Heart Uptake Research Articles

Pharmacokinetics and Biodistribution of 99mTc N-MPO in Healthy Human Volunteers

Tc N-MPO ([Tc N(MPO)(PNP5)]: HMPO = 2-mercaptopyridine N-oxide, and PNP5 = N-ethoxyethyl-N,N-bis[2-(bis(3-methoxypropyl)phosphino)ethyl]amine) is a new Tc radiotracer useful for myocardial perfusion imaging. This study was designed to determine its pharmacokinetics and biodistribution in healthy volunteers. Ten healthy volunteers were involved in this study. Each subject was administered approximately 925 MBq of Tc N-MPO under rest or stress conditions (n = 5 per group). Whole-body planar images were obtained at 10, 30, 60, 240, and 1440 minutes after injection. Organ uptake was quantified by region-of-interest analysis. The blood clearance and urine excretion kinetics were determined by collecting blood and urine samples at different time points. Tc N-MPO showed significant accumulation in myocardium with prolonged retention. At rest, its percentage of injected dose (%ID) uptake in the heart, lungs, and liver at 10 minutes after injection was 2.47% (0.64%), 1.84% (0.64%), and 20.88% (5.23%), respectively. The liver uptake decreased to 6.79%ID (1.60%ID) at 60 minutes after injection and 4.50%ID (1.86%ID) at 240 minutes after injection. Under stress conditions, the heart uptake was slightly increased (2.57%ID [0.21%ID]). The rapid liver clearance led to favorable heart-to-liver ratios, reaching values of 0.27%ID (0.07%ID) under rest condition and 0.28%ID (0.05%ID) under stress condition at 60 minutes after injection. Tc N-MPO demonstrates a highly favorable biodistribution in humans. The high heart uptake and the fast liver washout of Tc N-MPO will allow SPECT images of the left ventricle to be acquired as early as 10 minutes after injection.

Dynamic tracking of manganese uptake in mouse hearts by rapid multi-slice T1 mapping

Biomedical Engineering, Case Western Reserve University, Cleveland, Ohio, USA Figure 1 MSRLL pulse sequence. After 90° non-slice selective saturation pulses, 10 sequential multi-slice FLASH acquisitions separated by an interval τ were applied. Each block on the relaxation curve represents a multi-slice acquisition module. Jiang and Yu Journal of Cardiovascular Magnetic Resonance 2014, 16(Suppl 1):W35 http://www.jcmr-online.com/content/16/S1/W35

Chromatographic Separation and Utilization of Labeled 99mTc-Valsartan for Cardiac Imaging

A procedure was developed for preparing high radiochemical purity 99mTc-valsartan with yield of 98%. The complex was prepared by mixing of valsartan with SnCl2.2H2O solution and the pH of the mixture was adjusted to 8 then mixed with a freshly eluted 99mTc O4- (~400 MBq), shacked at room temperature for 30 min. The radiochemical yield and purity of the labeled product were determined individually by HPLC, paper chromatography and paper electrophoresis. Biodistribution studies were carried out in normal Albino Swiss mice at different time intervals after administration of 99mTc-valsartan. The labeled compound cleared from the systematic circulation within 2 h afteradministration, and the majority of organs showed significant decrease in the uptake of 99mTc-valsartan. The heart uptake of 99mTc-valsartan was sufficiently high for using this agent as myocardial imaging agent.

Circulating angiopoietin-like 4 links proteinuria with hypertriglyceridemia in nephrotic syndrome

The molecular link between proteinuria and hyperlipidemia in nephrotic syndrome is not known. We show in the present study that plasma angiopoietin-like 4 (Angptl4) links proteinuria with hypertriglyceridemia through two negative feedback loops. In previous studies in a rat model that mimics human minimal change disease, we observed localized secretion by podocytes of hyposialylated Angptl4, a pro-proteinuric form of the protein. But in this study we noted high serum levels of Angptl4 (presumably normosialylated based on a neutral isoelectric point) in other glomerular diseases as well. Circulating Angptl4 was secreted by extrarenal organs in response to an elevated plasma ratio of free fatty acids (FFAs) to albumin when proteinuria reached nephrotic range. In a systemic feedback loop, these circulating pools of Angptl4 reduced proteinuria by interacting with glomerular endothelial αvβ5 integrin. Blocking the Angptl4-β5 integrin interaction or global knockout of Angptl4 or β5 integrin delayed recovery from peak proteinuria in animal models. But at the same time, in a local feedback loop, the elevated extrarenal pools of Angptl4 reduced tissue FFA uptake in skeletal muscle, heart and adipose tissue, subsequently resulting in hypertriglyceridemia, by inhibiting lipoprotein lipase (LPL)-mediated hydrolysis of plasma triglycerides to FFAs. Injecting recombinant human ANGPTL4 modified at a key LPL interacting site into nephrotic Buffalo Mna and Zucker Diabetic Fatty rats reduced proteinuria through the systemic loop but, by bypassing the local loop, without increasing plasma triglyceride levels. These data show that increases in circulating Angptl4 in response to nephrotic-range proteinuria reduces the degree of this pathology, but at the cost of inducing hypertriglyceridemia, while also suggesting a possible therapy to treat these linked pathologies.

