Addition of lutein, lycopene, or β-carotene to LDL or serum in vitro: Effects on carotenoid distribution, LDL composition, and LDL oxidation

Carotenoids are dietary antioxidants transported with plasma lipoproteins, primarily low-density lipoprotein (LDL). In this study in vitro methods were used to increase the amounts of specific, individual carotenoids in LDL. By addition of carotenoid to isolated LDL or to serum, followed by (re)isolation of the lipoproteins, samples of LDL were enriched 4- to 150-fold with lutein, 2- to 15-fold with lycopene, or 3- to 25-fold with β-carotene. Enrichment with specific carotenoids was achieved without affecting the electrophoretic mobility of the lipoprotein, its cholesterol to protein ratio, or the levels of other cartenoids or -tocopherol. The distributions among lipoproteins of carotenoid added to serum were similar, but not identical, to the distributions of the endogenous carotenoids. In particular, for added lutein, a greater proportion was found in HDL, and for added β-carotene, more was found in very low-density lipoprotein (VLDL). We then studied the effect of enriching LDL with specific carotenoids on its susceptibility to oxidation by copper ions. Lutein, β-cryptoxanthin, lycopene, and β-carotene, the four major plasma carotenoids, and -tocopherol were destroyed before the formation of lipid peroxidation products. The rates of destruction of the individual carotenoids differed; lycopene was destroyed most rapidly and lutein most slowly. Upon oxidation of β-carotene-enriched LDL, the rates of destruction of β-carotene, lycopene, and lutein were slowed and the lag times before the initiation of lipid peroxidation increased from 19 to 65 min. Neither effect was observed in LDL enriched with lutein or lycopene. Thus, β-carotene was unique among the carotenoids studied in having a small, but significant effect on LDL oxidation in vitro.     

Why is glycated LDL more sensitive to oxidation than native LDL? A comparative study

It is well established that oxidative modification of low-density lipoprotein (LDL) plays a causal role in human atherogenesis and the risk of atherosclerosis is increased in patients with diabetes mellitus. To examine the influence of different agents which may influence LDL-glycation and oxidation, experiments including glycation with glucose, glucose 6-phosphate, metal chelators (EDTA) and antioxidants (BHT) were performed. The influence of time dependence on the glycation process and the alteration of the electrophoretic mobility of LDL under diverse glycation and/or oxidation conditions was also investigated. The formation of conjugated dienes and levels of lipid peroxides in these different LDL-modifications were estimated. The copper-induced oxidation of LDL in vitro was determined by measurement of thiobarbituric acid reactive substances (TBARS) and expressed as nmol MDA/mg of LDL protein. We found that glycated LDL is more prone to oxidation than native LDL. Using native LDL, the maximal oxidation effect was found to reach a value of 49.72 nmol MDA/mg protein after 8 h. The maximum oxidation of the 31 days, glycated LDL with glucose was 71.76 nmol MDA/mg protein amounting to 144.33% of the value found for native LDL. In the case of glucose 6-phosphate glycation, the maximum oxidation under the same conditions amounted to 173.77% of the value found for native LDL. To measure the extent of glycation, fluorescence of advanced glycation end products (AGEs) was determined (370 nm excitation and 440 nm emission). The most potent glycation agent was glucose 6-phosphate leading to the formation of very high amounts of AGEs. This process was promoted in the absence of EDTA, which prevents the oxidative cleavage of modified Amadori products (ketoamines) to AGEs. We therefore conclude that both processes, glycation and oxidation, result in the modification of LDL. The lower the glycation-rate (+/- EDTA) as measured by relative fluorescence units RFU (generation of AGEs), the lower the additional oxidation rate after glycation as measured by TBARS (generation of MDA equivalents). Glycation and/or oxidation change the electrophoretic mobility of LDL.     

Is the LDL Receptor Involved in Cortical Amyloid Protein Clearance?

This article puts forward the hypothesis that the Low Density Lipid Receptor (LDLR) is one of the molecules that is involved in the clearance of amyloid proteins in the brain and that it may play a role in Alzheimer’s Disease (AD) via its up-regulation by statins. The hypothesis is built on the following observations: a-statins (which have been shown to increase LDLR in astrocytes, see below) have a beneficial role in AD, b-defects in the LDL receptor gene are found in AD, c-molecules with similar structure to the LDLR have been shown to clear amyloid protein from the brain.     

