scholarly journals Longer-term regulation of pyruvate dehydrogenase kinase in cultured rat cardiac myocytes

1990 ◽  
Vol 267 (1) ◽  
pp. 245-247 ◽  
Author(s):  
D R Marchington ◽  
A L Kerbey ◽  
P J Randle
Keyword(s):  
Cyclic Amp ◽  
Cardiac Myocytes ◽  
Pyruvate Dehydrogenase ◽  
Kinase Activity ◽  
Pyruvate Dehydrogenase Kinase ◽  
Dibutyryl Cyclic Amp ◽  
Rat Cardiac Myocytes ◽  
Increased Activity

The increased activity of pyruvate dehydrogenase (PDH) kinase induced in hearts of rats by starvation for 48 h was maintained following preparation of cardiac myocytes, and it was also maintained, though at a decreased level, after 25 h of culture in medium 199. This loss of PDH kinase activity was not prevented by n-octanoate, dibutyryl cyclic AMP or glucagon. The PDH kinase activity of myocytes from fed rats was increased to that of starved rats after 25 h of culture with n-octanoate, dibutyryl cyclic AMP or both agents together.

scholarly journals Role of protein synthesis and of fatty acid metabolism in the longer-term regulation of pyruvate dehydrogenase kinase

1994 ◽  
Vol 300 (3) ◽  
pp. 659-664 ◽  
Author(s):  
D A Priestman ◽  
S C Mistry ◽  
A Halsall ◽  
P J Randle
Keyword(s):  
Cyclic Amp ◽  
Rat Liver ◽  
Pyruvate Dehydrogenase ◽  
Rat Heart ◽  
Kinase Activity ◽  
Specific Activity ◽  
Pyruvate Dehydrogenase Kinase ◽  
Rat Liver Mitochondria ◽  
Dibutyryl Cyclic Amp ◽  
Alpha Chain

Antibodies were raised in rabbits to free rat liver pyruvate dehydrogenase (PDH) kinase alpha-chain and shown to react with PDH kinase alpha-chain in rat heart and liver PDH complexes, in purified pig heart PDH complex and in bovine kidney dihydrolipoamide acetyltransferase-protein X-PDH kinase subcomplex. E.l.i.s.a for PDHE1 (pyruvate dehydrogenase) and PDH kinase have been developed and applied to assays of these proteins in extracts of rat liver and rat heart mitochondria; the measured immunoreactivities for PDHE1 (heart > liver) and for PDH kinase alpha-chain (liver > heart) paralleled known differences in PDH complex and PDH kinase activities respectively. The results of e.l.i.s.a of PDH kinase alpha-chain in extracts of rat liver mitochondria showed that the effects of starvation to increase PDH kinase activity in vivo, and the effects of dibutyryl cyclic AMP or palmitate to increase PDH kinase activity in hepatocytes cultured in vitro, are due largely (> 90%) to an increase in the specific activity of PDH kinase. The effect, in cultured hepatocytes, of dibutyryl cyclic AMP to increase PDH kinase activity was blocked by cycloheximide; the effect of palmitate was blocked by an inhibitor of carnitine palmitoyltransferase I (Etomoxir), but not by cycloheximide.

scholarly journals Longer-term regulation of pyruvate dehydrogenase kinase in cultured rat hepatocytes

1989 ◽  
Vol 257 (2) ◽  
pp. 487-491 ◽  
Author(s):  
D R Marchington ◽  
A L Kerbey ◽  
M G Giardina ◽  
A E Jones ◽  
P J Randle
Keyword(s):  
Fatty Acids ◽  
Cyclic Amp ◽  
Pyruvate Dehydrogenase ◽  
Kinase Activity ◽  
Rat Hepatocytes ◽  
Pyruvate Dehydrogenase Kinase ◽  
Activator Protein ◽  
Cultured Rat Hepatocytes ◽  
Culture Of Hepatocytes

