Hepatoprotective and cardioprotective effects of empagliflozin in spontaneously hypertensive rats fed a high-fat diet.

A combination of liver and heart dysfunction worsens the prognosis of human survival. The aim of this study was to investigate whether empagliflozin (a sodium-glucose transporter-2 inhibitor) has beneficial effects not only on cardiac and renal function but also on hepatic function. Adult (6-month-old) male spontaneously hypertensive rats (SHR) were fed a high-fat diet (60% fat) for four months to induce hepatic steatosis and mild heart failure. For the last two months, the rats were treated with empagliflozin (empa, 10 mg.kg-1.day-1 in the drinking water). Renal function and oral glucose tolerance test were analyzed in control (n=8), high-fat diet (SHR+HF, n=10), and empagliflozin-treated (SHR+HF+empa, n=9) SHR throughout the study. Metabolic parameters and echocardiography were evaluated at the end of the experiment. High-fat diet feeding increased body weight and visceral adiposity, liver triglyceride and cholesterol concentrations, and worsened glucose tolerance. Although the high-fat diet did not affect renal function, it significantly worsened cardiac function in a subset of SHR rats. Empagliflozin reduced body weight gain but not visceral fat deposition. It also improved glucose sensitivity and several metabolic parameters (plasma insulin, uric acid, and HDL cholesterol). In the liver, empagliflozin reduced ectopic lipid accumulation, lipoperoxidation, inflammation and pro-inflammatory HETEs, while increasing anti-inflammatory EETs. In addition, empagliflozin improved cardiac function (systolic, diastolic and pumping) independent of blood pressure. The results of our study suggest that hepatoprotection plays a decisive role in the beneficial effects of empagliflozin in preventing the progression of cardiac dysfunction induced by high-fat diet feeding.

Developmental Aspects of Cardiac Adaptation to Increased Workload.

The heart is capable of extensive adaptive growth in response to the demands of the body. When the heart is confronted with an increased workload over a prolonged period, it tends to cope with the situation by increasing its muscle mass. The adaptive growth response of the cardiac muscle changes significantly during phylogenetic and ontogenetic development. Cold-blooded animals maintain the ability for cardiomyocyte proliferation even in adults. On the other hand, the extent of proliferation during ontogenetic development in warm-blooded species shows significant temporal limitations: whereas fetal and neonatal cardiac myocytes express proliferative potential (hyperplasia), after birth proliferation declines and the heart grows almost exclusively by hypertrophy. It is, therefore, understandable that the regulation of the cardiac growth response to the increased workload also differs significantly during development. The pressure overload (aortic constriction) induced in animals before the switch from hyperplastic to hypertrophic growth leads to a specific type of left ventricular hypertrophy which, in contrast with the same stimulus applied in adulthood, is characterized by hyperplasia of cardiomyocytes, capillary angiogenesis and biogenesis of collagenous structures, proportional to the growth of myocytes. These studies suggest that timing may be of crucial importance in neonatal cardiac interventions in humans: early definitive repairs of selected congenital heart disease may be more beneficial for the long-term results of surgical treatment.

Development of ventricular trabeculae affects electrical conduction in the early endothermic heart.

BACKGROUND: The ventricular trabeculae play a role, among others, in the impulse spreading in ectothermic hearts. Despite the morphological similarity with the early developing hearts of endotherms, this trabecular function in mammalian and avian embryos was poorly addressed. RESULTS: We simulated impulse propagation inside the looping ventricle and revealed delayed apical activation in the heart with inhibited trabecular growth. This finding was corroborated by direct imaging of the endocardial surface showing early activation within the trabeculae implying preferential spreading of depolarization along with them. Targeting two crucial pathways of trabecular formation (Neuregulin/ErbB and Nkx2.5), we showed that trabecular development is also essential for proper conduction patterning. Persistence of the slow isotropic conduction likely contributed to the pumping failure in the trabeculae-deficient hearts. CONCLUSIONS: Our results showed the essential role of trabeculae in intraventricular impulse spreading and conduction patterning in the early endothermic heart. Lack of trabeculae leads to the failure of conduction parameters differentiation resulting in primitive ventricular activation with consequent impact on the cardiac pumping function.

