Marie-Louise Dubelaar

The microheterogeneity of human transferrins in biological fluids

The serum, cerebrospinal fluid, amniotic fluid and synovial fluid transferrins have been examined by isoelectric focusing in a pH gradient from 5-7. In all biological fluids the presence of transferrins with a varying content of sialic acid has been shown. Differences between the fluids with regard to transferrin have been noticed.

Aspects of fatty acid metabolism in vascular endothelial cells

Long-chain fatty acids are an important source of energy in vascular endothelium. Their oxidation is stimulated by carnitine and inhibited by blockage of the mitochondrial respiratory chain. Excess fatty acid can be reversibly stored as triacylglycerol in the cells. Cultured vascular endothelial cells, in contrast to cardiac vascular endothelium in the intact heart, take up and intracellularly degrade artificial chylomicrons (intralipid enriched with apolipoprotein C-II) but not natural chylomicrons. Fatty acids not bound to albumin, such as those generated from chylomicrons in the lipoprotein lipase reaction, although initially a good source of substrate for beta-oxidation, endanger heart function. Fatty acid excess initiates the breakdown of the endothelial barrier between the vascular lumen and interstitium; it may precipitate edema formation, lead to insufficient oxygenation and finally cause loss of heart function.

Effects of diets supplemented with lard fat or mackerel oil on plasma lipoprotein lipid concentrations and lipoprotein lipase activities in domestic swine

Levels of plasma lipoproteins and lipoprotein lipase activities in post-heparin serum were measured in 24-h fasted pigs which were fed a diet containing either 21 energy % mackerel oil or 21 energy % lard fat for 8 weeks. Lipoprotein fractionation was performed separately by density gradient ultracentrifugation and agarose gel chromatography. After 8 weeks levels of plasma triacylglycerol (-62%) and cholesterol (-55%) were lower in the mackerel oil than in the lard fat-fed animals. The triacylglycerol decline was exclusively due to the VLDL fraction, while cholesterol was reduced in all lipoprotein fractions (VLDL, IDL, LDL and HDL). Lipoprotein lipase activity in post-heparin serum, taken 6 h after a meal, was 31% decreased in mackerel oil-fed animals. The results support the hypothesis that regular intake of fish oil reduces VLDL secretion.

The source of malonyl-CoA in rat heart. The calcium paradox releases acetyl-CoA carboxylase and not propionyl-CoA carboxylase.

The formation of malonyl-CoA in rat heart is catalyzed by cytosolic acetyl-CoA carboxylase. The existence of this enzyme in heart is difficult to prove by the abundant occurrence of mitochondrial propionyl-CoA carboxylase, which is also able to catalyze the carboxylation of acetyl-CoA. We used the calcium paradox as a tool to separate cytosolic components from the remaining heart, and found that acetyl-CoA carboxylase activity was preferentially released, like lactate dehydrogenase and carnitine, while propionyl-CoA carboxylase was almost fully retained. Acetyl-CoA carboxylase activity was determined after activation by citrate ion and Mg2+. The activity decreased to 64% by 48 h of fasting.The formation of malonyl‐CoA in rat heart is catalyzed by cytosolic acetyl‐CoA carboxylase. The existence of this enzyme in heart is difficult to prove by the abundant occurrence of mitochondrial propionyl‐CoA carboxylase, which is also able to catalyze the carboxylation of acetyl‐CoA. We used the calcium paradox as a tool to separate cytosolic components from the remaining heart, and found that acetyl‐CoA carboxylase activity was preferentially released, like lactate dehydrogenase and carnitine, while propionyl‐CoA carboxylase was almost fully retained. Acetyl‐CoA carboxylase activity was determined after activation by citrate ion and Mg2+. The activity decreased to 64% by 48 h of fasting.

Protection by acyl-carnitines and phenylmethylsulfonyl fluoride of rat heart subjected to ischemia and reperfusion

Perfusion of rat hearts according to the Langendorff technique with micromolar concentrations of palmitoylcarnitine or millimolar concentrations of phenylmethylsulfonyl fluoride protect the heart from deterioration by reperfusion after total-ischemia. This is based on the retention of the cytosolic enzymes determined (lactate dehydrogenase, glycogen phosphorylase and glycogen synthase) and of myoglobin, as well as on the resumption of contractile activity. Palmitoylcarnitine, like phenylmethylsulfonyl fluoride, could protect through plasma membrane stabilization, since more hydrophilic compounds had no effect.

