Lipid synthesis in macrophages derived from the human cell line THP-1: modulation of the effects of native and oxidized chylomicron-remnant-like particles by oestrogen

2001 ◽  
Vol 101 (4) ◽  
pp. 403-413 ◽  
Author(s):  
Mariarosaria NAPOLITANO ◽  
Kelly V. BATT ◽  
Michael AVELLA ◽  
Elena BRAVO ◽  
Kathleen M. BOTHAM
Keyword(s):  
Cholesteryl Ester ◽  
Cell Line ◽  
Lipid Synthesis ◽  
Thiobarbituric Acid ◽  
Human Monocyte ◽  
Cholesterol Esterification ◽  
Chylomicron Remnant ◽  
Triacylglycerol Synthesis ◽  
Ester Formation ◽  
Chylomicron Remnants

The effects of native and oxidized chylomicron remnants on the synthesis of cholesteryl ester and triacylglycerol in macrophages, and the way that this is influenced by exposure of the cells to oestrogen, was investigated using the human monocyte cell line THP-1 and chylomicron-remnant-like particles containing human apolipoprotein E (CRLPs). Synthesis of the lipids was measured by the incorporation of [3H]oleate into cholesteryl ester and triacylglycerol. CRLPs (5-40μg of cholesterol/ml) containing either trilinolein or triolein as the triacylglycerol component caused a dose-dependent decrease in cholesteryl ester formation, while triacylglycerol production was unchanged. After oxidation of the CRLPs, the level of thiobarbituric acid-reactive substances was increased by 6.3-fold and 2.2-fold in particles containing trilinolein and triolein respectively. Furthermore, CRLPs containing oxidized trilinolein lost their ability to down-regulate cholesterol esterification, while CRLPs containing oxidized triolein did not. Both types of oxidized CRLPs decreased triacylglycerol synthesis. Treatment of the macrophages with 17β-oestradiol caused increases of approx. 94% and 34% in the synthesis of cholesteryl ester and triacylglycerol respectively in the absence of CRLPs. The differences between control and oestrogen-treated cells were abolished, however, when CRLPs (40μg of cholesterol/ml) were added to the incubations. In addition, in contrast with their lack of effect in control cells, CRLPs containing oxidized trilinolein decreased cholesterol esterification in oestrogen-treated cells by approx. 48%. These findings with CRLPs suggest that chylomicron remnants have significant effects on cholesteryl ester and triacylglycerol synthesis in macrophages, which may be modulated both by the oxidation state of the particles and by oestrogen.

scholarly journals Phospholipase A2 Mediates Apolipoprotein-Independent Uptake of Chylomicron Remnant-Like Particles by Human Macrophages

2012 ◽  
Vol 2012 ◽  
pp. 1-11 ◽  
Author(s):  
Mariarosaria Napolitano ◽  
Howard S. Kruth ◽  
Elena Bravo
Keyword(s):  
Phospholipase A2 ◽  
Cholesteryl Ester ◽  
Apolipoprotein E ◽  
Lipoprotein Lipase ◽  
Human Monocyte ◽  
Cholesteryl Ester Hydrolase ◽  
Chylomicron Remnant ◽  
Ester Hydrolase ◽  
Chylomicron Remnants ◽  
Cytosolic Pla2

Apolipoprotein E-receptor-mediated pathways are the main routes by which macrophages take up chylomicron remnants, but uptake may also be mediated by receptor-independent routes. To investigate these mechanisms, triacylglycerol (TG) accumulation induced by apolipoprotein-free chylomicron remnant-like particles (CRLPw/o) in human monocyte-derived macrophages was evaluated. Macrophage TG content increased about 5-fold after incubation with CRLPw/o, and this effect was not reduced by the inhibition of phagocytosis, macropinocytosis, apolipoprotein E function, or proteoglycan bridging. The role of lipases, including lipoprotein lipase, cholesteryl ester hydrolase, and secretory (sPLA2) and cytosolic phospholipase A2, was studied using [3H]TG-labelled CRLPw/o. Total cell radioactivity after incubation with [3H]TG CRLPw/o was reduced by 15–30% by inhibitors of lipoprotein lipase and cholesteryl ester hydrolase and by about 45% by inhibitors of sPLA2 and cytosolic PLA2. These results suggest that macrophage lipolytic enzymes mediate the internalization of postprandial TG-rich lipoproteins and that sPLA2and cytosolic PLA2, play a more important role than extracellular lipoprotein lipase-mediated TG hydrolysis.

