Structure and interactions of lipids in human plasma low density lipoproteins.

To study macrophage lipid droplet composition and the effects of TG on cholesteryl ester (CE) physical state, hydrolysis, and cholesterol efflux, a technique was developed to remove the majority of accumulated TG with minimal effect on CE content. THP-1 macrophages were incubated with acetylated LDL, and the accumulated TG was depleted by incubation with the acyl-CoA synthetase inhibitor triacsin D in the presence of albumin. Before TG removal, all cellular lipid droplets were isotropic as determined by polarizing light microscopy. When the TG concentration was reduced, anisotropic lipid droplets were visible, indicating a change in physical state, and suggesting that TG and CE originally accumulated in mixed lipid droplets. This change in physical state of lipid droplets was associated with slower rates of CE hydrolysis and cholesterol efflux. Although lipid droplets within the same cell had a similar physical state after TG depletion, there was considerable variability among cells in the physical state of their lipid droplets. In conclusion, THP-1 macrophages store accumulated CE and TG in mixed droplets, and the proportion of CE to TG varies among cells. Reducing accumulated TG altered CE physical state, which in turn affected hydrolysis of CE and cholesterol efflux. -Lada, A. T., M. C. Willingham, and R. W. St. Clair. Triglyceride depletion in THP-1 cells alters cholesteryl ester physical state and cholesterol efflux.

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confidence: 99%

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confidence: 99%

Triglyceride depletion in THP-1 cells alters cholesteryl ester physical state and cholesterol efflux

Lada

Clair

2002

“…LDL is a spherical particle (diameter 20-22 nm) composed of an internal core of cholesteryl esters containing small amounts of triglyceride, solubilized by a monolayer of phospholipid with small quantities of free cholesterol. Located in the monolayer is the receptor protein apoprotein B (apoB) which accounts for approximately 20% of the particle's weight (1). LDL is of interest in a variety of research areas, primarily atherosclerosis and lipid metabolism where the original studies defining receptor-dependent uptake were performed by Brown and Goldstein (2).…”

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confidence: 99%

“…Cells obtain cholesterol for membrane production from Abbreviations: apoB, apoprotein B; CO, cholesterol oleate; DFCS, delipidated fetal calf serum; FCS, fetal calf serum; PC, egg yolk phosphatidyl choline; PEPXsLDL, synthetic low density lipoprotein system containing PEPX, where X represents the peptide number; sLDL, synthetic low density lipoprotein; TO, triolein. 1 Present address: Division of Infection and Immunity, James Black Building, Glasgow University, Glasgow, G12 8QQ, UK. 2 To whom correspondence should be addressed.…”

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confidence: 99%

A synthetic low density lipoprotein particle capable of supporting U937 proliferation in vitro

Baillie¹,

Owens

Halbert

2002

A synthetic LDL (sLDL) has been prepared by combining a lipid microemulsion with amphipathic peptides containing the apoprotein B receptor domain. The biological properties of sLDL have been investigated using the U937 in vitro cell proliferation assay. sLDL exhibits a concentration dependent and saturable stimulation of U937 proliferation. By utilizing different amphipathic peptides, variable proliferation is achieved, indicating a specific interaction between sLDL and the U937 LDL receptor are possible. U937 proliferation is reduced by the addition of an anti-LDL receptor antibody, indicating that sLDL is assimilated via the LDL receptor pathway.The behavior of sLDL mimics that of native LDL, and this approach represents a viable technique for the production of an sLDL particle on a large scale for research and general application. -Baillie, G., M. D. Owens, and G. W. Halbert. A synthetic low density lipoprotein particle capable of supporting U937 proliferation in vitro.

“…Some of the IDL is taken up by the liver and the rest is converted to LDL, which is slowly removed from plasma. Early work using X-ray diffraction and calorimetry (4)(5)(6)(7)(8) showed that apoB was on the exterior part of LDL while the lipid core contained cholesteryl esters and triglycerides. Phospholipids were also identified to be exclusively on the surface whereas free cholesterol was distributed between core and surface (5,8).…”

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confidence: 99%

Morphology of sodium deoxycholate-solubilized apolipoprotein B-100 using negative stain and vitreous ice electron microscopy

Gantz

Walsh

Small

2000

The primary and secondary structures of apolipoprotein B-100 (apoB-100) are well established. Previous morphological studies have suggested that apoB is a long, flexible, threadlike molecule that encircles the low density lipoprotein (LDL) particle. Several large domain regions of the protein have been observed in frozen hydrated LDL and may be involved in anchoring of the protein to the lipid surface of LDL. Calorimetric studies of sodium deoxycholate (NaDC)-solubilized apoB indicated a similar number of independently melting domains. We therefore undertook a morphological study of NaDC-solubilized apoB-100 using negative stain and vitreous ice cryoelectron microscopy, a nonperturbing preservation technique. Negative staining experiments were performed in two ways: 1 ) grids were pulled through NaDC-containing buffer surfaces on which monolayers of apoB had been promoted, or 2 ) apoB molecules were allowed to diffuse onto carbon surfaces of grids that were floated on sample droplets. Vitrified molecules of apoB were obtained by plunging a thin fluid layer of protein adhered to a holey carbon-coated grid into supercooled ethane and by preserving the molecules in liquid nitrogen. The majority of molecules prepared in negative stain and vitreous ice were curved or arced and had alternating thin and thick regions. In negative stain, the apoB molecules lay on the grid perpendicular to the electron beam and had a mean length of 650 Å. In vitreous ice the molecules were randomly oriented and their images ranged from 160 to 650 Å in length. Vitrified molecules provided visualization of one or two beaded regions. Similar regions were observed in negative stain but the overall thickness was two to three times greater. Some vitrified molecules contained ribbon-like portions. Our study supports previously obtained data on molecule length but suggests that negative staining overestimates molecule width. These first images of vitrified NaDC-solubilized apoB-100 confirm the long, flexible, beaded thread morphology of the molecule and support the unique potential of this technique when coupled with proper molecule orientation and antibody labeling to correlate the tertiary structure of apoB seen in the intact particle with that of the isolated molecule. -Gantz, D. L., M. T. Walsh, and D. M. Small. Morphology of sodium deoxycholate-solubilized apolipoprotein B-100 using negative stain and vitreous ice electron microscopy.