Molecular system bioenergetics: regulation of substrate supply in response to heart energy demands

This review re-evaluates regulatory aspects of substrate supply in heart. In aerobic heart, the preferred substrates are always free fatty acids, and workload-induced increase in their oxidation is observed at unchanged global levels of ATP, phosphocreatine and AMP. Here, we evaluate the mechanisms of regulation of substrate supply for mitochondrial respiration in muscle cells, and show that a system approach is useful also for revealing mechanisms of feedback signalling within the network of substrate oxidation and particularly for explaining the role of malonyl-CoA in regulation of fatty acid oxidation in cardiac muscle. This approach shows that a key regulator of fatty acid oxidation is the energy demand. Alterations in malonyl-CoA would not be the reason for, but rather the consequence of, the increased fatty acid oxidation at elevated workloads, when the level of acetyl-CoA decreases due to shifts in the kinetics of the Krebs cycle. This would make malonyl-CoA a feedback regulator that allows acyl-CoA entry into mitochondrial matrix space only when it is needed. Regulation of malonyl-CoA levels by AMPK does not seem to work as a master on–off switch, but rather as a modulator of fatty acid import.     

Structural and functional organization of the upper esophageal sphincter

Using traditional anatomical and histological methods, the muscle envelope of the pharynx-esophagus junction was investigated in humans and dogs. In the upper (cranial) portion of the esophagus of man and dogs, an inferior anatomical sphincter was detected which histologically can be referred to the group of rhabdo-sphincters. The upper esophageal sphincter is a purely esophageal structure which in man is located at a distance of 25-30 cm from the maxillary incisors. In adult humans, it is 25-30 mm long and is situated obliquely to the long esophageal axis. The posterior semicircle of the sphincter is located higher than the anterior one. In the area of the upper esophageal sphincter the esophageal wall is of different thickness. Due to the muscle envelope and submucous membrane of the base, the right wall is 1.7-2.0 times thicker than the left, anterior or posterior wall. The data obtained from fiber esophagoscopy of patients and electromyography of the pharynx-esophagus junction of dogs have shown that the upper (cranial) esophageal sphincter control food passage from the pharynx to the esophagus and prevents food reflux to the laryngopharynx, protecting airways from aspiration.     

Exploring the Role of Peer Advice in Self-Regulated Learning: Metacognitive,Social, and Environmental Factors

This Conversation Starters article presents a selected research abstract from the 2016 Association of American Medical Colleges Northeast Region Group on Educational Affairs annual spring meeting. The abstract is paired with the integrative commentary of three experts who shared their thoughts stimulated by the pilot study. These thoughts explore the metacognitive, social, and environmental mechanisms whereby advice plays a role in self-regulated learning.     

Recombinant interferon gamma in hairy cell leukemia, multiple myeloma, and Waldenstrom's macroglobulinemia

Fifteen patients with multiple myeloma, five with hairy cell leukemia, and five with Waldenstrom's macroglobulinemia were treated with recombinant interferon gamma (rINF-gamma) to determine the antitumor activity of this agent. The rIFN-gamma was administered by daily intramuscular injection at doses ranging from 0.125 to 0.5 mg/m2. No responses were observed in patients with multiple myeloma, although in one patient the disease has remained stable for over 16 months. Minimal improvement in some hematologic indexes were observed in three of five patients with hairy cell leukemia. One partial remission and one minor response were documented in two of the five patients with Waldenstrom's macroglobulinemia. In five patients, an increase in normal serum immunoglobulins was observed. These results suggest that there is only minimal activity of rIFN-gamma as a single agent in neoplasms of B-cell origin.     

