Low density lipoprotein immunoapheresis does not increase plasma lipid peroxidation products in vivo.

Extracorporeal elimination of low density lipoprotein (LDL) is frequently used in drug-resistant hypercholesterolemia. LDL-immunoapheresis selectively removes LDL and lipoprotein(a) [Lp(a)] from plasma. Lipid peroxidation is one unwanted side effect, that occurs during extracorporeal plasma treatment. The purpose of this study was to investigate the effect of LDL immunoapheresis on lipid peroxidation. Before and after a single LDL-immunoapheresis treatment, plasma concentrations of lipid hydroperoxides, determined with two different spectophotometric assays, thiobarbituric acid-reacting substances (TBARS), determined spectrophotometrically and malondialdehyde (MDA), determined by an MDA-TBA/HPLC method, were measured in 13 hypercholesterolemic patients. In addition MDA was also determined in the eluate of the apheresis column. Before treatment, plasma cholesterol and LDL cholesterol concentrations were significantly higher in patients than in healthy control subjects, as were the lipid peroxidation products. LDL-immunoapheresis treatment of the patients led to significant decreases in total cholesterol (69+/-8%), LDL-cholesterol (79+/-7%), HDL-cholesterol (35+/-17%), triglycerides (38+/-21%), apolipoprotein-B (77+/-6%), apolipoprotein-A1 (25+/-5%) and Lp(a) concentrations (76+/-10%). Changes in plasma lipid peroxide concentrations (17+/-8 nmol/l before vs. 14+/-5 nmol/l after treatment) were not significant, neither were those in TBARS (3. 0+/-2.6 micromol/l vs. 2.3+/-1.3 micromol/l) or MDA concentrations (1.03+/-0.17 micromol/l vs. 1.0+/-0.20 micromol/l). Patients with high baseline values showed a decrease, whereas others did not. MDA was present (0.57+/-0.13 micromol/l) in the eluate of the apheresis column, suggesting that, along with LDL, lipid peroxidation products are also removed. From these results we conclude that a single LDL-immunoapheresis treatment effectively reduces LDL and Lp(a) in the absence of increases in plasma lipid peroxidation products.     

The evaluation of autoantibodies against oxidatively modified low-density lipoprotein (LDL), susceptibility of LDL to oxidation,serum lipids and lipid hydroperoxide levels,total antioxidant status,antioxidant enzyme activities,and endothelial dysfunction in patients with Behçet's disease

OBJECTIVES: Beh?et's disease is a multisystem disorder characterized by a chronic inflammation including acute attacks and remission periods. Decreased enzyme activity of the antioxidant system and increased levels of free radicals may have important roles in the damage of tissues observed in the disease period. In addition, the atherogenic tendency of serum lipid, lipoproteins, lipid peroxidation levels and endothelial dysfunction accompany the above mentioned findings. As a consequence of these events, different degrees of low density lipoprotein (LDL) oxidation occur in vivo, and then autoantibodies against oxidized-LDL(AuAb-oxLDL) are produced. DESIGN AND METHODS: Lipids, lipoproteins, lipid hydroperoxide, AuAb-oxLDL, total antioxidant status (TAS), serum-soluble intercellular adhesion molecule-1 (sICAM-1), plasminogen activator inhibitor 1 (PAI-1) levels in serum, the activities of antioxidant enzymes including glutathione peroxidase (GSH-Px), glutathione reductase (GR), superoxide dismutase (SOD), and catalase (CAT) in erythrocytes and plasma, were determined in 25 patients with Beh?et's disease and in 25 healthy volunteers. Also, susceptibility to copper-induced in vitro oxidation of LDL by using lag time, a measure of resistance to oxidation, oxidation rate and extent of oxidation, a measure of diene production in both groups, was studied. RESULTS: It was observed that lipid hydroperoxide and AuAb-oxLDL levels in patients with Beh?et's disease were significantly higher, but erythrocyte SOD, CAT, plasma GSH-Px activities, and TAS were significantly lower than those in healthy subjects. Susceptibility of LDL to oxidation in the patients was found to be increased. Total cholesterol, LDL-C and apo B levels and acute phase reactants were significantly higher, but HDL-C and apo AI levels were significantly lower, in patients when compared to healthy subjects. The levels of AuAb-oxLDL in patients were found to correlate with TAS, total cholesterol, LDL-C, lipid hydroperoxide and erythrocyte SOD activities (r = -0.62, p < 0.01; r = 0.64, p < 0.01; r = 0.55, p < 0.01; r = 0.81, p < 0.01; r = -0.63, p < 0.01, respectively). In addition, lipid hydroperoxide levels were found to correlate with total cholesterol, LDL-C and erythrocyte SOD activities (r = 0.45, p < 0.05; r = 0.45, p < 0.05; r = -0.46, p < 0.05, respectively). PAI-1 and sICAM-1 were found to be increased in the patients and correlated with AuAb-oxLDL and lipid hydroperoxide levels (r = 0.56, p < 0.01; r = 0.67, p < 0.01 and r = 0.59, p < 0.01; r = 0.61, p < 0.01, respectively). CONCLUSIONS: It was concluded that the observed increase of lipid, lipoproteins, lipid hydroperoxide, susceptibility of LDL to oxidation, autoantibodies against ox-LDL levels and decrease of antioxidant enzyme activities and total antioxidant status and increased secretion of endothelial derivated peptides including sICAM and PAI-1, and their interactions may indicate that there is a tendency to atherothrombotic events in patients with Beh?et's disease.     

