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Biomedical subjects

S N Mathur

Publications and source records attributed to S N Mathur.

At least 37 records · Page 2Linked to original sources
PubMed1994

Regulation of apolipoprotein B secretion by biliary lipids in CaCo-2 cells.

The regulation of apoB synthesis and secretion by lipids present within bile was investigated in CaCo-2 cells grown on semipermeable filters. Bile acids decreased the basolateral secretion of immunoreactive apoB. Taurocholic acid decreased the secretion of newly synthesized apoB by increasing the rate of apoB degradation, but had no effect on the synthesis and secretion of apoA-I or trichloroacetic acid-precipitable proteins. The calcium ionophore, A23187, decreased apoB secretion similar to that observed for taurocholate. The addition of the ionophore and taurocholate together did not cause a further decrease in apoB secretion. Cholesterol or its hydroxylated derivative, 25-hydroxycholesterol, did not alter secretion of immunoreactive or newly synthesized apoB. Phosphatidylcholine increased apoB synthesis and secretion without affecting the synthesis or secretion of apoA-I. Phosphatidylcholine also reversed the effect of A23187 on apoB secretion. When phosphatidylcholine was added to the basolateral medium, apoB secretion was not altered. ApoB secretion was not increased by phospholipids of other classes. Dioleoylphosphatidylcholine increased apoB secretion, whereas dipalmitoylphosphatidylcholine did not. Fatty acid-labeled phosphatidylcholine was not hydrolyzed in the apical medium. Only 2% of the added phosphatidylcholine was cell-associated, and of this, 80% of the label remained as phosphatidylcholine with most of the remainder in triacylglycerols, fatty acids, and phosphatidylethanolamine. The results suggest that bile acids decrease apoB secretion by increasing its rate of degradation. This effect may be related to their ionophoric property. Cholesterol flux does not regulate apoB secretion. Phosphatidylcholine, independent of triacylglycerol flux and independent of its hydrolysis, increases the secretion of apoB by increasing apoB synthesis. Luminal phosphatidylcholine may play a role in apoB secretion in the intestine.

Apolipoproteins B↗
PubMed1993

Effect of okadaic acid on apo B and apo A-I secretion by CaCo-2 cells.

The effect of protein phosphorylation on the synthesis and secretion of apo B and apo A-I by CaCo-2 cells was investigated. Okadaic acid, a potent inhibitor of protein serine/threonine phosphatases 1 and 2A, caused a significant increase in total cellular protein phosphorylation. Apo B-48 was phosphorylated in control cells and this was increased significantly in the presence of okadaic acid. Under the experimental conditions, the phosphorylation of apo B-100 or apo A-I was not observed. No evidence of tyrosine phosphorylation of apo B-100, B-48, or apo A-I was found. Okadaic acid did not change the amount of apo B mass within cells but apo B mass secreted into the basolateral medium was decreased by 40%. Apo A-I mass within cells or in the basolateral medium was unaffected by okadaic acid. Despite causing an 18% decrease in total protein synthesis, okadaic acid did not alter the rate of synthesis of apo B-100, apo B-48, or apo A-I. Cellular turnover of labeled apo B-100 in cells incubated with okadaic acid was similar to controls, whereas apo B-48 and apo A-I turnover were slowed by okadaic acid. Compared to controls, however, 1 microM okadaic acid caused a 75% and 50% decrease in the secretion of newly synthesized apo B-100 and apo B-48, respectively, while decreasing labeled apo A-I secretion by 35%. In contrast to apo A-I mRNA levels, which were not altered by okadaic acid, apo B mRNA levels were significantly decreased by the polyether fatty acid. Despite differences observed in the phosphorylation state of apo B-100 and apo B-48, okadaic acid decreased the secretion of both forms of apo B without altering their synthesis. Okadaic acid, by increasing cellular protein phosphorylation, significantly disrupts the secretory processing of apo B by CaCo-2 cells.

Apolipoprotein A-I↗
PubMed1993

Regulation of LDL receptor expression by luminal sterol flux in CaCo-2 cells.

