Effect of ethanol on phospholipid turnover and calcium mobilization in chick embryos

Effects of ethanol on [3H]inositol and [14C]choline incorporation into phosphatidylinositol (PI) and phosphatidylcholine (PC), free intrasynaptosomal Ca2+ ([Ca2+]i) and synaptosomal 45Ca2+ uptake, were investigated in the brain and heart of 17-day-old chick embryos to which a 10% ethanol solution had been injected on the 3rd day of embryogenesis. In brain synaptosomes, ethanol increased the incorporation of [3H]inositol and [14C]choline into PI and PC, increased [Ca2+]i, and decreased 45Ca2+ uptake. On the other hand, in heart synaptosomal membrane, ethanol decreased the incorporation of [3H]inositol and [14C]choline into PI and PC, decreased [Ca2+]i, and increased 45Ca2+ uptake. Ethanol stimulated in vitro [3H]inositol and [14C]choline incorporation into PI and PC in the brain and heart in both the control and ethanol-treated groups. However, addition of ethanol did not affect the release of 45Ca2+ from the synaptosomal membrane of either organ in either group. Addition of ethanol inhibited 45Ca2+ uptake in a dose-dependent manner in the brain but not in the heart. In both organs, there was a relationship between phospholipid turnover and [Ca2+]i after ethanol.     

Phosphatidylcholine metabolism in lung microsomes and lung surfactant of rats exposed intratracheally to coal fly ash

The effect of intratracheal administration of fly ash has been studied on lung microsomal and lung surfactant phosphatidylcholine (PC) metabolism in rats using [methyl-14C]choline and [methyl-14C]methionine. Fly-ash administration significantly increased total phospholipids, PC, phosphatidylethanolamine (PE), and phosphatidylglycerol (PG) of lung surfactant. Fly-ash administration stimulated the formation of lung microsomal PC (as measured by the incorporation of labeled precursors) both by the cytidine 5'-diphosphate (CDP)-choline pathway and by the N-methylation pathway, but this stimulation was fourfold higher in the latter case and only twofold higher in the former as compared to the control. Likewise, the secretion of PC formed by the N-methylation pathway was sixfold higher as compared to the control whereas secretion of PC formed by the CDP-choline pathway was only threefold higher as compared to the control. Fly-ash administration further increased total saturation and decreased unsaturation in PC, PE, and lysophosphatidylcholine (LPC) of lung and in PC, PE, LPC, and PG of lung surfactant as compared to the controls.     

Effect of ethanol on calcium-uptake and phospholipid turnover by stimulation of adrenoceptors and muscarinic receptors in mouse brain and heart synaptosomes

The effect of ethanol treatment on mouse brain and heart synaptosomal 45Ca uptake and the incorporation of [3H]inositol and [14C]choline into phosphatidylinositol (PI) and phosphatidylcholine (PC) were investigated. Ethanol in drinking water (15%) was given to mice for 3 weeks. The consumption of ethanol increased gradually during treatment but food intake was almost the same as control. The body weight of ethanol-treated mice was slightly less than that of control. The synaptosomal lipid peroxidation level of ethanol-treated mice was almost the same as control in the brain and heart. On the other hand, the synaptosomal glutathione level of ethanol-treated mice was higher than control in both brain and heart. The 45Ca uptake of brain and heart from ethanol-treated mice was 87% and 216% of control mice, respectively. Not only ethanol but also norepinephrine (NE), carbachol (Carb), or isoproterenol (IsoPro) added in vitro increased 45Ca uptake in all cases. The incorporation of [3H]inositol into PI in the brain and heart synaptosomes of ethanol-treated mice was 150% and 113% of control, respectively. The incorporation of [14C]choline into PC in the brain and heart of ethanol-treated mice was 104% and 125% of control, respectively. In vitro addition of ethanol, NE, Carb or IsoPro to brain synaptosomes increased the incorporation of [3H]inositol and [14C]choline into PI and PC, respectively, in both control and ethanol-treated mice. In the case of heart synaptosomes, NE and Carb increased the incorporation of [3H]inositol and [14C]choline into phospholipids in control mice but not ethanol-treated mice. However, IsoPro increased the incorporation by both control and ethanol-treated heart synaptosomes. These results suggest that alpha-adrenoceptors and the cholinergic system of the heart play important roles in modulating the toxic effects of ethanol.     