Myocardial development in crocodylians.

BACKGROUND: Recent reports confirmed the notion that there exists a rudimentary cardiac conduction system (CCS) in the crocodylian heart, and development of its ventricular part is linked to septation. We thus analyzed myocardial development with the emphasis on the CCS components and vascularization in two different crocodylian species. RESULTS: Using optical mapping and HNK-1 immunostaining, pacemaker activity was localized to the right-sided sinus venosus. The atrioventricular conduction was restricted to dorsal part of the atrioventricular canal. Within the ventricle, the impulse was propagated from base-to-apex initially by the trabeculae, later by the ventricular septum, in which strands of HNK-1 positivity were temporarily observed. Completion of ventricular septation correlated with transition of ventricular epicardial activation pattern to mature apex-to-base direction from two periapical foci. Despite a gradual thickening of the ventricular wall, no morphological differentiation of the Purkinje network was observed. Thin-walled coronary vessels with endothelium positive for QH1 obtained a smooth muscle coat after septation. Intramyocardial vessels were abundant especially in the rapidly thickening left ventricular wall. CONCLUSIONS: Most of the CCS components present in the homeiotherm hearts can be identified in the developing crocodylian heart, with a notable exception of the Purkinje network distinct from the trabeculae carneae.

Development and diseases of the coronary microvasculature and its communication with the myocardium.

We review the current understanding of formation and development of the coronary microvasculature which supplies oxygen and nutrients to the heart myocardium and removes waste. We emphasize the close relationship, mutual development, and communication between microvasculature endothelial cells and surrounding cardiomyocytes. The first part of the review is focused on formation of microvasculature during embryonic development. We summarize knowledge about establishing the heart microvasculature density based on diffusion distance. Then signaling mechanisms which are involved in forming the microvasculature are discussed. This includes details of cardiomyocyte-endothelial cell interactions involving hypoxia, VEGF, NOTCH, angiopoietin, PDGF, and other signaling factors. The microvasculature is understudied due to difficulties in its visualization. Therefore, currently available imaging methods to delineate the coronary microvasculature in development and in adults are discussed. The second part of the review is dedicated to the importance of the coronary vasculature in disease. Coronary microvasculature pathologies are present in many congenital heart diseases (CHD), especially in pulmonary atresia, and worsen outcomes. In CHDs, where the development of the myocardium is impaired, microvasculature is also affected. In adult patients coronary microvascular disease is one of the main causes of sudden cardiac death, especially in women. Coronary microvasculature pathologies affect myocardial ischemia and vice versa; myocardial pathologies such as cardiomyopathies are closely connected with coronary microvasculature dysfunction. Microvasculature inflammation also worsens the outcomes of COVID-19 disease. Our review stresses the importance of coronary microvasculature and provides an overview of its formation and signaling mechanisms and the importance of coronary vasculature pathologies in CHDs and adult diseases. This article is categorized under: Cardiovascular Diseases > Stem Cells and Development Congenital Diseases > Molecular and Cellular Physiology Cardiovascular Diseases > Molecular and Cellular Physiology.

Coating Ti6Al4V implants with nanocrystalline diamond functionalized with BMP-7 promotes extracellular matrix mineralization in vitro and faster osseointegration in vivo.

