High-energy Phosphate Production Research Articles

Fenofibrate modulates cardiac energy metabolism in moderate-severe aortic stenosis: a mechanistic double-blinded randomised controlled MR trial

Abstract Background Aortic stenosis (AS) is associated with impaired cardiac metabolism and high-energy phosphate production which can further exacerbate the progression to HF. Myocardial lipid accumulation and reduced production of high-energy phosphates have important implications for both systolic and diastolic heart function in AS. Peroxisome proliferator-activated receptor alpha (PPAR-alpha agonist, has the potential to optimise cardiac substrate metabolism, thereby, may be effective in improving cardiac dysfunction and functional capacity in AS. Methods In this prospective, randomized, double-blind, placebo-controlled study, patients with moderate-to-severe asymptomatic AS (n=72) were randomly assigned to Fenofibrate 200 mg daily or matching placebo for 6 months in 4:1 fashion. Of the total 72 participants, 59 received Fenofibrate and 13 received placebo. All subjects underwent cardiac magnetic resonance (CMR) and exercise testing at 0 and 6 months, with follow-up data available in 47 patients. The primary end point was change in myocardial triglyceride content (MTG) determined by proton magnetic resonance spectroscopy (1H-MRS). Secondary outcome measures included change in cardiac energetics (PCr/ATP) determined by phosphorus magnetic resonance spectroscopy (31P-MRS), left ventricular strain determined by myocardial tagging and exercise capacity i.e. peak VO2 assessment via cardiopulmonary exercise testing. Results Cardiac energetics (PCr/ATP) showed statistically significant improvement in the Fenofibrate group (0.14 ± 0.48; CI: - 0.03, 0.30; p=0.05) vs placebo group (0.11 ± 0.75; CI -0.42, 0.65; p=0.64). There was also significant reduction in MTG in the Fenofibrate group (mean change -0.40 ± 0.97; 95% CI -0.74, -0.06; p=0.02). MTG content also reduced in the placebo group (mean change -0.41 ± 0.26, 95% CI -1.10, 0.18, p 0.13), but the reduction was not statistically significant. Though no significant changes were seen in the systolic function parameters, there was a significant improvement in diastolic function in the fenofibrate group only (peak diastolic longitudinal strain rate 9.6, 95% CI: 1.13, 18.1; p = 0.02). Peak VO2 did not show any significant improvement in either group. Conclusions Fenofibrate works as a metabolic modulator upregulating fatty acid metabolism in pressure overload AS hearts. Such modulation is associated with an improvement in cardiac energetics and diastolic function in moderate-severe AS.Main results from baseline to 6 monthsBaseline demographic and CMR parameters

Mitochondrial Substrate-Level Phosphorylation as Energy Source for Glioblastoma: Review and Hypothesis.

Glioblastoma multiforme (GBM) is the most common and malignant of the primary adult brain cancers. Ultrastructural and biochemical evidence shows that GBM cells exhibit mitochondrial abnormalities incompatible with energy production through oxidative phosphorylation (OxPhos). Under such conditions, the mitochondrial F0-F1 ATP synthase operates in reverse at the expense of ATP hydrolysis to maintain a moderate membrane potential. Moreover, expression of the dimeric M2 isoform of pyruvate kinase in GBM results in diminished ATP output, precluding a significant ATP production from glycolysis. If ATP synthesis through both glycolysis and OxPhos was impeded, then where would GBM cells obtain high-energy phosphates for growth and invasion? Literature is reviewed suggesting that the succinate-CoA ligase reaction in the tricarboxylic acid cycle can substantiate sufficient ATP through mitochondrial substrate-level phosphorylation (mSLP) to maintain GBM growth when OxPhos is impaired. Production of high-energy phosphates would be supported by glutaminolysis—a hallmark of GBM metabolism—through the sequential conversion of glutamine → glutamate → alpha-ketoglutarate → succinyl CoA → succinate. Equally important, provision of ATP through mSLP would maintain the adenine nucleotide translocase in forward mode, thus preventing the reverse-operating F0-F1 ATP synthase from depleting cytosolic ATP reserves. Because glucose and glutamine are the primary fuels driving the rapid growth of GBM and most tumors for that matter, simultaneous restriction of these two substrates or inhibition of mSLP should diminish cancer viability, growth, and invasion.

Metabolic Modulation of Cardiac Metabolism in Heart Failure.