The regulation of expression of the intestinal low density lipoprotein (LDL) receptor by luminal (apical) sterol flux was investigated in the human intestinal cell line CaCo-2. Cells were cultured on semipermeable micropore filters, which separated an upper and lower well. To the apical media were added solutions containing either taurocholate micelles alone or micelles containing sterols. Because of an efflux of cholesterol, which occurred from cells incubated with micelles alone, LDL receptor mRNA levels increased threefold. With an influx of micellar sterols, receptor mRNA levels decreased in a dose-dependent manner. Synthesis and degradation of the LDL receptor were addressed by pulse-chase experiments. In cells incubated with micelles containing 25-hydroxycholesterol, the rate of receptor synthesis was significantly decreased, whereas the rate of receptor turnover remained unchanged. As assessed by immunoblots and steady-state labeling of proteins followed by immunoprecipitation of the LDL receptor, cells incubated with micellar 25-hydroxycholesterol contained substantially less receptor protein. These cells also bound and degraded less LDL. In contrast, in cells incubated with micelles alone, the rate of receptor synthesis was increased and cells contained more LDL receptor protein, although this was not reflected in an increased in LDL binding. The results suggest that LDL receptor expression in CaCo-2 cells is regulated by luminal sterol flux and that this regulation occurs at the level of transcription.

Cell Line↗
PubMed1993

Cholesterol and sphingomyelin syntheses are regulated independently in cultured human intestinal cells, CaCo-2: role of membrane cholesterol and sphingomyelin content.

There is a presumed association between cellular cholesterol and sphingomyelin metabolism. To study this relationship in the intestine, the activity of the rate controlling enzyme of sphingolipid synthesis, serine palmitoyltransferase (SPT), and the biosynthesis of long-chain bases were characterized in cultured human intestinal cells, CaCo-2. Cells were then incubated with substances known to alter cholesterol biosynthesis, and the effect of these mediators on SPT activity and long-chain base synthesis was determined and compared with their effects on HMG-CoA reductase activity and cholesterol synthesis. The polar sterol, 25-hydroxycholesterol, the squalene epoxide inhibitor, U18666A, and the inhibitor of HMG-CoA reductase, lovastatin, all significantly inhibited the synthesis of cholesterol without altering either SPT activity or long-chain base synthesis. Mevalonate, which increased cholesterol production 3-fold, also had no affect on SPT activity or sphingoid base synthesis. Serine, which significantly increased the synthesis of long-chain bases, did not alter cholesterol biosynthesis. Moreover, the suicide inhibitors of SPT, beta-chloroalanine and cycloserine, did not alter cholesterol synthesis while markedly decreasing long chain base synthesis. Cells were incubated with palmitic, oleic, linoleic, and eicosapentaenoic acids. Only palmitic acid, the preferred substrate for SPT, increased the production of long-chain bases. Both palmitic and oleic acids, however, increased the synthesis of cholesterol. Cells enriched in sphingomyelin had higher rates of synthesis of both cholesterol and long-chain bases compared to their controls. In contrast, cholesterol and long-chain base syntheses were significantly decreased in cells enriched in cholesterol. Control cells incubated with phospholipid liposomes alone had higher rates of synthesis of both lipids.(ABSTRACT TRUNCATED AT 250 WORDS)

Acyltransferases↗
PubMed1992

Sphingomyelin content of intestinal cell membranes regulates cholesterol absorption. Evidence for pancreatic and intestinal cell sphingomyelinase activity.

Micellar cholesterol uptake and secretion were investigated in the human intestinal cell line CaCo-2 following depletion of apical membrane sphingomyelin. The addition of exogenous sphingomyelinase, which hydrolysed 60% of prelabelled sphingomyelin, resulted in a 50% decrease in the uptake of cholesterol from bile salt micelles. The flux of membrane cholesterol into the cell by the hydrolysis of membrane sphingomyelin decreased the rate of cholesterol synthesis by 43% and inhibited hydroxymethylglutaryl-CoA reductase activity by 54%. Moreover, the rate of cholesterol esterification was increased 4-fold. Total cellular cholesterol mass was unchanged by the addition of sphingomyelinase; however, cholesteryl esters increased by 50% and the amount of unesterified cholesterol decreased significantly. The basolateral secretion of cholesterol mass was also decreased following sphingomyelin hydrolysis. Human pancreatic juice was found to contain neutral sphingomyelinase activity which required taurocholate for full expression. The presence of neutral sphingomyelinase activity was also documented in membranes prepared from CaCo-2 cells and in whole homogenates from human duodenal biopsies. The data suggest that the amount of sphingomyelin present in the apical membrane of the intestinal absorptive cell regulates cholesterol uptake from bile salt micelles. Sphingomyelinase activity within intestinal cells and in pancreatic juice could alter the sphingomyelin content of brush-border membranes of small intestinal absorptive cells and thus regulate the amount of cholesterol absorbed by the gut.

