Effect of Low-Density Lipoprotein (LDL) Antigen Source on an Enzyme-Linked Immunosorbent Assay for Autoantibodies against Oxidized LDL

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.

Download Full-text

Quantitation of plasma oxidatively modified low density lipoprotein by sandwich enzyme linked immunosorbent assay

Clinica Chimica Acta ◽

10.1016/0009-8981(93)90225-s ◽

1993 ◽

Vol 218 (1) ◽

pp. 97-103 ◽

Cited By ~ 2

Author(s):

Hualiang Wang ◽

Sicong Chen ◽

Xiantao Kong ◽

Xiaoli Wang ◽

Guoyuan Chang ◽

...

Keyword(s):

Immunosorbent Assay ◽

Low Density Lipoprotein ◽

Density Lipoprotein ◽

Enzyme Linked Immunosorbent Assay ◽

Low Density ◽

Oxidatively Modified

Download Full-text

Characterization and visualization of the low density lipoprotein (LDL) receptor by ligand blotting using anti-LDL-enzyme linked immunosorbent assay

Electrophoresis ◽

10.1002/elps.1150050610 ◽

1984 ◽

Vol 5 (6) ◽

pp. 372-373 ◽

Cited By ~ 2

Author(s):

Hans A. Dresel ◽

Gotthard Schettler

Keyword(s):

Immunosorbent Assay ◽

Low Density Lipoprotein ◽

Ldl Receptor ◽

Density Lipoprotein ◽

Enzyme Linked Immunosorbent Assay ◽

Low Density

Download Full-text

Development of Capture Assays for Different Modifications of Human Low-Density Lipoprotein

Clinical and Diagnostic Laboratory Immunology ◽

10.1128/cdli.12.1.68-75.2005 ◽

2005 ◽

Vol 12 (1) ◽

pp. 68-75 ◽

Cited By ~ 27

Author(s):

Gabriel Virella ◽

M. Brooks Derrick ◽

Virginia Pate ◽

Charlyne Chassereau ◽

Suzanne R. Thorpe ◽

...

Keyword(s):

Oxidized Ldl ◽

Low Density Lipoprotein ◽

Density Lipoprotein ◽

Gas Chromatography Mass Spectrometry ◽

Low Density ◽

Modified Ldl ◽

Capture Assay ◽

Carboxymethyl Lysine

ABSTRACT Antibodies to malondialdehyde (MDA)-modified low-density lipoprotein (LDL), copper-oxidized LDL (oxLDL), N ε(carboxymethyl) lysine (CML)-modified LDL, and advanced glycosylation end product (AGE)-modified LDL were obtained by immunization of rabbits with in vitro-modified human LDL preparations. After absorption of apolipoprotein B (ApoB) antibodies, we obtained antibodies specific for each modified lipoprotein with unique patterns of reactivity. MDA-LDL antibodies reacted strongly with MDA-LDL and also with oxLDL. CML-LDL antibodies reacted strongly with CML-LDL and also AGE-LDL. oxLDL antibodies reacted with oxLDL but not with MDA-LDL, and AGE-LDL antibodies reacted with AGE-LDL but not with CML-LDL. Capture assays were set with each antiserum, and we tested their ability to capture ApoB-containing lipoproteins isolated from precipitated immune complexes (IC) and from the supernatants remaining after IC precipitation (free lipoproteins). All antibodies captured lipoproteins contained in IC more effectively than free lipoproteins. Analysis of lipoproteins in IC by gas chromatography-mass spectrometry showed that they contained MDA-LDL and CML-LDL in significantly higher concentrations than free lipoproteins. A significant correlation (r = 0.706, P < 0.019) was obtained between the MDA concentrations determined by chemical analysis and by the capture assay of lipoproteins present in IC. In conclusion, we have developed capture assays for different LDL modifications in human ApoB/E lipoprotein-rich fractions isolated from precipitated IC. This approach obviates the interference of IC in previously reported modified LDL assays and allows determination of the degree of modification of LDL with greater accuracy.