LDL亚组分研究进展

根据低密度脂蛋白（low density lipoprotein,LDL）颗粒的不均一性,可以利用密度梯度超速离心法和梯度凝胶电泳法将其分成若干亚组分。近年来,对于LDL亚组分分离方法的研究取得了显著进展。除对上述两种基本实验方法进行改进外,有实验室采用Western印迹法对LDL颗粒进行分离。LDL亚组分分离方法的进步,使对LDL亚组分的认识更加深入：LDL亚组分的高度不均一性、氧化易感性及电负性等不同特性与动脉粥样硬化（atherosclerosis,AS）关系密切。LDL亚组分的研究为认识动脉粥样硬化及其相关疾病提供了重要的理论依据。     

α-tocopherol,ascorbic acid,and rutin inhibit synergistically the copper-promoted LDL oxidation and the cytotoxicity of oxidized LDL to cultured endothelial cells

Low-density lipoproteins (LDL) mildly oxidized by copper ions or UV radiations exhibit a cytotoxic effect to cultured endothelial cells. Rutin, a polyphenolic flavonoid, ascorbic acid, and α-tocopherol were able to inhibit the peroxidation of LDL and their subsequent cytotoxicity. The mixture of the three compounds (rutin/ascorbic acid/α-tocopherol, 4/4/1) exhibited a supra-additive antioxidant effect. The inhibition of the cytotoxic effect was well correlated with that of TBARS formation. Another important conclusion is that these antioxidants were able to prevent directly at the cellular level the cytotoxic effect of oxidized LDL, since cells preincubated with them were protected against the cytotoxic effect of previously oxidized LDL. The protective effect of antioxidants was limited because of their own toxicity. The antioxidant mixture permitted a maximal cytoprotective effect with relatively lower concentrations to be obtained and the cytotoxicity of high concentrations to be avoided. In conclusion, rutin, ascorbic acid, and α-tocopherol constitute two lines of defense in protecting cells against injury owing to oxidation of LDL (1) at the LDL level, by inhibiting the LDL oxidation and the subsequent cytotoxicity, and (2) at the cellular level, by protecting the cells directly, i.e., by increasing their resistance against the cytotoxic effect of oxidized LDL.     

Three-Dimensional cryoEM Reconstruction of Native LDL Particles to 16? Resolution at Physiological Body Temperature

Background

Low-density lipoprotein (LDL) particles, the major carriers of cholesterol in the human circulation, have a key role in cholesterol physiology and in the development of atherosclerosis. The most prominent structural components in LDL are the core-forming cholesteryl esters (CE) and the particle-encircling single copy of a huge, non-exchangeable protein, the apolipoprotein B-100 (apoB-100). The shape of native LDL particles and the conformation of native apoB-100 on the particles remain incompletely characterized at the physiological human body temperature (37°C).

Methodology/Principal Findings

To study native LDL particles, we applied cryo-electron microscopy to calculate 3D reconstructions of LDL particles in their hydrated state. Images of the particles vitrified at 6°C and 37°C resulted in reconstructions at ∼16 Å resolution at both temperatures. 3D variance map analysis revealed rigid and flexible domains of lipids and apoB-100 at both temperatures. The reconstructions showed less variability at 6°C than at 37°C, which reflected increased order of the core CE molecules, rather than decreased mobility of the apoB-100. Compact molecular packing of the core and order in a lipid-binding domain of apoB-100 were observed at 6°C, but not at 37°C. At 37°C we were able to highlight features in the LDL particles that are not clearly separable in 3D maps at 6°C. Segmentation of apoB-100 density, fitting of lipovitellin X-ray structure, and antibody mapping, jointly revealed the approximate locations of the individual domains of apoB-100 on the surface of native LDL particles.

Conclusions/Significance

Our study provides molecular background for further understanding of the link between structure and function of native LDL particles at physiological body temperature.     