Cells, Cultured↗
PubMed1992

Differential metabolism of hydroxyeicosatetraenoic acid isomers by mouse cerebromicrovascular endothelium.

Hydroxyeicosatetraenoic acid (HETE) derivatives of arachidonic acid are produced in the brain and have been implicated as pathologic mediators in various types of brain injury. To understand better their fate in the brain, particularly in cerebral microvessels, several HETEs were incubated with cultured mouse cerebromicrovascular endothelium for 1, 2, and 4 h, followed by HPLC analysis of medium and cellular lipids. 5(S)-, 8(RS)-, and 9(RS)-HETE were not metabolized by the cells, but were extensively incorporated, unmodified, into cell lipids. On the other hand, 11(RS)-, 12(S)-, and 15(S)-HETE were extensively metabolized and only minimally incorporated into cell lipids. Previously, the major 12-HETE metabolite was identified as 8-hydroxyhexadecatrienoic acid. In the present study, we identified the major 11-HETE metabolite as 7-hydroxyhexadecatrienoic acid and the major 15-HETE metabolite as 11-hydroxyhexadecatrienoic acid. omega-3 compounds, 15(S)- and 12(S)-hydroxyeicosapentaenoic acids (HEPE), were also metabolized to more polar compounds, but to a lesser extent than their tetraenoic acid, omega-6 counterparts. Comparison of 5-, 12-, and 15-HETE enantiomers revealed no differences in metabolism or incorporation between the R and S stereoisomers. These data suggest that many isomers of HETE and HEPE can be incorporated into cell lipids or metabolized by pathways that do not distinguish between enantiomers. These pathways, however, are sensitive to the position or number of double bonds and are selective based on the position of the hydroxyl group.

Animals↗
PubMed1992

Apolipoprotein B mRNA abundance is decreased by eicosapentaenoic acid in CaCo-2 cells. Effect on the synthesis and secretion of apolipoprotein B.

The regulation of apolipoprotein B (apo B) metabolism by eicosapentaenoic acid was investigated in CaCo-2 cells. Cells cultured on semipermeable membranes that separated an upper from a lower well were incubated for 48 hours with albumin alone or 1 mM eicosapentaenoic acid or oleic acid attached to albumin (4:1, mol/mol). Compared with cells incubated with oleic acid, cells incubated with eicosapentaenoic acid synthesized and secreted less [3H]glycerol-labeled triglycerides. Although both fatty acids increased cellular triglyceride mass compared with control cells, less triglycerides accumulated in cells incubated with the n-3 fatty acid. The secretion of triglyceride and apo B mass by cells incubated with eicosapentaenoic acid was less than that observed by cells incubated with oleate. The amount of apo B mass within cells, however, was not altered by either of the fatty acids and was similar to amounts found in control cells. Apo B mRNA abundance was decreased fourfold in cells exposed for 48 hours to eicosapentaenoic acid. In contrast, in cells incubated with oleic acid, apo B mRNA levels were not significantly altered. Pulse-chase experiments were performed to investigate the regulation of apo B synthesis and degradation by the fatty acids. In cells incubated with eicosapentaenoic acid, the synthesis and basolateral secretion of newly synthesized apo B-100 and apo B-48 were significantly less compared with control cells or cells incubated with oleic acid. In contrast, the synthesis and secretion of newly synthesized apo B in cells exposed to oleic acid were similar to control cells. Rates of apo A-I synthesis were similar in cells incubated with either of the fatty acids. Compared with control cells and cells incubated with eicosapentaenoic acid, the residence time of labeled apo B in cells incubated with oleic acid was prolonged. The percentage of newly synthesized apo B that was degraded was less in cells incubated with oleic acid. In contrast, residence times and the percentages of apo A-I and apo B-48 degraded were similar in control cells and cells incubated with the fatty acids. Thus, in CaCo-2 cells, compared with the effects of oleic acid, eicosapentaenoic acid impairs triglyceride transport in part by inhibiting apo B synthesis and secretion. The inhibition of apo B synthesis by eicosapentaenoic acid may be related to a decrease in gene transcription or a decrease in mRNA stability, as apo B mRNA levels were significantly decreased in cells incubated with this fatty acid.(ABSTRACT TRUNCATED AT 400 WORDS)

Apolipoprotein A-I↗
PubMed1991

Regulation of gene expression and synthesis and degradation of 3-hydroxy-3-methylglutaryl coenzyme A reductase by micellar cholesterolin CaCo-2 cells.