Download Full-text

Oxidative stress augments pulmonary hypertension in chronically hypoxic mice overexpressing the oxidized LDL receptor

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00169.2012 ◽

2013 ◽

Vol 305 (2) ◽

pp. H155-H162 ◽

Cited By ~ 24

Author(s):

Sayoko Ogura ◽

Tatsuo Shimosawa ◽

ShengYu Mu ◽

Takashi Sonobe ◽

Fumiko Kawakami-Mori ◽

...

Keyword(s):

Pulmonary Hypertension ◽

Vascular Remodeling ◽

Chronic Hypoxia ◽

Oxidized Ldl ◽

Low Density Lipoprotein ◽

Density Lipoprotein ◽

Ros Generation ◽

Low Density ◽

Ros Production ◽

Oxidized Low Density Lipoprotein

Chronic hypoxia is one of the main causes of pulmonary hypertension (PH) associated with ROS production. Lectin-like oxidized low-density lipoprotein receptor (LOX)-1 is known to be an endothelial receptor of oxidized low-density lipoprotein, which is assumed to play a role in the initiation of ROS generation. We investigated the role of LOX-1 and ROS generation in PH and vascular remodeling in LOX-1 transgenic (TG) mice. We maintained 8- to 10-wk-old male LOX-1 TG mice and wild-type (WT) mice in normoxia (room air) or hypoxia (10% O2 chambers) for 3 wk. Right ventricular (RV) systolic pressure (RVSP) was comparable between the two groups under normoxic conditions; however, chronic hypoxia significantly increased RVSP and RV hypertrophy in LOX-1 TG mice compared with WT mice. Medial wall thickness of the pulmonary arteries was significantly greater in LOX-1 TG mice than in WT mice. Furthermore, hypoxia enhanced ROS production and nitrotyrosine expression in LOX-1 TG mice, supporting the observed pathological changes. Administration of the NADPH oxidase inhibitor apocynin caused a significant reduction in PH and vascular remodeling in LOX-1 TG mice. Our results suggest that LOX-1-ROS generation induces the development and progression of PH.

Download Full-text

Monoclonal antibodies can precipitate low-density lipoprotein. I. Characterization and use in determining apolipoprotein B.

Clinical Chemistry ◽

10.1093/clinchem/31.10.1654 ◽

1985 ◽

Vol 31 (10) ◽

pp. 1654-1658 ◽

Cited By ~ 19

Author(s):

S Marcovina ◽

D France ◽

R A Phillips ◽

S J Mao

Keyword(s):

Monoclonal Antibodies ◽

Apolipoprotein B ◽

Polyclonal Antibodies ◽

Low Density Lipoprotein ◽

Density Lipoprotein ◽

Radial Immunodiffusion ◽

Enzyme Linked Immunosorbent Assay ◽

Low Density ◽

Apo B ◽

Blotting Technique

Abstract We produced 20 mouse monoclonal antibodies against human plasma low-density lipoprotein (LDL). Individually they failed to precipitate LDL in agarose gel by the double-immunodiffusion technique; collectively they did, or as few as two combined monoclonal antibodies could do so. To mimic polyclonal antibodies in determination of apolipoprotein B (apo B) by radial immunodiffusion, a combination of four particular monoclonal antibodies (clones A, B, C, and D) was necessary. We characterized these four clones with respect to temperature dependency, affinity, total binding to 125I-labeled LDL, and specificity to the different species of apolipoprotein B. Two monoclonal antibodies (B and C) bound 100% of 125I-labeled LDL; clones A and D bound 80% and 87%, respectively. All four clones bound maximally to LDL at 4 degrees C. The affinity constants for clones A, B, C, and D were 0.6, 2.1, 3.8, and 2.3 X 10(9) L/mol, respectively. By the Western blotting technique, the four monoclonal antibodies all reacted with the species B-100 and B-74 of apolipoprotein B, and to various degrees with B-48 and B-26. Radial immunodiffusion (chi) and direct enzyme-linked immunosorbent assay (y) with a mixture of the four monoclonal antibodies gave almost identical results for 70 patients: y = 0.921 chi-2.58; r = 0.933.