Evaluation of the atherogenic tendency of lipids and lipoprotein content and their relationships with oxidant-antioxidant system in patients with psoriasis

BACKGROUND: Psoriasis is a common chronic and recurrent inflammatory skin disease that can occur due to abnormalities in essential fatty acid metabolism, lymphokine secretion, free radical generation, lipid peroxidation and eicosanoid metabolism, and has been associated with increased frequency of cardiovascular events. The current study was designed to evaluate plasma lipids, susceptibility of LDL to oxidation and oxidant-antioxidant status and their relationships in patients with psoriasis. METHODS: The study group included 35 patients with psoriasis (18 females and 17 males), and 35 sex- and age-matched healthy volunteers (16 females and 19 males). From blood samples, their lipids, lipoproteins, acute phase reactants, lipid peroxidation products [lipid hydroperoxide (LHP) and malondialdehyde (MDA)], antioxidant enzymes [glutathione peroxidase (GSH-Px), glutathione reductase (GR), superoxide dismutase (SOD), catalase (CAT)], total antioxidant status (TAS) and autoantibodies against oxidized low-density lipoprotein (AuAb-oxLDL) levels were determined. Moreover, the susceptibility of copper-induced in vitro oxidation of LDL was examined. RESULTS: The mean levels of atherogenic lipids (total cholesterol [TC], triacylglycerol [TG] and LDL cholesterol [LDL-C]), acute-phase reactants (CRP, ESR, PMNLs, ceruloplasmin and fibrinogen) and lipid peroxidation products, AuAb-oxLDL levels in patients with psoriasis were found to be significantly higher than those of healthy subjects. On the other hand, TAS and antioxidant enzyme activities (CAT, SOD and GSH-Px in erythrocyte and SOD in plasma) were significantly lower when compared to healthy subjects. The lag times [t(lag)], a measure of resistance to oxidation of LDL, were also lower. The levels of AuAb-oxLDL in patients were correlated with TC, LDL-C, plasma LHP, erythrocyte MDA, oxidized LDL-MDA (oxLDL-MDA), fibrinogen, CRP, PMNL levels and plasma SOD activities (r = 0.69, P < 0.01; r = 0.64, P < 0.01; r = 0.38, P < 0.05; r = 0.65, P < 0.01; r = 0.34, P < 0.05; r = 0.34, P < 0.05; r = 0.53, P < 0.01, r = 0.34, P < 0.05; r = -0.67, P < 0.01, respectively). On the other hand, t(lag) was correlated negatively with the levels of VLDL-TG, VLDL-TC and LDL-TG but positively correlated with the levels of TAS in psoriatics (r = -0.49, P < 0.01; r = -0.49, P < 0.01, r = -0.65, P < 0.05; r = 0.37, P < 0.05). CONCLUSIONS: It was concluded that the psoriatic patients could be considered as a group with an increased atherosclerotic risk because of increased oxidant stress, decreased antioxidant capacity and susceptibility in lipid profile and lipoprotein content to atherogenicity.     