Effect of typical inducers of microsomal drug-metabolizing enzymes on phospholipid metabolism in rat liver.

The effect of administration of phenobarbital (PB), polychlorinated biphenyls(PCBs) or 3-methyl-cholanthrene (3-MC) on the metabolism of phospholipids in rat liver was studied by the i.p. injection of [³²P]orthophosphate or [Me-¹⁴C]choline chloride. The inducers were given to animals for 2 successive days PB had no significant effect on the incorporation rate of ³²Pi into liver microsomal phospholipid classes 48 hr after the first administration. The rate of incorporation of [¹⁴C]choline into phosphatidylcholine (PC) in both subcellular components of the liver and blood plasma decreased slightly at this experimental period. In addition, the ratio of [¹⁴C] sp. act. of phosphorylcholinc to that of microsomal phosphulipid was considerably higher on PB-trcated rats as compared with the ratio in control rats. These and previous findings strongly suggest that proliferation of liver endoplasmic reticulum (ER) membranes induced by PB would be accompanied by the stimulation of phospholipid synthesis at the early process of induction and subsequently followed by the decrease in turnover rate of microsomal phospholipids. The administration of PCBs, on the other hand, caused strong inhibition of both ³²Pi and ¹⁴C incorporation into PC in liver subcellular fractions. The secretion of PC from liver cells to blood plasma was also strongly depressed. In addition, phospholipid catabolizing activity was found to be depressed in the liver. These results indicate that hypertrophy of ER membranes in the liver after PCBs administration could be due to a depression of both secretion of lipoprotein from liver cells to plasma and catabolic activity toward membranous phospholipids in liver cells. The decrease in ³²Pi incorporation into liver microsomal PC was also observed in rats treated with 3-MC. There occurred a considerable accumulation of ¹⁴C activity in phosphorylcholine in the liver of rats treated with either PCBs or 3-MC, suggesting strongly that these drugs caused an inhibition of PC synthesis at the site of CDP-choline formation, namely the inhibition of the reaction catalyzed by cholinephosphate cytidylyltransferase.     

Altered lipid metabolism in 2,5-hexanedione-induced testicular atrophy and peripheral neuropathy in the rat

2,5-Hexanedione (HD) induces testicular atrophy and peripheral neuropathy in rats. Since altered lipid metabolism is frequently associated with these disease states, lipid metabolism was investigated in vitro in testes and sciatic nerves of rats fed 1% HD in the drinking water for 6 weeks. Testes from HD-treated rats were 30–60% smaller and weighed threefold less than testes from pair-fed control (PFC) rats. Compared to testes from PFC rats, testes from HD rats exhibited increased incorporation of [¹⁴C]acetate into phospholipids (344%), triacylglycerols (281%), and cholesteryl esters + hydrocarbons (246%) but decreased incorporation into free fatty acids (25%) and sterols + diacylglycerols (65%). The increased incorporation of [¹⁴C]acetate into phospholipids induced by HD reflected an approximate 300% increase into phosphatidyl choline, lysophosphatidyl choline, phosphatidyl serine + phosphatidyl inositol, and phosphatidyl ethanolamine and a disproportionate 800% increase into sphingomyelin. HD rats exhibited clinical signs of peripheral neuropathy, including everted and flat foot placement and hindlimb weakness; similar changes were not observed in PFC rats. In sciatic nerves, the incorporation of [¹⁴C]leucine was decreased into sterols + diacylglycerols (47%), digitoninprecipitable sterols (45%), and cholesteryl esters + hydrocarbons (40%) in HD compared to PFC rats; incorporation of [¹⁴C]leucine into free fatty acids, triacylglycerols, and phospholipids was similar in HD and PFC rats. In contrast to the testis and nerve, lipid metabolism in the liver was similar in HD and PFC rats. The concentrations of 2,5-hexanedione and 2,5-dimethylfuran, respectively, were 0.6 ± 0.3 and 6.5 ± 0.9 μg/g wet weight in the testes and 3.1 ± 0.4 and 3.0 ± 0.4 μg/g wet weight in the livers of HD rats. The data indicate that altered metabolism is associated with HD-induced testicular atrophy and distal axonopathy.     