Download Full-text

A monoclonal-antibody-based enzyme-linked immunosorbent assay of lipoprotein(a)

Clinical Chemistry ◽

10.1093/clinchem/36.2.192 ◽

1990 ◽

Vol 36 (2) ◽

pp. 192-197 ◽

Cited By ~ 30

Author(s):

W L Wong ◽

D L Eaton ◽

A Berloui ◽

B Fendly ◽

P E Hass

Keyword(s):

Monoclonal Antibody ◽

Immunosorbent Assay ◽

Clinical Laboratory ◽

Low Density Lipoprotein ◽

Density Lipoprotein ◽

Enzyme Linked Immunosorbent Assay ◽

Apo B ◽

Lipoprotein A ◽

Premature Coronary Heart Disease ◽

Large Populations

Abstract Lipoprotein(a) [Lp(a)] is a low-density lipoprotein (LDL)-like lipoprotein particle recently described as a risk factor for premature coronary heart disease, stroke, and atherosclerosis. Structurally, Lp(a) is similar to LDL in that it has comparable lipid composition and contains apolipoprotein B-100 (apo B-100). In addition, Lp(a) contains the glycoprotein apolipoprotein(a) [apo(a)], which is disulfide-linked to apo B-100. The recent awareness of a striking correlation between atherosclerosis and concentrations of Lp(a) in plasma prompted our development of an accurate quantitative assay for plasma Lp(a), a monoclonal-antibody-based enzyme-linked immunosorbent assay for Lp(a) that is shown to be sensitive, precise, and highly specific. The response to several isoforms of Lp(a) is linear, and as many as 80 samples can be quantified on one plate. This easily performed assay is suitable for use in the clinical laboratory and for screening large populations.

Download Full-text

Influence of gender on vasomotor effects of oxidized low-density lipoprotein in porcine coronary arteries

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.1997.272.6.h2577 ◽

1997 ◽

Vol 272 (6) ◽

pp. H2577-H2583 ◽

Cited By ~ 2

Author(s):

D. A. Cox ◽

M. L. Cohen

Keyword(s):

Coronary Arteries ◽

Oxidized Ldl ◽

Low Density Lipoprotein ◽

Density Lipoprotein ◽

Low Density ◽

Oxidized Low Density Lipoprotein ◽

Male And Female ◽

No Release ◽

Contraction And Relaxation ◽

Coronary Endothelium

This study compared 5-hydroxytryptamine (5-HT)-induced contraction and relaxation in coronary arteries from male and female pigs and compared the vasomotor effects of the atherosclerotic lipoprotein, oxidized low-density lipoprotein (LDL), in these tissues. 5-HT-induced contraction and endothelium-dependent relaxation were similar, as was sodium nitroprusside-induced relaxation, in coronary arteries from male and female pigs. These data suggest that there were no gender-related differences in 5-HT-induced contraction or 5-HT-mediated nitric oxide (NO) release from the coronary endothelium. In contrast, oxidized LDL (100 micrograms/ml) enhanced 5-HT-induced contraction to a greater extent in coronary arteries from male versus female pigs. Because oxidized LDL inhibited 5-HT-induced relaxation similarly in arteries from male and female animals, a greater effect of oxidized LDL on agonist-induced NO release in tissues from male pigs cannot explain the greater effect on 5-HT-induced contraction. Oxidized LDL contracted coronary arteries from males with a greater force than arteries from females when measured from baseline tone, suggesting that oxidized LDL inhibited basal NO release to a greater extent in coronary arteries from male pigs compared with females, an effect that may have participated in the greater enhancement of 5-HT-induced contraction that occurred in arteries from male pigs. These gender-related differences in the vasomotor effects of oxidized LDL may play an important role in the lesser incidence of cardiovascular disease in premenopausal females than in males and may provide insight into the cardioprotective effect of estrogen.

Download Full-text

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

Biochemical Journal ◽

10.1042/bj2780429 ◽

1991 ◽

Vol 278 (2) ◽

pp. 429-434 ◽

Cited By ~ 58

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.

Download Full-text