scholarly journals Time-course studies by neutron solution scattering and biochemical assays of the aggregation of human low-density lipoprotein during Cu2+-induced oxidation

1995 ◽  
Vol 310 (2) ◽  
pp. 417-426 ◽  
Author(s):  
D F Meyer ◽  
M O Mayans ◽  
P H E Groot ◽  
K E Suckling ◽  
K R Bruckdorfer ◽  
...  
Keyword(s):  
Protein Modification ◽  
Time Course ◽  
Oxidized Ldl ◽  
Density Lipoprotein ◽  
Protein Band ◽  
Peritoneal Macrophages ◽  
Distribution Functions ◽  
Oxidative Modification ◽  
Radius Of Gyration ◽  
Low Density

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)

scholarly journals Treatment of macrophages with oxidized low-density lipoprotein increases their intracellular glutathione content

1991 ◽  
Vol 278 (2) ◽  
pp. 429-434 ◽  
Author(s):  
V M Darley-Usmar ◽  
A Severn ◽  
V J O'Leary ◽  
M Rogers
Keyword(s):  
Oxidized Ldl ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Human Monocyte ◽  
Cell Types ◽  
Oxidative Modification ◽  
Glutathione Depletion ◽  
Lipid Phase ◽  
Low Density ◽  
Oxidized Low Density Lipoprotein

Macrophages derived from the human monocyte cell line THP-1 or isolated from the peritoneum of C3H/HEJ mice were incubated with oxidized low-density lipoprotein (LDL) and the total glutathione content (oxidized plus reduced) was measured. An initial depletion of glutathione was followed by an increase, such that after a period of 24 h the glutathione content has approximately doubled. This response required the oxidation of the lipid phase of the LDL molecule, since both native LDL and acetylated LDL had little effect on glutathione levels. The response of the cells to oxidized LDL was dependent on the extent of oxidative modification of the protein. It was also found that 4-hydroxynonenal had a similar effect on THP-1 cells, and we suggest that this or other aldehydes present in oxidized LDL causes the induction of glutathione synthesis in response to an initial oxidative stress and consequent glutathione depletion. In addition, we found that both cell types possess transferases and peroxidases capable of detoxifying aldehydes and peroxides. However, treatment of cells with oxidized LDL or 4-hydroxynonenal for a period of 24 h had no effect on the activities of these enzymes.

scholarly journals Endogenously produced lipoprotein lipase enhances the binding and cell association of native, mildly oxidized and moderately oxidized low-density lipoprotein in mouse peritoneal macrophages

1999 ◽  
Vol 343 (2) ◽  
pp. 347-353 ◽  
Author(s):  
Xiaosong WANG ◽  
Joachim GREILBERGER ◽  
Sanja LEVAK-FRANK ◽  
Robert ZIMMERMANN ◽  
Rudolf ZECHNER ◽  
...  
Keyword(s):  
Lipoprotein Lipase ◽  
Oxidized Ldl ◽  
Low Density Lipoprotein ◽  
Relative Proportion ◽  
Scavenger Receptor ◽  
Density Lipoprotein ◽  
Peritoneal Macrophages ◽  
Low Density ◽  
Cell Binding ◽  
Mouse Peritoneal Macrophages

It has been well established that purified lipoprotein lipase (LPL) can facilitate the cellular uptake of various native and modified lipoproteins when added exogenously to macrophages. Because activated macrophages express LPL endogenously, it was the aim of this study to investigate the effect of macrophage-produced LPL on the uptake of native low-density lipoprotein (LDL) and LDL that has been modified to various degrees by Cu2+-mediated oxidation. Cell binding and uptake of Eu3+-labelled native and oxidized LDL was determined in mouse peritoneal macrophages (MPM) from normal mice and induced mutant mice that lack LPL expression in MPM. We found that LPL expressed by MPM was able to increase cell binding and association of native LDL (by 121% and 101% respectively), mildly oxidized LDL (by 47% and 43%) and moderately oxidized LDL (by 30% and 22%). With increased levels of lipoprotein oxidation, the relative proportion of LPL-mediated LDL uptake decreased. This decrease was not due to weakened binding of LPL to oxidized LDL. The drastically increased uptake of highly oxidized LDL in MPM by scavenger-receptor-mediated pathways might dominate the simultaneous exogenous or endogenous LPL-mediated uptake of this lipoprotein. Competition experiments with positively charged poly(amino acids) furthermore suggested that histidine, arginine and lysine residues in LPL are important for the interaction between LDL and LPL. Our results imply that physiological levels of LPL produced by macrophages facilitate the uptake of native LDL as well as mildly and moderately oxidized LDL. This process might, in the micro-environment of arteries, contribute to the accumulation of macrophage lipids and the formation of foam cells.

scholarly journals GLYCATED LOW-DENSITY LIPOPROTEIN IN DIABETIC AND NON-DIABETIC PATIENTS

