Chlorpromazine increases the turnover of metabolically active phosphoinositides and elevates the steady-state level of phosphatidylinositol-4-phosphate in human platelets.

Non-permeabilizing concentrations (< 40 microM) of chlorpromazine (CPZ) increase the radioactivity of phosphatidylinositol-4-phosphate (PIP) in platelets pre-labelled with [32P]Pi, but the biochemical mechanisms underlying this increase are poorly understood. Incubation of [32P]Pi-labelled, gel-filtered platelets with 25 microM CPZ for 10 min increased: (1) the mass of PIP from 315 to 476 nmol/10(11) platelets but not the total inositol phospholipid mass, (2) the specific phosphodiester radioactivities in phosphatidylinositol (PI), PIP and phosphatidylinositol-4,5-bisphosphate (PIP2) by 34, 63 and 37%, respectively, and (3) the specific phosphomonoester radioactivities in PIP and PIP2 by 53 and 10%, respectively. In control platelets (no CPZ) the specific radioactivity of the phosphodiester was the same in PI, PIP and PIP2, and the specific radioactivity in the phosphomonoester in PIP and PIP2 was 55% of that of the gamma-phosphoryl in ATP, measured as metabolically active, actin-bound ADP. These results suggest that 55% of each of PI, PIP and PIP2 constitutes a metabolic pool which is labelled by 32P in the platelets, while the remainder is in a metabolically inactive pool and not labelled. CPZ has two major effects: (1) CPZ interferes with the kinase and phosphohydrolase reactions that maintain the steady-state level of PIP in the metabolic phosphoinositide pool, resulting in a 92% increase in the PIP level of this pool, and (2) CPZ causes synthesis (45% in 10 min) of new phosphodiester in the metabolically active phosphoinositides by tentative stimulation of the turnover of the phosphoinositide cycle, de novo phosphoinositide synthesis and/or diacylglycerol formation through phospholipases C and D. The marked alteration by CPZ of phosphoinositide metabolism may be part of the mechanism by which this drug effects its psychotropic action.     

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.     

Effects of various antihypertensive agents on lipid metabolism: alterations in the pattern of lipids synthesized from [14C]oleate in rat liver in vitro

The effects of five antihypertensive agents on lipid biosynthesis from [1-14C]oleate were studied in rat liver minces. At a level of 1 mM, propranolol and prazosin increased the incorporation of [14C]oleate into diglycerides and cholesteryl esters by two- to fourfold and increased total phospholipid labeling by 20-30%. Chlorthalidone and metoprolol at 1 mM also stimulated the incorporation of [14C]oleate into phospholipids and diglycerides (20-50%) but did not affect its incorporation into triglycerides or cholesteryl esters. All four of the compounds statistically significantly inhibited the incorporation of [14C]oleate into phosphatidylcholine by 12-37% but stimulated incorporation into phosphatidylinositol by 17-95%. Nadolol differed from the other compounds in that it did not show selective effects but rather inhibited the incorporation of [14C]oleate into all lipid classes by approximately 50%. The data are discussed in terms of possible mechanisms involved in the lipid synthesis patterns and suggest the possibility that plasma lipid/lipoprotein changes observed in patients undergoing antihypertensive therapy may reflect, in part, altered hepatic lipid synthesis.     

Effect of pentobarbital on regional brain phospholipid synthesis

Rats were given 45 mg/kg i.p. sodium pentobarbital 15 min prior to the intraventricular injection of 200 μCi [³²P]phosphoric acid and 50 μCi [³H]glycerol. The animals were sacrificed 1 hr later, subcellular fractions were prepared from four subcortical brain regions and phospholipids were extracted. Pentobarbital significantly increased the ratio of [³H]- and [³²P]-triphosphatidylinositol (TPI) to diphosphatidylinositol (DPI) in the microsomal but not synaptosomal fractions. The possible relationship of this change to nicotinic receptor activity is discussed. Pentobarbital specifically decreased ³²Pi but not [³H]glycerol incorporation into synaptosomal phosphatidylinositol (PI). Thus, pentobarbital induced the opposite of the “neurotransmitter effect” on PI turnover. Pentobarbital either decreased or had no effect on the incorporation of ³²Pi and [³H]glycerol into phosphatidylserine (PS), phosphatidylcholine (PC) and phosphatidylethanolamine (PE).     

