scholarly journals Quantification of malondialdehyde and 4-hydroxynonenal adducts to lysine residues in native and oxidized human low-density lipoprotein

1997 ◽  
Vol 322 (1) ◽  
pp. 317-325 ◽  
Author(s):  
Jesús R. REQUENA ◽  
Min Xin FU ◽  
Mahtab U. AHMED ◽  
Alicia J. JENKINS ◽  
Timothy J. LYONS ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Gas Chromatography ◽  
Oxidized Ldl ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Thiobarbituric Acid ◽  
Low Density ◽  
Lysine Residues

Malondialdehyde (MDA) and 4-hydroxynonenal (HNE) are major end-products of oxidation of polyunsaturated fatty acids, and are frequently measured as indicators of lipid peroxidation and oxidative stress in vivo. MDA forms Schiff-base adducts with lysine residues and cross-links proteins in vitro; HNE also reacts with lysines, primarily via a Michael addition reaction. We have developed methods using NaBH4 reduction to stabilize these adducts to conditions used for acid hydrolysis of protein, and have prepared reduced forms of lysine-MDA [3-(Nε-lysino)propan-1-ol (LM)], the lysine-MDA-lysine iminopropene cross-link [1,3-di(Nε-lysino)propane (LML)] and lysine-HNE [3-(Nε-lysino)-4-hydroxynonan-1-ol (LHNE)]. Gas chromatography/MS assays have been developed for quantification of the reduced compounds in protein. RNase incubated with MDA or HNE was used as a model for quantification of the adducts by gas chromatography/MS. There was excellent agreement between measurement of MDA bound to RNase as LM and LML, and as thiobarbituric acid-MDA adducts measured by HPLC; these adducts accounted for 70Ő80% of total lysine loss during the reaction with MDA. LM and LML (0.002Ő0.12mmol/mol of lysine) were also found in freshly isolated low-density lipoprotein (LDL) from healthy subjects. LHNE was measured in RNase treated with HNE, but was not detectable in native LDL. LM, LML and LHNE increased in concert with the formation of conjugated dienes during the copper-catalysed oxidation of LDL, but accounted for modification of < 1% of lysine residues in oxidized LDL. These results are the first report of direct chemical measurement of MDA and HNE adducts to lysine residues in LDL. LM, LML and LHNE should be useful as biomarkers of lipid peroxidative modification of protein and of oxidative stress in vitro and in vivo.

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.

RPE Cells Internalize Low-Density Lipoprotein (LDL) and Oxidized LDL (oxLDL) in Large Quantities In Vitro and In Vivo

2004 ◽  
Vol 45 (8) ◽  
pp. 2822 ◽  
Author(s):  
Nataliya Gordiyenko ◽  
Maria Campos ◽  
Jung Wha Lee ◽  
Robert N. Fariss ◽  
Jorge Sztein ◽  
...  
Keyword(s):  
Oxidized Ldl ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Low Density ◽  
Rpe Cells

Enhancement of macrophage survival and DNA synthesis by oxidized-low-density-lipoprotein (LDL)-derived lipids and by aggregates of lightly oxidized LDL

2001 ◽  
Vol 355 (1) ◽  
pp. 207-214 ◽  
Author(s):  
John A. HAMILTON ◽  
Wendy JESSUP ◽  
Andrew J. BROWN ◽  
Genevieve WHITTY
Keyword(s):  
Dna Synthesis ◽  
Foam Cell ◽  
Oxidized Ldl ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Low Density ◽  
Low Concentrations ◽  
Macrophage Colony