The influence of silybin from Silybum marianum (L.) Gaertn. on in vitro phosphatidyl choline biosynthesis in rat livers

In rat livers, removed 60 min after i.p. application of 150.6 mg/kg silybin-dihemisuccinate-di-Na-salt, in vitro incorporation of choline-(methyl-14C) into phosphatidyl choline by the postmitochondrial fraction of liver homogenates was enhanced in comparison with controls. These results are associated with increased activities of CTP-choline-phosphate cytidyltransferase. Enzyme activities were also enhanced after addition of 7.5 x 10(-6) mol/l silybin to incubation mixtures obtained from untreated rats.     

Sex difference in the development of fatty liver by orotic acid.

Effects of orotic acid on liver lipid accumulation and incorporation of methionine [methyl-14C] into liver phosphatidylcholine and protein, and into serum beta-lipoprotein were studied. Male and female rats of Wistar strain were fed a semisynthetic diet supplemented with 1 per cent orotic acid for 7 days. Feeding of orotic acid induced a marked fatty liver in female rats, but not in males. In female rats, radioactivity in liver phosphatidylcholine was significantly decreased by orotic acid, and that in liver protein was slightly decreased. In male rats, incorporation of methionine [methyl-14C] into liver phosphatidylcholine and protein was unchanged between the control and the rats fed orotic acid. Radioactivity in serum beta-lipoprotein was decreased to a greater extent in female rats than in males. These results suggest that sex difference in the development of fatty liver may be due to the difference in the effect of orotic acid on liver phosphatidylcholine biosynthesis.     

Effect of 4-(4'-chlorobenzyloxy)benzyl nicotinate (KCD-232) on cholesterol metabolism in rats

The effects of 4-(4'-chlorobenzyloxy)benzyl nicotinate (KCD-232), a new hypolipidemic agent, on serum cholesterol level and cholesterol biosynthesis were studied in normolipidemic rats. KCD-232 dose-dependently reduced the serum cholesterol level. The in vivo incorporation of [14C]-acetate and 3H from [3H]water into liver digitonin-precipitable sterols was inhibited by oral administration of KCD-232, while that of [14C]mevalonic acid into the sterols was not inhibited. Hepatic 3-hydroxy-3-methylglutaryl (HMG)-CoA reductase activity was suppressed significantly by the oral administration of the drug. A KCD-232 metabolite, 4-(4'-chlorobenzyloxy)benzoic acid (MII), strongly inhibited [14C]acetate incorporation and weakly inhibited [14C]mevalonic acid incorporation into the sterols in liver slices. MII also significantly inhibited the sterol synthetic rate measured with [3H]water and the HMG-CoA reductase activity in dispersed hepatocytes. MII and its CoA thioester (MII-CoA) inhibited the incorporation of [14C]acetate, [14C]acetyl-CoA and [14C]HMG-CoA into nonsaponifiable lipids in a cell-free enzyme system from rat liver. MII-CoA further showed a weak inhibition of [14C]-mevalonic acid incorporation into nonsaponifiable lipids in the system, while MII showed no effect on mevalonic acid incorporation. These results indicate that KCD-232 possesses a major inhibitory site for sterol synthesis on HMG-CoA reductase due to both MII and MII-CoA, and a possible second site of action beyond mevalonic acid due to MII-CoA. The latter inhibitory site, however, is considered to play a minor role in the inhibition of sterol synthesis in vivo.     

Evidence of differential effects produced by ethanol on specific phospholipid biosynthetic pathways in rat hepatocytes.