Metabolic control of contractile performance in isolated perfused rat heart. Analysis of experimental data by reaction:diffusion mathematical model

The intracellular mechanisms of regulation of energy fluxes and respiration in contracting heart cells were studied. For this, we investigated the workload dependencies of the rate of oxygen consumption and metabolic parameters in Langendorff-perfused isolated rat hearts.(31)P NMR spectroscopy was used to study the metabolic changes during transition from perfusion with glucose to that with pyruvate with and without active creatine kinase system. The experimental results showed that transition from perfusion with glucose to that with pyruvate increased the phosphocreatine content and stability of its level at increased workloads. Inhibition of creatine kinase reaction by 15-min infusion of iodoacetamide decreased the maximal developed tension and respiration rates by a factor of two.(31)P NMR data were analyzed by a mathematical model of compartmentalized energy transfer, which is independent from the restrictions of the classical concept of creatine kinase equilibrium. The analysis of experimental data by this model shows that metabolic stability-constant levels of phosphocreatine, ATP and inorganic phosphate-at increased energy fluxes is an inherent property of the compartmentalized system. This explains the observed substrate specificity by changes in mitochondrial membrane potential. The decreased maximal respiration rate and maximal work output of the heart with inhibited creatine kinase is well explained by the rise in myoplasmic ADP concentration. This activates the adenylate kinase reaction in the myofibrillar space and in the mitochondria to fulfil the energy transfer and signal transmission functions, usually performed by creatine kinase. The activity of this system, however, is not sufficient to maintain high enough energy fluxes. Therefore, there is a kinetic explanation for the decreased maximal respiration rate of the heart with inhibited creatine kinase: i.e. a kinetically induced switch from an efficient energy transfer pathway (PCr-CK system) to a non-efficient one (myokinase pathway) within the energy transfer network of the cell under conditions of low apparent affinity of mitochondria to ADP in vivo. This may result in a significant decrease in the thermodynamic affinity of compartmentalized ATPase systems and finally in heart failure.     

Intracellular energetic units in healthy and diseased hearts

BACKGROUND:

The present review examines the role of intra-cellular compartmentation of energy metabolism in vivo.

OBJECTIVE:

To compare the kinetics of the activation of mitochondrial respiration in skinned cardiac fibres by exogenous and endogenous adenine nucleotides in dependence of the modulation of cellular structure and contraction.

METHODS:

Saponin-permeabilized cardiac fibres or cells were analyzed using oxygraphy and confocal microscopy.

RESULTS:

Mitochondria respiration in fibres or cells was upregulated by cumulative additions of ADP to the medium with an apparent K_m of 200 μM to 300 μM. When respiration was stimulated by endogenous ADP produced by intracellular ATPases, a near maximum respiration rate was achieved at an ADP concentration of less than 20 μM in the medium. A powerful ADP-consuming system, consisting of pyruvate kinase and phosphoenolpyruvate, that totally suppressed the activation of respiration by exogenous ADP, failed to abolish the stimulation of respiration by endogenous ADP, but did inhibit respiration after the cells were treated with trypsin. The addition of up to 4 μM of free Ca²⁺ to the actively respiring fibres resulted in reversible hypercontraction associated with a decreased apparent K_m for exogenous ADP. These changes were fully abolished in fibres after the removal of myosin by KCl treatment.

CONCLUSIONS:

Mitochondria and ATPases, together with cytoskeletal proteins that establish the structural links between mitochondria and sarcomeres, form complexes – intracellular energetic units (ICEUs) – in cardiac cells. Within the ICEUs, the mitochondria and ATPases interact via specialized energy transfer systems, such as the creatine kinase- and adenylate kinase-phosphotransfer networks, and direct ATP channelling. Disintegration of the structure and function of ICEUs results in dyscompartmentation of adenine nucleotides and may represent a basis for cardiac diseases.     