To investigate whether, and by what mechanisms, luminal (dietary) cholesterol regulates cholesterol synthesis in human intestinal cells, HMG-CoA reductase activity, gene expression, synthesis, and degradation were investigated in CaCo-2 cells exposed to taurocholate micelles containing cholesterol. In cells incubated with cholesterol solubilized in 5 mM taurocholate and 30 microM monoolein, HMG-CoA reductase activity was decreased. 25-Hydroxycholesterol, delivered to the cells in the same manner as native cholesterol, was significantly more potent in inhibiting reductase activity and was used, therefore, to investigate mechanisms for sterol regulation. Cells incubated with taurocholate micelles without cholesterol lost cellular cholesterol into the medium causing an increase in HMG-CoA reductase activity and enzyme mass. Although steady-state levels of HMG-CoA reductase mRNA were increased under conditions of cholesterol efflux, synthesis rates of reductase protein were not increased. An increase in activity and enzyme mass in cells incubated with micelles alone, however, was accompanied by a significant decrease in the rate of degradation of reductase protein. In contrast, sterol influx from taurocholate micelles was associated with a marked decrease in HMG-CoA reductase activity and mass without altering mRNA levels except at high concentrations of the polar sterol which did decrease reductase mRNA levels by 50%. The absorption of apical sterol resulted in a significant decrease in the translational efficiency of reductase mRNA and a modest increase in the rate of degradation of the enzyme. Thus, although the primary function of the enterocyte is to transport luminal (dietary) cholesterol to other tissues of the body, apically derived cholesterol enters metabolic pools within the cell which regulates its own cholesterol synthesis. Dietary cholesterol, therefore, will regulate the contribution to the total body cholesterol pool of endogenously derived cholesterol from the intestine. The mechanism for this regulation of intestinal HMG-CoA reductase by luminal cholesterol occurs primarily at the post-transcriptional level.

Cell Line↗
PubMed1990

12-Hydroxyeicosatetraenoic acid is metabolized by beta-oxidation in mouse peritoneal macrophages. Identification of products and proposed pathway.

The products derived from the metabolism of 12-hydroxyeicosatetraenoic acid (12-HETE) by mouse peritoneal macrophages were characterized by high performance liquid chromatography (HPLC) and GC-mass spectrometry. HPLC analysis demonstrated two predominant polar products and several minor ones. The proportion and amounts of these products were dependent on the concentration of 12-HETE, the number of macrophages incubated with the monohydroxy fatty acid, and the time of incubation. The products identified by GC-mass spectrometry suggested that 12-HETE had undergone beta-oxidation. The intermediates identified were: 3,12-dihydroxy-5,8,10,14, 20:4; 10-hydroxy-3,6,8,12, 18:4; 3,10-dihydroxy-6,8,12, 18:3; 8-hydroxy-4,6,10, 16:3; 6-hydroxy-4,8, 14:2; and 4-hydroxy, 12:1. The major products, as identified by HPLC and GC-mass spectrometry, were 8-hydroxy-4,6,10, 16:3 and 4-hydroxy, 12:1. A minor product, 10-hydroxy-6,8,12, 18:3 was postulated to arise from either the isomerization and reduction of 10-hydroxy-3,6,8,12, 18:4 or from chain elongation of 8-hydroxy-4,6,10, 16:3. Inhibiting cyclooxygenase and lipoxygenase activities by ibuprofen and nordihydroguaiaretic acid, respectively, did not inhibit the formation of these products. 82% to 98% of 12-HETE was converted and released into the medium as products of beta-oxidation. The remainder was taken up into cellular lipids. beta-Oxidation of 12-HETE was decreased by only 12 and 21% after inhibiting mitochondrial fatty acid oxidation by 89 and 93% by 5 and 100 microM concentrations of the mitochondrial fatty acid oxidation inhibitor, methyl palmoxirate, respectively. It is thus postulated that the beta-oxidation of 12-HETE by mouse peritoneal macrophages occurs in peroxisomes.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
PubMed1990

Effect of eicosapentaenoic acid on triacylglycerol transport in CaCo-2 cells.

