Influence of high density lipoprotein on esterified cholesterol stores in macrophages and hepatoma cells.

Bernard, David W.; Rodríguez, Annabelle; Rothblat, George H.; Glick, Jane M.

doi:10.1161/01.atv.10.1.135

Cited by 76 publications

(34 citation statements)

References 39 publications

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“…This may be due to several factors. J774 macrophages hydrolyze stored CE slower than primary mouse macrophages, 29,49 and J774 macrophages are a dividing cell population in which FC accumulation may be moderated by dilution due to cell growth. Both of these characteristics may serve to limit the FC accumulation and thereby prevent a nucleating event.…”

Section: Cholesterol Crystal Formationmentioning

confidence: 97%

Effects of Intracellular Free Cholesterol Accumulation on Macrophage Viability

Kellner-Weibel

Jerome

Small

et al. 1998

ATVB

156

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Abstract-This study was designed to identify cellular responses associated with free cholesterol (FC) accumulation in model macrophage foam cells. Mouse peritoneal macrophages (MPMs) or J774 macrophages were loaded with cholesteryl esters using acetylated LDL and FC/phospholipid dispersions and were subsequently exposed to an acyl coenzyme A:cholesterol acyltransferase (ACAT) inhibitor. This treatment produced a rapid accumulation of cellular FC. The FC that accumulated due to ACAT inhibition was more readily available for efflux to 2-hydroxypropyl-␤-cyclodextrin (which removes cholesterol from the plasma membrane) than FC in untreated control cells. After a 3-hour exposure to an ACAT inhibitor, a significant increase in phospholipid synthesis was seen, followed by the leakage of LDH after 12 hours of treatment.

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Section: Cholesterol Crystal Formationmentioning

confidence: 97%

Effects of Intracellular Free Cholesterol Accumulation on Macrophage Viability

Kellner-Weibel

Jerome

Small

et al. 1998

ATVB

156

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“…In some experiments, acLDL was incubated with UCphospholipid liposomes (225 g UC per mL) and BSA (10 mg/mL) overnight at 37°C before use in experiments. 17 During the course of these experiments, we found that the addition of liposomes and BSA to the medium containing acLDL did not further increase esterified cholesterol (EC) accumulation in HMMs, and these agents were subsequently omitted from the incubation medium. The concentration of acLDL used in these studies was based on our previous work in human THP-1 macrophages and on the work of other investigators studying foam cell formation in HMMs.…”

Section: Lipid Enrichment Of Cellsmentioning

confidence: 99%

Novel Effects of the Acyl-Coenzyme A:Cholesterol Acyltransferase Inhibitor 58-035 on Foam Cell Development in Primary Human Monocyte–Derived Macrophages

Rodríguez¹,

Bachorik

Wee³

1999

ATVB

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Abstract-We examined the effect of acyl-coenzyme A:cholesterol acyltransferase (ACAT) inhibitors on intracellular cholesterol stores in primary human monocyte-derived macrophages (HMMs) during foam cell formation. HMMs were exposed to acetylated low density lipoprotein (acLDL, 500 g protein per mL) with or without 58-035 (1 to 10 g/mL) or CI-976 (2 g/mL) for 2 to 48 hours. Total cholesterol (TC) and esterified cholesterol (EC) mass was significantly lower while unesterified cholesterol (UC) increased slightly in cells incubated with acLDL plus ACAT inhibitors. Sterol mass was also measured in cells coincubated with acLDL (500 g protein per mL) with or without 58-035 (2 g/mL), high density lipoprotein (HDL, 400 g protein per mL), or HDLϩ58-035 for 48 hours. TC and EC were 23% and 55% lower, respectively (PϽ0.0004), while UC was 11% higher (PϽ0.04) in cells incubated with acLDL plus 58-035. In contrast, coincubation with HDL alone did not significantly affect TC, EC, or UC mass compared with acLDL alone. The effect of 58-035 could not be explained by cytotoxicity, because adenine release, secreted lactate dehydrogenase, glucose utilization, and cell protein were similar in cells exposed to acLDL regardless of the presence of 58-035. We investigated several potential mechanisms for the decreased TC mass, including increased UC efflux and decreased acLDL binding and uptake. Efflux was measured in cells exposed to [1,2-3 H]cholesteryl oleate-labeled acLDL, unlabeled control acLDL, and native untreated acLDL (500 g protein per mL) with or without 58-035 (5 g/mL) for 24 or 48 hours. UC efflux increased in a time-dependent manner from cells exposed to acLDL plus 58-035 compared with cells exposed to acLDL alone (PϽ0.04). High-affinity binding was measured in cells exposed to 125 I-acLDL (5 g protein per mL) with or without excess unlabeled acLDL (100 or 500 g protein per mL) for 4 hours at 4°C. Specific acLDL binding, uptake, and total degradation were significantly lower when 58-035 was present during cholesterol enrichment compared with cells exposed to acLDL alone (PϽ0.001). Unlike the effects of ACAT inhibitors on foam cell formation in rodent macrophages, these compounds lowered TC accumulation in HMMs during foam cell formation by limiting the uptake of acLDL and enhancing UC efflux. They may offer promise as drug therapies for atherosclerosis.