scholarly journals Phosphatidylcholine increases the secretion of triacylglycerol-rich lipoproteins by CaCo-2 cells

1996 ◽  
Vol 314 (2) ◽  
pp. 569-575 ◽  
Author(s):  
Satya N. MATHUR ◽  
Ella BORN ◽  
Shubha MURTHY ◽  
F. Jeffrey FIELD
Keyword(s):  
Cholesteryl Ester ◽  
Secretory Pathway ◽  
Cholesterol Acyltransferase ◽  
Lipid Synthesis ◽  
Brefeldin A ◽  
Mrna Levels ◽  
Ester Synthesis ◽  
Acyltransferase Activity ◽  
Apob Mrna ◽  
Triacylglycerol Synthesis

The regulation of lipid synthesis and secretion by phosphatidylcholine was investigated in CaCo-2 cells grown on semipermeable filters. In cells incubated with 1 mM taurocholate and 100–500 μM phosphatidylcholine, cholesteryl ester synthesis was decreased, triacylglycerol synthesis was increased modestly, whereas phospholipid synthesis was unaffected. Acyl-CoA–cholesterol acyltransferase activity was decreased secondary to a decrease in the substrate (cholesterol) supply. The basolateral secretion of newly synthesized triacylglycerol and triacylglycerol mass was significantly increased by phosphatidylcholine, whereas cellular triacylglycerol mass decreased. This effect was not specific for phosphatidylcholine as phosphatidylethanolamine and phosphatidylserine also increased the secretion of newly synthesized triacylglycerols. Dioleoylphosphatidylcholine was as effective as egg phosphatidylcholine in increasing triacylglycerol transport. Dipalmitoylphosphatidylcholine, in contrast, was without effect. Phosphatidylcholine also increased the basolateral secretion of apolipoprotein B (apoB) mass without altering apoB mRNA levels. Disruption of the Golgi apparatus by monensin or brefeldin A prevented the increase in apoB secretion by phosphatidylcholine. Compared with microsomes prepared from control cells, those from cells incubated with phosphatidylcholine contained more newly synthesized apoB. The percentage of new synthesized apoB isolated from the lumen of microsomes (as an estimate of apoB destined for secretion), however, was similar in the two preparations. Thus in CaCo-2 cells incubated with phosphatidylcholine, the transport of apoB and triacylglycerols is increased whereas cholesteryl ester synthesis and secretion are decreased. A normally functioning secretory pathway is required for phosphatidylcholine to increase triacylglycerol-rich lipoprotein secretion.

scholarly journals Lysophosphatidylcholine increases the secretion of cholesteryl ester-poor triacylglycerol-rich lipoproteins by CaCo-2 cells

1994 ◽  
Vol 304 (1) ◽  
pp. 35-42 ◽  
Author(s):  
F J Field ◽  
E Born ◽  
H Chen ◽  
S Murthy ◽  
S N Mathur
Keyword(s):  
Cholesteryl Ester ◽  
Cholesterol Esterification ◽  
Ester Synthesis ◽  
Triacylglycerol Synthesis ◽  
Apoprotein B ◽  
Apob Synthesis ◽  
Mass Accumulation

To address the effect of lysophosphatidylcholine on triacylglycerol transport in intestine, CaCo-2 cells, grown on semipermeable supports, were incubated with lysophosphatidylcholine solubilized in 1 mM taurocholate. [14C]Palmitoyllysophosphatidylcholine was readily taken up and incorporated predominantly into cellular phospholipids, particularly phosphatidylcholine. Twenty-five percent of the label was found in triacylglycerols. Compared with labelled cellular phospholipids, labelled triacylglycerols were preferentially secreted. Lysophosphatidylcholine caused a profound decrease in cholesteryl ester synthesis and secretion, whereas cellular triacylglycerol mass and triacylglycerol synthesis and secretion were increased. The effect was more pronounced with oleoyllysophosphatidylcholine than with either palmitoyl- or stearyl-lysophosphatidylcholine. Lysophosphatidylcholine increased the secretion of immunoreactive and newly-synthesized apoprotein B (apoB) without altering the rate of apoB synthesis. Thus, luminal lysophosphatidylcholine and/or its uptake decreases cholesterol esterification and secretion, but increases triacylglycerol synthesis and secretion, triacylglycerol mass accumulation and the secretion of apoB by CaCo-2 cells.