Heart failure (HF) is associated with metabolic changes that cause a progressive impairment of cardiac and skeletal muscle high-energy phosphate production. As a consequence of the impaired cardiac metabolism, other processes are activated in the failing heart that further exacerbate the progression of HF. The reduced production of high-energy phosphates has important implications for both systole and diastole in HF with both preserved and reduced left ventricular function. The aim of this review is to summarise the state-of-the-art on metabolic therapy in HF with a particular focus on trimetazidine. Metabolic agents optimise cardiac substrate metabolism without exerting negative haemodynamic effects. In particular, as studies with metabolic agents modulating cardiac metabolism have consistently demonstrated, this approach is effective in improving symptoms, functional capacity and prognosis in people with HF when added to optimal medical therapy. Therefore, the modulation of cardiac metabolism is an important therapeutic approach to the treatment of HF, especially in patients where it is of ischaemic or metabolic origin. Although further studies are needed, metabolic agents might be a new, effective strategy for the treatment of HF.

Innovative therapeutic approaches for the treatment of myocardial ischemia and angina

Cardiac energy metabolism is complex and has three main components: 1. Substrate utilisation, 2 Oxydative phosphorilation and 3. Transfer of high energy phosphates. Substrate utilization involves the cellular uptake of free fatty acids, glucose and amino acids, their breakdown by beta-oxidation and glycolysis, and the entry of Acetyl Coenzyme A into the Krebs cycle. Oxidative phosphorylation involves the production of energy by the mitochondrial respiratory chain and the production of high-energy phosphates. Until recent it was not known that it was possible to delimitate the infarct size through therapeutic interventions. Likely, the improvement to understand the mechanisms of cellular survivor occurs in that year, when Murry et al. discovered an intrinsic mechanism of myocardial protection, which they denominated as ischemic preconditioning. In that experiment, it was observed that four cycles of five minutes of ischemia, with intermittent reperfusion, before 40 min of ischemia period, resulted in reduction of 75% of the infarct size. Also, it was demonstrated by these researches that this protection it was not resulted by collateral overture and that protective effect of the ischemic preconditioning was abolished in the presence of prolonged ischemia, with duration of 3 h. Two phenomenons, the walk-through angina and the warm up phenomenon could represent a clinical manifestation of preconditioning. The phenomenon of walk-through angina was first related in the end of century XVIII in a patient that developed angina during the effort, but experimented the paradox disappearing of the pain with the exercise continuation. The exact molecular mechanisms involved in ischemic preconditioning are not fully elucidated. Therefore, due to the high prevalence of risk factors to cardiovascular diseases and the high morbidity and mortality associated with coronary artery disease, there is a needed to understand even better the cardiac metabolism during ischemia and the strategies to reduce the consequences of the ischemia, to preserve the viability of the ischemic myocardium.

The long and winding road to the mitochondrial pyruvate carrier

The extraction of energy and biosynthetic building blocks from fuel metabolism is a fundamental requisite for life. Through the action of cellular enzymes, complex carbon structures are broken down in reactions coupled to the production of high-energy phosphates as in ATP and GTP as well as electron carriers such as NADH and FADH2. These processes traverse across compartments inside the cell in order to access specific enzymes and environments. Pyruvate is the end product of cytosolic glycolysis and has a variety of possible fates, the major one being mitochondrial oxidation. While this metabolite has been known to cross the inner mitochondrial membrane for decades, it is only recently that proteins necessary for this activity have been identified. This review will chronicle more than 40 years of research interrogating this critical process and will discuss some of the possible implications of this discovery for cancer metabolism.

Trimetazidine improves exercise performance in patients with peripheral arterial disease

Trimetazidine, an inhibitor of free fatty acids (FFA) oxidation, shifts cardiac and muscle metabolism from FFA to glucose utilization. This effect results in a greater production of high energy phosphates and ultimately into an anti-ischemic effect. Whether the anti-ischemic cardiac effects of trimetazidine (TMZ) can be translated to skeletal muscle in patients with claudication is unknown. We investigated the effectiveness of TMZ on functional performance in patients with peripheral arterial disease (PAD) and claudication. One hundred patients with claudication were enrolled in a parallel, double-blind, 3 months study. Patients were randomized to receive TMZ or matching placebo and were included in a domiciliary exercise program, consisting in daily sessions of aerobic and isotonic exercise for at least five days a week. All patients underwent a treadmill test, evaluating maximal walking distance (MWD), and ankle–brachial index (ABI) at baseline and after 3 months. ABI was similar in the two groups at baseline and did not significantly change at the end of the study in either groups (0.83 + 0.04 vs 0.85 + 0.03, TMZ vs placebo, respectively). MWD improved in all patients with exercise training; however, a greater improvement in MWD was observed with TMZ compared to placebo (23% vs 14%, p < 0.0001). Physical training ameliorates functional performance in PAD. The adjunct of TMZ to exercise induces a greater improvement in MWD, suggesting that the inhibition of FFA oxidation improves functional capacity in patients with PAD and claudication.