The present study investigates the effect of an oxidized nanocrystalline diamond (O-NCD) coating functionalized with bone morphogenetic protein 7 (BMP-7) on human osteoblast maturation and extracellular matrix mineralization in vitro and on new bone formation in vivo. The chemical structure and the morphology of the NCD coating and the adhesion, thickness and morphology of the superimposed BMP-7 layer have also been assessed. The material analysis proved synthesis of a conformal diamond coating with a fine nanostructured morphology on the Ti6Al4V samples. The homogeneous nanostructured layer of BMP-7 on the NCD coating created by a physisorption method was confirmed by AFM. The osteogenic maturation of hFOB 1.19 cells in vitro was only slightly enhanced by the O-NCD coating alone without any increase in the mineralization of the matrix. Functionalization of the coating with BMP-7 resulted in more pronounced cell osteogenic maturation and increased extracellular matrix mineralization. Similar results were obtained in vivo from micro-CT and histological analyses of rabbit distal femurs with screws implanted for 4 or 12 weeks. While the O-NCD-coated implants alone promoted greater thickness of newly-formed bone in direct contact with the implant surface than the bare material, a further increase was induced by BMP-7. It can be therefore concluded that O-NCD coating functionalized with BMP-7 is a promising surface modification of metallic bone implants in order to improve their osseointegration.

The Tale-Tell Heart: Evolutionary tetrapod shift from aquatic to terrestrial life-style reflected in heart changes in axolotl (Ambystoma mexicanum).

BACKGROUND: During amphibian metamorphosis, the crucial moment lies in the rearrangement of the heart, reflecting the changes in circulatory demands. However, little is known about the exact shifts linked with this rearrangement. Here, we demonstrate such myocardial changes in axolotl (Ambystoma mexicanum) from the morphological and physiological point of view. RESULTS: Micro-CT and histological analysis showed changes in ventricular trabeculae organization, completion of the atrial septum and its connection to the atrioventricular valve. Based on Myosin Heavy Chain and Smooth Muscle Actin expression we distinguished metamorphosis-induced changes in myocardial differentiation at the ventricular trabeculae and atrioventricular canal. Using optical mapping, faster speed of conduction through the atrioventricular canal was demonstrated in metamorphic animals. No differences between the groups were observed in the heart rates, ventricular activation times, and activation patterns. CONCLUSIONS: Transition from aquatic to terrestrial life-style is reflected in the heart morphology and function. Rebuilding of the axolotl heart during metamorphosis was connected with reorganization of ventricular trabeculae, completion of the atrial septum and its connection to the atrioventricular valve, and acceleration of AV conduction.

Excess ischemic tachyarrhythmias trigger protection against myocardial infarction in hypertensive rats.

Increased level of C-reactive protein (CRP) is a risk factor for cardiovascular diseases, including myocardial infarction and hypertension. Here, we analyzed the effects of CRP overexpression on cardiac susceptibility to ischemia/reperfusion (I/R) injury in adult spontaneously hypertensive rats (SHR) expressing human CRP transgene (SHR-CRP). Using an in vivo model of coronary artery occlusion, we found that transgenic expression of CRP predisposed SHR-CRP to repeated and prolonged ventricular tachyarrhythmias. Excessive ischemic arrhythmias in SHR-CRP led to a significant reduction in infarct size (IS) compared with SHR. The proarrhythmic phenotype in SHR-CRP was associated with altered heart and plasma eicosanoids, myocardial composition of fatty acids (FAs) in phospholipids, and autonomic nervous system imbalance before ischemia. To explain unexpected IS-limiting effect in SHR-CRP, we performed metabolomic analysis of plasma before and after ischemia. We also determined cardiac ischemic tolerance in hearts subjected to remote ischemic perconditioning (RIPer) and in hearts ex vivo. Acute ischemia in SHR-CRP markedly increased plasma levels of multiple potent cardioprotective molecules that could reduce IS at reperfusion. RIPer provided IS-limiting effect in SHR that was comparable with myocardial infarction observed in naïve SHR-CRP. In hearts ex vivo, IS did not differ between the strains, suggesting that extra-cardiac factors play a crucial role in protection. Our study shows that transgenic expression of human CRP predisposes SHR-CRP to excess ischemic ventricular tachyarrhythmias associated with a drop of pump function that triggers myocardial salvage against lethal I/R injury likely mediated by protective substances released to blood from hypoxic organs and tissue at reperfusion.

Tissue clearing and imaging methods for cardiovascular development.

