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L Abas

Publications and source records attributed to L Abas.

7 recordsLinked to original sources
PubMed2000

Mitochondrial ATP production is necessary for activation of the extracellular-signal-regulated kinases during ischaemia/reperfusion in rat myocyte-derived H9c2 cells.

To search for the stimuli involved in activating the mitogen-activated protein kinases (MAPKs) during ischaemia and reperfusion, we simulated the event in a system in vitro conducive to continuous and non-invasive measurements of several major perturbations that occur at the time: O(2) tension, mitochondrial respiration and energy status. Using H9c2 cells (a clonal line derived from rat heart), we found that activation of the extracellular signal-regulated MAPKs (ERKs) on reoxygenation was abolished if the mitochondria were inhibited prior to and during reoxygenation. Re-introduction of O(2) per se is therefore not sufficient to activate the ERKs. Recovery and maintenance of cellular ATP levels by mitochondrial respiration is necessary, although ATP recovery alone is not sufficient. ERK activation by H(2)O(2), but not phorbol esters, was also sensitive to mitochondrial inhibition. Thus, reoxygenation and H(2)O(2)-mediated oxidative stress share a mechanism of ERK activation that is ATP- or mitochondrion-dependent, and this common feature suggests that the reoxygenation response is mediated by reactive oxygen species. A correlation between ERK activity and ATP levels was also found during the anoxic phase of ischaemia, an effect that was not due to substrate limitation for the kinases. Our results reveal the importance of cellular metabolism in ERK activation, and introduce ATP as a novel participant in the mechanisms underlying the ERK cascade.

Adenosine Triphosphate↗
PubMed2000

Intact mitochondrial electron transport function is essential for signalling by hydrogen peroxide in cardiac myocytes.

Oxidative stress has been proposed as a mediator of cardiac injury during ischemia and reperfusion. We examined the signalling events initiated by short-term exposure of cardiac myocytes to oxidative stress elicited by hydrogen peroxide. A potent stimulation of tyrosine phosphorylation was observed within 1 to 2 min exposure to 1 m m hydrogen peroxide. Within 5 min, the ERK mitogen-activated protein kinases (ERK MAPKs) were activated. This activation of ERK MAPKs was blocked by N-acetylcysteine (NAC), implicating a role for free radicals in the signalling events. NAC failed to inhibit ERK MAPK activation by the hypertrophic agent, phenylephrine, or hyperosmotic shock. Myxothiazol, an inhibitor of complex III of the mitochondrial electron transport chain, also inhibited ERK MAPK activation by hydrogen peroxide, but not by 12- O -tetradecanoylphorbol-13-acetate (TPA) or hyperosmotic shock. Myxothiazol completely inhibited the increase in tyrosine phosphorylated proteins observed with hydrogen peroxide treatment. A variety of inhibitors which act at different levels of the mitochondrial electron transport chain (rotenone, theonyltrifluoroacetone, antimycin A, cyanide) also inhibited activation of the ERK MAPKs by hydrogen peroxide but not TPA or hyperosmotic shock. These studies suggest a novel mechanism of regulation of the ERK MAPK pathway and oxidative stress signalling by hydrogen peroxide.

Acetylcysteine↗
PubMed1997

Fuel choices by human platelets in human plasma.

Despite the fact that homogeneous preparations of isolated cells are now being used very effectively to study a range of important biochemical questions, it is still not known what combination of fuels and energy-producing pathways is used by cells when offered the complex mixture characteristic of plasma or extracellular fluid. We have developed an in vitro system whereby highly purified and functional human platelets are incubated in human plasma that has been minimally modified from its native state. The concentration of platelets and fuels, and the complexity of fuels in the incubation are similar to those in vivo. The preparation thus represents a reasonable approximation of the physiological condition, considering the complex nature of the system being studied. Measurements carried out simultaneously during the incubation are rates of oxygen consumption, lactate production and fuel oxidation. The data allow the calculation of total ATP turnover, and contributions to this turnover by lactate production and the oxidation of individual fuels. Lactate production accounts for 24% of the ATP turnover. The oxidation of glucose and 3-hydroxybutyrate each account for under 5%, palmitate for 21%, oleate for 7% and acetate for 9%, leaving 32% of the ATP turnover as yet unaccounted for. The results confirm some previous measurements in the literature, but show that data collected under non-physiological experimental conditions can be misleading.