The human intestinal cell line, CaCo-2, was used to study the effect of the n-3 fatty acid, eicosapentaenoic acid, on triacylglycerol secretion. In cells incubated with 250 microM eicosapentaenoic acid, the incorporation of [3H]glycerol into triacylglycerols secreted into the medium was decreased by 58% compared to cells incubated with 250 microM oleic acid. The incorporation of [3H]glycerol into cellular triacylglycerols was decreased 32% in cells incubated with eicosapentaenoic acid. In cells preincubated with [3H]glycerol to label existing triacylglycerols, the rates of secretion of preformed triacylglycerols were similar in response to the addition of either fatty acid. Initial uptake rates of the n-3 fatty acid were higher than for oleic acid. Both eicosapentaenoic acid and oleic acid were minimally oxidized to CO2. Oleic acid was predominantly incorporated into cellular triacylglycerols (62% vs. 47%), whereas more eicosapentaenoic acid was incorporated into cellular phospholipids (46% vs. 30%). Phospholipids of microsomes prepared from cells incubated with eicosapentaenoic acid were enriched in this fatty acid. The rate of synthesis of triacylglycerol and diacylglycerol acyltransferase activities were significantly less in microsomes prepared from cells incubated with eicosapentaenoic acid. Triacylglycerol mass secreted by CaCo-2 cells incubated with either fatty acid was similar. In CaCo-2 cells, eicosapentaenoic acid decreases the synthesis and secretion of newly synthesized triacylglycerol without decreasing the secretion of triacylglycerol mass. Modification of microsomal membrane phospholipid fatty acid composition is associated with a decrease in microsomal triacylglycerol synthesis and diacylglycerol acyltransferase activities.

Biological Transport↗
PubMed1990

Regulation of cholesterol metabolism in the intestine.

The small intestine is a major site of cholesterol biosynthesis and lipoprotein degradation. It is also the organ responsible for absorbing dietary and endogenously produced biliary cholesterol. Cholesterol metabolism in the intestine is regulated by factors that will alter cellular cholesterol requirements. Thus, during increased cholesterol flux, which occurs by bile acid-faciliated cholesterol absorption or by lipoprotein-mediated uptake of cholesterol, cholesterol synthetic rates decrease and esterification rates increase. The mechanisms by which dietary fats regulate intestinal cholesterol metabolism are complex. Dietary fats alter membrane fatty acid composition. Simultaneously, they also promote lipoprotein secretion and alter cholesterol absorption. Intestinal 3-hydroxyl-3-methylglutaryl coenzyme. A reductase activity is regulated by enzyme phosphorylation-dephosphorylation. The regulation of acylcoenzyme A-cholesterol acyltransferase activity by this mechanism remains controversial. Data on hormone regulation of intestinal cholesterol metabolism are not conclusive, although progesterone seems to be a potent inhibitor of acylcoenzyme A-cholesterol acyltransferase activity in intestinal cell culture and isolated cells. In a manner similar to the regulation of cholesterol metabolism in other cells, the enterocyte responds appropriately to factors that alter cholesterol flux. Therefore, changes that occur in the rates of cholesterol synthesis and esterification will reflect the cholesterol requirements of the cell.

Animals↗
PubMed1989

Regulation of 12-hydroxyeicosatetraenoic acid synthesis by acetyl-LDL in mouse peritoneal macrophages.