Tissue imaging in 3D using visible light is limited and various clearing techniques were developed to increase imaging depth, but none provides universal solution for all tissues at all developmental stages. In this review, we focus on different tissue clearing methods for 3D imaging of heart and vasculature, based on chemical composition (solvent-based, simple immersion, hyperhydration, and hydrogel embedding techniques). We discuss in detail compatibility of various tissue clearing techniques with visualization methods: fluorescence preservation, immunohistochemistry, nuclear staining, and fluorescent dyes vascular perfusion. We also discuss myocardium visualization using autofluorescence, tissue shrinking, and expansion. Then we overview imaging methods used to study cardiovascular system and live imaging. We discuss heart and vessels segmentation methods and image analysis. The review covers the whole process of cardiovascular system 3D imaging, starting from tissue clearing and its compatibility with various visualization methods to the types of imaging methods and resulting image analysis.

Gap Junctional Communication via Connexin43 between Purkinje Fibers and Working Myocytes Explains the Epicardial Activation Pattern in the Postnatal Mouse Left Ventricle.

The mammalian ventricular myocardium forms a functional syncytium due to flow of electrical current mediated in part by gap junctions localized within intercalated disks. The connexin (Cx) subunit of gap junctions have direct and indirect roles in conduction of electrical impulse from the cardiac pacemaker via the cardiac conduction system (CCS) to working myocytes. Cx43 is the dominant isoform in these channels. We have studied the distribution of Cx43 junctions between the CCS and working myocytes in a transgenic mouse model, which had the His-Purkinje portion of the CCS labeled with green fluorescence protein. The highest number of such connections was found in a region about one-third of ventricular length above the apex, and it correlated with the peak proportion of Purkinje fibers (PFs) to the ventricular myocardium. At this location, on the septal surface of the left ventricle, the insulated left bundle branch split into the uninsulated network of PFs that continued to the free wall anteriorly and posteriorly. The second peak of PF abundance was present in the ventricular apex. Epicardial activation maps correspondingly placed the site of the first activation in the apical region, while some hearts presented more highly located breakthrough sites. Taken together, these results increase our understanding of the physiological pattern of ventricular activation and its morphological underpinning through detailed CCS anatomy and distribution of its gap junctional coupling to the working myocardium.

The formation of the atrioventricular conduction axis is linked in development to ventricular septation.

During development, the ventricles of mammals and birds acquire a specialized pattern of electrical activation with the formation of the atrioventricular conduction system (AVCS), which coincides with the completion of ventricular septation. We investigated whether AVCS formation coincides with ventricular septation in developing Siamese crocodiles (Crocodylus siamensis). Comparisons were made with Amazon toadhead turtle (Mesoclemmys heliostemma) with a partial septum only and no AVCS (negative control) and with chicken (Gallus gallus) (septum and AVCS, positive control). Optical mapping of the electrical impulse in the crocodile and chicken showed a similar developmental specialization that coincided with full ventricular septation, whereas in the turtle the ventricular activation remained primitive. Co-localization of neural marker human natural killer-1 (HNK-1) and cardiomyocyte marker anti-myosin heavy chain (MF20) identified the AVCS on top of the ventricular septum in the crocodile and chicken only. AVCS formation is correlated with ventricular septation in both evolution and development.

HIF-1α is required for development of the sympathetic nervous system.

The molecular mechanisms regulating sympathetic innervation of the heart during embryogenesis and its importance for cardiac development and function remain to be fully elucidated. We generated mice in which conditional knockout (CKO) of the Hif1a gene encoding the transcription factor hypoxia-inducible factor 1α (HIF-1α) is mediated by an Islet1-Cre transgene expressed in the cardiac outflow tract, right ventricle and atrium, pharyngeal mesoderm, peripheral neurons, and hindlimbs. These Hif1aCKO mice demonstrate significantly decreased perinatal survival and impaired left ventricular function. The absence of HIF-1α impaired the survival and proliferation of preganglionic and postganglionic neurons of the sympathetic system, respectively. These defects resulted in hypoplasia of the sympathetic ganglion chain and decreased sympathetic innervation of the Hif1aCKO heart, which was associated with decreased cardiac contractility. The number of chromaffin cells in the adrenal medulla was also decreased, indicating a broad dependence on HIF-1α for development of the sympathetic nervous system.