scholarly journals Chylomicron-remnant uptake by freshly isolated hepatocytes. Effect of heparin and of hepatic triacylglycerol lipase

1989 ◽  
Vol 258 (2) ◽  
pp. 587-594 ◽  
Author(s):  
F Sultan ◽  
D Lagrange ◽  
X Le Liepvre ◽  
S Griglio
Keyword(s):  
Cholesteryl Ester ◽  
Hepatic Lipase ◽  
Isolated Hepatocytes ◽  
Chylomicron Remnant ◽  
Triacylglycerol Lipase ◽  
Cell Pellet ◽  
Chylomicron Remnants ◽  
Dose Response Study ◽  
Cholesteryl Ester Accumulation

Chylomicron remnants labelled biologically with [3H]cholesterol were efficiently taken up by freshly isolated hepatocytes during a 3 h incubation in Krebs bicarbonate medium. Their [3H]cholesteryl ester was hydrolysed (74% net hydrolysis), and 0.1 mM-chloroquine could partially inhibit this hydrolysis, provided that hepatocytes were first preincubated for 2 h 30 min at 37 degrees C. This hydrolysis was also measured in preincubated cells with remnants double-labelled (3H and 14C) on their free cholesterol moiety; [3H]cholesterol arising from [3H]cholesteryl ester hydrolysis was recovered in the free [3H]cholesterol pool. A dose-response study showed saturation of remnant uptake at 180 micrograms of remnant protein/10(7) cells. Heparin (10 units/ml) increased remnant uptake by 63% (P less than 0.01), [3H]cholesteryl ester accumulation in the cell pellet by 110% (P less than 0.025) and hepatic lipase activity secreted in the medium by 2.4-fold (P less than 0.01) and by 3.3-fold (P less than 0.01) at the end of the preincubation and incubation periods respectively. Addition of 100 munits of semi-purified hepatic lipase preparation/flask stimulated remnant uptake by 44-69%, and [3H]cholesteryl ester accumulation in the presence of chloroquine by 2.1-fold (P less than 0.025). When hepatic lipase was incubated solely with the remnants, it decreased their triacylglycerol and phospholipid contents by 24% and 26% respectively. Thus freshly isolated hepatocytes may be used to study chylomicron-remnant uptake. Hepatic lipase, which seems to underly the stimulating effect of heparin, facilitates remnant uptake in vitro, and this could be mediated by at least one (or both) of its hydrolytic properties.

scholarly journals Binding, interiorization and degradation of cholesterol ester-labelled chylomicron-remnant particles by rat hepatocyte monolayers

1977 ◽  
Vol 168 (3) ◽  
pp. 483-494 ◽  
Author(s):  
Claes-Henrik Florén ◽  
Åke Nilsson
Keyword(s):  
Cell Surface ◽  
Cholesteryl Ester ◽  
Time Course ◽  
Inhibitory Effect ◽  
Metabolic Inhibitors ◽  
High Density Lipoproteins ◽  
Chylomicron Remnant ◽  
Very Low Density Lipoproteins ◽  
Chylomicron Remnants ◽  
Hydrolysis Of