The activities of pyruvate dehydrogenase (PDH) kinase and of PDH kinase activator protein (KAP) were increased 2-2.4-fold during 25 h of culture of hepatocytes from fed rats with glucagon plus n-octanoate. PDH kinase activity in hepatocytes from starved rats (initially 2.2 x fed control) fell during 25 h of culture in medium 199 (to 1.5 x fed control), but was maintained by glucagon plus octanoate. Dibutyryl or 8-bromo cyclic AMP increased PDH kinase activity 2-2.2-fold in hepatocytes from fed rats, but phenylephrine and isoproterenol (isoprenaline) were without effect. Insulin blocked the action of glucagon to increase PDH kinase activity and decreased the effect of octanoate and octanoate plus glucagon. It is suggested that the effects of starvation to increase activities of PDH kinase and of KAP in liver are mediated by alterations in circulating concentrations of glucagon, fatty acids and insulin and in hepatic cyclic AMP.

Inhibition of pyruvate dehydrogenase kinase by pyruvate in cultured cardiac myocytes

1997 ◽  
Vol 25 (1) ◽  
pp. 101S-101S ◽  
Author(s):  
DAVID A. PRIESTMAN ◽  
KAREN A. ORFALI ◽  
MARY C. SUGDEN
Keyword(s):  
Cardiac Myocytes ◽  
Pyruvate Dehydrogenase ◽  
Pyruvate Dehydrogenase Kinase

Abstract 331: Recovery of Mitochondrial Bioenergetics after Hypoxia via Regulating Pyruvate Dehydrogenase Kinase and Adenosine Monophosphate-activated Kinase

2014 ◽  
Vol 115 (suppl_1) ◽  
Author(s):  
Jessica M Toli ◽  
Minzhen He ◽  
Carolyn Suzuki ◽  
Maha Abdellatif
Keyword(s):  
Fatty Acid ◽  
Oxidative Phosphorylation ◽  
Cardiac Myocytes ◽  
Pyruvate Dehydrogenase ◽  
Pyruvate Dehydrogenase Kinase ◽  
Neonatal Rat ◽  
Acid Oxidation ◽  
Peroxisome Proliferator ◽  
Mitochondrial Bioenergetics ◽  
Peroxisome Proliferator Activated Receptor

Mitochondrial quality control is critical for the survival of cardiac myocytes during stress. The purpose of this study was to examine the effect of metabolic substrates and regulators of metabolism on mitochondrial bioenergetics, as an indicator of mitochondrial quality, and how these factors might influence the recovery of the cell’s bioenergetics after hypoxia/ischemia. By monitoring oxygen consumption rates (OCR), in real-time, in live neonatal rat myocytes and human cardiac myocyte-differentiated induced pluripotent stem cells, we found that both cell types can maintain basal OCR efficiently with any metabolic substrate; however, the neonatal cells require both glucose and fatty acid, while the human adult cells require fatty acid only, for mounting maximum reserve respiratory capacity (RRC). Our data also show that subjecting cardiac myocytes to hypoxia results in a reduction of the cells’ basal OCR and oxidative phosphorylation, and exhausts the RRC, which is accompanied by an increase in pyruvate dehydrogenase kinase (Pdk) 1 and 4. Except for normalization of Pdk1 levels, there was little or no recovery of these parameters after reoxygenation. We, thus, hypothesized, that inhibition of Pdks may help recovery of the cell’s bioenergetics. Indeed, our results show that by inhibiting Pdks with dichloroacetate (DCA) before or after hypoxia, the cells’ bioenergetics, including OCR, oxidative phosphorylation, and RRC in neonatal myocytes, and RRC in the human myocytes fully recover within 24 h. On the other hand, activating AMP-activated kinase (AMPK) resulted in delayed (96 h) improvement of the cells’ RRC that was accompanied by an increase in peroxisome proliferator-activated receptor gamma, coactivator 1α (3.5x), peroxisome proliferator-activated receptor-α (2x), and mitochondrial number (2x). These results led us to conclude that compromised mitochondrial quality can be rescued through mechanisms that regulate glucose or fatty acid oxidation by either inhibiting Pdks or activating AMPK, respectively, in rodent and human myocytes.

