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

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.

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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

Clinical Science ◽

10.1042/cs1010403 ◽

2001 ◽

Vol 101 (4) ◽

pp. 403-413 ◽

Cited By ~ 7

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.

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Cholesteryl ester hydrolase in human monocyte/macrophage: cloning, sequencing, and expression of full-length cDNA

Physiological Genomics ◽

10.1152/physiolgenomics.2000.2.1.1 ◽

2000 ◽

Vol 2 (1) ◽

pp. 1-8 ◽

Cited By ~ 70

Author(s):

SHOBHA GHOSH

Keyword(s):

Cholesteryl Ester ◽

Peripheral Blood ◽

Human Peripheral Blood ◽

Human Monocyte ◽

Cholesteryl Ester Hydrolase ◽

Full Length ◽

Human Macrophage ◽

Full Length Cdna ◽

Ester Hydrolase ◽

Sequencing And Expression

Ghosh, Shobha. Cholesteryl ester hydrolase in human monocyte/macrophage: cloning, sequencing, and expression of full-length cDNA. Physiol. Genomics 2: 1–8, 2000.—The sensitive technique of RT-PCR was used to identify cholesteryl ester hydrolase (CEH) expressed in human macrophages. This enzyme is thought to regulate the availability of intracellular free cholesterol for efflux. The expected 667-bp product was obtained starting with RNA from human peripheral blood and THP-1 monocytes and macrophages. The cDNA for human macrophage CEH was then cloned by PCR-based screening of a λ-gt11 cDNA library. The full-length cDNA was sequenced and found to exhibit 76% homology (at the nucleotide and conceptually translated protein level) to hepatic CEH, an enzyme shown to be involved in hepatic cholesterol homeostasis and regulated by cholesterol at the transcription level via sterol response elements in the proximal promoter. Identification of the conserved catalytic triad (Ser221, His468, and Glu354) and the SEDCLY motif places human macrophage CEH in the family of carboxylesterases. A greater than 20-fold increase in CEH activity was observed when COS-1 and COS-7 cells were transiently transfected with an eukaryotic expression vector, pcDNA3.1/V5/His-TOPO, containing the cDNA for human macrophage CEH. Using this full-length cDNA as a probe, a 2.2-kb transcript was identified by Northern blot analysis of total RNA from human peripheral blood and THP-1 macrophages. Overexpression of human macrophage CEH resulted in an impairment of upregulation of low-density lipoprotein (LDL) receptor mRNA in Chinese hamster ovary (CHO-K1) cells grown in cholesterol-deficient environment. These data identify the human macrophage CEH, demonstrate its expression in human peripheral blood macrophage and human macrophage cell line, THP-1, and suggest its role in the intracellular cholesteryl ester metabolism.

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