Simon Eaton

Multiple biochemical effects in the pathogenesis of alcoholic fatty liver

The pathogenesis of alcoholic fatty liver is unknown, but several causes have been proposed based on biochemical findings. These include the metabolism of alcohol leading to a shift in the cytosolic [NAD+]/[NADH] ratio to reduction, which in turn causes a direct inhibition of β‐oxidation and enhanced triacylglycerol formation via the [glycerol‐3‐phosphate]/[dihydroxyacetone phosphate] ratio. There are also chronic effects of ethanol on hepatic enzyme activities. Thus, increased activity of phosphatidate phosphohydrolase, an increased amount of fatty acid binding protein, decreased secretion of very low‐density lipoprotein and impairment of the respiratory chain as a result of decreased protein synthesis or decreased amounts of ubiquinone could all lead to fat accumulation and steatosis. The interplay of each of these with nutritional and genetic factors would then lead to the heterogeneity of the severity and characteristics of the steatosis observed in human alcoholics.

Plasma coenzyme Q(10) in children and adolescents undergoing doxorubicin therapy.

The objective of this study was to test the hypothesis that doxorubicin treatment for cancer in childhood and adolescence causes a dose-related decrease in the concentration of plasma coenzyme Q(10). The concentration of plasma coenzyme Q(10) was measured before and after administration of doxorubicin in six patients, and before and after chemotherapy in six patients undergoing treatments that did not include doxorubicin. There was a significant increase in the concentration of plasma coenzyme Q(10) in post-treatment samples compared to pre-treatment samples in patients treated with doxorubicin (P=0.008; n=32), whereas there were no significant changes in plasma coenzyme Q(10) concentrations in patients treated with chemotherapy that did not include doxorubicin. (P=0.770; n=30). We hypothesise that the increase in plasma coenzyme Q(10) that was observed in patients undergoing doxorubicin treatment is due to release of coenzyme Q(10) from apoptotic or necrotic cardiac tissue. We conclude that the cardiotoxicity due to doxorubicin therapy does not involve acute myocardial depletion of coenzyme Q(10).

Control of mitochondrial beta-oxidation: sensitivity of the trifunctional protein to [NAD(+)]/[NADH] and [acetyl-CoA]/[CoA]

Isolated human mitochondrial trifunctional protein was incubated with 2-hexadecenoyl-CoA, CoA and NAD+ and the resultant CoA esters measured. Steady state with respect to the concentrations of the intermediates 3-hydroxyhexadecanoyl-CoA and 3-ketohexadecanoyl-CoA and the rate of formation of the product tetradecanoyl-CoA was reached within 4 min. Flux was greatly enhanced by the addition of Tween 20 (0.2% v/v) which stimulated 3-ketoacyl-CoA thiolase activity by over 7-fold. When 3-ketoacyl-CoA thiolase was not stimulated, 3-hydroxyhexadecanoyl-CoA was the prominent CoA ester accumulated, presumably due to inhibition of 3-hydroxyacyl-CoA dehydrogenase activity by accumulated 3-ketoacyl-CoA, analogous to the inhibition of short-chain 3-hydroxyacyl-CoA dehydrogenase by 3-ketoacyl-CoA. When [NAD+]/[NADH] was varied at a fixed total [NAD++NADH], the overall flux was only inhibited by [NAD+]/[NADH] less than 1. In contrast, when [acetyl-CoA]/[CoA] was varied at a fixed total [CoA], much greater sensitivity was observed.

Intermediates of myocardial mitochondrial β-oxidation: possible channelling of NADH and of CoA esters

Adult rat heart mitochondria were isolated and incubated with [U-14C]hexadecanoyl-CoA or unlabelled hexadecanoyl-CoA. The accumulating CoA and carnitine esters and [NAD+]/[NADH] ratio were measured by HPLC or tandem mass spectrometry. Despite minimal changes in the intramitochondrial [NAD+]/[NADH] ratio, 2, 3-unsaturated and 3-hydroxyacyl esters were observed as well as saturated acyl-CoA and acylcarnitine esters. In addition to acetylcarnitine, significant amounts of butyryl-, hexanoyl-, octanoyl- and decanoylcarnitines were detected and measured. Rat myocardial beta-oxidation is subject to control at the level of 3-hydroxyacyl-CoA dehydrogenase but this control is not due to a simple lack of oxidised NAD. We hypothesise a pool of NAD in contact between the trifunctional protein of beta-oxidation and complex I of the respiratory chain, the turnover of which is responsible for some of the control of beta-oxidation flux. In addition, short- and medium-chain acylcarnitine esters were detected whereas only small amounts of long-chain acylcarnitines were present. This may imply the presence of a mitochondrial carnitine octanoyl transferase or may reflect channelling of long-chain CoA esters so that they are not available for carnitine palmitoyl transferase II activity.

Tissue Specific Differences in Intramitochondrial Control of β-Oxidation

It has become clear that there are important tissue specific differences in the control and regulation of oxidation and that these differences may largely reside with the entry of acyl moieties into the mitochondrion by the carnitine palmitoyltransferase (CPT) /translocase system. CPT I has wide tissue-specific variations in sensitivity to its physiological inhibitor, malonyl-CoA,1 which is probably due to the presence of different isozymes. However, it is possible that there are additional intramitochondrial controls which vary amongst tissues. The individual reactions of the oxidation of long-chain fatty acids within the mitochondrion have been known for many years although their functional and topological relationship is less well known. Recently, it has been found that the long-chain activities of three of the enzymes reside on a single membrane-bound protein (the trifunctional protein,4,5) and, in addition, there exists a fourth acyl-CoA dehydrogenase which is similarly membrane bound. Hence, the possibility of functional organization of oxidation enzyme activities, associated with the inner mitochondrial membrane, must be considered. The organization of oxidation enzymes within the mitochondrion has previously been postulated as a result of the classic studies of Stanley and Tubbs, amongst others, leading to the “leaky hosepipe” model of oxidation. Their work also led to the view that the acyl-CoA dehydrogenases were the “rate-limiting step” for oxidation, as only saturated acyl groups accumulated under well oxygenated conditions. These workers measured acyl groups resulting from hydrolysis of CoA and car-