a combination of (1)H MRI, (19)F MRI and (19)F MRS. The cells took up the targeted emulsion to a greater extent than nontargeted emulsion. The targeted emulsion was internalized into large 1-7 µm diameter vesicles in the perinuclear region, whereas nontargeted emulsion ended up in 1-4 µm diameter vesicles, which were more evenly distributed in the cytoplasm. Association of the targeted emulsion with the cells resulted in different proton longitudinal relaxivity values, r(1), for targeted and control nanoparticles, prohibiting unambiguous quantification of local contrast agent concentration. Upon cellular association, the fluorine R(1) was constant with concentration, while the fluorine R(2) increased nonlinearly with concentration. Even though the fluorine relaxation rate was not constant, the (19)F MRI and (19)F MRS signals for both targeted nanoparticles and controls were linear and quantifiable as function of nanoparticle concentration.

Small animal cardiovascular MR imaging and spectroscopy

The use of MR imaging and spectroscopy for studying cardiovascular disease processes in small animals has increased tremendously over the past decade. This is the result of the remarkable advances in MR technologies and the increased availability of genetically modified mice. MR techniques provide a window on the entire timeline of cardiovascular disease development, ranging from subtle early changes in myocardial metabolism that often mark disease onset to severe myocardial dysfunction associated with end-stage heart failure. MR imaging and spectroscopy techniques play an important role in basic cardiovascular research and in cardiovascular disease diagnosis and therapy follow-up. This is due to the broad range of functional, structural and metabolic parameters that can be quantified by MR under in vivo conditions non-invasively. This review describes the spectrum of MR techniques that are employed in small animal cardiovascular disease research and how the technological challenges resulting from the small dimensions of heart and blood vessels as well as high heart and respiratory rates, particularly in mice, are tackled.

Accelerated high-frame-rate mouse heart cine-MRI using compressed sensing reconstruction

We introduce a new protocol to obtain very high‐frame‐rate cinematographic (Cine) MRI movies of the beating mouse heart within a reasonable measurement time. The method is based on a self‐gated accelerated fast low‐angle shot (FLASH) acquisition and compressed sensing reconstruction. Key to our approach is that we exploit the stochastic nature of the retrospective triggering acquisition scheme to produce an undersampled and random k–t space filling that allows for compressed sensing reconstruction and acceleration. As a standard, a self‐gated FLASH sequence with a total acquisition time of 10 min was used to produce single‐slice Cine movies of seven mouse hearts with 90 frames per cardiac cycle. Two times (2×) and three times (3×) k–t space undersampled Cine movies were produced from 2.5‐ and 1.5‐min data acquisitions, respectively. The accelerated 90‐frame Cine movies of mouse hearts were successfully reconstructed with a compressed sensing algorithm. The movies had high image quality and the undersampling artifacts were effectively removed. Left ventricular functional parameters, i.e. end‐systolic and end‐diastolic lumen surface areas and early‐to‐late filling rate ratio as a parameter to evaluate diastolic function, derived from the standard and accelerated Cine movies, were nearly identical. Copyright

High frame rate retrospectively triggered Cine MRI for assessment of murine diastolic function

To assess left ventricular (LV) diastolic function in mice with Cine MRI, a high frame rate (>60 frames per cardiac cycle) is required. For conventional electrocardiography‐triggered Cine MRI, the frame rate is inversely proportional to the pulse repetition time (TR). However, TR cannot be lowered at will to increase the frame rate because of gradient hardware, spatial resolution, and signal‐to‐noise limitations. To overcome these limitations associated with electrocardiography‐triggered Cine MRI, in this paper, we introduce a retrospectively triggered Cine MRI protocol capable of producing high‐resolution high frame rate Cine MRI of the mouse heart for addressing left ventricular diastolic function. Simulations were performed to investigate the influence of MRI sequence parameters and the k‐space filling trajectory in relation to the desired number of frames per cardiac cycle. An optimized protocol was applied in vivo and compared with electrocardiography‐triggered Cine for which a high‐frame rate could only be achieved by several interleaved acquisitions. Retrospective high frame rate Cine MRI proved superior to the interleaved electrocardiography‐triggered protocols. High spatial‐resolution Cine movies with frames rates up to 80 frames per cardiac cycle were obtained in 25 min. Analysis of left ventricular filling rate curves allowed accurate determination of early and late filling rates and revealed subtle impairments in left ventricular diastolic function of diabetic mice in comparison with nondiabetic mice. Magn Reson Med, 2013.

Diabetic db/db mice do not develop heart failure upon pressure overload:A longitudinal in vivo PET, MRI, and MRS study on cardiac metabolic, structural, and functional adaptations