2019 ◽  
pp. 123-126
Author(s):  
OMAR ABDULWAHID AL-ANI ◽  
ABDURRAHMAN AL-BAZZAZ
Keyword(s):  
Oxidized Ldl ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Oxidative Modification ◽  
Diabetic Group ◽  
Enzyme Linked Immunosorbent Assay ◽  
Diabetic Patients ◽  
Low Density ◽  
Serum Samples ◽  
Diabetes Center

Objective: The importance of measuring the blood level of modified low-density lipoprotein (LDL) molecules is an effective method of identifying people at risk of coronary atherosclerosis; this is because, in the early stages of atherosclerosis, lipolysis and oxidative modification have a role in promoting the uptake of these lipids through macrophages; therefore, this research aims to measure the level of glycated LDL (Gly-LDL) in the blood and its association with metabolic parameters of diabetic patients (diabetes mellitus) and non-diabetic (hyperlipidemia). Methods: At a University Diabetes Center in Riyadh, we using routine automatic analysis methods, fasting serum samples were analyzed for 31 patients with Type-2 diabetes and 31 non-diabetic patients for LDL, high-density lipoprotein (HDL), total cholesterol, glycated hemoglobin, glucose, and triglycerides (TG), and using enzyme-linked immunosorbent assay to analyze Gly-LDL for the same sample. Results: The level of serum Gly-LDL in non-diabetic was higher than in diabetic patients (p=0.037). Gly-LDL level correlated significantly with LDL in the diabetic group (p=0.035) and was insignificant with other parameters; moreover, it is significantly correlated with HDL (p=0.048), TG (p=0.035), and very LDL (p=0.03) in the non-diabetic group and insignificant with other parameters. Conclusion: Measuring rates of Gly-LDL can be used in the early detection of cardiovascular disease, especially in people with diabetes, as they are more susceptible to modified and oxidized LDL.

scholarly journals Detection of new epitopes formed upon oxidation of low-density lipoprotein, lipoprotein (a) and very-low-density lipoprotein. Use of an antiserum against 4-hydroxynonenal-modified low-density lipoprotein

1990 ◽  
Vol 265 (2) ◽  
pp. 605-608 ◽  
Author(s):  
G Jürgens ◽  
A Ashy ◽  
H Esterbauer
Keyword(s):  
Unsaturated Fatty Acids ◽  
Oxidized Ldl ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Oxidative Modification ◽  
Lipid Hydroperoxides ◽  
Very Low Density Lipoprotein ◽  
Low Density ◽  
Dependent Manner ◽  
Lipoprotein A

4-Hydroxynonenal (HNE) is a major aldehydic propagation product formed during peroxidation of unsaturated fatty acids. The aldehyde was used to modify freshly prepared human low-density lipoprotein (LDL). A polyclonal antiserum was raised in the rabbit and absorbed with freshly prepared LDL. The antiserum did not react with human LDL, but reacted with CuCl2-oxidized LDL and in a dose-dependent manner with LDL, modified with 1, 2 and 3 mM-HNE, in the double-diffusion analysis. LDL treated with 4 mM of hexanal or hepta-2,4-dienal or 4-hydroxyhexenal or malonaldehyde (4 or 20 mM) did not react with the antiserum. However, LDL modified with 4 mM-4-hydroxyoctenal showed a very weak reaction. Lipoprotein (a) and very-low-density lipoprotein were revealed for the first time to undergo oxidative modification initiated by CuCl2. This was evidenced by the generation of lipid hydroperoxides and thiobarbituric acid-reactive substances, as well as by a marked increase in the electrophoretic mobility. After oxidation these two lipoproteins also reacted positively with the antiserum against HNE-modified LDL.

scholarly journals Oxidation of low-density lipoprotein by hypochlorite causes aggregation that is mediated by modification of lysine residues rather than lipid oxidation

1994 ◽  
Vol 302 (1) ◽  
pp. 297-304 ◽  
Author(s):  
L J Hazell ◽  
J J M van den Berg ◽  
R Stocker
Keyword(s):  
Lipid Peroxidation ◽  
Lipid Oxidation ◽  
Oxidized Ldl ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Oxidative Modification ◽  
Size Exclusion ◽  
Lipid Hydroperoxides ◽  
Low Density ◽  
Lysine Residues

Peroxidation of low-density lipoprotein (LDL) lipid is generally thought to represent the initial step in a series of modification reactions that ultimately transform the protein moiety of the lipoprotein into a form recognized by receptors different from those that bind native LDL. Uptake of LDL via these alternative receptors can lead to the formation of lipid-laden cells, which are typical for the early stages of atherogenesis. We have studied the oxidative modification of LDL by hypochlorite (-OCl), a powerful oxidant produced from H2O2 and chloride via the action of myeloperoxidase which is released from activated neutrophils and monocytes. Exposure of LDL to reagent or enzymically generated -OCl at 4 or 37 degrees C resulted in immediate and preferential oxidation of amino acid residues of apolipoprotein B-100, the single protein associated with LDL. Lysine residues quantitatively represented the major target and, like tryptophan, were oxidized to approximately the same extent with reagent or enzymically generated -OCl. In contrast, LDL lipid oxidation was less favoured than protein oxidation, as judged by the amounts of lipid hydroperoxides, chlorohydrins, cholesterol or fatty acid oxidation products formed. Treatment with -OCl caused aggregation of LDL, as shown by an increased turbidity of the oxidized LDL solution and elution from a size-exclusion h.p.l.c. column of high-molecular-mass LDL complexes. Chemical modification of lysine residues before oxidation with -OCl prevented aggregation, while it enhanced the extent of lipid peroxidation. Treatment of LDL with -OCl also caused the formation of carbonyl groups and release of ammonia; both these modifications were inhibited by lysine-residue modification before oxidation. These results demonstrate that aggregation reactions are dependent on initial lysine oxidation by -OCl, followed by deamination and carbonyl formation, but do not involve lipid (per)oxidation. We propose that the observed -OCl-mediated aggregation of LDL is caused, at least in part, by cross-linking of apoproteins by Schiff-base formation independently of lipid peroxidation.