Epoxyeicosatrienoic acid analog EET-B attenuates post-myocardial infarction remodeling in spontaneously hypertensive rats.

Epoxyeicosatrienoic acids (EETs) and their synthetic analogs have cardiovascular protective effects. Here, we investigated the action of a novel EET analog EET-B on the progression of post-myocardial infarction (MI) heart failure in spontaneously hypertensive rats (SHR). Adult male SHR were divided into vehicle- and EET-B (10 mg/kg/day; p.o., 9 weeks)-treated groups. After 2 weeks of treatment, rats were subjected to 30-min left coronary artery occlusion or sham operation. Systolic blood pressure (SBP) and echocardiography (ECHO) measurements were performed at the beginning of study, 4 days before, and 7 weeks after MI. At the end of the study, tissue samples were collected for histological and biochemical analyses. We demonstrated that EET-B treatment did not affect blood pressure and cardiac parameters in SHR prior to MI. Fractional shortening (FS) was decreased to 18.4 ± 1.0% in vehicle-treated MI rats compared with corresponding sham (30.6 ± 1.0%) 7 weeks following MI induction. In infarcted SHR hearts, EET-B treatment improved FS (23.7 ± 0.7%), markedly increased heme oxygenase-1 (HO-1) immunopositivity in cardiomyocytes and reduced cardiac inflammation and fibrosis (by 13 and 19%, respectively). In conclusion, these findings suggest that EET analog EET-B has beneficial therapeutic actions to reduce cardiac remodeling in SHR subjected to MI.

Haloperidol affects coupling between QT and RR intervals in guinea pig isolated heart.

Prolonged QT interval is an independent risk factor for development of ventricular arrhythmias. Haloperidol is one of the drugs inducing QT prolongation. Previous studies showed that haloperidol affects not only QT duration but also heart rate (RR interval). The present work focused on relationship between QT and RR and its changes under acute and chronic haloperidol administration. The study included 14 male guinea pigs divided into control and haloperidol-treated group. After 21-days administration of haloperidol or vehiculum, electrograms in isolated hearts were recorded. QT/RR and dQT/dRR coupling were calculated. Chronic haloperidol administration significantly decreases the coupling between QT and RR. Acute haloperidol exposure significantly decreases the dQT/dRR coupling in both treated and untreated guinea pig hearts. Flatter QT/RR relationship reveals a lack of QT adaptation to increased heart rate. It should be emphasized that in such situation ECG recording will not show significant QT prolongation evaluated according to clinical rules. However, if QT interval does not adapt to increased heart rate sufficiently, the risk of ventricular arrhythmias may be increased despite practically normal QT interval length. The results are supported by findings in biochemical analyses, which proved eligibility of the used model.

Novel approaches to study coronary vasculature development in mice.

BACKGROUND: Coronary artery development is an intensely studied field. Mice are a popular genetic model for developmental studies, but there is no widely accepted protocol for high-throughput, high-resolution imaging of their developmental and adult coronary artery anatomy. RESULTS: Using tissue-clearing protocols and confocal microscopy, we have analyzed embryonic and juvenile mouse hearts in Cx40:GFP knock-in models with a special focus on septal artery development. We found that the septal artery, which supplies the interventricular septum, was initially formed as an arterial plexus that connected to both the left and right coronary arteries. During development, the plexus was remodeled into a single tube, which then remained connected only to the right coronary artery. Since optical imaging became limited at postnatal stages, it was supplemented with injection techniques using India ink and Microfil; the latter was subsequently analyzed with micro-CT to visualize the anatomy of coronary vessels in 3D. CONCLUSIONS: The techniques described here enable us to study the finer details of coronary artery development in mice and can, therefore, be implemented to study the pathogenesis of coronary malformations in various mouse models. Developmental Dynamics 247:1018-1027, 2018. © 2018 Wiley Periodicals, Inc.

Trabecular Architecture Determines Impulse Propagation Through the Early Embryonic Mouse Heart.