Early detection of iron overload in the heart: a key role for MRI!

Hereditary haemochromatosis is autosomal recessive disorder associated with the HLA-A locus on the short arm of chromosome 6 [1]. It is the most common genetic disorder worldwide [2]. About 56,000 babies are born each year with thalassaemia major, of whom 30,000 require lifelong transfusions to survive [3]. The disease is very prevalent in European countries along the Mediterranean Sea and in South East Asia. In other countries it becomes a problem because of immigration from these areas. Excessive iron deposition occurs mostly in the pancreas, pituitary, but also in the heart. In normal subjects, absorption of iron from the gut is inversely correlated with iron stores. In homozygous hereditary haemochromatosis there is increased absorption of iron, which is not regulated by iron stores [4]. This leads to progressive iron accumulation because there is no normal mechanism for excretion of excess iron once it has been absorbed into the body. The clinical manifestations of iron overload include liver disease in 75% of cases, weakness and lethargy in 74%, skin hyperpigmentation in 70%, diabetes mellitus in 48%, arthralgia in 44%, impotence in 45%, electrocardiographic abnormalities in 31% and heart failure and conduction disturbances in 15% [5, 6]. Heart failure remains a major cause of death in these patients, frequently people younger than 35 years [7]. In the majority of patients iron induced cardiomyopathy can be reversed by intense chelation therapy with desferrioxamine and deferiprone [3, 8]. At this moment T2* magnetic resonance imaging is the best way for early detection of iron overload in patients with haemochromatosis [9–11]. T2* is an unique magnetic resonance parameter measured in milliseconds and shortened in relation to the level iron in the heart. The parameter can be used for screening haemochromatosis patients and to guide chelation therapy in those with established cardiac haemochromatosis [8]. The paper of Mavrogeni et al. [12] in this issue of the International Journal of Cardiac Imaging shows, that heart failure patients with severe iron overload indicated by a T2* value less than 10 ms had a lower exercise capacity than patients with a T2* value higher than 10 ms. The authors also show, that there was a excellent relation of the extent of iron overload measured by T2* and exercise tolerance in thalassemia patients. As shown by others, thalassemia patients present prolonged phase II on-transient oxygen kinetics during submaximal, constant workload exercise, compared with healthy controls, possibly suggesting a slower rate of high energy phosphate production and utilization and reduced oxidative capacity of myocytes [13]. This could also account for their significantly limited exercise tolerance. Reduction of exercise tolerance is also s highly associated with the limited functional status of peripheral muscles in these patients [14]. This study shows again that T2* MRI plays a crucial role in screening cardiac haemochromatosis and guidance of chelation therapy. Early detection of cardiac iron load and early initiation of treatment are crucial for survival of these patients. Recently the chelator deferasirox has been shown to be more effective in better at improving myocardial iron [15]. This could improve outcome in thalassemia patients. As shown, early effects of therapy can only be detected with dedicated MRI sequences. Unfortunately these MRI scanners are not always available in those areas, where the prevalence of haemachromatosis patients is extremely high.

Impaired oxygen kinetics in beta‐thalassaemia major patients

Beta-thalassaemia major (TM) affects oxygen flow and utilization and reduces patients' exercise capacity. The aim of this study was to assess phase I and phase II oxygen kinetics during submaximal exercise test in thalassaemics and make possible considerations about the pathophysiology of the energy-producing mechanisms and their expected exercise limitation. Twelve TM patients with no clinical evidence of cardiac or respiratory disease and 10 healthy subjects performed incremental, symptom-limited cardiopulmonary exercise testing (CPET) and submaximal, constant workload CPET. Oxygen uptake (VO2), carbon dioxide output and ventilation were measured breath-by-breath. Peak VO2 was reduced in TM patients (22.3 +/- 7.4 vs. 28.8 +/- 4.8 mL kg(-1) min(-1), P < 0.05) as was anaerobic threshold (13.1 +/- 2.7 vs. 17.4 +/- 2.6 mL kg(-1) min(-1), P = 0.002). There was no difference in oxygen cost of work at peak exercise (11.7 +/- 1.9 vs. 12.6 +/- 1.9 mL min(-1) W(-1) for patients and controls respectively, P = ns). Phase I duration was similar in TM patients and controls (24.6 +/- 7.3 vs. 23.3 +/- 6.6 s respectively, P = ns) whereas phase II time constant in patients was significantly prolonged (42.8 +/- 12.0 vs. 32.0 +/- 9.8 s, P < 0.05). TM patients present prolonged phase II on-transient oxygen kinetics during submaximal, constant workload exercise, compared with healthy controls, possibly suggesting a slower rate of high energy phosphate production and utilization and reduced oxidative capacity of myocytes; the latter could also account for their significantly limited exercise tolerance.