Distinct effects of LDL apheresis by hemoperfusion (DALI) and heparin-induced extracorporeal precipitation (HELP) on leukocyte respiratory burst activity of patients with familial hypercholesterolemia

Hypercholesterolemia and oxidative stress are major risk factors in atherogenesis. In the last years, lipid apheresis has been established as an effective clinical therapy by lowering not only elevated plasma low-density lipoprotein (LDL) levels but also by reducing the incidence of cardiovascular events. The aim of the present study was to investigate peripheral leukocyte oxidant generation in patients with familial hypercholesterolemia (FH) undergoing regular LDL apheresis. The activity state of leukocytes was estimated prior to, immediately after, and 2 days after LDL apheresis carried out by two distinct techniques: hemoperfusion with the DALI system and heparin-induced extracorporeal LDL precipitation (HELP). Oxidant generating activity was measured by chemiluminescence (CL) in whole blood and isolated polymorphonuclear leukocytes (PMNL). The results of our study show increased baseline respiratory burst activities in FH patients as compared to healthy controls. Apheresis with the HELP system was followed by increases in leukocyte count, zymosan-induced whole blood CL, and plasma PMNL elastase levels. The DALI technique caused no changes in leukocyte count and elastase levels and decreased whole blood CL activity. Two days after lipid removal the observed changes returned to pre-apheresis levels. Leukocyte activity parameters before and after apheresis did not correlate with the corresponding plasma levels of triglycerides, total cholesterol, and LDL cholesterol, suggesting that different handling in the framework of both apheresis techniques rather than lipid profile changes during therapy accounted for leukocyte activity modulation.     

Acute and long-term effects of low-density lipoprotein (LDL)-apheresis on oxidative damage to LDL and reducing capacity of erythrocytes in patients with severe familial hypercholesterolaemia

Several studies have suggested that the oxidative modification of low-density lipoprotein (LDL) could play a key role in the early stages of atherosclerosis. The susceptibility of LDL to oxidation has been found to be greater in patients with coronary heart disease. Familial hypercholesterolaemia (FH) is a powerful clinical model in which to study the predictive role of LDL in atherogenesis. LDL-apheresis is a treatment that is able to decrease lipid levels in plasma. This study was aimed at investigating the reducing capacity of erythrocytes and the in vitro susceptibility to oxidation of LDL isolated from patients with homozygous, heterozygous and double-heterozygous FH, who were treated fortnightly with LDL-apheresis or left untreated. In 14 FH patients, at baseline and after a cycle of treatment, the susceptibility of LDL to oxidative modification was analysed by studying the kinetics of conjugate diene formation. Plasma hydroperoxides, polyunsaturated fatty acid content, LDL electrophoretic mobility on agarose, the titre of auto-antibodies against oxidized LDL and serum paraoxonase activity were also measured. Furthermore, in order to evaluate a potential relationship between LDL oxidation and redox status, erythrocyte GSH and ATP levels were determined in FH patients treated regularly or never treated previously by LDL-apheresis. Unlike in the control group, the oxidative status of LDL in all FH patients was modified by LDL-apheresis, as revealed by the higher negative charge and the increase in levels of hydroperoxides and antibodies against oxidized LDL in the plasma. Our findings suggest both an acute effect and a long-term effect of LDL-apheresis in FH patients treated with dextran sulphate cellulose apheresis. The acute effect of LDL-apheresis on the susceptibility to oxidation of plasma and LDL was demonstrated by significant decreases in plasma hydroperoxide content, total LDL concentration and polyunsaturated fatty acid content. The increased resistance of LDL to oxidation was shown by prolongation of the lag time (P<0.05) in samples after a single cycle of treatment. The long-term effect of LDL-apheresis was demonstrated by the comparable values for lag phases (obtained from the kinetics of conjugate diene formation) in patients under active treatment and controls. Compared with healthy controls and untreated patients, the erythrocyte GSH content was significantly higher (P相似文献   

Review: the oxidant/antioxidant balance during regular low density lipoprotein apheresis.