Assessment of Global Cardiac Uptake of Radiolabeled Iron Oxide Nanoparticles in Apolipoprotein-E-Deficient Mice: Implications for Imaging Cardiovascular Inflammation

Atherosclerosis is a leading cause of death in industrialized countries and is characterized by the accumulation of lipids and inflammatory cells, including macrophages, in blood vessel walls. Therefore, the ability to image macrophages could help identify plaques that are precursors of acute thrombotic events. Previous research has shown that long-circulating nanoparticles could be used to detect macrophages within atherosclerotic plaques of the aorta. By conducting this study, we investigated whether global cardiac uptake of radiolabeled nanoparticles could allow assessment of total macrophage burden in the coronary arteries. Dextran-coated iron oxide nanoparticles (IONPs) were labeled with iodine-125 via Bolton-Hunter (sulfosuccinimidyl-3-[4-hydroxyphenyl]propionate) method. IONPs were characterized by means of dynamic light scattering and transmission electronic microscopy. Biodistribution studies were performed in healthy and atherosclerotic mice. Additionally, digital autoradiography of hearts from both healthy and atherosclerotic mice was performed to assess regional and global atherosclerotic burden. The [(125)I]IONPs exhibited high radiolabel stability and long blood circulation, which eventually led to high heart uptake in apoE -/- mice when compared with healthy controls. Furthermore, digital autoradiography showed substantially enhanced emission of signals from the hearts of atherosclerotic mice, while no or minimal cardiac signals were detected in healthy mice. This preparation showed adequate physical-chemical properties for in vivo studies, such as small size (∼30 nm), good radiolabel stability, and long circulation time. There was also significant accumulation in the heart of apoE-/- mice compared with that of healthy control animals. These findings suggest that radiolabeled dextran-coated iron oxide nanoparticles may have potential to become a useful tool to detect macrophages in the atherosclerosis plaques of coronary arteries; however, these preliminary findings should be confirmed by further studies in a larger scale in various atherosclerosis models.

Abstract 9907: Cardiac Adiponectin Resistance Develops Early After MI and Contributes to Heart Failure Progression

Deficiency of adiponectin (APN), a cardioprotective adipokine, as a risk factor of cardiovascular disease is well-defined. Recent clinical studies report that plasma APN levels are increased in heart failure (HF) patients, suggesting APN resistance might be another unrecognized risk factor. However, direct evidence supporting cardiac APN resistance during HF progression is currently lacking. Adult male mice were subjected to HF by coronary artery occlusion. The biological functions of APN were determined during the course of HF development (0-6 weeks). Plasma APN levels were reduced 3 days post-MI (P<0.01), gradually recovered thereafter, and reached a level greater than control 14 days post-MI (P<0.05). Consistent with APN level reduction, cardiac AMPK phosphorylation (a readout of APN intracellular signaling) was decreased 3 days post-MI (P<0.05). However, cardiac AMPK phosphorylation remained lower than control 14 days post-MI (P<0.01), despite significantly increased plasma APN levels at this time, suggesting reduced cardiac response to endogenous APN. APN administration via osmotic pump beginning 14 days post-MI for the remaining duration of MI did not protect against MI/R injury, in contrast to numerous previous studies demonstrating the cardioprotective effects of APN in acute MI/R. At the molecular level, APN treatment significantly increased both GLUT4 membrane recruitment and 18 F-FDG uptake in the normal heart (P<0.01). However, these responses were dramatically reduced 14-days post-MI. To gain more direct evidence demonstrating impaired cardiomyocyte response to APN during HF development, left ventricular cardiomyocytes were isolated from normal or MI mice (14 days), and incubated with human recombinant APN. APN-induced AMPK phosphorylation and PKA activation were significantly reduced in CM from MI mice compared to control (P<0.01). Together, these results demonstrate that APN resistance develops 2 weeks after MI, a critical period when the transition from adaptive to pathological remodeling occurs. APN resistance renders a potent endogenous cardioprotective molecule ineffective, thus facilitating HF progression. Interventions restoring cardiac APN response may be a novel approach against post-MI HF development.