Acute effect of antidiabetic 1,4‐dihydropyridine compound cerebrocrast on cardiac function and glucose metabolism in the isolated, perfused normal rat heart

Diabetes mellitus (DM) is an important cardiovascular risk factor and is associated with abnormalities in endothelial and vascular smooth muscle cell function, evoked by chronic hyperglycemia and hyperlipidemia. Chronic insulin deficiency or resistance is marked by decreases in the intensity of glucose transport, glucose phosphorylation, and glucose oxidation, plus decreases in ATP levels in cardiac myocytes. It is important to search for new agents that promote glucose consumption in the heart and partially inhibit extensive fatty acid beta-oxidation observed in diabetic, ischemia. When the oxygen supply for myocardium is decreased, the heart accumulates potentially toxic intermediates of fatty acid beta-oxidation, that is, long-chain acylcarnitine and long-chain acyl-CoA metabolites. Exogenous glucose and heart glycogen become an important compensatory source of energy. Therefore we studied the effect of the antidiabetic 1,4-dihydropyridine compound cerebrocrast at concentrations from 10(-10) M to 10(-7) M on isolated rat hearts using the method of Langendorff, on physiological parameters and energy metabolism. Cerebrocrast at concentrations from 10(-10) M to 10(-7) M has a negative inotropic effect on the rat heart. It inhibits L-type Ca(2+)channels thereby diminishing the cellular Ca(2+) supply, reducing contractile activity, and oxygen consumption, that normally favors enhanced glucose uptake, metabolism, and production of high-energy phosphates (ATP content) in myocardium. Cerebrocrast decreases heart rate and left ventricular (LV) systolic pressure; at concentrations of 10(-10) M and 10(-9) M it evokes short-term vasodilatation of coronary arteries. Increase of ATP content in the myocytes induced by cerebrocrast has a ubiquitous role. It can preserve the integrity of the cell plasma membranes, maintain normal cellular function, and inhibit release of lactate dehydrogenase (LDH) from cells that is associated with diabetes and heart ischemia. Administration of cerebrocrast together with insulin shows that both compounds only slightly enhance glucose uptake in myocardium, but significantly normalize the rate of contraction and relaxation ( +/- dp/dt). The effect of insulin on coronary flow is more pronounced by administration of insulin together with cerebrocrast at a concentration of 10(-7) M. Cerebrocrast may promote a shift of glucose consumption from aerobic to anerobic conditions (through the negative inotropic properties), and may be very significant in prevention of cardiac ischemic episodes.

Heart failure: a model of cardiac and skeletal muscle energetic failure

Chronic heart failure (CHF), the new epidemic in cardiology, is characterized by energetic failure of both cardiac and skeletal muscles. The failing heart wastes energy due to anatomical changes that include cavity enlargement, altered geometry, tachycardia, mitral insufficiency and abnormal loading, while skeletal muscle undergoes atrophy. Cardiac and skeletal muscles also have altered high-energy phosphate production and handling in CHF. Nevertheless, there are differences in the phenotype of myocardial and skeletal muscle myopathy in CHF: cardiomyocytes have a lower mitochondrial oxidative capacity, abnormal substrate utilisation and intracellular signalling but a maintained oxidative profile; in skeletal muscle, by contrast, mitochondrial failure is less clear, and there is altered microvascular reactivity, fibre type shifts and abnormalities in the enzymatic systems involved in energy distribution. Underlying these phenotypic abnormalities are changes in gene regulation in both cardiac and skeletal muscle cells. Here, we review the latest advances in cardiac and skeletal muscle energetic research and argue that energetic failure could be taken as a unifying mechanism leading to contractile failure, ultimately resulting in skeletal muscle energetic failure, exertional fatigue and death.