Cardioprotection by ischemic preconditioning preserves mitochondrial function and functional coupling between adenine nucleotide translocase and creatine kinase

M. N. Laclau, S. Boudina, J. B. Thambo, L. Tariosse, G. Gouverneur, S. Bonoron-Adèle, V. A. Saks, K. D. Garlid and P. Dos Santos. Cardioprotection by Ischemic Preconditioning Preserves Mitochondrial Function and Functional Coupling Between Adenine Nucleotide Translocase and Creatine Kinase. Journal of Molecular and Cellular Cardiology (2001) 33, 947-956. This study investigates the effect of ischemic preconditioning on mitochondrial function, including functional coupling between the adenine nucleotide translocase and mitochondrial creatine kinase, which is among the first reactions to be altered in ischemia. Three groups of Langendorff-perfused rat hearts were studied: a control group, a group subjected to 30 min ischemia followed by 15 min reperfusion, and a group subjected to ischemic preconditioning prior to 30 min ischemia and 15 min reperfusion. Ischemic preconditioning significantly delayed the onset and amplitude of contracture during ischemia, decreased enzymatic release, and improved the recovery of heart contractile function after reperfusion. Mitochondrial function was assessed in permeabilized skinned fibers. The protective effect of preconditioning was associated with preservation of mitochondrial function, as evidenced by maintenance of the high K(1/2)for ADP in regulation of mitochondrial respiration and V(max)of respiration, the near absence of respiratory stimulation by exogenous cytochrome c, and preservation of functional coupling between mitochondrial creatine kinase and adenine nucleotide translocase. These data suggest that ischemic preconditioning preserves the structure-function of the intermembrane space, perhaps by opening the mitochondrial ATP-sensitive K(+)channel. The consequence is preservation of energy transfer processes from mitochondria to ATP-utilizing sites in the cytosol. Both of these factors may contribute to cardioprotection and better functional recovery of preconditioned hearts.     

Tubulin binding blocks mitochondrial voltage-dependent anion channel and regulates respiration

Regulation of mitochondrial outer membrane (MOM) permeability has dual importance: in normal metabolite and energy exchange between mitochondria and cytoplasm and thus in control of respiration, and in apoptosis by release of apoptogenic factors into the cytosol. However, the mechanism of this regulation, dependent on the voltage-dependent anion channel (VDAC), the major channel of MOM, remains controversial. A long-standing puzzle is that in permeabilized cells, adenine nucleotide translocase (ANT) is less accessible to cytosolic ADP than in isolated mitochondria. We solve this puzzle by finding a missing player in the regulation of MOM permeability: the cytoskeletal protein tubulin. We show that nanomolar concentrations of dimeric tubulin induce voltage-sensitive reversible closure of VDAC reconstituted into planar phospholipid membranes. Tubulin strikingly increases VDAC voltage sensitivity and at physiological salt conditions could induce VDAC closure at <10 mV transmembrane potentials. Experiments with isolated mitochondria confirm these findings. Tubulin added to isolated mitochondria decreases ADP availability to ANT, partially restoring the low MOM permeability (high apparent K_m for ADP) found in permeabilized cells. Our findings suggest a previously unknown mechanism of regulation of mitochondrial energetics, governed by VDAC and tubulin at the mitochondria–cytosol interface. This tubulin–VDAC interaction requires tubulin anionic C-terminal tail (CTT) peptides. The significance of this interaction may be reflected in the evolutionary conservation of length and anionic charge in CTT throughout eukaryotes, despite wide changes in the exact sequence. Additionally, tubulins that have lost significant length or anionic character are only found in cells that do not have mitochondria.     

Results of prospective cytological diagnoses of cervical intraepithelial neoplasia

The value of prospective cytologic diagnosis in cervical intraepithelial neoplasia (CIN) is analysed by means of ROC (receiver operating characteristic) curves. Examined was a group of 160 women with subsequent cone biopsies. The result of a prospective cytologic diagnosis in moderate dysplasia, severe dysplasia and carcinoma in situ is analysed separately, taking into consideration the particular frequencies in the studied group. The most reliable prospective cytologic diagnosis is in moderate dysplasia, followed by carcinoma in situ a slightly more reliable prospective diagnosis than that in severe dysplasia.