Studies on the mechanism of alteration by propranolol and mepacrine of the metabolism of phosphoinositides and other glycerolipids in the rabbit iris muscle

We have investigated the effects and mechanism of action of propranolol and mepacrine, two drugs with local anesthetic-like properties, on phospholipid metabolism in rabbit iris and iris microsomal and soluble fractions. In the iris, propranolol, like mepacrine [A. A. Abdel-Latif and J. P. Smith, Biochim, biophys. Acta 711, 478 (1982)], stimulated the incorporation of [14C]arachidonic acid ( [14C]AA) into phosphatidic acid (PA), CDP-diacylglycerol (CDP-DG), phosphatidylinositol (PI), the polyphosphoinositides (poly PI) and DG, and it inhibited that of phosphatidylcholine (PC), phosphatidylethanolamine (PE), triacylglycerol (TG) and the prostaglandins. Similarly, mepacrine, like propranolol [A. A. Abdel-Latif and J. P. Smith, Biochem. Pharmac. 25, 1697 (1976)], altered the incorporation of [14C]oleic acid, [3H]glycerol, 32Pi and [14C]choline into glycerolipids of the iris. Time-course studies in iris muscle prelabeled with [14C]AA showed an initial decrease in the production of DG and a corresponding increase in that of PA by the drugs, followed by an increase in accumulation of DG at longer time intervals (60-90 min). The above findings are in accord with the hypothesis that these drugs redirect glycerolipid synthesis by inhibiting PA phosphohydrolase. Propranolol and mepacrine stimulated the activities of DG kinase and phosphoinositide kinases and inhibited that of DG cholinephosphotransferase. The drugs had little effect on the activity of DG acyltransferase. It is concluded that propranolol and mepacrine redirect glycerolipid metabolism in the iris by exerting multiple effects on the enzymes involved in phospholipid biosynthesis. We suggest that these drugs could exert their local anesthetic-like effects by effecting an increase in the synthesis of the acidic phospholipids (PA, PI and the poly PI) and subsequently the binding of Ca2+- to the cell plasma membrane.     

Effect of desipramine on inositol phosphate formation and inositol phospholipids in rat brain and human platelets.

To examine the mechanism of action of antidepressant drugs, we studied the effect of desipramine (DMI) in vitro on agonist-stimulated inositol phosphate formation and inositol phospholipids in rat brain and human platelets. We observed that DMI inhibited thrombin-stimulated 3H-inositol bisphosphate (IP2) and 3H-inositol trisphosphate (IP3) but not 3H-inositol monophosphate (IP1) formation in human platelets. DMI also inhibited norepinephrine (NE) and serotonin (5-HT) stimulated 3H-IP1 formation in rat cerebral cortex. DMI increased levels of all three 3H-inositol phospholipids, 3H-phosphatidyl inositol (PI), 3H-PI-4-phosphate (PIP), and 3H-PI 4,5-bisphosphate (PIP2), in both platelets and rat cortex. The decreased formation of inositol phosphates and increased levels of [3H]-PI, [3H]-PIP, and [3H]-PIP2 by DMI appears to be due to the inhibition of the enzyme phospholipase C rather than its effects on receptors. It is thus possible that interaction of tricyclic antidepressant drugs with the PI-signaling system may be related to their mechanism of action.     