The mechanism for the regulation of 12-hydroxyeicosatetraenoic acid (12-HETE) production by cholesterol-rich macrophages was investigated. beta-VLDL and acetyl-LDL, lipoproteins which result in cholesterol accumulation in macrophages, stimulated 12-HETE secretion. Lipoproteins which do not induce cholesterol accumulation, such as low- and high-density lipoproteins, did not. Cell-free homogenates from cholesterol-rich macrophages had significantly more 12-lipoxygenase activity than homogenates from unmodified cells. Preincubating homogenates prepared from unmodified macrophages with acetyl-LDL, LDL or multilamellar liposomes containing total lipids from acetyl-LDL but not apoproteins significantly increased 12-lipoxygenase activity. This stimulatory effect was caused by the phospholipid moiety of the lipoprotein. 12-HETE synthesis was not increased in macrophages enriched 6-fold in unesterified cholesterol. Acetyl-LDL stimulated 12-HETE synthesis in macrophages in which cholesteryl ester accumulation was prevented by inhibiting acylcoenzyme A:cholesterol acyltransferase activity. When binding of acetyl-LDL to its receptor was decreased by increasing concentrations of dextran sulfate, or when lysosomal metabolism of the lipoprotein was prevented by chloroquine, 12-HETE production significantly decreased. Moreover, the combination of inhibiting acetyl-LDL binding and degradation completely blocked the stimulation of 12-HETE synthesis by acetyl-LDL. The data indicate that acetyl-LDL must enter the macrophage and be partially degraded to regulate 12-HETE synthesis. The regulation is independent of cholesterol accumulation but is related to the entering lipoprotein phospholipid.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
PubMed1989

Effects of phospholipid or cholesterol enrichment of rat intestinal brush border membrane on membrane order and transport of calcium.

Calcium uptake by brush border membrane vesicles from rat small intestine measured under initial rate conditions comprises both saturable and nonsaturable components. Because the brush border is a lipid bilayer and may be sensitive to changes in membrane lipid, vesicles were treated with liposomes to enrich phospholipid (PL) or cholesterol (C) content above that of the control (Reference) vesicle. The effects of the changes in lipid composition on membrane fluidity were determined from fluorescence anisotropy (r) of diphenylhexatriene. Compared with Reference vesicles, liposome-treated vesicles showed decreased Vmax for saturable and KD for nonsaturable uptakes. Liposome treatment changed vesicle phospholipid composition compared with Reference vesicles. Liposome-treated vesicles had similar phospholipid composition but differed in greater cholesterol content of C- compared with PL-vesicles. Mean Vmax and KD were lower in C- than PL-vesicles, but the difference did not reach statistical significance, although fluidity was significantly lower in C- than PL-vesicles. The mechanism of inhibition of saturable calcium uptake in PL- and C-vesicles was uncompetitive. Thus, lipid composition is crucial for determining calcium uptake: any change from native lipid composition decreased transport. Fluidity, measured by the conventional probe diphenylhexatriene, did not correlate with calcium uptake by Reference compared with liposome-treated vesicles.

Animals↗
PubMed1989

Decreased prostaglandin production by cholesterol-rich macrophages.

The regulation of prostaglandin production by macrophages enriched in cholesterol was examined. Mouse peritoneal macrophages were incubated for 18 h with 25 micrograms/ml of human acetyl-LDL (low density lipoprotein) and trace amounts of labeled arachidonic acid. After cholesterol enrichment, the cells were incubated with phorbol 12-myristate 13-acetate (PMA), calcium ionophore, or zymosan to stimulate endogenous arachidonic acid metabolism. A high performance liquid chromatography profile of the eicosanoids released revealed no qualitative differences between unmodified and modified macrophages. Cholesterol-rich cells, however, released less prostacyclin (PGI2) and prostaglandin E2 (PGE2) compared to unmodified cells, and products from the lipoxygenase pathway became the predominant metabolites. A decrease in the synthesis of PGI2 and PGE2 by cholesterol-rich macrophages was confirmed by radioimmunoassay and radiolabeled experiments. The activity of prostaglandin synthetase was modestly increased in the cholesterol-modified macrophages compared to controls. As an estimation of phospholipase activity, the release of labeled arachidonic acid from membrane phospholipids, however, was significantly decreased in cholesterol-rich macrophages. The phosphatidylinositol fraction was particularly resistant to arachidonate release in response to calcium ionophore and PMA in the modified cells. The measurement of membrane phospholipid fatty acid composition before and after calcium ionophore supported the observation that less arachidonate was released by cholesterol-enriched cells in response to the ionophore. Based on these observations, we propose that prostaglandin synthesis from endogenous arachidonate stores is decreased in the cholesterol-rich macrophage. A decrease in agonist-induced activation of the phospholipase activity is proposed as a mechanism for this effect.

Animals↗
PubMed1989

Inhibition of acylcoenzyme A:cholesterol acyltransferase activity in CaCo-2 cells results in intracellular triglyceride accumulation.

