The Antibiotic Doxycycline Impairs Cardiac Mitochondrial and Contractile Function.

Tetracycline antibiotics act by inhibiting bacterial protein translation. Given the bacterial ancestry of mitochondria, we tested the hypothesis that doxycycline-which belongs to the tetracycline class-reduces mitochondrial function, and results in cardiac contractile dysfunction in cultured H9C2 cardiomyoblasts, adult rat cardiomyocytes, in Drosophila and in mice. Ampicillin and carbenicillin were used as control antibiotics since these do not interfere with mitochondrial translation. In line with its specific inhibitory effect on mitochondrial translation, doxycycline caused a mitonuclear protein imbalance in doxycycline-treated H9C2 cells, reduced maximal mitochondrial respiration, particularly with complex I substrates, and mitochondria appeared fragmented. Flux measurements using stable isotope tracers showed a shift away from OXPHOS towards glycolysis after doxycycline exposure. Cardiac contractility measurements in adult cardiomyocytes and Drosophila melanogaster hearts showed an increased diastolic calcium concentration, and a higher arrhythmicity index. Systolic and diastolic dysfunction were observed after exposure to doxycycline. Mice treated with doxycycline showed mitochondrial complex I dysfunction, reduced OXPHOS capacity and impaired diastolic function. Doxycycline exacerbated diastolic dysfunction and reduced ejection fraction in a diabetes mouse model vulnerable for metabolic derangements. We therefore conclude that doxycycline impairs mitochondrial function and causes cardiac dysfunction.

An iterative sparse deconvolution method for simultaneous multicolor 19 F-MRI of multiple contrast agents.

PURPOSE: 19 F-MRI is gaining widespread interest for cell tracking and quantification of immune and inflammatory cells in vivo. Different fluorinated compounds can be discriminated based on their characteristic MR spectra, allowing in vivo imaging of multiple 19 F compounds simultaneously, so-called multicolor 19 F-MRI. We introduce a method for multicolor 19 F-MRI using an iterative sparse deconvolution method to separate different 19 F compounds and remove chemical shift artifacts arising from multiple resonances. METHODS: The method employs cycling of the readout gradient direction to alternate the spatial orientation of the off-resonance chemical shift artifacts, which are subsequently removed by iterative sparse deconvolution. Noise robustness and separation was investigated by numerical simulations. Mixtures of fluorinated oils (PFCE and PFOB) were measured on a 7T MR scanner to identify the relation between 19 F signal intensity and compound concentration. The method was validated in a mouse model after intramuscular injection of fluorine probes, as well as after intravascular injection. RESULTS: Numerical simulations show efficient separation of 19 F compounds, even at low signal-to-noise ratio. Reliable chemical shift artifact removal and separation of PFCE and PFOB signals was achieved in phantoms and in vivo. Signal intensities correlated excellently to the relative 19 F compound concentrations (r-2 = 0.966/0.990 for PFOB/PFCE). CONCLUSIONS: The method requires minimal sequence adaptation and is therefore easily implemented on different MRI systems. Simulations, phantom experiments, and in-vivo measurements in mice showed effective separation and removal of chemical shift artifacts below noise level. We foresee applicability for simultaneous in-vivo imaging of 19 F-containing fluorine probes or for detection of 19 F-labeled cell populations.

Emerging Magnetic Resonance Imaging Techniques for Atherosclerosis Imaging.

Atherosclerosis is a prevalent disease affecting a large portion of the population at one point in their lives. There is an unmet need for noninvasive diagnostics to identify and characterize at-risk plaque phenotypes noninvasively and in vivo, to improve the stratification of patients with cardiovascular disease, and for treatment evaluation. Magnetic resonance imaging is uniquely positioned to address these diagnostic needs. However, currently available magnetic resonance imaging methods for vessel wall imaging lack sufficient discriminative and predictive power to guide the individual patient needs. To address this challenge, physicists are pushing the boundaries of magnetic resonance atherosclerosis imaging to increase image resolution, provide improved quantitative evaluation of plaque constituents, and obtain readouts of disease activity such as inflammation. Here, we review some of these important developments, with specific focus on emerging applications using high-field magnetic resonance imaging, the use of quantitative relaxation parameter mapping for improved plaque characterization, and novel 19F magnetic resonance imaging technology to image plaque inflammation.