Metabolic Therapy for Heart Disease: Impact of Trimetazidine

Coronary artery disease remains the major contributor to the development of heart failure. Despite the massive efforts targeted toward preventative therapy and life style changes, the incidence of coronary artery disease continues to be high, shows little or no signs of abatement, and remains the leading health economic burden in developed countries and, for that matter, worldwide. There is no doubt that the aggressive use of thrombolytic therapy and primary coronary angioplasty has had a major impact on limiting the sequelae of acute myocardial infarction. There is also little doubt that routine coronary revascularization through coronary artery bypass surgery or coronary angioplasty with attendant coronary stent deployment has had a positive impact, albeit transient, on limiting symptoms of coronary artery disease. Despite all of these successes of the twentieth century, however, chronic stable angina pectoris remains a major clinical problem that must be dealt with by cardiologists, internists and primary care physicians alike as we move into the next millenium. Many of the patients that present with stable angina pectoris are not candidates for further revascularization, or, in this the age of rationed health care, even if eligible, must “stand in line” to have the procedure performed. Needless to say that most, if not all, of these patients are candidates for the development of congestive heart failure (CHF), truly the disease of the next millenium. The myocardium depends on oxygen to support high energy phosphate production by oxidative phosphorylation. The latter is the only metabolic process that can generate sufficient chemical energy to fuel the continuous needs of the contractile machinery of the heart. Accordingly, when the amount of oxygen delivered to the myocardium is insufficient to meet the requirements for mitochondrial respiration, the production of high energy phosphates falls and lactate, the end product of anaerobic glycolysis, starts to accumulate. The interruption of oxygen supply to the myocardium or ischemia, regardless of etiology, leads to the clinical symptom of angina pectoris. Ischemia can be defined as an imbalance between the supply of oxygenated blood to the myocardium and the oxygen requirements of the myocardium. When the supply of oxygen cannot meet the demands, as is the case when coronary artery disease is present, contractile function diminishes along with clinical manifestation of anginal pain. The latter the result of stimulation of cardiac sensory afferent nerve fibers by “noxious stimuli” produced in the myocardium. The cardiac sensory afferents nerves are stimulated by a rise in interstitial adenosine and K+, both of which increase when oxygen delivery and aerobic ATP synthesis are reduced [1]. Electrocardiographically, ischemia is reflected through ST-segment elevation or depression on the electrocardiogram with the latter believed to reflect delayed repolarization in the ischemic zone, and has recently been attributed to KATP channels [2], which presumably open in response to the fall in ATP. Taken together, both angina and ST-segment changes during ischemia are arguably the result of a failure to regenerate ATP, and as such, are of metabolic origin.

Supplementation of Nucleoside-Nucleotide Mixture Enhances Functional Recovery and Energy Metabolism Following Long-Time Hypothermic Heart Preservation

OG-VI, a well-balanced mixture of nucleoside and nucleotides, has been demonstrated to have a favorable effect on energy metabolism. In this study, we tested the hypothesis that addition of OG-VI to the University of Wisconsin solution can improve the cardiac functional recovery following long-time hypothermic preservation. Forty-two male Wistar rats were randomized into four groups. After 30-min of isolated working heart perfusion, the rat hearts were arrested with St. Thomas cardioplegic solution and preserved at 4 degrees C in saline, OG-VI, UW, and UW+OG-VI, respectively. After 12-h of preservation, the hearts were reperfused for 60-min during which the recovery of cardiac functions were monitored continuously. Myocardial adenine nucleotides were analyzed using high-performance liquid chromomatograph. In the UW+OG-VI group, the recovery of cardiac output, coronary flow, aortic flow, rate-pressure product, left ventricle stroke volume, and stroke work were significantly higher than other groups (P < 0.05). Furthermore, all phosphate high-energy compounds were significantly higher in the UW+OG-VI group than in the other groups (P < 0.05). Coronary vascular resistance and myocardial wet/dry weight ratio were obviously lower in the UW+OG-VI group, compared to the other groups (P < 0.05). Heart function was better recovered when nucleoside-nucleotide mixture was added to UW solution during long-time hypothermic rat heart preservation. The mechanism is not totally clear, but enhancement of high-energy phosphate production is possible.

Sepsis and Myocardial Depression in a Young Woman

Severe sepsis may be associated with depression of myocardial function, attributed to various inflammatory mediators. Myocardial dysfunction in sepsis is characterized by biventricular failure and complicates usual therapy with high-volume fluid resuscitation and vasopressors. However, in patients who survive septic shock, myocardial dysfunction can improve rapidly. We describe a young woman with septic shock due to Streptococcus pneumoniae, complicated by severe but reversible biventricular dysfunction.