阴离子型LDL吸附材料概述

动脉粥样硬化是导致心血管疾病发生的最重要因素。血液中低密度脂蛋白(LDL)浓度过高是引起动脉粥样硬化的主要原因。体外去除法是目前降低LDL浓度最有效的方法之一,吸附材料是影响LDL体外去除法降低LDL浓度效果的关键。阴离子型吸附材料是一种常用的吸附材料,因选材广泛、吸附效果佳备受关注,由发挥吸附功能的阴离子化合物配基和承载配基的载体基材组成,通过阴离子所带的负电荷与带正电的LDL产生特异性吸附。根据配基分子大小,阴离子型吸附材料主要分为大分子阴离子型和小分子阴离子型吸附材料,本文总结了国内外的阴离子型吸附材料主要研究现状及发展趋势。     

LDL的氧化修饰和氧化修饰LDL的组成和结构变化

与低密度脂蛋白(LDL)相比,氧化修饰LDL(O-LDL)的组成、结构和生物学特性发生了深刻的变化,而组成和结构的改变是生物学特性改变的基础.本文根据最近文献资料.结合我们实验室的工作.对LDL的氧化修饰、O-LDL的组成、结构改变,以及它们的机理作一简要综述.     

5-Lipoxygenase contributes to PPARγ activation in macrophages in response to apoptotic cells

Macrophage polarization to an anti-inflammatory phenotype upon contact with apoptotic cells is a contributing hallmark to immune suppression during the late phase of sepsis. Although the peroxisome proliferator-activated receptor γ (PPARγ) supports this macrophage phenotype switch, it remains elusive how apoptotic cells activate PPARγ. Assuming that a molecule causing PPARγ activation in macrophages originates in the cell membrane of apoptotic cells we analyzed lipid rafts from apoptotic, necrotic, and living human Jurkat T cells which showed the presence of 5-lipoxygenase (5-LO) in lipid rafts of apoptotic cells only. Incubating macrophages with lipid rafts of apoptotic, but not necrotic or living cells, induced PPAR responsive element (PPRE)-driven mRuby reporter gene expression in RAW 264.7 macrophages stably transduced with a 4xPPRE containing vector. Experiments with lipid rafts of apoptotic murine EL4 T cells revealed similar results. To verify the involvement of 5-LO in activating PPARγ in macrophages, Jurkat T cells were incubated with the 5-LO inhibitor MK-866 prior to induction of apoptosis, which failed to induce mRuby expression. Similar results were obtained with lipid rafts of apoptotic EL4 T cells preexposed to the 5-LO inhibitors zileuton and CJ-13610. Interestingly, Jurkat T cells overexpressing 5-LO failed to activate PPARγ in macrophages, while their 5-LO overexpressing apoptotic counterparts did. Our results suggest that during apoptosis 5-LO gets associated with lipid rafts and synthesizes ligands that in turn stimulate PPARγ in macrophages.     

Oxidized low-density lipoprotein (oxLDL) plasma levels and oxLDL to LDL ratio — Are they real oxidative stress markers in dialyzed patients?

AimsDyslipidemia and oxidative stress are commonly present in patients during maintenance dialysis treatment. However, the significance of oxidized LDL (oxLDL) as a marker of oxidative stress in uremia is still unresolved. The aim of this study was to establish the role of oxLDL and oxLDL/LDL ratio as markers of lipoprotein abnormalities and oxidative stress in the dialyzed patients.Main methodsPlasma oxLDL level was measured by ELISA, and oxLDL/LDL ratio was calculated in 106 dialyzed patients and 20 controls. The linkages between oxLDL, oxLDL/LDL ratio and lipid profile and oxidative stress markers malondialdehyde (MDA) and Cu/Zn superoxide dismutase (Cu/Zn SOD) levels were also analyzed.Key findingsOxLDL levels and oxLDL/LDL ratio were similar in hemodialyzed patients and controls, whereas these parameters were lower in peritoneally dialyzed patients when compared to healthy individuals. In contrast, both MDA and Cu/Zn SOD levels were significantly higher in uremics than in controls. oxLDL and oxLDL/LDL ratio positively correlated with lipid profile (except of HDL), whereas there were no positive associations between these parameters and both MDA and Cu/Zn SOD. Multiple regression analysis confirmed that increased oxLDL/HDL and TC/HDL ratios and total cholesterol levels are the parameters which independently predicted oxLDL in dialyzed patients. In the case of oxLDL/LDL ratio, the independent variables were oxLDL/HDL ratio, total cholesterol and HDL levels.SignificanceoxLDL levels and oxLDL/LDL ratio seem to be the markers of lipoprotein abnormalities rather than the markers of oxidative stress in the population of dialyzed patients.     