3-Hydroxybutyric Acid↗
PubMed1996

Carbohydrate metabolism in human platelets in a low glucose medium under aerobic conditions.

The metabolism of human platelets has been the subject of investigation for at least three decades, at the level of basic metabolism, and because of the increasing requirement for platelet storage. Platelets are relatively active metabolically and are typical cells in terms of fuels and metabolic pathways. They contain glycogen and utilize glucose and demonstrate aerobic glycolysis and carbohydrate oxidation. Both glycolysis and carbohydrate oxidation contribute significantly to total ATP turnover, so platelets are an ideal system in which to study the partitioning of carbohydrate metabolism between the two available fuels and the two available pathways, in the presence of adequate oxygen. We have designed a system whereby we can study carbohydrate metabolism in relatively pure human platelets, under sterile conditions, over long periods. The system enables us to determine total ATP turnover and, with the aid of a mathematical model, the contribution to this turnover of glycolysis and the oxidation of glucose/glycogen and lactate. When glucose and glycogen are present, most of the glucose and glycogen utilised is converted to lactate, but lactate is being oxidised at this time. When glucose/glycogen stores are exhausted lactate oxidation continues and increases with the result that carbohydrate oxidation accounts for 41% of total ATP turnover over 48 h.

Adenosine Triphosphate↗
PubMed1995

A complete platelet system for metabolic studies: pure, defined, and viable over 8 hours.

We have been using a human platelet preparation to investigate various aspects of cellular metabolism. These experiments require a pure, quantitated platelet preparation that is viable for up to 8 h. The purity of the preparation has previously been characterized and we have now assessed its viability over 8 h and developed a simple spectrophotometric counting method. This method of counting results in estimations that are within 6% of those from an electronic cell counter (Coulter). Pure platelets stored in a Hepes physiological saline were resuspended periodically in autologous plasma to assess function over 24 h. The platelets consumed oxygen, changed shape, aggregated, and released dense granule ATP at all times. There was a slow and linear decrease in the various processes with time, ranging from 1.7% per hour for oxygen consumption to 3.8% per hour for percentage aggregation. The preparation is therefore not stable, but the changes with time are small and defined and it represents a useful system for the study of platelet and cell biology.

Adenosine Triphosphate↗
PubMed1995

The Pasteur effect in human platelets: implications for storage and metabolic control.

The Pasteur effect and the associated acidosis have long been considered a major cause of platelet death during storage. We have investigated this phenomenon using a defined platelet preparation and a system whereby the oxidative and glycolytic contributions to total ATP production can be measured over a range of oxygen concentrations from saturating (pO2 = 158 mmHg) to anoxic (pO2 = 0 mmHg). Platelets do not show a Pasteur effect until the pO2 decreases to < 2.0 mmHg, whereupon lactate production increases 1.5-fold. The Pasteur effect is therefore not a likely cause of platelet death during storage where pO2 in a storage bag typically drops to no less than 50 mmHg. The data also have implications for the role of oxygen diffusion in oxidative metabolism, and for the compensatory nature of the Pasteur effect. As platelets are relatively small cells, and the onset of the Pasteur effect occurs at a relatively low oxygen concentration, diffusion may limit the rate of oxygen consumption in most other (larger) cells. The Pasteur effect is only fully compensative if the P/O2 ratio used for the calculations is lower than the conventional one. Since recent research strongly suggests that the conventional P/O2 ratio is too high, examples of fully compensative Pasteur effects may be more common than the literature suggests.

Adenosine Triphosphate↗