Selective modifications in the de novo biosynthesis of retinal phospholipids and glycerides by propranolol or phentolamine

The effects of propranolol or phentolamine on the metabolism of phospholipids, diacylglycerol, and triacylglycerol were studied in the bovine retina in vitro. Lipid labeling was followed during short-term incubation of intact bovine retinas with [U-₁₄C]glycerol and [1-¹⁴C]palmitic acid. Each of these precursors was recovered in the appropriate lipid moiety. Most of the [¹⁴C]glycerol appeared progressively in triacylglycerol (TG) through the sequence from phosphatidic acid (PA) to diacylglycerol (DG). Labeled palmitate appeared in much lower quantities than labeled glycerol in all glycerolipids except phosphatidylcholine (PC). Propranolol and phentolamine greatly enhanced the [¹⁴'C]glycerol specific activities of PA, phosphatidylinositol (PI), and phosphatidylserine (PS), whereas labeling in other glycerolipids was much lower than in controls. The labeling in TG with both precursors was found to be less than 50% of the control values; however, a late increase in DG labeling was observed. The effects of these drugs on broken cell preparations were also described, although lipid synthesis from labeled glycerol in these preparations was only 9% that of intact retinas. It appeared that an amphiphilic cationic structure was necessary to produce these drug effects; propranolol glycol, the hydrophobic moiety of propranolol, did not elicit the same effects. It is suggested that, among other changes, the drugs inhibited phosphatidate phosphohydrolase and redirected the flux predominantly toward PI. Support for the proposed multiple lipid effects elicited by these drugs was provided by the dual changes found in the labeling of DG.     

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.     

Increase of phosphatidylinositol arachidonic acid incorporation induced by mepacrine

In view of the fact that mepacrine (Mp) is usually used as an inhibitor of the endogenous phospholipase A2, and since this enzyme produces the release of arachidonic acid (AA) from membrane phospholipids, we studied the effect of different concentrations of Mp on the mobilization of [1-14C]AA in rat renomedullary phospholipids. During the acylation period, 0.1 mM Mp did not produce any significant change in the incorporation of [1-14C]AA into phosphatidylcholine (PC) and phosphatidylethanolamine (PE), and only a slight increase in phosphatidylinositol (PI). Higher concentrations of Mp (0.5 to 1.0 mM) produced a decrease of radioactivity in PE and PC with an increase in PI. Using prelabeled slices, a dose-dependent decrease in the 14C-radioactivity in PE and PC was observed, with a parallel increase in PI. This effect of Mp persisted even in the presence of a physiological activator of phospholipase A2, bradykinin (BK). No change in the net amount of phospholipids was observed at any of the Mp concentrations used. The results of this study show that Mp, at concentrations generally used to inhibit phospholipase A2, produced a transfer of arachidonic acid from PE and PC to PI, rather than a blockade in the release of AA from membrane phospholipids.     

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.     

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.     

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.     

Muscarinic receptor stimulated phosphoinositide turnover in cardiac atrial tissue

The properties of muscarinic agonist stimulated phosphoinositide turnover in canine atrial slices were investigated. In slices prelabeled with 32Pi, carbachol stimulated a 20-30% decrease of 32P-labeled phosphatidylinositol 4'-phosphate (PIP) and phosphatidylinositol 4',5'-bisphosphate (PIP2) content within 10-15 sec. This was followed by a resynthesis of these lipids to control levels after 30 sec. Carbachol-stimulated PIP and PIP2 turnover was followed by a relatively slower increase in 32P incorporation into phosphatidylinositol (PI) and phosphatidic acid (PA) which was maximal after 5-10 min. Carbachol increased 32P-labeling of PA and PI in most regions of right and left atria with equal effectiveness. Muscarinic receptor stimulated increases in PA and PI labeling showed high specificity for certain muscarinic agonists and, unlike most tissues, this muscarinic receptor mediated phospholipid effect was dependent on extracellular calcium. Carbachol did not increase 32P incorporation into PA and PI if Mn2+, Co2+, Mg2+, or La3+ was substituted for extracellular Ca2+. Unlike other muscarinic agonists, acetylcholine increased 32P incorporation into phosphatidylcholine in addition to PA and PI. Low concentrations of calcium channel blockers, verapamil, nifedipine or diltiazem, did not block carbachol-stimulated changes in PA and PI labeling in the presence of Ca2+; however, higher concentrations (greater than or equal to 10 microM) of verapamil increased PA and PI labeling. Ouabain enhanced carbachol-stimulated 32P incorporation into PA but attenuated incorporation into PI. The mechanisms associated with the actions of these agents on phospholipid metabolism and their possible physiological significance are discussed.     