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“…After 3 days, the cells were washed and labeled with [ 3 H]cholesterol in the presence of 1 μg/mL Sandoz 58035, an inhibitor of intracellular cholesterol esterification, 28 during an overnight incubation in MEM containing 2.5% FBS. ACAT inhibitor compound 58035 was present at 1 μg/mL of media in cell labeling, cell equilibration, and cellular efflux stages.…”

Section: Cholesterol Flux Studiesmentioning

confidence: 99%

“…28 To generate cells that were both UC and EC enriched, Sandoz compound 58035 was omitted 28 during both cholesterol enrichment of the cells and the subsequent incubations with cholesterol acceptors.…”

Section: Cholesterol Flux Studiesmentioning

confidence: 99%

Remodeling and Shuttling

Rodrigueza

Williams

Rothblat

et al. 1997

ATVB

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In normal physiology, cells are exposed to cholesterol acceptors of different sizes simultaneously. The current study examined the possible interactions between two different classes of acceptors, one large (large unilamellar phospholipid vesicles, LUVs) and one small (HDL or other small acceptors), added separately or in combination to Fu5AH rat hepatoma cells. During a 24-hour incubation, LUVs of palmitoyl-oleoyl phosphatidylcholine at 1 mg phospholipid (PL) per milliliter extracted ≈20% of cellular unesterified cholesterol (UC) label and mass in a slow, continuous fashion (half-time [t½] for UC efflux was ≈50 hours) and human HDL 3 at 25 μg PL per milliliter extracted ≈15% cellular UC label with no change in cellular cholesterol mass (t½ of ≈8 hours). In contrast, the combination of LUVs and HDL 3 extracted over 90% of UC label (t½ of ≈4 hours) and ≈50% of the UC mass, indicating synergy. To explain this synergy, specific particle interactions were examined, namely, remodeling, in which the two acceptors alter each other's composition and thus the ability to mobilize cellular cholesterol, and shuttling, in which the small acceptor ferries cholesterol from cells to the large acceptor. To examine remodeling, LUVs and HDL were coincubated and reisolated before application to cells. This HDL became UC depleted, PL enriched, and lost a small amount of apolipoprotein A-I. Compared with equivalent numbers of control HDL particles, remodeled HDL caused faster efflux (t½ ≈4 hours) and exhibited a greater capacity to sequester cellular cholesterol over 24 hours (≈38% versus ≈15% for control HDL), consistent with their enrichment in PL. Remodeled LUVs still extracted ≈20% of cellular UC. Thus, remodeling accounted for some but not all of the synergy between LUVs and HDL. To examine shuttling, several approaches were used. First, reisolation of particles after an 8-hour exposure to cells revealed that HDL contained very little of the cellular UC label. The label was found almost entirely with the LUVs, suggesting that LUVs continuously stripped the HDL of cellular UC. Second, bidirectional flux studies demonstrated that LUVs blocked the influx of HDL UC label into cells, while the rate of efflux of cellular UC was maintained. These kinetic effects explained the massive net loss of cellular UC to LUVs with HDL. Third, cyclodextrin, an artificial small acceptor that does not acquire PL and hence does not become remodeled, exhibitedCorrespondence to Dr Michael C. Phillips, Department of Biochemistry, Medical College of Pennsylvania and Hahnemann University, 2900 Queen Ln, Philadelphia, PA 19129. Portions of this work were published in abstract form in Abstracts Submitted to the Council on Arteriosclerosis for the 68th Scientific Sesssions of the American Heart Association. 1995:54. HHS Public Access Author Manuscript Author ManuscriptAuthor ManuscriptAuthor Manuscript substantial synergy with LUVs, supporting shuttling. Thus, the presence of large and small acceptors together can overcome intrinsic deficiencies in...

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Influence of high density lipoprotein on esterified cholesterol stores in macrophages and hepatoma cells.

Cited by 76 publications

References 39 publications

Effects of Intracellular Free Cholesterol Accumulation on Macrophage Viability

Effects of Intracellular Free Cholesterol Accumulation on Macrophage Viability

Novel Effects of the Acyl-Coenzyme A:Cholesterol Acyltransferase Inhibitor 58-035 on Foam Cell Development in Primary Human Monocyte–Derived Macrophages

Remodeling and Shuttling

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