Low density lipoprotein (LDL) apheresis is a safe procedure to treat severe hypercholesterolemia in patients with chronic heart disease (CHD). However, both hypercholesterolemia and extracorporeal treatment have been associated with oxidative stress. Even though LDL lowering has been proven to reduce CHD, the oxidative modification of LDL has been suggested to render these lipoproteins more atherogenic. It is therefore important to know whether LDL apheresis is safe with respect to oxidative stress including LDL oxidation. The contact of living cells such as leukocytes with artificial surfaces during extracorporeal treatment induces the liberation of various chemokines and cytokines as well as oxygen-derived radicals also known as respiratory burst. These effects justify the consideration of leukocyte activation resulting from extracorporeal treatment as an inflammatory reaction. In extracorporeal circuits such as those used for hemodialysis, the release of oxygen radicals has been shown and depends on the fiber material used in the dialyzer membranes. Reactive oxygen radicals can interact with different cell components such as carbohydrates, DNA, proteins, and lipids. Antioxidants in the form of low molecular weight molecules such as glutathione or radical scavenging enzymes such as superoxide dismutase offer protection against the damaging effects of prooxidants. The disturbed balance between prooxidants and antioxidants is considered as oxidative stress. Therefore, either an increase in oxygen radical formation or a decrease of antioxidants will lead to oxidative stress. During LDL apheresis, a decrease of low molecular weight antioxidants has been reported. In contrast, we have observed an increase in plasma glutathione concentrations but no severe reduction in the activity of antioxidant enzymes in plasma, red cells, or granulocytes, which may explain the lack of plasma lipid peroxidation shown during this kind of extracorporeal treatment. In addition, LDL isolated at the end of apheresis procedures are more resistant to oxidation. These findings suggest that LDL apheresis is safe with respect to radical mediated injury.     

Incides of lipid peroxidation in patients with hypercholesterolemia and hypertriglyceridemia

Peroxidation of lipoproteins has received attention recently in connection with its possible initiation and propagation of atherosclerosis. We studied indices of lipid peroxidation in plasma of 30 patients with either hypercholesterolemia or both hypercholesterolemia and hypertriglyceridemia, and compared them with those of 19 healthy subjects. In the patients lipid hydroperoxides measured both iodometrically and as thiobarbituric acid-reactive substances (TBA-RS) were significantly increased, while concentrations of thiol groups remained unchanged. There was no correlation between concentrations of hydroperoxides and TBA-RS, possibly because the two methods assessed different breakdown products of lipid peroxidation. Lipid hydroperoxide levels, but not those of TBA-RS were correlated with LDL-cholesterol and triglyceride levels. In 6 patients administration of vitamin E at a daily dosage of 300 mg for 4 to 6 weeks depressed elevated hydroperoxides by 33%, and TBA-RS by 44% on average.     

Lipid peroxidation in hemodialysis patients: effect of vitamin C supplementation

OBJECTIVES: Renal failure is associated with several metabolic disturbances and increasing evidences support a role of oxidative stress and impaired antioxidant defence in the pathologic mechanisms that may contribute to accelerated atherogenesis in these patients. Aim of the study was to further investigate the relationship between oxidative stress and chronic renal failure. DESIGN AND METHODS: We compared the paraoxonase (PON1) activity, the levels of lipid hydroperoxides and AGE adducts in plasma of hemodialysis patients before and after intravenous administration of vitamin C. RESULTS: An increase in lipid hydroperoxides, AGE adducts and a decrease in the activity of PON1 were observed in patients with respect to controls. The comparison before and after supplementation with vitamin C showed an increase of PON1 activity and a decrease of AGE and lipid hydroperoxides levels. CONCLUSIONS: The results provide further evidence that lipid peroxidation and impairment of antioxidant system in plasma of patients may play a role in renal disease and suggest that evaluation of PON1 activity could represents an useful approach to monitor antioxidant treatment and new dialysis therapies.     