Human atherosclerotic plaque contains a partially characterized range of normal and oxidized lipids formed mainly from free and esterified cholesterol and phospholipids, some of which can be located in macrophage-derived ‘foam’ cells. Oxidation of low-density lipoprotein (LDL) is often considered as an important event leading to subsequent foam-cell development, which may also include enhanced cell survival and/or proliferation. The active component(s) in oxidized LDL (ox.LDL) causing macrophage proliferation is debated. We report here that the lipid component of ox.LDL can promote macrophage survival and DNA synthesis, the latter response showing a synergistic effect in the presence of low concentrations of macrophage colony-stimulating factor. 7-Ketocholesterol showed some stimulation of macrophage DNA synthesis whereas hypochlorite-oxidized (i.e. apolipoprotein B-oxidized) LDL did not. Plaque-derived lipids could enhance macrophage survival. It has not been proven that LDL in lesions is oxidized sufficiently to be the dominant source of sterols in vivo or to be able to induce macrophage growth in vitro or in vivo; it has been suggested that aggregation of modified LDL in vivo is an important step in the deposition of intracellular lipid. We found that aggregation of lightly oxidized LDL potentiated dramatically its ability to stimulate macrophage DNA synthesis, indicating that extensive oxidation of LDL is not required for this response in vitro and perhaps in vivo.

scholarly journals Reductions of copper ion-mediated low-density lipoprotein (LDL) oxidations of trypsin inhibitors, the sweet potato root major proteins, and LDL binding capacities

2020 ◽  
Vol 61 (1) ◽  
Author(s):  
Yeh-Lin Lu ◽  
Chia-Jung Lee ◽  
Shyr-Yi Lin ◽  
Wen-Chi Hou
Keyword(s):  
Sweet Potato ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Trypsin Inhibitors ◽  
Water Soluble ◽  
Affinity Column ◽  
Low Density ◽  
Native Page

Abstract Background The root major proteins of sweet potato trypsin inhibitors (SPTIs) or named sporamin, estimated for 60 to 80% water-soluble proteins, exhibited many biological activities. The human low-density lipoprotein (LDL) showed to form in vivo complex with endogenous oxidized alpha-1-antitrypsin. Little is known concerning the interactions between SPTIs and LDL in vitro. Results The thiobarbituric-acid-reactive-substance (TBARS) assays were used to monitor 0.1 mM Cu2+-mediated low-density lipoprotein (LDL) oxidations during 24-h reactions with or without SPTIs additions. The protein stains in native PAGE gels were used to identify the bindings between native or reduced forms of SPTIs or soybean TIs and LDL, or oxidized LDL (oxLDL). It was found that the SPTIs additions showed to reduce LDL oxidations in the first 6-h and then gradually decreased the capacities of anti-LDL oxidations. The protein stains in native PAGE gels showed more intense LDL bands in the presence of SPTIs, and 0.5-h and 1-h reached the highest one. The SPTIs also bound to the oxLDL, and low pH condition (pH 2.0) might break the interactions revealed by HPLC. The LDL or oxLDL adsorbed onto self-prepared SPTIs-affinity column and some components were eluted by 0.2 M KCl (pH 2.0). The native or reduced SPTIs or soybean TIs showed different binding capacities toward LDL and oxLDL in vitro. Conclusion The SPTIs might be useful in developing functional foods as antioxidant and nutrient supplements, and the physiological roles of SPTIs-LDL and SPTIs-oxLDL complex in vivo will investigate further using animal models.

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

2005 ◽  
Vol 12 (1) ◽  
pp. 68-75 ◽  
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.

scholarly journals Inhibition of in vitro macrophage-induced low density lipoprotein oxidation by thyroid compounds

2003 ◽  
Vol 177 (1) ◽  
pp. 137-146 ◽  
Author(s):  
L Oziol ◽  
P Faure ◽  
N Bertrand ◽  
P Chomard
Keyword(s):  
Cell Viability ◽  
Antioxidant Capacity ◽  
Density Lipoprotein ◽  
Thiobarbituric Acid ◽  
Human Monocyte ◽  
Ldl Oxidation ◽  
Low Density ◽  
Redox Systems