Mesencephalic dopamine neurons interfacing the shell of nucleus accumbens and the dorsolateral striatum in the rat.

Parallel corticostriatonigral circuits have been proposed that separately process motor, cognitive, and emotional-motivational information. Functional integration requires that interactions exist between neurons participating in these circuits. This makes it imperative to study the complex anatomical substrate underlying corticostriatonigral circuits. It has previously been proposed that dopaminergic neurons in the ventral mesencephalon may play a role in this circuit interaction. Therefore, we studied in rats convergence of basal ganglia circuits by depositing an anterograde neuroanatomical tracer into the ventral striatum together with a retrograde fluorescent tracer ipsilaterally in the dorsolateral striatum. In the mesencephalon, using confocal microscopy, we looked for possible appositions of anterogradely labeled fibers and retrogradely labeled neurons, "enhancing" the latter via intracellular injection of Lucifer Yellow. Tyrosine hydroxylase (TH) immunofluorescence served to identify dopaminergic neurons. In neurophysiological experiments, we combined orthodromic stimulation in the medial ventral striatum with recording from ventral mesencephalic neurons characterized by antidromic stimulation from the dorsal striatum. We observed terminal fields of anterogradely labeled fibers that overlap populations of retrogradely labeled nigrostriatal cell bodies in the substantia nigra pars compacta and lateral ventral tegmental area (VTA), with numerous close appositions between boutons of anterogradely labeled fibers and nigrostriatal, TH-immunopositive neurons. Neurophysiological stimulation in the medial ventral striatum caused inhibition of dopaminergic nigrostriatal neurons projecting to the ventrolateral striatal territory. Responding nigrostriatal neurons were located in the medial substantia nigra and adjacent VTA. Our results strongly suggest a functional link between ventromedial, emotional-motivational striatum, and the sensorimotor dorsal striatum via dopaminergic nigrostriatal neurons.

Selective subepicardial localization of monocyte subsets in response to progressive coronary artery constriction.

Following myocardial infarction and atherosclerotic lesion development, monocytes contribute to myocardial protection and repair, while also partaking in myocardial ischemic injury. The balance of proinflammatory and reparative monocyte subsets is crucial in governing these therapeutic and pathological outcomes. Myocardial ischemic damage displays heterogeneity across the myocardium, whereby the subendocardium shows greatest vulnerability to ischemic damage. In this study we examined the transmural distribution of monocyte subsets in response to gradual coronary artery occlusion. CD14(+) monocytes were isolated from peripheral blood of New Zealand White rabbits and divided into two subgroups based on the expression of CD62L. We employed a rabbit model of progressive coronary artery obstruction to induce chronic myocardial ischemia and reinfused fluorescently labeled autologous monocytes. The distribution of fluorescently labeled autologous monocytes was examined with a high-resolution three-dimensional imaging cryomicrotome. The subepicardial layer contained the largest infiltration of both monocyte subgroups, with a significantly greater proportion of CD14(+)CD62L(+) monocytes at the time when the ischemic area was at a maximum. By targeting CD13(+) angiogenic vessels, we confirmed the presence of angiogenesis in epicardial and midmyocardial regions. These myocardial regions demonstrated the highest level of infiltration of both monocyte subsets. Furthermore, CD14(+)CD62L(+) monocytes showed significantly greater migration towards monocyte chemoattractant protein-1, greater adhesive capacity, and higher expression of C-C chemokine receptor type-2 relative to CD14(+)CD62L(-) monocytes. In conclusion, we note selective subepicardial distribution of monocyte subpopulations, with changes in proportion depending on the time after onset of coronary narrowing. Selective homing is supported by divergent migratory properties of each respective monocyte subgroup.

How to 19F MRI: applications, technique, and getting started.