Salusin-β accelerates inflammatory responses in vascular endothelial cells via NF-κB signaling in LDL receptor-deficient mice in vivo and HUVECs in vitro

The bioactive peptide salusin-β is highly expressed in human atheromas; additionally, infusion of antiserum against salusin-β suppresses the development of atherosclerosis in atherogenic mice. This study examined the roles of salusin-β in vascular inflammation during atherogenesis. Infusion of antiserum against salusin-β attenuated the induction of VCAM-1, monocyte chemoattractant protein (MCP)-1, and IL-1β and as well as nuclear translocation of NF-κB in aortic endothelial cells (ECs) of LDL receptor-deficient mice, which led to the prevention of monocyte adhesion to aortic ECs. In vitro experiments indicated that salusin-β directly enhances the expression levels of proinflammatory molecules, including VCAM-1, MCP-1, IL-1β, and NADPH oxidase 2, as well as THP-1 monocyte adhesion to cultured human umbilical vein ECs (HUVECs). Both salusin-β-induced VCAM-1 induction and monocyte/HUVEC adhesion were suppressed by pharmacological inhibitors of NF-κB, e.g., Bay 11-7682 and curcumin. Furthermore, the VCAM-1 induction was significantly prevented by the phosphatidylinositol 3-kinase (PI3K) inhibitor LY-294002, whereas it was accelerated by the ERK inhibitor, U-0126. Treatment of HUVECs with salusin-β, but not with salusin-α, accelerated oxidative stress and nuclear translocation of NF-κB as well as phosphorylation and degradation of IκB-α, an endogenous inhibitor of NF-κB. Thus, salusin-β enhanced monocyte adhesion to vascular ECs through NF-κB-mediated inflammatory responses in ECs, which can be modified by PI3K or ERK signals. These findings are suggestive of a novel role of salusin-β in atherogenesis.     

Myeloperoxidase-mediated LDL oxidation and endothelial cell toxicity of oxidized LDL: attenuation by (−)-epicatechin

Recent data suggest an inverse epidemiological association between intake of flavanol-rich cocoa products and cardiac mortality. Potential beneficial effect of cocoa may be attributed to flavanol-mediated improvement of endothelial function, as well as to enhancement of bioavailability and bioactivity of nitric oxide in vivo. ( ? )-Epicatechin is one bioactive flavanol found in cocoa. This review deals with protective actions of ( ? )-epicatechin on two key processes in atherogenesis, oxidation of LDL and damage to endothelial cell by oxidized LDL (oxLDL), with emphasis on data from this laboratory. ( ? )-Epicatechin not only abrogates or attenuates LDL oxidation but also counteracts deleterious actions of oxLDL on vascular endothelial cells. These protective actions are only partially shared by other vasoprotective agents such as vitamins C and E or aspirin. Thus, ( ? )-epicatechin appears to be a pleiotropic protectant for both LDL and endothelial cells.     

UN system efforts to support the response to AIDS in China

INTRODUCTION China has made considerable progress in the response to the AIDS epidemic in the past years, particularly in terms of commitment by the national leadership, the establish- ment of a supportive policy framework, improved under- standing of the…     

Myosins in melanocytes: to move or not to move?

The actin network has been implicated in the intracellular transport and positioning of the melanosomes, organelles that are specialized in the biosynthesis and the storage of melanin. It contributes also to molecular mechanisms that underlie the intracellular membrane dynamics and thereby can control the biogenesis of melanosomes. Two mechanisms for actin‐based movements have been identified: one is dependent on the motors associated to actin namely the myosins; the other is dependent on actin polymerization. This review will focus on to the role of the actin cytoskeleton and myosins in the transport and in the biogenesis of melanosomes. Myosins involved in membrane traffic are largely seen as transporters of organelles or membrane vesicles containing cargos along the actin networks. Yet increasing evidence suggests that some of the myosins contribute to the dynamics of internal membrane by using other mechanisms. The role of the myosins and the different molecular mechanisms by which they contribute or may contribute to the distribution, the movement and the biogenesis of the melanosomes in epidermal melanocytes and retinal pigmented epithelial (RPE) cells will be discussed.