The activity of acylcoenzyme A:cholesterol acyltransferase (ACAT) in CaCo-2 cells was inhibited by the ACAT inhibitor, 58-035. The inhibitory effect of this acylamide was specific for cholesterol esterification catalyzed by ACAT; the rates of triglyceride, phospholipid, and cholesterol synthesis were not inhibited by this agent. Cholesteryl esters were depleted in CaCo-2 cells 24 hr after inhibition of ACAT activity, whereas the unesterified cholesterol content increased by 56% after 96 hr. Moreover, inhibiting ACAT activity with 58-035 resulted in a time-dependent 2.5-fold increase in intracellular triglycerides. This accumulation of triglycerides in CaCo-2 cells was associated with a 37% increase in triglyceride synthesis by 96 hr in the presence of 58-035. Triglyceride-rich lipoprotein secretion (d less than 1.006 g/ml) was not affected by inhibiting ACAT activity for up to 6 hr. However, triglyceride-rich lipoprotein secretion was significantly decreased in CaCo-2 cells that were preincubated with 58-035 for 24 to 96 hr. Lipoproteins of density less than 1.006 g/ml that were isolated from CaCo-2 cells incubated with the ACAT inhibitor were deficient in cholesteryl esters and triglycerides compared to lipoproteins isolated from control cells. The data suggest that triglycerides accumulate in CaCo-2 cells in which ACAT activity has been inhibited by 58-035. This accumulation of triglycerides is associated with a modest increase in triglyceride synthesis and a decrease in triglyceride secretion. Altering intracellular cholesterol pools by regulating ACAT activity in the gut could result in the decrease of triglyceride transport and/or the secretion of triglyceride-rich lipoprotein particles of abnormal composition.

Acyl Coenzyme A↗
PubMed1989

Effect of phenylbutazone on acute hemorrhagic pancreatitis in dogs.

Experiments were conducted on 24 mongrel dogs to study the effect of phenylbutazone on acute experimental pancreatitis. Necrotico-hemorrhagic pancreatitis was produced by local infiltration of autologous bile. The severity of pancreatitis was assessed by biochemical estimation and histopathological examination. Pretreatment with phenylbutazone reduced the severity of pancreatitis, both biochemically and histologically (total score 6.0 +/- 1.52 in the test group vs 8.33 +/- 1.80 in the control group; p less than 0.01).

Animals↗
PubMed1988

Incorporation of lipoxygenase products into cholesteryl esters by acyl-CoA:cholesterol acyltransferase in cholesterol-rich macrophages.

Macrophages which were incubated with acetylated low-density lipoproteins, resulting in cholesteryl ester accumulation, incorporated the monohydroxyeicosatetraenoic acids (5-, 15-, and 12-HETEs) into cholesteryl esters. The esterification of these hydroxy fatty acids to cholesterol by total membrane preparations of cholesterol-rich macrophages was dependent on the synthesis of the fatty acyl-CoA derivative, and was catalysed by acyl-CoA:cholesterol acyltransferase (ACAT). Stimulation of membrane ACAT activity by 25-hydroxycholesterol increased the synthesis of cholesteryl 12-HETE by 40%. In contrast, inhibiting ACAT activity by progesterone and compound 58-035 decreased cholesteryl 12-HETE production by 60% and 90% respectively. Although 5-, 15- and 12-HETE were esterified to cholesterol by ACAT, these monohydroxy fatty acids were less optimal as substrates compared with oleic acid or arachidonic acid. The hydrolysis and release of 12-HETE and the other monohydroxyeicosatetraenoic acids from intracellular cholesteryl esters and phospholipids occurred at a faster rate than for the more conventional fatty acids, oleate and arachidonate. Cholesteryl esters which contain hydroxy fatty acids therefore provide only a transient storage for lipoxygenase products, as these fatty acids are released into the medium as readily as hydroxy fatty acids found in phospholipids and triacylglycerols. The data provide evidence, for the first time, of an ACAT-dependent esterification of the lipoxygenase products 5-, 15- and 12-HETEs to cholesterol in the macrophage-derived foam cell. The channelling of these monohydroxy fatty acids to cholesteryl esters provides a mechanism which can alter the amount of lipoxygenase products incorporated into cellular phospholipids, thus averting deleterious changes to cell membranes. ACAT, by catalysing the esterification of monohydroxyeicosatetraenoic acids to cholesterol, could play a key role in regulating the amount of lipoxygenase products in the pericellular space of the cholesterol-enriched macrophage.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