Effect of chlorpromazine on protein, phospholipid and RNA synthesis or degradation in rat liver

Chlorpromazine (CPZ) administration to rats (25 mg/kg ip) significantly stimulated (14C)-leucine incorporation to liver microsomal proteins, (14C)-orotic acid incorporation to liver RNA and (32P) incorporation to liver microsomal lipids. CPZ administration did not modify the decay of radioactivity of liver microsomal lipids prelabeled with (32P) or that of microsomal proteins prelabeled with [(14C)-guanidino]-arginine. Results suggest that CPZ administration stimulates protein, phospholipids and RNA synthesis but does not affect their degradation. The relation of these findings with CPZ preventive effects on CCl4-induced liver injury is discussed.     

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.     

Cationic amphiphilic drugs as a potential tool for modifying phospholipids of tumor cells. An in vitro study of chlorpromazine effects on Krebs II ascites cells

[³²P]orthophosphate and [U-¹⁴C]glycerol incorporation into Krebs ascites cell lipids was studied in vitro in the presence of chlorpromazine (CPZ)¹. At concentrations not exceeding 0.1 mM, the drug produced no cell damage within 3 hr incubation. Under these conditions, CPZ inhibited the incorporation of [³²P]orthophosphate into phosphatidylcholine and phosphatidylethanolamine and of [U-¹⁴C]glycerol into phosphatidylcholine and triglycerides, in a dose-dependent manner. On the other hand, synthesis of phosphatidic acid and phosphatidylglycerol was greatly enhanced, whereas phosphatidylinositol showed no major change. These results are compatible with an inhibition of phosphatidate phosphohydrolase, redirecting phospholipid biosynthesis towards the anionic classes. In vitro treatment of cells for 3 hr induced profound changes of phospholipid composition, which displayed a relative increase of phosphatidylglycerol and phosphatidic acid at the expense of phosphatidylcholine and phosphatidylethanolamine. The use of amphipathic cationic drugs can thus offer an interesting model for studying the relationship between cell proliferation and membrane phospholipid biosynthesis.     

Effect of polychlorinated biphenyls (PCBs) administration on phospholipid biosynthesis in rat liver

The effect of PCBs or phenobarbital on the biosynthesis of phospholipids in hepatic endoplasmic reticulum of rats was studied by the intraperitoneal injection of [³²P]orthophosphate, [Me?¹⁴ C]choline or [2?³H]glycerol. Significant increases in liver microsomal phospholipid content after the administration of either PCBs or phenobarbital indicated the actual proliferation of endoplasmic reticulum membranes. The rate of both [³²P] and [¹⁴C] incorporations into microsomal choline-containing phospholipids, such as phosphatidylcholine, sphingomyelin and lysophosphatidylcholine, was reduced to one fifth by PCBs administration compared with control animals. The incorporation of [³²P]orthophosphate into phosphatidylethanolamine or other phospholipid classes was less or not affected, respectively, by PCBs administration. The specific inhibitory effect of PCBs on the incorporation into cholinecontaining phospholipids was not observed when [2?³-H]glycerol was used as a precursor. Phenobarbital administration, however, increased significantly the rate of [³²P] incorporation into liver phospholipids, especially phosphatidylcholine. It is suggested that the increase in microsomal phospholipid content by PCBs administration is not due to the stimulation of synthesis but to the inhibition of the catabolism of membrane phospholipids and that the increase in content caused by phenobarbital is due at least in part, to the stimulation of synthesis. The possible site(s) of PCBs-induced inhibition of phospholipid biosynthesis in rat liver is discussed.     