The Friedewald formula underestimates LDL cholesterol at low concentrations.

Due to recent advances in the treatment of hypercholesterolemia, low density lipoprotein (LDL) cholesterol concentrations below 2.6 mmol/l have become attainable. In general, LDL cholesterol is determined indirectly according to Friedewald. We examined the performance of the Friedewald formula at low concentrations of LDL cholesterol in comparison with a beta-quantification method. We analyzed 176 samples from individuals treated by LDL apheresis with a mean LDL cholesterol concentration of 3.07 mmol/l and found that the Friedewald formula underestimated LDL cholesterol with a bias of -18.5%, -14.5%, -7.3%, and -3.8% at mean LDL cholesterol levels of 1.58, 2.4, 3.49, and 4.67 mmol/l, respectively. Thus, the lower the LDL cholesterol concentration was, the greater the negative bias. We conclude that the Friedewald formula may not be reliable at low LDL cholesterol concentrations produced by LDL apheresis. This finding may also be of relevance to the monitoring of patients being treated with lipid lowering drugs.     

Lecithin–cholesterol acyltransferase activity in normocholesterolaemic and hypercholesterolaemic roosters: modulation by lipid apheresis

Lipid apheresis, a recently described procedure for the elimination of lipid but not apolipoproteins from plasma, was applied to normocholesterolaemic and hypercholesterolaemic roosters. Lipid apheresis resulted in an immediate reduction in plasma unesterified cholesterol concentration, which was sustained for 150 min. The reduction in unesterified cholesterol concentration was higher in the normocholesterolaemic animals than in the hypercholesterolaemic animals. Lipid apheresis induced changes in the ratio of plasma unesterified to total cholesterol in normocholesterolaemic animals but not in hypercholesterolaemic animals. In hypercholesterolaemic animals, lecithin–cholesterol acyltransferase (LCAT) activity was not affected by lipid apheresis, whereas in normocholesterolaemic animals LCAT activity was acutely reduced for 150 min after lipid apheresis. Saturated LCAT kinetics occurred in the hypercholesterolaemic animals but not in the normocholesterolaemic animals. LCAT obeyed Michaelis–Menten kinetics. After lipid apheresis, there was a pool of unesterified cholesterol that was available as substrate for LCAT to a greater extent in hypercholesterolaemic animals than in normocholesterolaemic animals. These observations may have important implications for lipid apheresis as a treatment for atherosclerosis.     

Lecithin–cholesterol acyltransferase activity in normocholesterolaemic and hypercholesterolaemic roosters: modulation by lipid apheresis

Lipid apheresis, a recently described procedure for the elimination of lipid but not apolipoproteins from plasma, was applied to normocholesterolaemic and hypercholesterolaemic roosters. Lipid apheresis resulted in an immediate reduction in plasma unesterified cholesterol concentration, which was sustained for 150 min. The reduction in unesterified cholesterol concentration was higher in the normocholesterolaemic animals than in the hypercholesterolaemic animals. Lipid apheresis induced changes in the ratio of plasma unesterified to total cholesterol in normocholesterolaemic animals but not in hypercholesterolaemic animals. In hypercholesterolaemic animals, lecithin–cholesterol acyltransferase (LCAT) activity was not affected by lipid apheresis, whereas in normocholesterolaemic animals LCAT activity was acutely reduced for 150 min after lipid apheresis. Saturated LCAT kinetics occurred in the hypercholesterolaemic animals but not in the normocholesterolaemic animals. LCAT obeyed Michaelis–Menten kinetics. After lipid apheresis, there was a pool of unesterified cholesterol that was available as substrate for LCAT to a greater extent in hypercholesterolaemic animals than in normocholesterolaemic animals. These observations may have important implications for lipid apheresis as a treatment for atherosclerosis.     