TPPP/p25: from unfolded protein to misfolding disease: prediction and experiments

TPPP/p25, the first representative of a new protein family, identified as a brain-specific unfolded protein induces aberrant microtubule assemblies in vitro, suppresses mitosis in Drosophila embryo and is accumulated in inclusion bodies of human pathological brain tissues. In this paper, we present prediction and additional experimental data that validate TPPP/p25 to be a new member of the “intrinsically unstructured” protein family. The comparison of these characteristics with that of α-synuclein and tau, involved also in neurodegenerative diseases, suggested that although the primary sequences of these proteins are entirely different, there are similarities in their well-defined unstructured segments interrupted by “stabilization centres”, phosphorylation and tubulin binding motives. SK-N-MC neuroblastoma cells were transfected with pEGFP-TPPP/p25 construct and a stable clone denoted K4 was selected and used to establish the effect of this unstructured protein on the energy state/metabolism of the cells. Our data by analyzing the mitochondrial membrane polarization by fluorescence microscopy revealed that the high-energy phosphate production in K4 clone is not damaged by the TPPP/p25 expression. Biochemical analysis with cell homogenates provided quantitative data that the ATP level increased 1.5-fold and the activities of hexokinase, glucosephosphate isomerase, phosphofructokinase, triosephosphate isomerase and glyceraldehyde-3-phosphate dehydrogenase were 1.2 to 2.0-fold higher in K4 as compared to the control. Our modelling using these data and rate equations of the individual enzymes suggests that the TPPP/p25 expression stimulates glucose metabolism. At pathological conditions TPPP/p25 is localized in inclusion bodies in multiple system atrophy, it tightly co-localizes with α-synuclein, partially with tubulin and not with vimentin. The previous and the present studies obtained with immunohistochemistry with pathological human brain tissues rendered it possible to classify among pathological inclusions on the basis of immunolabelling of TPPP/p25, and suggest this protein to be a potential linkage between Parkinson’s and Alzheimer’s diseases.

Peroxisome proliferator activator receptors (PPAR), insulin resistance, and cardiomyopathy: friends or foes for the diabetic patient with heart failure?

Diabetes is a risk factor for coronary atherosclerosis, myocardial infarction, and ischemic cardiomyopathy. Insulin resistance is associated with left ventricular (LV) hypertrophy and hypertensive cardiomyopathy. Even in the absence of coronary artery disease or hypertension, "diabetic cardiomyopathy" can develop because of myocardial autonomic dysfunction or impaired coronary flow reserve. The relationship between insulin resistance and cardiomyopathy is bidirectional. Systemic and myocardial glucose uptake is compromised in heart failure independent of etiology. These abnormalities are associated with cellular deficits of insulin signaling. Insulin resistance in heart failure can be detrimental, because transcriptional shifts in metabolic gene expression favor glucose over fat as a substrate for high-energy phosphate production. Although preexisting diabetes accelerates this process of "metabolic death," insulin resistance can also develop secondary to cardiomyopathy-associated overabundance of neurohormones and cytokines. Insulin resistance and fatty acid excess are potential therapeutic targets in heart failure, striving for efficient myocardial substrate utilization. Peroxisome proliferator activator receptor gamma (PPARgamma) agonists are antidiabetic agents with antilipemic and insulin-sensitizing activity. Experimental studies suggest salutary effects in limiting infarct size, attenuating myocardial reperfusion injury, inhibiting hypertrophic signaling and vascular antiinflammatory actions through cytokine inhibition. However, clinical applicability in diabetic patients experiencing heart failure has been hampered because of increased edema and even fewer reports of exacerbation associated with these compounds. Evidence to date argues for peripheral mechanisms of edema unrelated to central hemodynamics. Nevertheless, they are currently contraindicated in New York Heart Association (NYHA) III-IV patients, particularly in combination with insulin. Investigations are underway to decipher mechanisms, risks, and benefits of PPARgamma agonists, as well as the role of the structurally related PPARalpha receptor on cardiovascular metabolism and function.

Nutritional supplementation with mixed essential amino acids enhances myocyte survival, preserving mitochondrial functional capacity during ischemia-reperfusion injury