Re(I) and 99mTc(I) tricarbonyl complexes with ether-containing pyrazolyl-based chelators: Chemistry, biodistribution and metabolism

Tris(pyrazolyl)methane chelators, L1–L3, containing one or two ether groups at different positions of the azole rings, were synthesized and fully characterized. These chelators enabled the synthesis of fac-[99mTc(CO)3{HC[4-(ROCH2)pz]3}]+ (R = Me (Tc1), Et (Tc2)) and fac-[99mTc(CO)3{HC[3,5-(EtOCH2)2pz]3}]+ (Tc3) which were identified by HPLC in comparison with the rhenium counterparts. The evaluation of Tc1–Tc3 in CD-1 mice has shown that the number and/or nature of the ether groups greatly influence the biodistribution profile, pharmacokinetics and metabolic stability of these complexes. Tc1 and Tc2, bearing a unique ether substituent at the 4-position of the pyrazolyl ring, undergo metabolic transformation in vivo while Tc3 is not metabolized. The metabolization of Tc1 and Tc2 enhanced their rate of excretion but, most probably, also justify their negligible heart uptake in contrast with the high heart uptake of congener non-metabolizable complexes (99mTc-DMEOP and 99mTc-TMEOP), which have recently emerged as potential myocardial imaging agents.The attempts made to identify the metabolites of Tc1 and Tc2 have shown that the metabolization of these compounds must involve the ether functions with probable formation of carboxylic acid derivatives. A comparative study with the congener fac-[99mTc(CO)3{[4-(MeOCH2)pz](CH2)2NH(CH2)2NH2}]+ (Tc6) led us to confirm the formation of such type of metabolites. In fact, Tc6 is also metabolized in mice with formation of fac-[99mTc(CO)3{[4-(HOCH2)pz](CH2)2NH(CH2)2NH2}]+ (Tc7) and fac-[99mTc(CO)3{[4-(HOOC)pz](CH2)2NH(CH2)2NH2}]+ (Tc8), which were chemically identified by HPLC in comparison with the Re congeners (Re7 and Re8).

LMBD1 Protein Serves as a Specific Adaptor for Insulin Receptor Internalization

Energy homeostasis is crucial for maintaining normally functioning cells; disturbances in this balance often cause various diseases. The limb region 1 (LMBR1) domain containing 1 gene (lmbrd1) encodes the LMBD1 protein that possesses 9 putative transmembrane domains. LMBD1 has been suggested to be involved in the lysosome in aiding the export of cobalamin. In this study, we determined that LMBD1 plays a regulatory role in the plasma membrane. A micro-positron emission tomography analysis showed that a single-allele knock-out of lmbrd1 increased the (18)F-fluorodeoxyglucose uptake in murine hearts. In addition, the knockdown of lmbrd1 resulted in an up-regulated signaling of the insulin receptor (IR) and its downstream signaling molecule, Akt. Confocal and live total internal reflection fluorescence microscopy showed that LMBD1 co-localized and co-internalized with clathrin and the IR, but not with the transferrin receptor. The results of the mutation analysis and phenotypic rescue experiments indicate that LMBD1 interacts with adaptor protein-2 and is involved in the unique clathrin-mediated endocytosis of the IR. LMBD1 selectively interacts with the IR. The knockdown of lmbrd1 attenuated IR endocytosis, resulting in the perturbation of the IR recycling pathway and consequential enhancement of the IR signaling cascade. In summary, LMBD1 plays an imperative role in mediating and regulating the endocytosis of the IR.

Evaluation of 18F-labeled BODIPY dye as potential PET agents for myocardial perfusion imaging