1. The aim of the present study was to investigate the effects of ethanol in vitro on the phospholipid biosynthetic pathways in hepatocytes isolated from the rat. We have used [methyl-14C]-choline, [1-3H]-ethanolamine and L-[3-3H]-serine as exogenous precursors of the corresponding phospholipids, phosphatidylcholine (PC), phosphatidylethanolamine (PE) and phosphatidylserine (PS). 2. Incubation of hepatocytes in the presence of ethanol significantly alters the incorporation of radiolabel from [14C]-choline and [3H]-ethanolamine into the metabolic intermediates and the final products of the CDP-choline and CDP-ethanolamine pathways. Radioactivity in the metabolic intermediates of both pathways was significantly decreased and the amount of label in PE was reduced whilst that of PC was not modified. 3. In the presence of 4-methylpyrazole, an inhibitor of alcohol dehydrogenase (ADH) activity, ethanol produces a reduction in the label of choline phosphate, ethanolamine phosphate and a significant decrease in the amount of PC and PE radiolabel. 4. On the other hand, ethanol increases the incorporation of serine into phosphatidylserine, phosphatidylethanolamine and phosphatidylcholine, although this effect is observed only in the absence of 4-methylpyrazole, indicating that this alteration is produced by some metabolite generated as a consequence of hepatic alcohol metabolism. 5. Ethanol also interferes with the methylation of phosphatidylethanolamine produced via the CDP-ethanolamine pathway but it does not alter phosphatidylethanolamine methylation when this phospholipid is produced by mitochondrial phosphatidylserine decarboxylation, suggesting the existence of different intramembrane pools of phosphatidylethanolamine, which may exhibit different sensitivity to alcohol. 6. Our results indicate that ethanol exerts two different effects on phospholipid metabolism in hepatocytes: a stimulatory effect on the incorporation of exogenous substrates into different phospholipids probably related to an alteration in the availability of lipogenic substrates as a consequence of ethanol metabolism, and another inhibitory effect produced by ethanol per se, which can be observed only when ethanol metabolism is inhibited by the presence of a specific inhibitor of alcohol dehydrogenase activity.     

The effect of ethanol on phospholipid metabolism in rat pancreas

The phospholipid effect involves agonist-induced breakdown of phosphatidyl inositol (or polyinositides) generating second messengers followed by increased incorporation of 32P during the resynthetic phase of the cycle. Ethanol, an aetiological factor in pancreatitis, has been shown to have various effects on pancreatic secretion. In this study ethanol decreased the incorporation of 32P into phosphatidyl inositol but had no effect on the stimulated breakdown of prelabelled phosphatidyl inositol. However, in addition to recycling of phosphatidyl inositol stimulation of pancreatic tissue results in increased incorporation of precursors into other phospholipids. Cholecystokinin increased the incorporation of both [U-14C] glucose and 32P into phosphatidyl ethanolamine 3-fold but had no effect on 32P incorporation into phosphatidyl choline. As well as increased incorporation of 32P into phosphatidyl inositol (8-fold) cholecystokinin also increased the incorporation of [U-14C] glucose into phosphatidyl inositol (4-5-fold) implying significant de novo synthesis of 1,2 diacyl glycerol in addition to the currently accepted recycling of the 1,2 diacyl glycerol back to phosphatidyl inositol. Ethanol caused an inhibition of 32P incorporation into total phospholipid of rat pancreas during basal and stimulated conditions. When individual phospholipids were separated ethanol was found to decrease the incorporation of 32P into phosphatidyl choline under basal conditions and into all phospholipids during cholecystokinin stimulation. With [U-14C] glucose as the precursor, ethanol inhibited its incorporation into phosphatidyl choline only. Ethanol did not alter the total 32P radioactivity in the aqueous phase of the pancreatic extract nor the percent incorporated into nucleotides. This excluded decreased uptake of 32P and incorporation into nucleotides as a mechanism for the differential inhibition of 32P versus [U-14C] glucose incorporation into phospholipids other than phosphatidyl choline under stimulated conditions.     

In vivo studies on the effect of garlic oil in mice

The in vivo effect of garlic oil on protein, glycogen and lipid metabolism as well as on oxidative phosphorylation has been studied in liver and kidneys of mice. The incorporation of [¹⁴C] leucine into proteins of the liver is virtually unchanged by treatment with garlic oil whereas the labelling of proteins of the kidneys is reduced. Garlic oil enhances the incorporation of [¹⁴C] glucose into hepatic glycogen, produces a temporary rise in blood glucose levels and depletes liver glycogen. Renal glycogen levels and incorporation of [¹⁴C] glucose into renal glycogen remain unchanged following garlic oil treatment. The compound has little or no effect on the incorporation of [¹⁴C] acetate into hepatic and renal lipids. Garlic oil impairs partially the oxidative phosphorylation associated with the oxidation of ascorbate in hepatic mitochondria without affecting noticeably the P/O ratio during the oxidation of glutamate and succinate.     

Effect of phenobarbital on methyl transfer between methylated drugs and hepatic microsomal phospholipids.