Magnetic resonance imaging (MRI) plays a significant role in the routine imaging workflow, providing both anatomical and functional information. 19F MRI is an evolving imaging modality where instead of 1H, 19F nuclei are excited. As the signal from endogenous 19F in the body is negligible, exogenous 19F signals obtained by 19F radiofrequency coils are exceptionally specific. Highly fluorinated agents targeting particular biological processes (i.e., the presence of immune cells) have been visualised using 19F MRI, highlighting its potential for non-invasive and longitudinal molecular imaging. This article aims to provide both a broad overview of the various applications of 19F MRI, with cancer imaging as a focus, as well as a practical guide to 19F imaging. We will discuss the essential elements of a 19F system and address common pitfalls during acquisition. Last but not least, we will highlight future perspectives that will enhance the role of this modality. While not an exhaustive exploration of all 19F literature, we endeavour to encapsulate the broad themes of the field and introduce the world of 19F molecular imaging to newcomers. 19F MRI bridges several domains, imaging, physics, chemistry, and biology, necessitating multidisciplinary teams to be able to harness this technology effectively. As further technical developments allow for greater sensitivity, we envision that 19F MRI can help unlock insight into biological processes non-invasively and longitudinally.

Longitudinal CMR assessment of cardiac global longitudinal strain and hemodynamic forces in a mouse model of heart failure.

To longitudinally assess left ventricle (LV) global longitudinal strain (GLS) and hemodynamic forces during the early stages of cardiac dysfunction in a mouse model of heart failure with preserved ejection fraction (HFpEF). Cardiac MRI measurements were performed in control mice (n = 6), and db/db mice (n = 7), whereby animals were scanned four times between the age of 11-15 weeks. After the first scan, the db/db animals received a doxycycline intervention to accelerate progression of HFpEF. Systolic function was evaluated based on a series of prospectively ECG-triggered short-axis CINE images acquired from base to apex. Cardiac GLS and hemodynamic forces values were evaluated based on high frame rate retrospectively gated 2-, 3-, and 4-chamber long-axis CINE images. Ejection fraction (EF) was not different between control and db/db animals, despite that cardiac output, as well as end systolic and end diastolic volume were significantly higher in control animals. Whereas GLS parameters were not significantly different between groups, hemodynamic force root mean square (RMS) values, as well as average hemodynamic forces and the ratio between hemodynamic forces in the inferolateral-anteroseptal and apical-basal direction were lower in db/db mice compared to controls. More importantly, hemodynamic forces parameters showed a significant interaction effect between time and group. Our results indicated that hemodynamic forces parameters were the only functional outcome measure that showed distinct temporal differences between groups. As such, changes in hemodynamic forces reflect early alterations in cardiac function which can be of added value in (pre)clinical research on HFpEF.

Quantification of Mouse Heart Left Ventricular Function, Myocardial Strain, and Hemodynamic Forces by Cardiovascular Magnetic Resonance Imaging.

Mouse models have contributed significantly to understanding genetic and physiological factors involved in healthy cardiac function, how perturbations result in pathology, and how myocardial diseases may be treated. Cardiovascular magnetic resonance imaging (CMR) has become an indispensable tool for a comprehensive in vivo assessment of cardiac anatomy and function. This protocol shows detailed measurements of mouse heart left ventricular function, myocardial strain, and hemodynamic forces using 7-Tesla CMR. First, animal preparation and positioning in the scanner are demonstrated. Survey scans are performed for planning imaging slices in various short- and long-axis views. A series of prospective ECG-triggered short-axis (SA) movies (or CINE images) are acquired covering the heart from apex to base, capturing end-systolic and end-diastolic phases. Subsequently, single-slice, retrospectively gated CINE images are acquired in a midventricular SA view, and in 2-, 3-, and 4-chamber views, to be reconstructed into high-temporal resolution CINE images using custom-built and open-source software. CINE images are subsequently analyzed using dedicated CMR image analysis software. Delineating endomyocardial and epicardial borders in SA end-systolic and end-diastolic CINE images allows for the calculation of end-systolic and end-diastolic volumes, ejection fraction, and cardiac output. The midventricular SA CINE images are delineated for all cardiac time frames to extract a detailed volume-time curve. Its time derivative allows for the calculation of the diastolic function as the ratio of the early filling and atrial contraction waves. Finally, left ventricular endocardial walls in the 2-, 3-, and 4-chamber views are delineated using feature-tracking, from which longitudinal myocardial strain parameters and left ventricular hemodynamic forces are calculated. In conclusion, this protocol provides detailed in vivo quantification of the mouse cardiac parameters, which can be used to study temporal alterations in cardiac function in various mouse models of heart disease.