Despite the great potential of positron emission tomography/computed tomography (PET/CT) in cardiovascular disease imaging, one of the major limitations is the availability of PET probes with desirable half-lives and reasonable cost. In this report, we hypothesized that lipophilic cationic BODIPY dye could be selectively accumulated in cardiac muscle, possibly for the development of novel PET myocardial perfusion imaging (MPI) probes. A (18)F-labeled BODIPY dye ([(18)F]1) was synthesized efficiently through a fluoride exchange reaction catalyzed by the Lewis acid tin chloride (SnCl₄). The compound was first evaluated by a cellular uptake assay in vitro, followed by biodistribution and microPET imaging studies in vivo. [(18)F]1 was obtained in more than 90% labeling yield, with >98% radiochemical purity. The HEK-293 cellular uptake assay showed that the preferential uptake of [(18)F]1 could be related to the cell membrane potential. The biodistribution data demonstrated high levels of [(18)F]1 accumulation in the heart. In the biodistribution study in mice, the radioactivity uptake in the heart, blood, liver and lung was 3.01 ± 0.44, 0.39 ± 0.09, 0.69 ± 0.07, 1.71 ± 0.27%ID/g, respectively, at 3h post-injection (p.i.). The heart-to-lung and heart-to-liver ratios are 1.76 ± 0.14 and 4.37 ± 0.51 at 3h p.i., respectively. Volume-of-interest analysis of the microPET images correlated well with the biodistribution studies in mice. The heart was clearly visualized in normal rats, with 0.72 ± 0.18, 0.69 ± 0.18, 0.67 ± 0.20 and 0.59 ± 0.17%ID/g uptake at 0.5, 1, 2 and 4h p.i., respectively. (18)F-labeled BODIPY dye showed good heart uptake and heart-to-blood and heart-to-lung contrast. A (18)F-labeled BODIPY dyes may represent a new category of cationic PET agents for myocardial perfusion imaging.

Adrenergic pathway activation enhances brown adipose tissue metabolism: A [18F]FDG PET/CT study in mice

Pharmacologic approaches to study brown adipocyte activation in vivo with a potential of being translational to humans are desired. The aim of this study was to examine pre- and postsynaptic targeting of adrenergic system for enhancing brown adipose tissue (BAT) metabolism quantifiable by [(18)F]fluoro-2-deoxyglucose ([(18)F]FDG) positron emission tomography (PET)/computed tomography (CT) in mice. A β₃-adrenoreceptor selective agonist (CL 316243), an adenylyl cyclase enzyme activator (forskolin) and a potent blocker of presynaptic norepinephrine transporter (atomoxetine), were injected through the tail vein of Swiss Webster mice 30 minutes before intravenous (iv) administration of [(18)F]FDG. The mice were placed on the PET/CT bed for 30 min PET acquisition followed by 10 min CT acquisition for attenuation correction and anatomical delineation of PET images. Activated interscapular (IBAT), cervical, periaortic and intercostal BAT were observed in 3-dimentional analysis of [(18)F]FDG PET images. CL 316243 increased the total [(18)F]FDG standard uptake value (SUV) of IBAT 5-fold greater compared to that in placebo-treated mice. It also increased the [(18)F]FDG SUV of white adipose tissue (2.4-fold), and muscle (2.7-fold), as compared to the control. There was no significant difference in heart, brain, spleen and liver uptakes between groups. Forskolin increased [(18)F]FDG SUV of IBAT 1.9-fold greater than that in placebo-treated mice. It also increased the [(18)F]FDG SUV of white adipose tissue (2.2-fold) and heart (5.4-fold) compared to control. There was no significant difference in muscle, brain, spleen, and liver uptakes between groups. Atomoxetine increased [(18)F]FDG SUV of IBAT 1.7-fold greater than that in placebo-treated mice. There were no significant differences in all other organs compared to placebo-treated mice except liver (1.6 fold increase). A positive correlation between SUV levels of IBAT and CT Hounsfield unit (HU) (R(2)=0.55, p<0.001) and between CT HU levels of IBAT and liver (R(2)=0.69, p<0.006) was observed. The three pharmacologic approaches reported here enhanced BAT metabolism by targeting different sites in adrenergic system as measured by [(18)F]FDG PET/CT.

Synthesis, characterization and pre-clinical evaluation of99mTc-tricarbonyl complexes as potential myocardial perfusion imaging agents

Myocardial perfusion imaging is an established Nuclear Medicine investigation. Current myocardial perfusion imaging agents sestamibi and tetrofosmin have number of drawbacks; low heart uptake coupled with uptake into the surrounding tissues leads to a poorer image quality. There is a need for continued research into designing and evaluating potentially superior myocardial imaging agents. Tri-carbonyl-technetium and rhenium complexes were prepared by combination with mono-dentate and bi-dentate ligands. Complexes were characterized by HPLC, MAS, nuclear magnetic resonance, infrared, single-crystal X-ray diffraction and partition coefficient determinations. (99m) Tc(CO)3 complexes were administered intravenously to Sprague Dawley rats, and tissue distribution studies were carried out at 15 min and 1 h p.i. Radiochemical purity was assessed as >90%. 1-10-phenanthroline, 2,2'-bipyridine and imidazole complexes gave the highest heart uptake. The percentage injected dose per gram (n = 3) at 1 h for 1-10-phenanthroline/imidazole was blood 0.21 ± 0.01, heart 1.12 ± 0.11, kidney 3.61 ± 1.13, liver 0.62 ± 0.06, lung 0.28 ± 0.12, spleen 0.24 ± 0.05, small intestine contents 1.87 ± 0.92; and for 2,2'-bipyridine /imidazole was blood 0.23 ± 0.02, heart 1.07 ± 0.18, kidney 3.31 ± 1.28, liver 0.56 ± 0.09, lung 0.14 ± 0.02, spleen 0.2 ± 0.1, small intestine content 1.05 ± 0.48. Further investigation to evaluate more complexes based on 1,10-phenanthroline, 2,2'-bipyridine and imidazole derivatives could potentially lead to agents with an increased heart uptake and faster clearance from the liver and gastrointestinal tract.