The effect of phenobarbital on the incorporation of the label from N-[¹⁴C-Me]nicotine and [¹⁴C]formaldehyde into hepatic phospholipids of the rat has been studied. ¹⁴C was utilized for the formation of methylated phospholipids from both precursors. Phenobarbital elicited no significant action either on the synthesis of total hepatic phospholipids or on the incorporation of radioactivity into the total or individual liver phospholipid fractions. However, this treatment increased phospholipid content and the uptake of the label from nicotine into microsomal phospholipids. Phenobarbital raised microsomal phosphatidylethanolamine, -choline (PC), -serine (PS), and lysophosphatidylcholine contents and the incorporation of ¹⁴C-labeled methyl groups from nicotine into PC and PS fractions. Radioactivity from [¹⁴C]formaldehyde was also incorporated into hepatic phospholipids. Phenobarbital however, had no significant effect on the incorporation either into total or microsomal phospholipids. Comparing the utilization of ¹⁴C for synthesis of liver microsomal phospholipids from N-[¹⁴C-Me]nicotine or [¹⁴C]formaldehyde with the natural methyl donor, l-[¹⁴C]-Me]methionine, greater amounts were taken up from methionine than nicotine or formaldehyde. The methyl group of nicotine was probably incorporated into phospholipids via the metabolic pool; the enhancing effect of phenobarbital on this process was associated with increased metabolism and with increased methyl transfer into methyl group containing microsomal phospholipids.     

Effect of 3-amino-1,2,4-triazole on in vivo formation of liver triglyceride

The mechanics of the decrease in the liver triglyceride (TG) level caused by the administration of 3-amino-1,2,4-triazole (AT) were studied in vivo. The incorporation of [¹⁴C]acetate (10 μCi/100 g, i.p.) into liver TG of rats injected with AT (100 mg/100 g, i.p.) decreased about 55 per cent when compared with the control group. Incorporation of [¹⁴C]palmitate into liver and serum TG, did not differ in the AT and control group. Incorporation of [¹⁴C]acetate into liver TG of rats administered ethanol (600 mg/100 g, p.o.) was 3 times that of the control, but with AT-pretreatment incorporation was 50 per cent less when compared with that of ethanol group. Serum free fatty acids of rats injected with adrenalin (200 μg/100 g, i.m.) increased two times above the control group. The increase was not affected by pretreatment with AT.     

Effects of S-8527 (1,1-bis(4'-(1'-carboxy-1'-methylpropoxy)phenyl) cyclohexane), a new hypolipidemic compound, on cholesterol and lipoprotein metabolism in rats.

The effects of S-8527 (1,1-bis[4'-(1Prime;-carboxy-1Prime;-methylpropoxy)phenyl]cyclohexane on cholesterol and lipoprotein metabolism were examined and compared with those of clofibrate in rats under various experimental conditions. When rats were given a daily oral dose of S-8527 for 8 days, the incorporation of [¹⁴C]acetate into liver cholesterol was not inhibited at the dose of 30 mg/kg of S-8527. which was reported to decrease the serum cholesterol significantly, but the higher dose (300 mg/kg) of S-8527 decreased the incorporation of [¹⁴C]acetate into liver cholesterol. Under these experimental conditions, clofibrate (300 mg/kg) caused a decrease in labeled cholesterol in the liver. Oral doses of S-8527 or clofibrate for 8 days did not affect the incorporation of [¹⁴C]mevalonate into liver cholesterol. Also, when the drugs were added to normal rat liver slices, the effects of S-8527 were not so marked as those seen with clofibrate. Oral doses of S-8527 (30 mg/kg) or clofibrate (300 mg/kg) for 8 days decreased the incorporation of [¹⁴C]lcucine into the protein of serum lipoproteins. S-8527 and clofibrate did not affect the body retention of radioactivity after the injection of labeled cholesterol into rats. In view of the above results, it is conceivable that S-8527 primarily inhibits either the secretion of lipoproteins containing cholesterol into plasma or the formation of lipoproteins containing cholesterol in the liver, or both, and secondarily interferes with the biosynthesis of liver cholesterol.     