1. The cholesteryl ester of isolated chylomicron-remnant particles was efficiently degraded by hepatocyte monolayers. The degradation was sensitive to metabolic inhibitors. 2. With increasing amounts of remnant cholesteryl ester the rate of uptake approached saturation and conformed to a linear double-reciprocal plot. The Vmax. was determined as 80ng of cholesteryl ester/h per mg of protein and the apparent Km as 1.4μg of cholesteryl ester per mg of protein. The time course for the uptake and hydrolysis suggested that binding of particles to the cell surface preceded the degradation. 3. Cholesteryl esters of native chylomicrons were degraded to a much smaller extent and their presence had only a small inhibitory effect on the degradation of chylomicron remnants. Intestinal very-low-density lipoproteins were degraded somewhat faster than chylomicrons, and caused more inhibition of remnant degradation. Rat high-density lipoproteins inhibited the hydrolysis of remnant cholesteryl ester by up to 50%, but had less influence on the amount of cholesteryl ester that was bound to the cells. Serum decreased both the uptake and hydrolysis, whereas d=1.21 infranatant had no effect. 4. The cholesteryl ester hydrolysis after the uptake by the cells was inhibited by chloroquine and by colchicine. Only 28–36% of the unhydrolysed cholesteryl ester could be released from these cells by trypsin treatment, indicating that the major portion was truly intracellular. The particles that could be released from the cell surface by trypsin and those remaining in the medium had the same triacylglycerol/cholesteryl ester ratio as the added remnant particles. Significant amounts of denser particles were thus not formed during contact with the cell surface. 5. The presence of heparin, as well as preincubation of the cells with heparin, increased the uptake of chylomicron remnants. This effect was most marked in the presence of serum. A much smaller proportion of the other serum lipoproteins was taken up, and this proportion was not increased by heparin.

scholarly journals Chylomicron-remnant clearance in homozygote and heterozygote Watanabe-heritable-hyperlipidaemic rabbits is defective. Lack of evidence for an independent chylomicron-remnant receptor

1991 ◽  
Vol 276 (2) ◽  
pp. 381-386 ◽  
Author(s):  
A Bowler ◽  
T G Redgrave ◽  
J C L Mamo
Keyword(s):  
Cholesteryl Ester ◽  
Plasma Cholesterol ◽  
Low Density Lipoproteins ◽  
Low Density ◽  
Lipid Emulsions ◽  
Chylomicron Remnant ◽  
Unesterified Cholesterol ◽  
Apolipoprotein B100 ◽  
Plasma Triacylglycerol ◽  
Chylomicron Remnants

Lymph chylomicrons radiolabelled in triacylglycerol and cholesteryl ester were injected into control and Watanabe heritable-hyperlipidaemic (WHHL) rabbits. Clearance of chylomicrons was slower in heterozygote and homozygote WHHL rabbits. Slower remnant clearance in WHHL rabbits was confirmed by monitoring the clearance from plasma of preformed chylomicron remnants. Use of chylomicron-like lipid emulsions injected into control and WHHL rabbits also confirmed the defect in remnant clearance in heterozygote WHHL and homozygote WHHL groups. Clearance from plasma of emulsion triolein was delayed in both WHHL groups compared with controls, owing to slower remnant clearance. The clearance from plasma of radioiodinated rabbit low-density lipoproteins (LDL) in heterozygote WHHL rabbits was the same as control rabbits. Defective chylomicron-remnant removal but normal LDL clearance in the heterozygote WHHL corresponded to elevated concentrations of plasma triacylglycerol and normal concentrations of plasma cholesterol. Receptor versus non-receptor clearances of chylomicron remnants were studied by comparing the clearance of emulsions with and without unesterified cholesterol respectively. Unlike control rabbits, there were no significant differences in the clearances of the two emulsion types in either the homozygote or heterozygote WHHL rabbits, indicating that the apolipoprotein-B100/E receptor is the primary route for clearance of chylomicron remnants from plasma.

Binding and uptake of chylomicron remnants by primary and THP-1 human monocyte-derived macrophages: determination of binding proteins

2001 ◽  
Vol 101 (2) ◽  
pp. 111-119 ◽  
Author(s):  
Caryn L. ELSEGOOD ◽  
Sebely PAL ◽  
Paul D. ROACH ◽  
John C. L. MAMO
Keyword(s):  
Binding Site ◽  
Foam Cell ◽  
Ldl Receptor ◽  
Human Monocyte ◽  
Foam Cell Formation ◽  
Chylomicron Remnant ◽  
Surface Binding ◽  
Monocytic Cell ◽  
Monocytic Cell Line ◽  
Chylomicron Remnants