Cardiac lipoprotein lipase: effects of lipopolysaccharide and tumor necrosis factor

SummaryLipopolysaccharide (LPS), the active principle of certain endotoxins, protein-free perfused in rat hearts leads in 3 h to a considerable loss of lipoprotein lipase (LPL) activity. In the presence of albumin LPS has virtually no effect. Tumor necrosis factor (TNF) added instead of LPS had no effects on LPL activity during 3 hin vitro perfusion.LPS injected into rats intravenously leads within 3 h to severe toxic phenomena amongst which increased capillary permeability. This was visualized as increased rate of interstitial fluid formation in Langendorff hearts mounted 3 h after rats had been treated with LPS. LPL activity did not decline in 3 h lasting endotoxemia. Six hours after LPS injection, however, cardiac LPL activity was considerably lowered, although immunoblotting and immunohistochemistry still showed LPL protein to be present. These date indicate the presence of a considerable pool of inactive LPL protein in addition to active LPL, that can be released in the presence of heparin. The LPL activity is lowered by LPS injection after a lag phase of at least 3 h, while capillary endothelial cells are influenced more rapidly. The relatively late expression of TNF toxicity in cardiomyocytes of the intact heart is discussed.

Lipoprotein lipases and stress hormones: studies with glucocorticoids and cholera toxin

The intravenous injection of cholera toxin in rats 17 h prior to experimentation results in increased levels of insulin and corticosterone in the blood. This is accompanied by a rise in lipoprotein lipase activity in muscle and a decrease in adipose tissue. Pre- and postheparin blood levels of the enzyme are increased, representing the higher overall muscle activity. Hepatic lipase is decreased by cholera toxin treatment. These enzyme changes are accompanied by increased levels of non-esterified fatty acids, ketone bodies and unesterified cholesterol in the blood, whereas triacylglycerol levels are lowered. The lipoprotein triacylglycerol secretion is not affected by cholera toxin, suggesting increased triacylglycerol removal from the blood. On the other hand the unesterified cholesterol removal may be decreased due to the decreased hepatic lipase activity. Administration of excess glucocorticoid 2 days prior to blood and tissue sampling also resulted in a rise in lipoprotein lipase, a decrease in hepatic lipase activity and an increase of non-esterified fatty acids. In contrast to the effect of cholera toxin, the triacylglycerol levels were increased. Adrenalectomy, whether by inhibition of 11-beta-steroid hydroxylase or by surgical intervention, did not abolish the choleratoxin effects. It is concluded that corticosterone increase is not essential to the cholera toxin effects. Corticosterone itself probably causes an increase of cyclic AMP and/or Ca2+ levels, as is discussed.

The effect of L-carnitine on force development of the latissimus dorsi muscle in dogs.

Using the latissimus dorsi (LD) muscle of the dog in situ, the effect of carnitine was tested for increase of force in the first period after stimulation. Carnitine administration resulted in an increase of force of 31 ± 6% (mean ± SEM). It is hypothesized that, during muscle stimulation, a relative carnitine deficiency occurs in cells of the vascular compartment. The previously observed lesser effect of carnitine in the trained muscle than in the untrained muscle is in line with this hypothesis, since the number of capillaries is known to increase by training. Also in agreement with this hypothesis is the observation that carnitine increased flow during exercise of the muscle.

Effects of tumor necrosis factor (TNF) on lipolytic activities of rat heart

SummaryTumor Necrosis Factor (TNF) inhibits lipoprotein lipase activity in cultured myocytes and in the Langendorff rat heart after 3 h perfusion with TNF of glucocorticoid-pretreated rats. TNF acutely stimulates glyc(ogen)olysis and concomitantly endogenous lipolysis. The latter was significantly increased only when rats had been pretreated with glucocorticoid or fed a trierucate-rich diet. Under these conditions, contractile activity of the Langendorff hearts was acutely increased by TNF The mechanism of the actue increase of contractile function and the accompanying increased glycolytic and lipolytic activities, by TNF, may be explained by increased cytosolic Ca2+ and cAMP levels.

Substrates for energy metabolism in the heart: the role of the interstitial compartment

Evidence is presented that, as in cardiomyocytes, vascular endothelial cells use fatty acids, in addition to glucose, as a respiratory fuel. Attention is focused on the cardiac interstitium, lined by vascular cells and cardiomyocytes, which may be enriched with metabolic products from these cells. Also, certain proteins are present in the interstitial fluid (Qi) such as plasma proteins and fatty acid binding protein (FABP). However, the concentration of FABP is so low in Qi that albumin is more important to shuttle long chain fatty acids in the interstitial fluid between cardiomyocytes and the vascular compartment. Under hypoxic conditions (hypo)xanthine, lactate and fatty acids may be expected to accumulate in the interstitium, as well as proteins from adjacent cells, such as xanthine oxidase from endothelial cells. This enzyme, acting upon the elevated level of (hypo)xanthine, giving rise to O2-., may be involved in the damage of the ischaemic heart. The significance of the interstitium in ischaemia and in fibrosis following long standing cardiac lipidosis is briefly discussed, as well as the possible mechanisms involved in fatty acid transport in the heart.