Transport and metabolism of double-labelled CDPcholine in mammalian tissues

Double-labelled [methyl-14C,5-3H]CDPcholine has been synthesized and subjected to a pharmacokinetic analysis in several biological systems. In transport experiments with intact human erythrocytes no incorporation of radioactivity is observable. On the other hand the results obtained with perfused rat liver suggest a rapid cleavage of the pyrophosphate bridge of the molecule, followed by a rapid uptake of the hydrolytic products. The plasma half-lives of intravenously injected CDPcholine and of its metabolites have been evaluated within 60 sec range. Renal and fecal excretion of the injected radioactivity is negligible: only 2.5% of administered 14C- and 6.5% of the 3H- is excreted up to 48 hr after administration. Liver and kidney are the major CDPcholine metabolizing organs, characterized by a fast and extensive uptake of choline metabolites, followed by a slow release; conversely the rate of uptake of both 3H and 14C-labelled moieties by rat brain is significantly slower, reaching a steady-state level after 10 hr. The characterization of the labelled compounds detectable in the investigated organs provides some insights on the metabolism of the drug: the 3H-cytidine moiety in all the examined organs appears to be incorporated into the nucleic acid fraction via the cytidine nucleotide pool; the [14C]choline moiety of the molecule is in part converted, at the mitochondrial level, into betaine which accounts for about 60% of the total 14C-radioactivity associated with liver and kidney 30 min after administration; [14C]betaine in turn acts as methyl donor to homocysteine yielding [14C]methionine subsequently incorporated into proteins; the time dependent increase in labelled phospholipids is indicative of a recycling of the choline methyl-groups in this lipid fraction via CDPcholine and/or S-adenosylmethionine; the rather extensive amount of labelled methionine detectable in brain probably arises from its uptake from the blood stream, since the enzyme catalyzing the conversion of betaine into methionine is lacking in brain.     

Effects of dl-propranolol on the synthesis of glycerolipids by rabbit iris muscle

Propranolol (0.03?0.3 mM), an amphiphilic cationic drug which is used therapeutically as a β-blocker, was found to alter significantly the incorporation of [¹⁴C]glucose, [¹⁴C]glycerol, [¹⁴C]acetate, ³²Pi, [³H]cytidine, [³H]inositol, [¹⁴C]choline, [¹⁴C]ethanolamine and [¹⁴C]serine into phospholipids of the iris muscle. Furthermore, it was found to exert a stimulatory effect on the [¹⁴C]serine incorporation into phosphatidylserine of the muscle and microsomes. In contrast, sotalol, another β-blocker-but lacking the hydrophobicity of propranolol-exerted no effect on lipid metabolism. Whereas norepinephrine stimulated only the turnover of the phosphate moiety of phosphatidic acid and phosphatidylinositol, in general propranolol caused the following changes: (a) it stimulated by 2- to 6-fold the labelling of phosphatidic acid and phosphatidylinositol from [¹⁴C]glucose, [¹⁴C]glycerol, [¹⁴C]acetate, ³²Pi and [³H]inositol, (b) it increased by 5- and 38-fold the incorporation of ³²Pi and [³H]cytidine, respectively into CDP-diglyceride, (c) it inhibited appreciably the incorporation of [¹⁴C]glucose, [¹⁴C]glycerol, [¹⁴C]acetate and ³²Pi into phosphatidylcholine and phosphatidylethanoalmine. However, while it inhibited significantly the [¹⁴C]choline incorporation into the former, it stimulated by 60 per cent the ethanolamine incorporation into the latter phospholipid. These results indicate that propranolol probably redirects phospholipid synthesis de novo, by inhibiting phosphatidate phosphohydrolase, such that the increase obtained in the biosynthesis of phosphatidylinositol is accompanied by a corresponding decrease in the synthesis of phosphatidylcholine and phosphatidylethanolamine.Propranolol also caused a 250 per cent increase in the [¹⁴C]serine incorporation into phosphatidylserine of the iris muscle and 28 per cent increase in that of microsomes. The drug appears to stimulate the Ca²⁺ -uptake by muscle and microsomes, which in turn could act to stimulate the Ca²⁺-catalyzed base-exchange reaction.In addition the metabolic pathways involved in the biosynthesis of the major phospholipids of the iris, a smooth muscle, are reported for the first time. These pathways were found to be essentially similar to those reported for other tissues.     

Evidence for a different metabolic behaviour of cytidine diphosphate choline after oral and intravenous administration to rats

Radioactivity plasma decay was studied in rats after intravenous and oral administration of cytidine diphosphate [methyl-14C]choline at doses of 25 and 300 mg/kg. The kinetics fitted well with a two compartment open model and showed a long lasting elimination phase with a half-life ranging from 2.0 to 2.6 days for the two doses and the two administration routes. Absorption of cytidine diphosphate choline radioactivity was complete after oral treatment with the low dose and accounted for 94.5% of the dose when 300 mg/kg of cytidine diphosphate [methyl-14C]choline were administered. However the distribution of radioactivity in tissues, urine and expired air suggest metabolic differences, at least from a quantitative point of view, between the oral and intravenous treatments. In particular, the higher excretion of radioactivity associated with trimethylamine in urine found when cytidine diphosphate [methyl-14C]choline was given orally, suggest that the compound may be metabolized, at least in part, previous to its gastrointestinal absorption.     