scholarly journals Non-oxidative modification of native low-density lipoprotein by oxidized low-density lipoprotein

1996 ◽  
Vol 316 (2) ◽  
pp. 377-380 ◽  
Author(s):  
Min YANG ◽  
David S. LEAKE ◽  
Catherine A. RICE-EVANS
Keyword(s):  
Oxidized Ldl ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Oxidative Modification ◽  
Low Density ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Process Studies

The oxidative modification of low-density lipoprotein (LDL) has been implicated in the pathogenesis of atherosclerosis, although little is known as yet about the precise mechanism of oxidation in vivo. The studies presented here demonstrate that, in the absence of cells or transition metals, oxidized LDL can modify native LDL through co-incubation in vitro such as to increase its net negative charge, in a concentration-dependent manner. The interaction is not inhibited by peroxyl radical scavengers or metal chelators, precluding the possibility that the modification of native LDL by oxidized LDL is through an oxidative process. Studies with radioiodinated oxidized LDL showed no transfer of radioactivity to the native LDL, demonstrating that fragmentation of protein and the transfer of some of the fragments does not account for the modified charge on the native LDL particle. The adjacency of native to oxidized LDL in the arterial wall may be a potential mechanism by which the altered recognition properties of the apolipoprotein B-100 may arise rapidly without oxidation or extensive modification of the native LDL lipid itself.

scholarly journals Oxidation of low-density lipoprotein with hypochlorite causes transformation of the lipoprotein into a high-uptake form for macrophages

1993 ◽  
Vol 290 (1) ◽  
pp. 165-172 ◽  
Author(s):  
L J Hazell ◽  
R Stocker
Keyword(s):  
Oxidized Ldl ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Fold Increase ◽  
Oxidative Modification ◽  
Lipid Hydroperoxides ◽  
Low Density ◽  
High Uptake ◽  
Apoprotein B ◽  
Lysine Residues

Oxidation of low-density lipoprotein (LDL) lipid is thought to represent the initial step in a series of oxidative modification reactions that ultimately transform this lipoprotein into an atherogenic high-uptake form that can cause lipid accumulation in cells. We have studied the effects of hypochlorite, a powerful oxidant released by activated monocytes and neutrophils, on isolated LDL. Exposure of LDL to reagent hypochlorite (NaOCl) at 4 degrees C resulted in immediate and preferential oxidation of amino acid residues of apoprotein B-100, the single protein associated with LDL. Neither lipoprotein lipid nor LDL-associated antioxidants, except ubiquinol-10, represented major targets for this oxidant. Even when high concentrations of NaOCl were used, only low levels of lipid hydroperoxides could be detected with the highly sensitive h.p.l.c. post-column chemiluminescence detection method. Lysine residues of apoprotein B-100 quantitatively represented the major target, scavenging some 68% of the NaOCl added, with tryptophan and cysteine together accounting for an additional 10% of the oxidant. Concomitant with the loss of LDL's amino groups, chloramines were formed and the anionic surface charge of the lipoprotein particle increased, indicated by a 3-4-fold increase in electrophoretic mobility above that of native LDL on agarose gels. While both these changes could be initially reversed by physiological reductants such as ascorbic acid and methionine, incubation of the NaOCl-modified LDL at 37 degrees C resulted in increasing resistance of the modified lysine residues against reductive reversal. Exposure of mouse peritoneal macrophages to NaOCl-oxidized LDL resulted in increased intracellular concentrations of cholesterol and cholesteryl esters. These findings suggest that lipid-soluble antioxidants associated with LDL do not efficiently protect the lipoprotein against oxidative damage mediated by hypochlorite, and that extensive lipid oxidation is not a necessary requirement for oxidative LDL modification that leads to a high-uptake form of the lipoprotein.

scholarly journals Time course-changes in phosphatidylcholine profile during oxidative modification of low-density lipoprotein

2014 ◽  
Vol 13 (1) ◽  
pp. 48 ◽  
Author(s):  
Naoko Sasabe ◽  
Yuka Keyamura ◽  
Takashi Obama ◽  
Nozomi Inoue ◽  
Yukihiro Masuko ◽  
...  
Keyword(s):  
Time Course ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Oxidative Modification ◽  
Low Density ◽  
Course Changes
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