Aims Heart failure is associated with altered myocardial substrate metabolism and impaired cardiac energetics. Comorbidities like diabetes may influence the metabolic adaptations during heart failure development. We quantified to what extent changes in substrate preference, lipid accumulation, and energy status predict the longitudinal development of hypertrophy and failure in the non-diabetic and the diabetic heart. Methods and results Transverse aortic constriction (TAC) was performed in non-diabetic (db/+) and diabetic (db/db) mice to induce pressure overload. Magnetic resonance imaging, 31P magnetic resonance spectroscopy (MRS), 1H MRS, and 18F-fluorodeoxyglucose-positron emission tomography (PET) were applied to measure cardiac function, energy status, lipid content, and glucose uptake, respectively. In vivo measurements were complemented with ex vivo techniques of high-resolution respirometry, proteomics, and western blotting to elucidate the underlying molecular pathways. In non-diabetic mice, TAC induced progressive cardiac hypertrophy and dysfunction, which correlated with increased protein kinase D-1 (PKD1) phosphorylation and increased glucose uptake. These changes in glucose utilization preceded a reduction in cardiac energy status. At baseline, compared with non-diabetic mice, diabetic mice showed normal cardiac function, higher lipid content and mitochondrial capacity for fatty acid oxidation, and lower PKD1 phosphorylation, glucose uptake, and energetics. Interestingly, TAC affected cardiac function only mildly in diabetic mice, which was accompanied by normalization of phosphorylated PKD1, glucose uptake, and cardiac energy status. Conclusion The cardiac metabolic adaptations in diabetic mice seem to prevent the heart from failing upon pressure overload, suggesting that restoring the balance between glucose and fatty acid utilization is beneficial for cardiac function.

In vivo proton T1 relaxation times of mouse myocardial metabolites at 9.4 T

Proton magnetic resonance spectroscopy (1H‐MRS) for quantitative in vivo assessment of mouse myocardial metabolism requires accurate acquisition timing to minimize motion artifacts and corrections for T1‐dependent partial saturation effects. In this study, mouse myocardial water and metabolite T1 relaxation time constants were quantified.

In vivo mouse myocardial (31)P MRS using three-dimensional image-selected in vivo spectroscopy (3D ISIS): technical considerations and biochemical validations.

31P MRS provides a unique non‐invasive window into myocardial energy homeostasis. Mouse models of cardiac disease are widely used in preclinical studies, but the application of 31P MRS in the in vivo mouse heart has been limited. The small‐sized, fast‐beating mouse heart imposes challenges regarding localized signal acquisition devoid of contamination with signal originating from surrounding tissues. Here, we report the implementation and validation of three‐dimensional image‐selected in vivo spectroscopy (3D ISIS) for localized 31P MRS of the in vivo mouse heart at 9.4 T. Cardiac 31P MR spectra were acquired in vivo in healthy mice (n = 9) and in transverse aortic constricted (TAC) mice (n = 8) using respiratory‐gated, cardiac‐triggered 3D ISIS. Localization and potential signal contamination were assessed with 31P MRS experiments in the anterior myocardial wall, liver, skeletal muscle and blood. For healthy hearts, results were validated against ex vivo biochemical assays. Effects of isoflurane anesthesia were assessed by measuring in vivo hemodynamics and blood gases. The myocardial energy status, assessed via the phosphocreatine (PCr) to adenosine 5′‐triphosphate (ATP) ratio, was approximately 25% lower in TAC mice compared with controls (0.76 ± 0.13 versus 1.00 ± 0.15; P < 0.01). Localization with one‐dimensional (1D) ISIS resulted in two‐fold higher PCr/ATP ratios than measured with 3D ISIS, because of the high PCr levels of chest skeletal muscle that contaminate the 1D ISIS measurements. Ex vivo determinations of the myocardial PCr/ATP ratio (0.94 ± 0.24; n = 8) confirmed the in vivo observations in control mice. Heart rate (497 ± 76 beats/min), mean arterial pressure (90 ± 3.3 mmHg) and blood oxygen saturation (96.2 ± 0.6%) during the experimental conditions of in vivo 31P MRS were within the normal physiological range. Our results show that respiratory‐gated, cardiac‐triggered 3D ISIS allows for non‐invasive assessments of in vivo mouse myocardial energy homeostasis with 31P MRS under physiological conditions. Copyright

High Fibroblast Growth Factor 23 concentrations in experimental renal failure impair calcium handling in cardiomyocytes

The overwhelming majority of patients with chronic kidney disease (CKD) die prematurely before reaching end‐stage renal disease, mainly due to cardiovascular causes, of which heart failure is the predominant clinical presentation. We hypothesized that CKD‐induced increases of plasma FGF23 impair cardiac diastolic and systolic function. To test this, mice were subjected to 5/6 nephrectomy (5/6Nx) or were injected with FGF23 for seven consecutive days. Six weeks after surgery, plasma FGF23 was higher in 5/6Nx mice compared to sham mice (720 ± 31 vs. 256 ± 3 pg/mL, respectively, P = 0.034). In cardiomyocytes isolated from both 5/6Nx and FGF23 injected animals the rise of cytosolic calcium during systole was slowed (−13% and −19%, respectively) as was the decay of cytosolic calcium during diastole (−15% and −21%, respectively) compared to controls. Furthermore, both groups had similarly decreased peak cytosolic calcium content during systole. Despite lower cytosolic calcium contents in CKD or FGF23 pretreated animals, no changes were observed in contractile parameters of cardiomyocytes between the groups. Expression of calcium handling proteins and cardiac troponin I phosphorylation were similar between groups. Blood pressure, the heart weight:tibia length ratio, α‐MHC/β‐MHC ratio and ANF mRNA expression, and systolic and diastolic function as measured by MRI did not differ between groups. In conclusion, the rapid, CKD‐induced rise in plasma FGF23 and the similar decrease in cardiomyocyte calcium transients in modeled kidney disease and following 1‐week treatment with FGF23 indicate that FGF23 partly mediates cardiomyocyte dysfunction in CKD.