Rat Imaging and In Vivo Stability Studies using [11C]-Dimethyl-Diphenyl Ammonium, a Candidate Agent for PET-Myocardial Perfusion Imaging

PET myocardial perfusion imaging (MPI) holds several advantages over SPECT for diagnosing coronary artery disease. The short half-lives of prevailing PET-MPI agents hamper wider clinical application of PET in nuclear cardiology; prompting the development of novel PET-MPI agents. We have previously reported on the potential of radiolabeled ammonium salts, and particularly on that of [(11)C]dimethyl-diphenyl-ammonium ([(11)C]DMDPA), for cardiac PET imaging. This study was designed to improve the radiosynthesis and increase the yield of [(11)C]DMDPA, characterize more meticulously the kinetics of radioactivity distribution after its injection via micro-PET/CT studies, and further explore its potential for PET-MPI. The radiosynthetic procedure of [(11)C]DMDPA was improved with respect to the previously reported one. The kinetics of radioactivity distribution following injection of [(11)C]DMDPA were investigated in juvenile and young adult male SD rats using microPET/CT, and compared to those of [(13)N]NH3. Furthermore, the metabolic fate of [(11)C]DMDPA in vivo was examined after its injection into rats. Following a radiosynthesis time of 25-27 min, 11.9 ± 1.1 GBq of [(11)C]DMDPA was obtained, with a 43.7% ± 4.3% radiochemical yield (n = 7). Time activity curves calculated after administration of [(11)C]DMDPA indicated rapid, high and sustained radioactivity uptake in hearts of both juvenile and young adult rats, having a two-fold higher cardiac radioactivity uptake compared to [(13)N]NH3. Accordingly, at all time points after injection to both juvenile and young adult rats, image quality of the left ventricle was higher with [(11)C]DMDPA compared to [(13)N]NH3. In vivo stability studies of [(11)C]DMDPA indicate that no radioactive metabolites could be detected in plasma, liver and urine samples of rats up to 20 min after injection, suggesting that [(11)C]DMDPA is metabolically stable in vivo. This study further illustrates that [(11)C]DMDPA holds, at least in part, essential qualities required from a PET-MPI probe. Owing to the improved radiosynthetic procedure reported herein, [(11)C]DMDPA can be produced in sufficient amounts for clinical use.

O.12 Peptide-enhanced uptake and bioactivity of antisense oligonucleotides in muscle and heart of DMD and DM1 mouse models

RNA-modulation by antisense oligonucleotides (AONs) represents an interesting therapeutic approach for different neuromuscular disorders, such as Duchenne muscular dystrophy (DMD) and myotonic dystrophy type 1 (DM1). DMD is caused by out of frame mutations in the DMD gene. AON-mediated exon skipping, aimed to restore the disrupted reading frame of the dystrophin transcript, is currently in (pre-) clinical development. In DM1, DM protein kinase transcripts contain a toxic expanded (CUG) n repeat stretch. Suppression of these toxic transcripts by AONs is currently being explored as a potential molecular intervention for DM1. For both diseases, efficient body-wide delivery of the AONs to muscle tissue and heart would enhance their therapeutic effect. The lack of dystrophin in DMD muscle results in more permeable muscle fibers, which, in the mdx mouse model, was demonstrated to promote AON uptake in muscle but to a lesser extent in heart. As the muscle fiber membranes are not impaired in DM1, various delivery-enhancing complexes and ligands are being explored to obtain sufficient muscle and heart uptake of AONs. Here, we present results with a 7-amino acid linear peptide that, conjugated to 2’-O-methyl phosphorothioate AONs, seems to enhance their uptake and bioactivity in muscle and, particularly, heart, in both DMD and DM1 mouse models.

