Differences in Structural Changes and Pathophysiological Effects of Low-Density Lipoprotein Particles upon Accumulation of Acylhydroperoxy Derivatives in Their Outer Phospholipid Monolayer or upon Modification of Apoprotein B-100 by Natural Dicarbonyls.

Nanoparticles of the lipid-transporting system of the organism, low-density lipoproteins (LDL) of blood plasma, are prone to free radical peroxidation with formation of their main modified forms - oxidized LDL itself (containing hydroperoxy-acyls in phospholipids of the outer layer of particles) and dicarbonyl-modified LDL (apoprotein B-100 in which chemically modified via the Maillard reaction). Based on the study of free radical oxidation kinetics of LDLs, it was found that the existing in the literature designation of "oxidized lipoproteins" is incorrect because it does not reveal the nature of oxidative modification of LDLs. It was shown in this study that the "atherogenic" LDLs (particles of which are actively captured by the cultured macrophages) are not the oxidized LDL (in which LOOH-derivatives of phospholipids are formed by enzymatic oxidation by C-15 lipoxygenase of rabbit reticulocytes), but dicarbonyl-modified LDLs. Important role of the dicarbonyl-modified LDLs in the molecular mechanisms of atherogenesis and endothelial dysfunction is discussed.

Clearance and Utilization of Dicarbonyl-Modified LDL in Monkeys and Humans.

The kinetics of elimination of various dicarbonyl-modified low-density lipoproteins from the bloodstream of Macaca mulatta monkeys were investigated. The low-density lipoproteins (LDL) in the monkey blood plasma were isolated by density gradient ultracentrifugation and labeled in vitro with the fluorescent dye FITC; thereupon, they were modified with different natural low molecular-weight dicarbonyls: malondialdehyde (MDA), glyoxal, or methylglyoxal. The control native FITC-labeled LDL and dicarbonyl-modified FITC-labeled LDL were injected into the monkey's ulnar vein; thereafter, blood samples were taken at fixed time intervals during 24 h. The plasma level of FITC-labeled LDL was determined with spectrofluorimetry. The study established that glyoxal- and monkeysglyoxal-labeled LDL circulated in monkey virtually at the same time as native (non-modified) LDL. In contrast, MDA-modified LDL disappeared from the blood extremely rapidly. Administration of the PCSK9 inhibitor involocumab (which increases LDL utilization) to patients with coronary heart disease (CHD) was found to significantly reduce levels of MDA-modified LDL.

Adsorption of Acylhydroperoxy-Derivatives of Phospholipids from Biomembranes by Blood Plasma Lipoproteins.

It has been established that acylhydroperoxy derivatives of phospholipids from oxidized rat liver mitochondria are captured predominantly by LDL particles but not by HDL during co-incubation with blood plasma lipoproteins, which refutes the previously suggested hypothesis about the involvement of HDL in the reverse transport of oxidized phospholipids and confirms the possibility of different mechanisms of lipohydroperoxide accumulation in LDL during oxidative stress.

Dicarbonyl-Modified Low-Density Lipoproteins Are Key Inducers of LOX-1 and NOX1 Gene Expression in the Cultured Human Umbilical Vein Endotheliocytes.

Expression of LOX-1 and NOX1 genes in the human umbilical vein endotheliocytes (HUVECs) cultured in the presence of low-density lipoproteins (LDL) modified with various natural dicarbonyls was investigated for the first time. It was found that among the investigated dicarbonyl-modified LDLs (malondialdehyde (MDA)-modified LDLs, glyoxal-modified LDLs, and methylglyoxal-modified LDLs), the MDA-modified LDLs caused the greatest induction of the LOX-1 and NOX1 genes, as well as of the genes of antioxidant enzymes and genes of proapoptotic factors in HUVECs. Key role of the dicarbonyl-modified LDLs in the molecular mechanisms of vascular wall damage and endothelial dysfunction is discussed.

Antioxidant Properties of Galanin and Its N-Terminal Fragments in in vitro and in vivo Oxidative Stress Modeling.