Predictability of low-density lipoprotein levels during apheretic treatment of hypercholesterolemia

The efficiency and efficacy of low-density lipoprotein (LDL) apheresis performed with a dextran sulphate cellulose (DSC) regenerating unit were tested in five familial hypercholesterolaemic patients. LDL apheresis was repeated four times at both bi-weekly and weekly intervals, processing one plasma volume each time. The efficiency of the procedure (i.e., the extent of lipoprotein removal) was nearly identical with both schedules. Efficacy parameters, i.e., decreases of plasma total and LDL cholesterol (TC and LDL-C) and apo B, were highly correlated (r greater than 0.96) with preapheresis levels, allowing an accurate prediction of the absolute lipid removal in the single individual. Plasma triglycerides, high-density lipoprotein cholesterol, apo A-I and apo A-II recovered rather rapidly, reaching 91-96% of the pre-apheresis values in 48 hours; the recovery of TC, LDL-C and apo B was much slower, with a relatively rapid early phase (80% recovery after about 7 days) followed by a successive slower rise. This pattern was highly reproducible in the single patient, allowing the definition of a simple mathematical model for an accurate (error less than 20%) prediction of the individual process. Based on this model one can design the treatment schedule necessary to maintain lipid levels within the desired range in the single individual. The hypolipidaemic efficacy of DSC apheresis appears, otherwise, not to be dependent upon the procedure per se, but on other individual factors, e.g. the amount of removable lipoproteins and the rate of lipid recovery; both can be predicted with sufficient accuracy.     

Formation of non-cyclooxygenase-derived prostanoids (F2-isoprostanes) in plasma and low density lipoprotein exposed to oxidative stress in vitro.

F2-isoprostanes are prostaglandin F2-like compounds that are known to be formed in vivo by free radical oxidation of arachidonyl-containing lipids, and their plasma levels have been suggested as indicators of in vivo oxidative stress. As oxidation of LDL, a likely causal factor in atherosclerosis, involves lipid peroxidation, we investigated whether F2-isoprostanes are formed in plasma and LDL exposed to oxidative stress, and how F2-isoprostane formation is related to endogenous antioxidant status. In plasma exposed to aqueous peroxyl radicals, lipid hydroperoxides and esterified F2-isoprostanes were formed simultaneously after endogenous ascorbate and ubiquinol-10 had been exhausted, despite the continued presence of urate, alpha-tocopherol, beta-carotene, and lycopene. In isolated LDL exposed to aqueous peroxyl radicals or Cu2+, consumption of endogenous ubiquinol-10 and alpha-tocopherol was followed by rapid formation and subsequent breakdown of lipid hydroperoxides and esterified F2-isoprostanes, and a continuous increase in LDL's electronegativity, indicative of atherogenic modification. In Cu(2+)-exposed LDL, the decrease in esterified F2-isoprostane levels was paralleled by the appearance of free F2-isoprostanes, suggesting that hydrolysis by an LDL-associated activity had occurred. Our data suggest that F2-isoprostanes are useful markers of LDL oxidation in vivo. As F2-isoprostanes are potent vasoconstrictors and can modulate platelet aggregation, their formation in LDL demonstrated here may also have important implications for the etiology of cardiovascular disease.     

Evolocumab and lipoprotein apheresis combination therapy may have synergic effects to reduce low‐density lipoprotein cholesterol levels in heterozygous familial hypercholesterolemia: A case report