Oxidized low density lipoproteins (LDL) are highly suspected of initiating the atherosclerosis process. Thyroid hormones and structural analogues have been reported to protect LDL from lipid peroxidation induced by Cu2+ or the free radical generator 2,2'-azobis-'2-amidinopropane' dihydrochloride in vitro. We have examined the effects of thyroid compounds on macrophage-induced LDL oxidation. Human monocyte-derived macrophages (differentiated U937 cells) were incubated for 24 h with LDL and different concentrations (0-20 microM) of 3,5,3'-triiodo-l -thyronine (T3), 3,5,3',5'-tetraiodo-L-thyronine (T4), 3,3',5'-tri-iodo-l -thyronine (rT3), the T3 acetic derivative (3,5,3'-tri-iodothyroacetic acid; TA3) or L-thyronine (T0) (experiment 1). Cells were also preincubated for 24 h with 1 or 10 microM of the compounds, washed twice, then incubated again for 24 h with LDL (experiment 2). Oxidation was evaluated by measurement of thiobarbituric acid-reactive substances (TBARS) and cell viability by lactate deshydrogenase release. In experiment 1, T0 had no effect, whereas the other compounds decreased LDL TBARS production, but T3 and TA3 were less active than T4 and rT3 (IC50: 11.0 +/- 2.6 and 8.1 +/- 0.8 vs 1.4 +/- 0.5 and 0.9 +/- 0.3 microM respectively). In experiment 2, the compounds at 1 microM had no effect; at 10 microM, T3 and rT3 slightly reduced LDL TBARS production, whereas TA3 and T4 inhibited it by about 50% and 70% respectively. TBARS released by the cells were also highly decreased by T3, T4, rT3 and TA3 in experiment 1, but only by T3 (30%) and T4 (70%) in experiment 2. Cell viability was not affected by the compounds except slightly by TA3 at 10 microM. The data suggested that the physico-chemical antioxidant capacity of thyroid compounds was modulated by their action on the intracellular redox systems of macrophage. Overall cellular effects of T3 led to a reduction of its antioxidant capacity whereas those of T4 increased it. Thus T4 might protect LDL against cellular oxidation in vivo more than T3.

scholarly journals Sagunja-Tang Improves Lipid Related Disease in a Postmenopausal Rat Model and HepG2 Cells

2015 ◽  
Vol 2015 ◽  
pp. 1-13 ◽  
Author(s):  
Hiroe Go ◽  
Jin Ah Ryuk ◽  
Hye Won Lee ◽  
In Sil Park ◽  
Ki-Jung Kil ◽  
...  
Keyword(s):  
Lipid Accumulation ◽  
Hepg2 Cells ◽  
Cholesterol Synthesis ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Low Density ◽  
Protein Levels ◽  
Related Disease

The present study was conducted to investigate the effect of Sagunja-tang on the lipid related disease in a rat model of menopausal hyperlipidemia and lipid accumulation in methyl-β-cyclodextrin-induced HepG2 cells. Inin vivostudy using menopausal hyperlipidemia rats, Sagunja-tang reduced retroperitoneal and perirenal fat, serum lipids, atherogenic index, cardiac risk factor, media thickness, and nonalcoholic steatohepatitis score, when compared to menopausal hyperlipidemia control rats. In HepG2 cells, Sagunja-tang significantly decreased the lipid accumulation, total cholesterol levels, and low-density/very-low-density lipoprotein levels. Moreover, Sagunja-tang reversed the methyl-β-cyclodextrin-induced decrease in the protein levels of critical molecule involved in cholesterol synthesis, sterol regulatory element binding protein-2, and low-density lipoprotein receptor and inhibited protein levels of 3-hydroxy-3-methylglutaryl coenzyme A reductase as well as activity. Phosphorylation level of AMP-activated protein kinase was stimulated by Sagunja-tang. These results suggest that Sagunja-tang has effect on inhibiting hepatic lipid accumulation through regulation of cholesterol synthesis and AMPK activityin vitro. These observations support the idea that Sagunja-tang is bioavailable bothin vivoandin vitroand could be developed as a preventive and therapeutic agent of hyperlipidemia in postmenopausal females.