Capillary Endothelial Fatty Acid Binding Proteins 4 and 5 Play a Critical Role in Fatty Acid Uptake in Heart and Skeletal Muscle

Fatty acids (FAs) are the major substrate for energy production in the heart. Here, we hypothesize that capillary endothelial fatty acid binding protein 4 (FABP4) and FABP5 play an important role in providing sufficient FAs to the myocardium. Both FABP4/5 were abundantly expressed in capillary endothelium in the heart and skeletal muscle. The uptake of a FA analogue, 125I-15-(p-iodophenyl)-3-(R,S)-methyl pentadecanoic acid, was significantly reduced in these tissues in double-knockout (DKO) mice for FABP4/5 compared with wild-type mice. In contrast, the uptake of a glucose analogue, 18F-fluorodeoxyglucose, was remarkably increased in DKO mice. The expression of transcripts for the oxidative catabolism of FAs was reduced during fasting, whereas transcripts for the glycolytic pathway were not altered in DKO hearts. Notably, metabolome analysis revealed that phosphocreatine and ADP levels were significantly lower in DKO hearts, whereas ATP content was kept at a normal level. The protein expression levels of the glucose transporter Glut4 and the phosphorylated form of phosphofructokinase-2 were increased in DKO hearts, whereas the phosphorylation of insulin receptor-β and Akt was comparable between wild-type and DKO hearts during fasting, suggesting that a dramatic increase in glucose usage during fasting is insulin independent and is at least partly attributed to the post-transcriptional and allosteric regulation of key proteins that regulate glucose uptake and glycolysis. Capillary endothelial FABP4/5 are required for FA transport into FA-consuming tissues that include the heart. These findings identify FABP4/5 as promising targets for controlling the metabolism of energy substrates in FA-consuming organs that have muscle-type continuous capillary.

Abstract 277: Acute Cardiac Insulin Resistance During Cardiopulmonary Bypass in Mini-pigs: Role of Hexosamine Biosynthesis and Protein O-Glycosylation

Our previous study as well as others has demonstrated a strong positive correlation between cardiac insulin signaling and myocardial function in ischemic/reperfused hearts. This study was designed to determine whether cardiac insulin signaling is impaired during cardiopulmonary bypass (CPB) and the possible mechanisms involved. Twelve male mini-pigs were anesthetized and subjected to CPB for 30 min. Blood samples and left ventricle biopsies were taken at pre-bypass (control), aortic cross clamp (AXC) 5 min and AXC release 120 min. Compared with sham-operated group, both blood glucose and insulin levels went up at 120 min after AXC release. Glucose uptake in heart dropped significantly as measured by attenuated coronary arterio-venous glucose difference and reduced cardiac 18 F-fluorodeoxyglucose uptake by Positron Emission Tomography imaging. Furthermore, myocardial insulin signaling was blunted as manifested by decreased phosphorylation of IRS-1, Akt and GSK-3beta (P&lt;0.01, n=4). Interestingly, these changes were associated with a substantial increase of GFAT activity (the rate-limiting enzyme for hexosamine biosynthesis) and a significant increase in protein O-GlcNAcylation in cardiac tissues (P&lt;0.05, n=6). Moreover, pretreatment with Alloxan, an inhibitor of O-GlcNAc-transferase, blocked both the impaired GSK-3beta phosphorylation and the increase in O-GlcNAcylation. Enhancement of cardiac insulin signaling with glucose-insulin-potassium before AXC decreased protein O-GlcNAcylation and ameliorated myocardial dysfunction after CPB. These data indicate the existence of acute myocardial insulin resistance during CPB which is attributable to elevated hexosamine biosynthesis and protein O-glycosylation.

Whole-body pharmacokinetics of HDAC inhibitor drugs, butyric acid, valproic acid and 4-phenylbutyric acid measured with carbon-11 labeled analogs by PET

The fatty acids, n-butyric acid (BA), 4-phenylbutyric acid (PBA) and valproic acid (VPA, 2-propylpentanoic acid) have been used for many years in the treatment of a variety of CNS and peripheral organ diseases including cancer. New information that these drugs alter epigenetic processes through their inhibition of histone deacetylases (HDACs) has renewed interest in their biodistribution and pharmacokinetics and the relationship of these properties to their therapeutic and side effect profiles. In order to determine the pharmacokinetics and biodistribution of these drugs in primates, we synthesized their carbon-11 labeled analogues and performed dynamic positron emission tomography (PET) in six female baboons over 90 min. The carbon-11 labeled carboxylic acids were prepared by using (11)CO2 and the appropriate Grignard reagents. [(11)C]BA was metabolized rapidly (only 20% of the total carbon-11 in plasma was parent compound at 5 min post injection) whereas for VPA and PBA 98% and 85% of the radioactivity were the unmetabolized compound at 30 min after their administration respectively. The brain uptake of all three carboxylic acids was very low (<0.006%ID/cc, BA>VPA>PBA), which is consistent with the need for very high doses for therapeutic efficacy. Most of the radioactivity was excreted through the kidneys and accumulated in the bladder. However, the organ biodistribution between the drugs differed. [(11)C]BA showed relatively high uptake in spleen and pancreas whereas [(11)C]PBA showed high uptake in liver and heart. Notably, [(11)C]VPA showed exceptionally high heart uptake possibly due to its involvement in lipid metabolism. The unique biodistribution of each of these drugs may be of relevance in understanding their therapeutic and side effect profile including their teratogenic effects.