Increased hepatic pyruvate dehydrogenase kinase activity in fed hyperthyroid rats: studies in vivo and with cultured hepatocytes

1996 ◽  
Vol 119 (2) ◽  
pp. 219-224 ◽  
Author(s):  
Mary C. Sugden ◽  
Lee G.D. Fryer ◽  
David A. Priestman ◽  
Karen A. Orfali ◽  
Mark J. Holness
Keyword(s):  
Pyruvate Dehydrogenase ◽  
Kinase Activity ◽  
Pyruvate Dehydrogenase Kinase ◽  
Cultured Hepatocytes

scholarly journals Receptor-mediated gonadotropin action in the ovary. Regulatory role of cyclic nucleotide phosphodiesterase(s) in intracellular adenosine 3′:5′-cyclic monophosphate turnover and gonadotropin-stimulated progesterone production by rat ovarian cells

1979 ◽  
Vol 180 (1) ◽  
pp. 201-211 ◽  
Author(s):  
Salman Azhar ◽  
K. M. Jairam Menon
Keyword(s):  
Cyclic Amp ◽  
Kinase Activity ◽  
Phosphodiesterase Inhibitors ◽  
Protein Kinase Activity ◽  
Dibutyryl Cyclic Amp ◽  
Steroid Production ◽  
Progesterone Production ◽  
Ovarian Cells ◽  
Cyclic Amp Accumulation

The regulatory role of cyclic nucleotide phosphodiesterase(s) and cyclic AMP metabolism in relation to progesterone production by gonadotropins has been studied in isolated rat ovarian cells. Low concentrations of choriogonadotropin (0.4–5ng/ml) increased steroid production without any detectable increase in cyclic AMP, when experiments were carried out in the absence of phosphodiesterase inhibitors. The concentration of choriogonadotropin (10ng/ml) that stimulated progesterone synthesis maximally resulted in a minimal increase in cyclic AMP accumulation and choriogonadotropin binding. Choriogonadotropin at a concentration of 10ng/ml and higher, however, significantly stimulated protein kinase activity and reached a maximum between 250 and 1000ng of hormone/ml. Higher concentrations (50–2500ng/ml) of choriogonadotropin caused an increase in endogenous cyclic AMP, and this increase preceded the increase in steroid synthesis. Analysis of dose–response relationships of gonadotropin-stimulated cyclic AMP accumulation, progesterone production and protein kinase activity revealed a correlation between these responses over a wide concentration range when experiments were performed in the presence of 3-isobutyl-1-methylxanthine. The phosphodiesterase inhibitors papaverine, theophylline and 3-isobutyl-1-methylxanthine each stimulated steroid production in a dose-dependent manner. Incubation of ovarian cells with dibutyryl cyclic AMP or 8-bromo cyclic AMP mimicked the steroidogenic action of gonadotropins and this effect was dependent on both incubation time and nucleotide concentration. Maximum stimulation was obtained with 2mm-dibutyryl cyclic AMP and 8-bromo cyclic AMP, and this increase was close to that produced by a maximally stimulating dose of choriogonadotropin. Other 8-substituted derivatives such as 8-hydroxy cyclic AMP and 8-isopropylthio cyclic AMP, which were less susceptible to phosphodiesterase action, also effectively stimulated steroidogenesis. The uptake and metabolism of cyclic [3H]AMP in ovarian cells was also studied in relation to steroidogenesis. When ovarian cells were incubated for 2h in the presence of increasing concentrations of cyclic [3H]AMP, the radioactivity associated with the cells increased almost linearly up to 250μm-cyclic [3H]AMP concentration in the incubation medium. The 3H label in the cellular extract was recovered mainly in the forms ATP, ADP, AMP, adenosine and inosine, with cyclic AMP accounting for less than 1% of the total tissue radioactivity. Incubation of cyclic AMP in vitro with ovarian cells resulted in a rapid breakdown of the nucleotide in the medium. The degradation products in the medium have been identified as AMP, adenosine and inosine. The rapid degradation of cyclic AMP by phosphodiesterase(s) makes it difficult to correlate changes in cyclic AMP concentrations with steroidogenesis. These observations thus provide an explanation for the previously observed lack of cyclic AMP accumulation under conditions in which low doses of choriogonadotropin stimulated steroidogenesis without any detectable changes in cyclic AMP accumulation.