Lipid modification in mouse peritoneal macrophages after chronic cadmium exposure

The effect of cadmium (Cd) exposure through drinking water on lipid status in mouse peritoneal macrophages (pM) was studied. After 2 months, adult male Balb/c mice that had drunk water with 15 ppm of Cd, showed tissue damage mediated by oxidative stress, as assessed by serum measuring of tissue damage and lipoperoxidation indicators. Resident pM obtained from Cd-exposed mice showed diminution in total lipids, total cholesterol, free cholesterol/esterified cholesterol ratio (FC/EC) and phospholipids in relation to control pM. On a percentage basis, the phospholipid composition showed that phosphatidylcholine (PC) and phosphatidylglycerol decreased, phosphatidylethanolamine (PE) increased, while phosphatidylinositol, sphingomyeline and phosphatidylserine did not change. The incorporation in vitro of [14C]-methyl-choline and [14C]-phosphorylcholine, as well as the activity of regulatory enzyme CTP-phosphocholine cytidylyltransferase, decreased in PC after 60 min. The incorporation of [14C]-linoleic acid increased after 1 h and the incorporation of [14C]-ethanolamine increased after 90 min in PC. The incorporation in vitro of [3H]-cholesterol in total lipids decreased after 120 min of incubation. Besides, the stearic acid and arachidonic acid content increased, while the contents of palmitoleic acid and linoleic acid decreased. Chronic Cd exposure alters the lipid composition in resident pM of Balb/c mice.     

Effects of NTE-122, a novel acyl-CoA:cholesterol acyltransferase inhibitor, on cholesterol esterification and secretions of apolipoprotein B-containing lipoprotein and bile acids in HepG2

We studied the effect of NTE-122 (trans-1,4-bis[[1-cyclohexyl-3-(4-dimethylamino phenyl) ureido]methyl]cyclohexane), a novel acyl-CoA:cholesterol acyltransferase (ACAT) inhibitor, on intracellular cholesterol esterification and the secretion of apolipoprotein B100 (apoB)-containing lipoprotein and bile acids in the human hepatoma cell line HepG2. NTE-122 markably inhibited [3H]oleate incorporation into cholesteryl esters in HepG2 cells incubated with 5 microg/ml 25-hydroxycholesterol as a stimulus for ACAT (IC50=6.0 nM). On the other hand, NTE-122 did not affect [3H]oleate incorporation into triglycerides and phospholipids and [14C]acetate incorporation into cholesterol. The stimulation of ACAT by 25-hydroxycholesterol caused significant increases in the secretion of radiolabeled cholesteryl esters, radiolabeled triglycerides and apoB mass. NTE-122 pronouncedly inhibited the secretion of radiolabeled cholesteryl esters in proportion to the inhibition of cellular cholesterol esterification, and it significantly reduced the secretion of radiolabeled triglycerides and apoB mass in HepG2 cells incubated with 25-hydroxycholesterol. Furthermore, NTE-122 increased the secretion of bile acids synthesized from [14C]-cholesterol. These results suggest that NTE-122 is capable of exhibiting anti-hyperlipidemic effects by reducing both the cholesterol content and the amount of secreted very low-density lipoprotein and enhancing the excretion of bile acid from the liver.     

Mobilization of arachidonic acid from diacyl and ether-linked phospholipids in FMLP stimulated alveolar macrophages

Exposure of [1 14C]AA labeled guinea-pig alveolar macrophages to FMLP for 15 min induced an extensive mobilization of AA from phospholipids. PC and PI mainly contributed to the AA release, and labeled PE remained unchanged. Analysis of ether-linked phospholipids showed a significant breakdown of labeled diacyl and alkyl-acyl PC and an increase in labeled alkenyl-acyl PE.