Cardiac Expression of Human Type 2 Iodothyronine Deiodinase Increases Glucose Metabolism and Protects Against Doxorubicin-induced Cardiac Dysfunction in Male Mice

Altered glucose metabolism in the heart is an important characteristic of cardiovascular and metabolic disease. Because thyroid hormones have major effects on peripheral metabolism, we examined the metabolic effects of heart-selective increase in T3 using transgenic mice expressing human type 2 iodothyronine deiodinase (D2) under the control of the α-myosin heavy chain promoter (MHC-D2). Hyperinsulinemic-euglycemic clamps showed normal whole-body glucose disposal but increased hepatic insulin action in MHC-D2 mice as compared to wild-type (WT) littermates. Insulin-stimulated glucose uptake in heart was not altered, but basal myocardial glucose metabolism was increased by more than two-fold in MHC-D2 mice. Myocardial lipid levels were also elevated in MHC-D2 mice, suggesting an overall up-regulation of cardiac metabolism in these mice. The effects of doxorubicin (DOX) treatment on cardiac function and structure were examined using M-mode echocardiography. DOX treatment caused a significant reduction in ventricular fractional shortening and resulted in more than 50% death in WT mice. In contrast, MHC-D2 mice showed increased survival rate after DOX treatment, and this was associated with a six-fold increase in myocardial glucose metabolism and improved cardiac function. Myocardial activity and expression of AMPK, GLUT1, and Akt were also elevated in MHC-D2 and WT mice following DOX treatment. Thus, our findings indicate an important role of thyroid hormone in cardiac metabolism and further suggest a protective role of glucose utilization in DOX-mediated cardiac dysfunction.

Comment on Minamimoto: incidental focal FDG uptake in heart is a lighthouse for considering cardiac screening

Dear Sir,An interesting article has recently been published [1] on theclinical significance of focal cardiac FDG uptake inpatients with unknown cardiac disease studied for unre-lated purposes. The topic is of great interest because thepossibility to identify unexpected cardiac disease duringPET studies could allow an early diagnosis and reduceunfavorable events. The paper suggests a correlationbetween focal cardiac FDG uptake and the likelihood ofcoronary artery disease (CAD) in patients without cardiacsymptoms and unknown cardiac disease. Apart from theinteresting analysis and the effort to identify the clinicalsignificance of focal uptakes, we agree and highlight theremarks and concerns reported by the authors in the paper.The myocardium derives energy from many sources suchas free fatty acid (FFAs), glucose, pyruvate, ketone bodies,lactate, and amino acids [2–4]. In fasting and resting con-dition, heart metabolism is mainly based on FFAs oxida-tion, the myocardial FDG uptake should be negligible,glycolysis contributes only about 30 % of substrate butdietary and hormonal conditions can modify this situation[5]. In the fasting state, the rate of myocardial FFAs oxi-dation is high, inhibiting glycolysis and glucose oxidationwith consequently low levels of glucose and insulin, while,after glucose load or during hyperglycemic condition,plasma concentrations of glucose and insulin rise reducingperipheral lipolysis, lowering plasma concentrations ofFFAs and hence reducing their availability to the myo-cardium. Glucose then becomes the dominant substrate formyocardial energy production [5, 6]. However, the myo-cardial FDG uptake in the fasting healthy subjects variesunder the clinical fasting conditions [7, 8] and increasedFDG uptake in the myocardium does not significantlycorrelate with the blood glucose level, age, or duration offasting in non-diabetic patients [7, 9]. Only in the uncon-trolled diabetic patients with high levels of blood glucosethere is absent or faint myocardial FDG uptake. Myocardialischemia strikingly alters myocardial substrate metabolism[5]. As blood flow and oxygen supply decline, oxidativemetabolism decreases but still remains the dominant sourceof residual ATP production accounting for more than 90 %[10]. Ischemia is also associated with increased glycolysisbut becomes uncoupled from glucose oxidation with excessproduction of pyruvate and lactate which impair glycolysis[4, 11]. In the normal myocardium FDG distribution pat-terns could be classified into three types (absent to faintuptake; regional uptake; diffuse uptake), but they couldhave no specific meaning [7]. Moreover, even in the sameindividual and under similar fasting conditions, the myo-cardial FDG uptake is neither stable nor reproducible andthe transition from the intense FDG uptake of a dominantlyglycolytic myocardial metabolism to the absent FDGuptake of a dominantly FFAs metabolism is not entirelyuniform either temporally or regionally [6]. In daily clini-cal practise, despite adequate fasting period, the pattern ofmyocardial FDG heart uptake varies a lot, making difficultthe differential diagnosis between physiological and path-ological uptakes. For these reasons the results of