Increased pyruvate dehydrogenase kinase activity in response to sepsis

1991 ◽  
Vol 260 (5) ◽  
pp. E669-E674 ◽  
Author(s):  
T. C. Vary
Keyword(s):  
Skeletal Muscle ◽  
Pyruvate Dehydrogenase ◽  
Kinase Activity ◽  
Pyruvate Dehydrogenase Complex ◽  
Pyruvate Dehydrogenase Kinase ◽  
Sterile Inflammation ◽  
Stable Factor ◽  
Skeletal Muscle Mitochondria ◽  
Dependent Inactivation ◽  
Significant Difference

The effect of sterile inflammation and sepsis on the proportion of active pyruvate dehydrogenase complex (PDH) in mitochondria isolated from skeletal muscle has been investigated. The proportion of active PDH in mitochondria isolated from septic animals was significantly reduced compared with control under all incubation conditions examined, even in the presence of inhibitors of the PDH kinase. There was no significant difference between control and sterile inflammation in any of the incubations examined. The rate constant for ATP-dependent inactivation of the PDH complex in mitochondrial extracts from control animals was -0.42 min-1 (r = 0.993; P less than 0.001) and was not altered in mitochondrial extracts from sterile inflammatory animals (-0.43 min-1; r = 0.999; P less than 0.001). However, rate constants for inactivation in septic animals was significantly increased over twofold to -1.08 min-1 (r = 0.987; P less than 0.001) (P less than 0.001 vs. control or sterile inflammation). In the presence of inhibitors of the PDH kinase reaction (2.5 mM pyruvate or 1 mM dichloroacetate), inactivation of PDH after addition of ATP was significantly greater in mitochondrial extracts from septic than either control or sterile inflammatory animals. These results suggest that sepsis, but not sterile inflammation, induces a stable factor in skeletal muscle mitochondria that increased PDH kinase activity.

scholarly journals Interaction between the individual isoenzymes of pyruvate dehydrogenase kinase and the inner lipoyl-bearing domain of transacetylase component of pyruvate dehydrogenase complex

2002 ◽  
Vol 366 (1) ◽  
pp. 129-136 ◽  
Author(s):  
Alina TUGANOVA ◽  
Igor BOULATNIKOV ◽  
Kirill M. POPOV
Keyword(s):  
Pyruvate Dehydrogenase ◽  
Kinase Activity ◽  
Pyruvate Dehydrogenase Complex ◽  
Tight Binding ◽  
Enzymic Activity ◽  
Pyruvate Dehydrogenase Kinase ◽  
Size Exclusion ◽  
Apparent Dissociation Constant ◽  
The Individual ◽  
Dehydrogenase Complex

Protein—protein interactions play an important role in the regulation of enzymic activity of pyruvate dehydrogenase kinase (PDK). It is generally believed that the binding of PDK to the inner lipoyl-bearing domain L2 of the transacetylase component E2 of pyruvate dehydrogenase complex largely determines the level of kinase activity. In the present study, we characterized the interaction between the individual isoenzymes of PDK (PDK1—PDK4) and monomeric L2 domain of human E2, as well as the effect of this interaction on kinase activity. It was found that PDK isoenzymes are markedly different with respect to their affinities for L2. PDK3 demonstrated a very tight binding, which persisted during isolation of PDK3—L2 complexes using size-exclusion chromatography. Binding of PDK1 and PDK2 was readily reversible with the apparent dissociation constant of approx. 10μM for both isoenzymes. PDK4 had a greatly reduced capacity for L2 binding (relative order PDK3>PDK1 = PDK2>PDK4). Monomeric L2 domain alone had very little effect on the activities of either PDK1 or PDK2. In contrast, L2 caused a 3-fold increase in PDK3 activity and approx. 37% increase in PDK4 activity. These results strongly suggest that the interactions between the individual isoenzymes of PDK and L2 domain are isoenzyme-specific and might be among the major factors that determine the level of kinase activity of particular isoenzyme towards the pyruvate dehydrogenase complex.

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