In patients undergoing coronary surgery, the uptake of amino acids, which has been shown to correlate with oxygen consumption, is a mechanism of cardiac adaptation to the iatrogenic ischemia-reperfusion injury associated with cardioplegic arrest. Based on these premises, we sought to determine whether oral supplementation with mixed amino acids may protect the rat heart exposed to ischemia-reperfusion and to address whether this hypothesized cardioprotection is achieved, at least in part, through preservation of the energy-producing properties of mitochondria. Sprague–Dawley rats were fed (by enteral route) a liquid diet, with or without mixed essential amino acids (daily dose of 1 g/kg) for 30 days. Hearts from anesthetized rats were perfused by the Langendorff method and randomized to 3 groups. The control group was perfused with buffer for 60 minutes; the ischemia-reperfusion control and the amino acid–treated groups were exposed to 35 minutes of ischemia, followed by 60 or 120 minutes of reperfusion. Amino acid supplements minimized infarct size (22 ± 1.8% vs 33 ± 2.5%; p <0.05) and occurrence of cardiomyocyte apoptosis, as assessed by co-localization of terminal deoxynucleotidyl transferase–mediated dUTP nick end labeling (TUNEL) and caspase-3–positive staining (p <0.01). Long-term treatment with amino acids also reduced the proportion of cardiomyocytes exhibiting immunostaining for cleaved caspase-9 (p <0.01) but was ineffective on processing of caspase-8. Similar results were obtained in the whole heart by caspase activity assays (p <0.01). The lessened activation of caspase-9 detected in amino acid-treated hearts paralleled a strong reduction in mitochondrial release of cytochrome c. Adenosine triphosphate (ATP) content and rate of ATP production in isolated mitochondria were reduced by >75% in control hearts after 2 hours of reperfusion (p <0.05 vs control hearts); these values returned toward those of the control group in hearts supplemented with amino acids (p <0.01). Finally, the oxygen consumption rate in myocardial skinned bundles was markedly reduced in ischemia-reperfusion control hearts and almost normalized in amino acid-treated hearts (approximately 20% and 93% of the value for normoxic hearts; p <0.01). These results suggest that oral amino acid supplementation attenuates the extent of ischemia-reperfusion injury in the rat heart, through preservation of the mitochondria-generated production of high-energy phosphates.

Metabolic mapping in mammalian retina: a biochemical and 3H-2-deoxyglucose autoradiographic study

It has long been known that mammalian retinas metabolize glucose aerobically to lactic acid and carbon dioxide. The classical view holds that glucose is the primary substrate for energy metabolism in all retinal cells, and that photoreceptor cells have the highest rates of glycolysis and respiration. A different and more recent view is that the Müller cells are the principal, if not sole aerobic producers of lactate, which then serves as the primary fuel for the mitochondria in photoreceptor cells and other retinal neurons. In this paper, we have examined these two competing hypotheses in rat and guinea pig retinas by identifying the cellular sites of glucose uptake and phosphorylation via hexokinase by means of autoradiographic studies with 3H-2-deoxyglucose ( 3H-2DG). The rat retina serves as a vascular model and the guinea pig retina serves as an avascular model. Rat and guinea pig eyecups were incubated in oxygenated, bicarbonate-buffered media containing glucose in the presence of labeled and unlabeled 2DG. Biochemical measurements of lactate production and ATP content were made on rat retinas incubated with different concentrations of glucose and 2DG in order to establish the optimal condition for conducting the autoradiographic studies with 3H-2DG. The optimal substrate concentrations were 1 m m glucose and 0·25 m m 2DG. Results showed that following incubation of dark-adapted rat eyecups for 1 hr in media containing 1 m m glucose/0·25 m m 2DG and supplemented with 3H-2DG, the label was distributed throughout all the layers of the retina, from the ganglion cell layer to the retinal pigment epithelium, with denser label associated with the outer retina (photoreceptors) relative to the density of label in the inner retina, as evaluated by counts of silver grains in individual retinal layers. Exposure of rat eyecups to light did not alter the relative distribution of label, but did increase total grain counts by 70%. However, uptake of labeled 2DG, as measured by scintillation counting of radioactivity in trichloroacetic acid extracts, was not significantly different between light- and dark-adapted rat retinas. In guinea pig eyecups, labeled 2DG was distributed throughout all the retinal layers. Addition of 10 m m lactate or pyruvate to the glucose/2DG media produced no measurable change in the density or distribution of label in the eyecups. Measurements of the activity of hexokinase in rat retinas revealed that this enzyme was present in both the mitochondrial and cytosolic fractions. The present results suggest that as long as the availability of ambient glucose is adequate, retinal neurons use glucose, rather than glial-derived lactate, as the major substrate for the production of high energy phosphates.

Vanadate improves cardiac function and myocardial energy metabolism in diabetic rat hearts.