Effects of an antitumor agent, ascofuranone, on the macromolecular syntheses of intact cells

Ascofuranone (AF) has antitumor protective property on experimental tumors. We examined the action of AF on lymphoma L5178Y to explore the mechanism of the antitumor activity. AF completely prevented the growth of L5178Y at 25 micrograms/ml cytostatically. The compound exhibited general inhibitory effects on the macromolecular syntheses. Among them, protein synthesis was most severely inhibited by AF and to the same extent as by cycloheximide. AF, however, did not affect protein synthesis by cell-free system even at 2 mg/ml. Although AF inhibited the incorporation of [14C]acetate into total acid precipitable products only slightly, the synthetic pattern of simple lipids from [14C]acetate was significantly changed. Especially, the incorporation of [14C]acetate into squalene was almost completely blocked at 25 micrograms/ml. The incorporation of [14C]acetate into triglyceride was inhibited and that into cholesterol was enhanced. Concerning the diglycerides, the incorporation of [14C]acetate was enhanced and that of [3H]glycerol was inhibited. The incorporation of [3H]glycerol and [3H]mevalonate into the intact cell was significantly inhibited as compared with [14C]acetate. As those effects were not observed with cycloheximide, they were suggested to be characteristic of AF. AF inhibited hypotonic hemolysis. In contrast, hemolysis by deoxycholate was stimulated. Possible mechanism of the antitumor activity of AF is discussed.     

Effect of T-2 toxin administration to rats on lipid metabolism in liver

The effect of oral dosing of rats with 1.5 mg T-2 toxin/kg body weight daily for 4 days on metabolism of liver lipids was studied. T-2 toxin significantly elevated total liver lipids, triglycerides, free cholesterol, total phospholipids and phosphatidyl choline, whereas the level of sphingomyelin + lysophosphatidyl ethanolamine was reduced. No change in the esterified cholesterol and phosphatidyl ethanolamine contents was observed. Incorporation of [1-14C]acetate into liver lipids, esterified cholesterol, triglycerides, free cholesterol and phosphatidyl ethanolamine was reduced in T-2 toxin-treated animals, implying reduced lipogenesis. Increased lipids in liver in T-2 toxin-treated rats are possibly due to an impaired secretion of lipids from the liver.     

Effect of vasoactive intestinal peptide on pulmonary surfactants phospholipid synthesis in lung explants

AIM: To investigate the effect of vasoactive intestinal peptide (VIP) on pulmonary surfactants (PS) phospholipidsynthesis in cultured lung explants. METHODS: Lung explants were cultured with serum-free medium, [methyl^3H]choline incorporation, total phospholipid, phosphatidylcholine, activity of choline-phosphate cytidylyltransferase(CCT) and CCTα mRNA level in lung explants were determined. RESULTS: (1) VIP (10^-10-10^-7 mol/L) for 16 hpromoted [methyl-^3H]choline incorporation in dose dependence and VIP (10.8 mol/L) for 2 h-16 h promoted [methyl-^3H]choline incorporation in time dependence. (2) VIP (10-8 mol/L) enhanced the contents of total phospholipidsand phosphatidylcholine in lung explants. (3) VIP (10^-10-10-7 mol/L) elevated microsomal CCT activity of lungexplants in dose dependence. (4) VIP (10.8 mol/L) increased expression of CCTα mRNA in lung explants andalveolar type II cells (ATII). (5) [D-P-C1-Phe(6)-Leu(17)]-VIP (10^-6 mol/L), a VIP receptors antagonist, abolishedthe increase of [^3H]choline incorporation, microsomal CCT activity and CCTα mRNA level induced by VIP (10-8mol/L) in lung explants. CONCLUSION: VIP could enhance synthesis of phosphatidylcholine, the major compo-nent of pulmonary surfactants by enhancing microsomal CCT activity and CCTα mRNA level via VIP receptor-mediated pathway.