Most embryonic ventricular cardiomyocytes are quite uniform, in contrast to the adult heart, where the specialized ventricular conduction system is molecularly and functionally distinct from the working myocardium. We thus hypothesized that the preferential conduction pathway within the embryonic ventricle could be dictated by trabecular geometry. Mouse embryonic hearts of the Nkx2.5:eGFP strain between ED9.5 and ED14.5 were cleared and imaged whole mount by confocal microscopy, and reconstructed in 3D at 3.4 µm isotropic voxel size. The local orientation of the trabeculae, responsible for the anisotropic spreading of the signal, was characterized using spatially homogenized tensors (3 × 3 matrices) calculated from the trabecular skeleton. Activation maps were simulated assuming constant speed of spreading along the trabeculae. The results were compared with experimentally obtained epicardial activation maps generated by optical mapping with a voltage-sensitive dye. Simulated impulse propagation starting from the top of interventricular septum revealed the first epicardial breakthrough at the interventricular grove, similar to experimentally obtained activation maps. Likewise, ectopic activation from the left ventricular base perpendicular to dominant trabecular orientation resulted in isotropic and slower impulse spreading on the ventricular surface in both simulated and experimental conditions. We conclude that in the embryonic pre-septation heart, the geometry of the A-V connections and trabecular network is sufficient to explain impulse propagation and ventricular activation patterns.

Effect of increased left ventricle mass on ischemia assessment in electrocardiographic signals: rabbit isolated heart study.

BACKGROUND: Detailed quantitative analysis of the effect of left ventricle (LV) hypertrophy on myocardial ischemia manifestation in ECG is still missing. The associations between both phenomena can be studied in animal models. In this study, rabbit isolated hearts with spontaneously increased LV mass were used to evaluate the effect of such LV alteration on ischemia detection criteria and performance. METHODS: Electrophysiological effects of increased LV mass were evaluated on sixteen New Zealand rabbit isolated hearts under non-ischemic and ischemic conditions by analysis of various electrogram (EG) parameters. To reveal hearts with increased LV mass, LV weight/heart weight ratio was proposed. Standard paired and unpaired statistical tests and receiver operating characteristics analysis were used to compare data derived from different groups of animals, monitor EG parameters during global ischemia and evaluate their ability to discriminate between unchanged and increased LV as well as non-ischemic and ischemic state. RESULTS: Successful evaluation of both increased LV mass and ischemia is lead-dependent. Particularly, maximal deviation of QRS and area under QRS associated with anterolateral heart wall respond significantly to even early phase (the 1st-3rd min) of ischemia. Besides ischemia, these parameters reflect increased LV mass as well (with sensitivity reaching approx. 80%). However, the sensitivity of the parameters to both phenomena may lead to misinterpretations, when inappropriate criteria for ischemia detection are selected. Particularly, use of cut-off-based criteria defined from control group for ischemia detection in hearts with increased LV mass may result in dramatic reduction (approx. 15%) of detection specificity due to increased number of false positives. Nevertheless, criteria adjusted to particular experimental group allow achieving ischemia detection sensitivity of 89-100% and specificity of 94-100%, respectively. CONCLUSIONS: It was shown that response of the heart to myocardial ischemia can be successfully evaluated only when taking into account heart-related factors (such as LV mass) and other methodological aspects (such as recording electrodes position, selected EG parameters, cut-off criteria, etc.). Results of this study might be helpful for developing new clinical diagnostic strategies in order to improve myocardial ischemia detection in patients with LV hypertrophy.

Calcium versus strontium handling by the heart muscle.

Calcium plays a crucial role in numerous processes in living systems, from both intracellular and intercellular signalling to blood clotting. Calcium can be replaced by strontium in various intracellular processes due to high level of their similarity and strontium thus may serve as a valuable tool for different experimental studies. On the other hand, strontium is also used in clinical medicine and is commonly taken to the human body with food and water. The negative cardiac side effects of strontium therapy of osteoporosis and bone metastases are well known, but still not fully explained. This fact explains enhanced interest in this element and its impact on human body. This article reviews effects of calcium and strontium on several biochemical and physiological processes, with special emphasis on cardiac muscle.