Vanadium mimicking the metabolic effects of insulin is known to decrease serum glucose levels and to influence glucose metabolism in diabetes mellitus. However, it is unclear whether vanadium ameliorates the metabolic disorder in diabetic hearts causing myocardial dysfunction. The purpose of this study was to assess the effects of vanadium on cardiac performance and energy metabolism in diabetic rat hearts. Four groups of Wistar rats were studied: untreated control rats (group C, n = 8). vanadate-treated rats (group V, n = 10), untreated diabetic rats (group DM, n = 9) induced by streptozotocin. and vanadate-treated diabetic rats (group DMV, n = 8). Vanadate-treated rats drank a 1.5 mM sodium orthovanadate (Na3VO4) solution during a 4 week diabetic condition. Hearts were perfused with Krebs-Henseleit buffer after the diabetic duration. After the maximum left ventricular dP/dt and cardiac efficiency were calculated, the myocardial contents of ATP and creatine phosphate (P-Cr) and myocardial energy metabolism were assessed by cytosolic phosphorylation potential. Peak positive and negative dP/dt, and cardiac efficiency decreased significantly in group DM compared with group C, while there were no significant differences between groups C and DMV. The myocardial contents of ATP (micromol/g wet heart) and P-Cr (micromol/g wet heart), and cytosolic phosphorylation potential (M(-1)) increased from 2.72 +/- 0.46. 1.45 +/- 0.58. and 3,530 +/- 1,220 in group DM to 3.88 +/- 0.76, 3.81 +/- 1.36, and 11,200 +/- 2,400 in group DMV, respectively. It is concluded that vanadium restored the production of high energy phosphates in the myocardium and improved myocardial dysfunction by regulating metabolic processes in diabetic rat hearts.

The Cytoplasm

Until the last decade or so, many intracellular signaling pathways were viewed as processes in which second messengers diffused uniformly through a well-mixed milieu of the cell’s cytoplasm to reach their targets. Although it was recognized that the cell’s interior was compartmentalized, this compartmentalization was believed to be largely defined by internal membranes, such as the nuclear envelope, endoplasmic reticulum (ER), sarcoplasmic reticulum (SR), and mitochondria. But like the joke about the person who has lost his keys in the dark but looks for them under the street lamp because the light is better, this view of the cytoplasm as a well-mixed milieu was less of a proven fact than a simplifying assumption. Over the last decade, advances in subcellular imaging have dramatically upset this view, so that now a high degree of compartmentalization of signaling pathways within the cytoplasm is considered the norm rather than the exception. It is now clear that the cytoplasm has a highly organized cytoskeleton and sophisticated molecular trafficking mechanisms that direct and tether proteins into macroaggregates at specific locations to facilitate localized signaling. The cytoplasm is now viewed as a system of microdomains with restricted diffusion (eg, hierarchical Ca2+ signaling) and direct channeling of substrates to enzymes (eg, protein kinases/phosphatases cascades). In the field of metabolism, subcellular compartmentation of energy production has been a well-accepted fact ever since mitochondria were identified as the engines driving aerobic high-energy phosphate production. In addition, glycolytic enzymes complexes are well-known to be associated with specific intracellular structures, such as the SR.1 On the flip side, however, energy consumption by the cell has traditionally been viewed as a fairly democratic process, with high-energy phosphates freely diffusing throughout the cytoplasm to be consumed wherever they are needed. In tissues with high energy requirements, such as muscle, the creatine …

Fructose-1,6-diphosphate and a glucose-free solution enhances functional recovery in hypothermic heart preservation

Background Fructose-1,6-diphosphate (FDP) has been shown to protect tissue during hypoxia under various ischemic conditions, including isolated heart perfusion. We tested the hypothesis that adding FDP to St. Thomas solution can extend hypothermic heart preservation time. Methods Sixteen adult Sprague-Dawley rats were used. Under general anesthesia, the hearts were removed and preserved at 4°C in St. Thomas solution (30 ml/kg) for 12 hours. FDP (5 mM) was added to the St. Thomas solution in the study group ( n = 8), whereas no FDP was used in the control group ( n = 10). The hearts were reperfused after 12 hours of preservation using a working heart model. Results In the study group, cardiac output ranged from 13.00 ± 2.34 to 17.66 ± 1.71 ml/min, maximum aortic flow was 3.40 ± 1.99 to 9.26 ± 1.72 ml/min, left ventricular stroke volume ranged from 0.074 ± 0.014 to 0.092 ± 0.009 ml, left ventricular stroke work ranged from 6.22 ± 0.39 to 7.95 ± 0.44 ml/mmHg, and maximum left ventricular generated power was 14.38 ± 2.94 to 20.16 ± 2.49 Joules/min. All of these parameters were higher than those in the control group ( p < 0.001). Coronary vascular resistance and myocardial tissue wet/dry weight ratio were lower in the study group than in the control group ( p < 0.05). Conclusions Heart function was better preserved when FDP was added to St. Thomas solution during hypothermic rat heart preservation. The mechanism is not totally clear, but enhancement of high-energy phosphate production during ischemia is possible. Key words: heart, procurement, hypothermia, fructose-1,6-diphosphate.