Antioxidant properties of rat galanin GWTLNSAGYLLGPHAIDNHRSFSDKHGLT-NH2 (Gal), N-terminal fragment of galanin (2-15 aa) WTLNSAGYLLGPHA (G1), and its modified analogue WTLNSAGYLLGPßAH (G2) were studied in vivo in the rat model of regional myocardial ischemia and reperfusion and in vitro in the process of Cu2+-induced free radical oxidation of human blood plasma low-density lipoproteins. Intravenous administration of G1, G2, and Gal to rats after ischemia induction reduced the infarction size and activities of the necrosis markers, creatine kinase-MB and lactate dehydrogenase, in blood plasma at the end of reperfusion. G1, G2, and Gal reduced formation of the spin adducts of hydroxyl radicals in the interstitium of the area at risk during reperfusion, moreover, G2 and Gal also reduced formation of the secondary products of lipid peroxidation in the reperfused myocardium. It was shown in the in vivo experiments and in the in vitro model system that the ability of galanin peptides to reduce formation of ROS and attenuate lipid peroxidation during myocardial reperfusion injury was not associated directly with their effects on activities of the antioxidant enzymes of the heart: Cu,Zn-superoxide dismutase, catalase, and glutathione peroxidase. The peptides G1, G2, and Gal at concentrations of 0.01 and 0.1 mM inhibited Cu2+-induced free radical oxidation of human low-density lipoproteins in vitro. The results of oxidative stress modeling demonstrated that the natural and synthetic agonists of galanin receptors reduced formation of the short-lived ROS in the reperfused myocardium, as well as of lipid radicals in blood plasma. Thus, galanin receptors could be a promising therapeutic target for cardiovascular diseases.

Aldehyde inhibition of antioxidant enzymes in the blood of diabetic patients.

BACKGROUND: The aim of the present study was to examine the effect of aldehyde modification on antioxidant enzyme activity in diabetic patients. METHODS: The activity of commercially available antioxidant enzymes (catalase, glutathione peroxidase [GPx], and Cu,Zn-superoxide dismutase [SOD]) was determined in vitro prior to and after aldehyde modification. The activity of erythrocyte Cu,Zn-SOD was assayed in blood drawn from healthy donors, diabetic patients with decompensated carbohydrate metabolism, and diabetic patients after glucose-lowering therapy. RESULTS: In vitro aldehyde modification had no effect on catalase activity, but diminished GPx and Cu,Zn-SOD activity. In diabetic patients with decompensated carbohydrate metabolism, glucose-lowering therapy significantly increased Cu,Zn-SOD activity, the effect being especially pronounced after administration of metformin. CONCLUSIONS: It is likely that metformin antagonizes the aldehyde-induced inhibition of erythrocyte Cu,Zn-SOD in diabetic patients more effectively than sulfonylurea drugs.

Antioxidants decreases the intensification of low density lipoprotein in vivo peroxidation during therapy with statins.

The oxidative modification of low density lipoprotein (LDL) is thought to play an important role in atherogenesis. Drugs of beta-hydroxy-beta-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors (statins) family are usually used as a very effective lipid-lowering preparations but they simultaneously block biosynthesis of both cholesterol and ubiquinone Q10 (coenzyme Q), which is an intermediate electron carrier in the mitochondrial respiratory chain. It is known that reduced form of ubiquinone Q10 acts in the human LDL as very effective natural antioxidant. Daily per os administration of HMG-CoA reductase inhibitor simvastatin to rats for 30 day had no effect on high-energy phosphates (adenosin triphosphate, creatine phosphate) content in liver but decreased a level of these substances in myocardium. We study the Cu2+-mediated susceptibility of human LDL to oxidation and the levels of free radical products of LDL lipoperoxidation in LDL particles from patients with atherosclerosis after 3 months treatment with natural antioxidants vitamin E as well as during 6 months administration of HMG-CoA reductase inhibitors such as pravastatin and cerivastatin in monotherapy and in combination with natural antioxidant ubiquinone Q10 or synthetic antioxidant probucol in a double-blind placebo-controlled trials. The 3 months of natural antioxidant vitamin E administration (400 mg daily) to patients did not increase the susceptibility of LDL to oxidation. On the other hand, synthetic antioxidant probucol during long-time period of treatment (3-6 months) in low-dose (250 mg daily) doesn't change the lipid metabolism parameters in the blood of patients but their high antioxidant activity was observed. Really, after oxidation of probucol-contained LDL by C-15 animal lipoxygenase in these particles we identified the electron spin resonance signal of probucol phenoxyl radical that suggests the interaction of LDL-associated probucol with lipid radicals in vivo. We observed that 6 months treatment of patients with pravastatine (40 mg daily) or cerivastatin (0.4 mg daily) was followed by sufficiently accumulation of LDL lipoperoxides in vivo. In contrast, the 6 months therapy with pravastatin in combination with ubiquinone Q10 (60 mg daily) sharply decreased the LDL initial lipoperoxides level whereas during treatment with cerivastatin in combination with probucol (250 mg daily) the LDL lipoperoxides concentration was maintained on an invariable level. Therefore, antioxidants may be very effective in the prevention of atherogenic oxidative modification of LDL during HMG-CoA reductase inhibitors therapy.