The binding and uptake of chylomicron remnants by human macrophages was studied in order to resolve paradoxical observations that have described the putative mechanisms by which postprandial lipoproteins induce foam cell formation. Chylomicron remnants bound to human monocyte-derived macrophages (HMMs) and to the transformed monocytic cell line THP-1 with high affinity (Kd of approx. 5.5 µg of chylomicron remnant protein/ml). Binding was found to be saturable for both cell types, and was strongly inhibited in the presence of unlabelled chylomicron remnants. Ligand blot studies with colloidal-gold-labelled chylomicron remnants identified two cell surface binding sites on both HMMs and THP-1 cells, with molecular masses of approx. 128 kDa and 43 kDa. The high-molecular-mass binding site was found to be the low-density lipoprotein (LDL) receptor, based on the strong inhibition of chylomicron remnant binding in the presence of unlabelled LDL, Fab2 antibody fragments to the LDL receptor or calcium chelators. Competition studies suggested that, in HMMs, the LDL receptor appeared to facilitate approximately half of the total chylomicron remnant uptake. In contrast, the LDL receptor was not significantly involved in macrophage uptake of chylomicron remnants by THP-1 cells. The identity of the 43 kDa binding site is presently unknown, but, importantly, expression was not inhibited as a consequence of sterol loading, which was induced by incubating HMMs and THP-1 cells with 25-hydroxycholesterol. In contrast, the expression of the LDL receptor was substantially attenuated following lipid loading. Collectively, our data suggest that, while the macrophage LDL receptor can bind chylomicron remnants and facilitate uptake in non-lipid-loaded HMMs, other sterol-insensitive sites are responsible for the unabated uptake of chylomicron remnants by macrophages. We propose that the 43 kDa macrophage chylomicron remnant binding protein may be a candidate for the sterol loading of macrophages.

Lipid synthesis in macrophages derived from the human cell line THP-1: modulation of the effects of native and oxidized chylomicron-remnant-like particles by oestrogen

2001 ◽  
Vol 101 (4) ◽  
pp. 403 ◽  
Author(s):  
Mariarosaria NAPOLITANO ◽  
Kelly V. BATT ◽  
Michael AVELLA ◽  
Elena BRAVO ◽  
Kathleen M. BOTHAM
Keyword(s):  
Cell Line ◽  
Human Cell ◽  
Lipid Synthesis ◽  
Human Cell Line ◽  
Chylomicron Remnant

scholarly journals Regulation of the metabolism of lipoprotein-proteoglycan complexes in human monocyte-derived macrophages

1994 ◽  
Vol 301 (3) ◽  
pp. 675-681 ◽  
Author(s):  
P Vijayagopal
Keyword(s):  
Cholesteryl Ester ◽  
Conditioned Medium ◽  
Density Lipoprotein ◽  
Human Monocyte ◽  
Cholesterol Esterification ◽  
Human Aorta ◽  
Ester Synthesis ◽  
Fibrous Plaque

Studies were performed to evaluate the effect of several factors on the metabolism of lipoprotein-proteoglycan complexes in human monocyte-derived macrophages. In vivo apoB-lipoprotein-proteoglycan complex was isolated from human aorta fibrous-plaque lesions and low-density lipoprotein (LDL)-proteoglycan complex was formed in vitro. Degradation of LDL-proteoglycan complex and cholesteryl ester synthesis mediated by the in vivo and in vitro complexes were lowest in freshly isolated monocytes. With the maturation of monocytes into macrophages, there was a dramatic rise in both. The degradation of the complex and the resultant stimulation of cholesterol esterification increased significantly with increasing cell density. Preincubation of macrophages in medium containing lipoprotein cholesterol did not down-regulate the subsequent degradation of LDL-proteoglycan complex. Macrophage-conditioned medium had a profound stimulatory effect on the degradation of LDL-proteoglycan complex and cholesterol esterification by mature macrophages and freshly isolated monocytes. The conditioned medium lost its stimulatory activity after boiling, dialysis and trypsin digestion. Macrophage activation with phorbol ester and bacterial lipopolysaccharide resulted in a marked suppression of the binding and degradation of the complex, as well as the complex-mediated cholesteryl ester synthesis. These results demonstrate that several factors regulate the metabolism of lipoprotein-proteoglycan complexes in human monocyte-derived macrophages.

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