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Characterization of the structure of polydisperse human low-density lipoprotein by neutron scattering.

ABSTRACT: Low-density lipoproteins (LDL) in plasma are constructed from a single molecule of apolipoprotein B-100 (M(r) 512000) in association with lipid (approximate M(r) 2-3 x 10(6)). The gross structure was studied using an updated pulsed-neutron camera LOQ with an area detector to establish the basis for the interpretation of structural changes seen during dynamic studies of LDL oxidation. Neutron-scattering data for LDL in 100% 2H2O buffers emphasize their external appearance. Guinier analysis on a continuous-flux neutron camera D17 revealed pronounced concentration-dependences in the radius of gyration, RG, and the intensity of forward scattering, I(0) (equivalent to the M(r) of LDL) between 0.5 and 11 mg of LDL protein/ml. LDL preparations from different donors gave different RG values. When extrapolated to zero concentration, RG values ranged between 8.3 and 10.6 nm and were linearly correlated with M(r), which is consistent with a spherical structure. The distance-distribution function P(r) in real space showed a single maximum at 9.1-10.9 nm, which is just under half the observed maximum dimension of 23.1 +/- 1.2 nm expected for a spherical structure. The neutron radial-density function p(r) exhibited a plateau of high and featureless density at the centre of LDL. LDL can be modelled by a polydisperse assembly of spheres with two internal densities and a mean radius close to 10.0 nm in a normal distribution of radii with a standard deviation of 2.0 nm. The data are consistent with recent electron-microscopy and ultracentrifugation data.(ABSTRACT TRUNCATED AT 250 WORDS)

SUBMITTER: Meyer DF

PROVIDER: S-EPMC1135910 | biostudies-other | 1995 Sep

REPOSITORIES: biostudies-other

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Project description:The oxidative modification of low-density lipoproteins (LDL) is recognized to be a key event in the development of atherosclerotic plaques on artery walls. The characteristics of LDL oxidized by cells of the artery wall can be imitated by the addition of Cu2+ ions to initiate lipid peroxidation in LDL. Neutron scattering of LDL in 2H2O buffers enables the time course of changes in the gross structure of LDL during oxidation to be continuously monitored under conditions close to physiological. Oxidation of LDL [2 mg of apolipoprotein B (apoB) protein/ml] was studied in the presence of 6.4, 25.6 and 51.2 mumol of Cu2+/g of apoB by incubation at 37 degrees C for up to 70 h. Neutron Guinier analyses showed that the radius of gyration RG (indicative of size) and the forward-scattered intensity at zero angle I(0) (indicative of M(r)) continuously increased during oxidation, indicating that LDL had aggregated. Both the rate of aggregation and the change in RG and I(0) values after 10 and 50 h increased with Cu2+ concentration. Distance-distribution functions P(r) showed that, within 4 h, the maximum dimension of LDL increased from 23 to 55 nm. The P(r) curves of oxidatively modified LDL exhibited two peaks at 10-12 nm and 26 nm. The 10-12 nm peak corresponds to native LDL, and the 26 nm peak is assigned to the initial formation of LDL dimers and trimers and their progression to form higher oligomers. The growth of the 26 nm peak depended on Cu2+ concentration. Particle-size-distribution functions Dv(r) suggested that the polydisperse spherical structure of LDL ceased to exist after 30 h, at which point the LDL samples underwent a phase separation. Related, but not identical, changes in the I(Q) and P(r) curves were observed when native LDL was self-aggregated by brief vortexing. Parallel assessment of LDL protein modification by SDS/PAGE showed increased aggregation and degradation of apoB with increased Cu2+ concentrations, and that the main apoB protein band had diminished after 2-8 h, depending on the amount of Cu2+ added. The uptake and degradation of oxidized 125I-labelled LDL by mouse peritoneal macrophages occurred maximally within the first 10 h, and increased in proportion to the Cu2+ concentration. ApoB protein broke down within the first 10 h of oxidation, and this is the period when scavenger receptors on macrophages can recognize and internalize oxidized LDL. Within 10 h, the protein-lipid interactions responsible for the spherical LDL structure became destabilized by protein fragmentation.(ABSTRACT TRUNCATED AT 400 WORDS)

Project description:Background MEDI6012 is recombinant human lecithin cholesterol acyltransferase, the rate-limiting enzyme in reverse cholesterol transport. Infusions of lecithin cholesterol acyltransferase have the potential to enhance reverse cholesterol transport and benefit patients with coronary heart disease. The purpose of this study was to test the safety, pharmacokinetic, and pharmacodynamic profile of MEDI6012. Methods and Results This phase 2a double-blind study randomized 48 subjects with stable coronary heart disease on a statin to a single dose of MEDI6012 or placebo (6:2) (NCT02601560) with ascending doses administered intravenously (24, 80, 240, and 800 mg) and subcutaneously (80 and 600 mg). MEDI6012 demonstrated rates of treatment-emergent adverse events that were similar to those of placebo. Dose-dependent increases in high-density lipoprotein cholesterol were observed with area under the concentration-time curves from 0 to 96 hours of 728, 1640, 3035, and 5318 should be: mg·h/mL in the intravenous dose groups and 422 and 2845 mg·h/mL in the subcutaneous dose groups. Peak mean high-density lipoprotein cholesterol percent change was 31.4%, 71.4%, 125%, and 177.8% in the intravenous dose groups and 18.3% and 111.2% in the subcutaneous dose groups, and was accompanied by increases in endogenous apoA1 (apolipoprotein A1) and non-ATP-binding cassette transporter A1 cholesterol efflux capacity. Decreases in apoB (apolipoprotein B) were observed across all dose levels and decreases in atherogenic small low-density lipoprotein particles by 41%, 88%, and 79% at the 80-, 240-, and 800-mg IV doses, respectively. Conclusions MEDI6012 demonstrated an acceptable safety profile and increased high-density lipoprotein cholesterol, endogenous apoA1, and non-ATP-binding cassette transporter A1 cholesterol efflux capacity while reducing the number of atherogenic low-density lipoprotein particles. These findings are supportive of enhanced reverse cholesterol transport and a functional high-density lipoprotein phenotype. Registration URL: https://www.clinicaltrials.gov; Unique identifier: NCT02601560.

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Characterization of the structure of polydisperse human low-density lipoprotein by neutron scattering.

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