Moderate hypothermia during ex vivo machine perfusion promotes recovery of hearts donated after cardiocirculatory death†.

OBJECTIVES: To establish the optimal machine perfusion temperature for recovery of hearts in a rodent model of donation after declaration of cardiocirculatory death (DCD). METHODS: Hearts from male Lewis rats (n = 14/group) were subjected to 25 min of in situ warm (37°C) ischaemia to simulate DCD. They were then explanted and reperfused with diluted autologous blood for 60 min at 20, 25, 30, 33 or 37°C, after which they were stored at 0-4°C in Custodiol preservation solution for 240 min. Fresh-excised and cold-stored ischaemic hearts were used as controls. The viability of the different groups was assessed by comparing heart rate and left ventricular contractility in a Langendorff circuit, as well as perfusate levels of troponin-t and creatine kinase (CK), and myocardial levels of adenosine triphosphate (ATP) and reduced glutathione. RESULTS: During ex vivo reperfusion, hearts in all groups resumed beating within minutes. The mean heart rate was highest in the 37°C group at 154.72 ± 33.01 beats × min(-1) (bpm), and declined in proportion to temperature to 39.72 ± 5.53 bpm at 20°C. Troponin-t levels were highest in the 37°C group (79.49 ± 20.79 µg/l), the values were significantly lower in all other reconditioned groups with a minimum of 12.472 ± 7.08 µg/l in the 20°C group (P < 0.0001). Tissue ATP levels ranged from 4.32 ± 1.71 µmol/g at 33°C to 4.59 ± 1.41 µmol/g at 30°C, all significantly higher than the mean ATP level of 1.41 ± 0.93 µmol/g in untreated ischaemic hearts (P < 0.0001). During Langendorff assessment, the mean heart rate and contractility of all groups were higher than those of cold-stored ischaemic hearts (P < 0.0001), yet not significantly different from those of fresh controls. The perfusate levels of troponin-t and CK, and myocardial levels of reduced-glutathione and ATP were not significantly different between groups. CONCLUSION: Our results suggest that mild hypothermia during ex vivo reperfusion improves recovery of ischaemic hearts in a rodent DCD model.

Mechanistic Role of MicroRNAs in Coupling Lipid Metabolism and Atherosclerosis.

MicroRNAs (miRNAs, miRs) represent a group of powerful and versatile posttranscriptional regulators of gene expression being involved in the fine control of a plethora of physiological and pathological processes. Besides their well-established crucial roles in the regulation of cell cycle, embryogenesis or tumorigenesis, these tiny molecules have also been shown to participate in the regulation of lipid metabolism. In particular, miRs orchestrate cholesterol and fatty acids synthesis, transport, and degradation and low-density and high-density lipoprotein (LDL and HDL) formation. It is thus not surprising that they have also been reported to affect the development and progression of several lipid metabolism-related disorders including liver steatosis and atherosclerosis. Mounting evidence suggests that miRs might represent important "posttranscriptional hubs" of lipid metabolism, which means that one miR usually targets 3'-untranslated regions of various mRNAs that are involved in different steps of one precise metabolic/signaling pathway, e.g., one miR targets mRNAs of enzymes important for cholesterol synthesis, degradation, and transport. Therefore, changes in the levels of one key miR affect various steps of one pathway, which is thereby promoted or inhibited. This makes miRs potent future diagnostic and even therapeutic tools for personalized medicine. Within this chapter, the most prominent microRNAs involved in lipid metabolism, e.g., miR-27a/b, miR-33/33*, miR-122, miR-144, or miR-223, and their intracellular and extracellular functions will be extensively discussed, in particular focusing on their mechanistic role in the pathophysiology of atherosclerosis. Special emphasis will be given on miR-122, the first microRNA currently in clinical trials for the treatment of hepatitis C and on miR-223, the most abundant miR in lipoprotein particles.