scholarly journals Cholesterol-Lowering Effects of Lactobacilli and Bifidobacteria Probiotics in vitro and in vivo

2018 ◽  
pp. 1-12
Author(s):  
Shahenda, M. Elaby ◽  
Asmaa A. Salem ◽  
Jehan, B. Ali ◽  
A. F. Abdel-Salam
Keyword(s):  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Cholesterol Content ◽  
Lipoprotein Cholesterol ◽  
Atherogenic Index ◽  
Low Density ◽  
Low Density Lipoprotein Cholesterol ◽  
Cholesterol Lowering

Two lactobacilli strains; Lactobacillus acidophilus ATCC 20079 and Lactobacillus plantarum ATCC 20179 and two bifidobacteria strains; Bifidobacterium bifidum GSGG 5286 and Bifidobacterium longum ATCC 15707 were studied their abilities to reduce the cholesterol content in vitro. It was investigated that the in vivo cholesterol-lowering effect of L. plantarum ATCC 20179, B. bifidum GSGG 5286 and mixture of both probiotics (L. plantarum ATCC20179 and B. bifidum GSGG5286) on hyperlipidaemic rats for 8 weeks. All lactobacilli and bifidobacteria strains assimilate the cholesterol content in laboratory media. It was observed the highest assimilation of cholesterol was in L. plantarum ATCC 20179 and B. bifidum GSGG 5286 strains. In vivo, L. plantarum ATCC 20179  group was more effective in improving serum lipid profile levels [total cholesterol (TC), triglycerides (TG), low density lipoprotein – cholesterol (LDL-C), high density lipoprotein – cholesterol                   (HDL-C), very low density lipoprotein – cholesterol (VLDL-C) and Atherogenic Index (AI)],                      liver enzyme activities (ALT, AST and ALP),  malonaldehyde (MDA), hydrogen peroxide (H2O2) and total antioxidants capacity (TAC) levels than mixed-organisms and B. bifidum groups, respectively of hyperlipidaemic rats. It was concluded that L. plantarum ATCC 20179 showed more                     favourable results than B. bifidum GSGG 5286 in relation to cardiovascular risk factors in hyperlipidaemic rats.

Folate: In Vitro and in Vivo Effects on VLDL and LDL Oxidation

2007 ◽  
Vol 77 (1) ◽  
pp. 66-72 ◽  
Author(s):  
McEneny ◽  
Couston ◽  
McKibben ◽  
Young ◽  
Woodside
Keyword(s):  
Folic Acid ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Folic Acid Supplementation ◽  
Serum Folate ◽  
Ldl Oxidation ◽  
Low Density ◽  
Acid Supplementation

Raised total homocysteine (tHcy) levels may be involved in the etiology of cardiovascular disease and can lead to damage of vascular endothelial cells and arterial wall matrix. Folic acid supplementation can help negate these detrimental effects by reducing tHcy. Recent evidence has suggested an additional anti-atherogenic property of folate in protecting lipoproteins against oxidation. This study utilized both an in vitro and in vivo approach. In vitro: Very-low-density lipoprotein (VLDL) and low density lipoprotein (LDL) were isolated by rapid ultracentrifugation and then oxidized in the presence of increasing concentrations (0→ μmol/L) of either folic acid or 5-methyltetrahydrofolate (5-MTHF). In vivo: Twelve female subjects were supplemented with folic acid (1 mg/day), and the pre- and post-VLDL and LDL isolates subjected to oxidation. In vitro: 5-MTHF, but not folic acid, significantly increased the resistance of VLDL and LDL to oxidation. In vivo: Following folic acid supplementation, tHcy decreased, serum folate increased, and both VLDL and LDL displayed a significant increase in their resistance to oxidation. These results indicated that in vitro, only the active form of folate, 5-MTHF, had antioxidant properties. In vivo results demonstrated that folic acid supplementation reduced tHcy and protected both VLDL and LDL against oxidation. These findings provide further support for the use of folic acid supplements to aid in the prevention of atherosclerosis.

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