Cardiac-specific adipose triglyceride lipase overexpression protects from cardiac steatosis and dilated cardiomyopathy following diet-induced obesity

Although obesity increases the risk of developing cardiomyopathy, the mechanisms underlying the development of this cardiomyopathy are incompletely understood. As obesity is also associated with increased intramyocardial triacylglycerol (TAG) deposition, also referred to as cardiac steatosis, we hypothesized that alterations in myocardial TAG metabolism and excess TAG accumulation contribute to obesity-induced cardiomyopathy. To test if increased TAG catabolism could ameliorate obesity-induced cardiac steatosis and dysfunction, we utilized wild-type (WT) mice and mice with cardiomyocyte-specific overexpression of adipose triglyceride lipase (MHC-ATGL mice), which regulates cardiac TAG hydrolysis. WT and MHC-ATGL mice were fed either regular chow (13.5 kcal% fat) or high fat-high sucrose (HFHS; 45 kcal% fat and 17 kcal% sucrose) diet for 16 weeks to induce obesity and mice were subsequently studied at the physiological, biochemical and molecular level. Obese MHC-ATGL mice were protected from increased intramyocardial TAG accumulation, despite similar increases in body weight and systemic insulin resistance as obese WT mice. Importantly, analysis of in vivo cardiac function using transthoracic echocardiography showed that ATGL overexpression protected from obesity-induced systolic and diastolic dysfunction and ventricular dilatation. Ex vivo working heart perfusions revealed impaired cardiac glucose oxidation following obesity in both WT and MHC-ATGL mice, which was consistent with similar impaired cardiac insulin signaling between genotypes. However, hearts from obese MHC-ATGL mice exhibited reduced reliance on palmitate oxidation when compared with the obese WT, which was accompanied by decreased expression of proteins involved in fatty acid uptake, storage and oxidation in MHC-ATGL hearts. These findings suggest that cardiomyocyte-specific ATGL overexpression was sufficient to prevent cardiac steatosis and decrease fatty acid utilization following HFHS diet feeding, leading to protection against obesity-induced cardiac dysfunction.

Assessment of the 18F-Labeled PET Tracer LMI1195 for Imaging Norepinephrine Handling in Rat Hearts

A novel (18)F-labeled tracer, LMI1195 (N-[3-bromo-4-(3-(18)F-fluoro-propoxy)-benzyl]-guanidine), is being developed for sympathetic nerve imaging; its high specificity for neural uptake-1 mechanism has previously been demonstrated in cell associative studies and in rabbit and nonhuman primate studies assessing heart uptake. The aim of this study was to investigate the mechanisms of (18)F-LMI1195 cardiac uptake in the rat, which is known to contain norepinephrine uptake mechanisms beyond uptake-1. Tracer accumulation in the heart was studied over time after intravenous administration of (18)F-LMI1195 in healthy male Wistar rats by quantitative in vivo PET imaging. The uptake mechanism was assessed by pretreatment with the nonselective norepinephrine uptake-1 and norepinephrine uptake-2 inhibitor phenoxybenzamine (50 mg/kg intravenously; n = 4), the selective norepinephrine uptake-1 inhibitor desipramine (2 mg/kg intravenously; n = 4), or saline control (intravenously; n = 4). (18)F-LMI1195 produced high and sustained heart uptake allowing clear delineation of the left ventricular wall over 60 min after tracer administration. Pretreatment with phenoxybenzamine markedly reduced the (18)F-LMI1195 cardiac uptake when compared with controls. In contrast, there was preserved (18)F-LMI1195 uptake after desipramine pretreatment. In rats, cardiac uptake of (18)F-LMI1195 was significantly inhibited by phenoxybenzamine but not desipramine, suggesting (18)F-LMI1195 is a substrate for the uptake-2 mechanism and is consistent with the rat heart having a dominant level of the mechanism.