A 49 years old woman (weight 68 kg, BMI 27.3 kg/m²) with heterozygous familial hypercholesterolemia (HeFH) and multiple statin intolerance with muscle aches and creatine kinase elevation, presented at the Outpatient Lipid Clinic of Verona University Hospital in May 2015. Hypercholesterolemia was firstly diagnosed during adolescence, followed in adulthood by a diagnosis of Cogan's syndrome, a rheumatologic disorder characterized by corneal and inner ear inflammation. No xanthomas, corneal arcus, or vascular bruits were detectable at physical examination. Screening for macrovascular complications did not reveal relevant damages. Ongoing medical therapy included salicylic acid, methylprednisolone, methotrexate, and protonic‐pump inhibitor. In the absence of specific lipid‐lowering therapy, plasma lipid levels at first visit were: total‐cholesterol = 522 mg/dL, LDL‐cholesterol = 434 mg/dL, HDL‐cholesterol = 84 mg/dL, triglycerides = 120 mg/dL, Lp(a) = 13 mg/dL. On December 2015, evolocumab 140 mg sc every 2 weeks was initiated. After a 24‐week treatment, the LDL‐cholesterol levels decreased by an average of 21.2% to 342 ± 22 mg/dL (mean ± SD). On May 2016, LDL‐apheresis (H.E.L.P.system) was started as add‐on therapy. Compared to the average levels obtained during the evolocumab monotherapy period, the LDL‐cholesterol was reduced by 49.4%, thus reaching an inter‐apheresis level (mean ± SD) of 173 ± 37 mg/dL. This report suggests that a combination therapy with evolocumab and lipoprotein‐apheresis may have synergic effects on circulating lipid levels. Its relevance as a highly effective treatment option for hyperlipidemia in HeFH patients warrants further investigation in larger datasets.     

Determination of lipid hydroperoxides in serum iodometry and high performance liquid chromatography compared.

It is postulated that lipid peroxidation plays a role in the pathogenesis of a variety of diseases. Efforts have therefore been made to develop reliable and practicable procedures for quantifying lipid peroxidation products such as lipid hydroperoxides in biological specimens. An iodometric cholesterol colour reagent (Merck, Darmstadt, Germany) can be used to measure lipid hydroperoxides in isolated low density lipoproteins without lipid extraction. This method has been validated with respect to its analytical performance and suitability for serum samples by comparing it with a high performance liquid chromatography technique. The method was found to have acceptable performance characteristics with aqueous fatty acid hydroperoxide solutions (linoleic acid) and isolated low density lipoproteins, but it cannot be applied to native serum samples without extraction of lipids.     

Lack of plasma lipid peroxidation during LDL-apheresis by heparin-induced extracorporeal LDL-precipitation

Abstract. The heparin-induced extracorporeal precipitation of low density lipoproteins (HELP) is a well established clinical apheresis procedure to markedly reduce cholesterol levels. The biocompatibility of this filter system was investigated by the determination of lipid peroxidation products. Both lipid hydro-peroxides and thiobarbituric acid reactive substances (TBARS) were determined before, during and after 38 aphereses in 21 patients undergoing regular HELP treatment. Although HELP patients had significantly elevated TBARS compared to 93 healthy controls (3.13 ±0.64 vs. l.66±0.50 μmol L^-1; P <0.01), no significant differences were observed compared to either 104 patients suffering from angiographically confirmed coronary heart disease (3.42 ±0.81 μmol L^-1; P > 0.05 vs. HELP patients) or 38 aged-matched hyperlipidaemic patients (3.30 ±0.75 μmol L^-1; P >0.05 vs. HELP patients), neither of which were included in the HELP programme. No lipid hydroperoxides were detected in the plasma of HELP patients either before or after the extracorporeal treatment. After the LDL-apheresis TBARS were significantly decreased (2.60 ± 0.52 μmol L^-1) compared to the values before the treatment ( P <0.01). There was no evidence for the formation of lipid hydroperoxides within the HELP circuit. It is suggested, therefore, that plasma lipids are not oxidized by the HELP procedure.     

Effects of LDL-immunoapheresis on plasma concentrations of vitamin E and carotenoids in patients with familial hypercholesterolemia

Recently very potent extracorporeal cholesterol-lowering treatment options have become available for patients with hypercholesterolemia. LDL immunoapheresis treatment selectively removes LDL and lipoprotein(a) from the circulation. Since LDL is the major carrier of lipophilic antioxidants in plasma, the purpose of the present study was to assess the effects of a single LDL apheresis treatment on plasma concentrations of tocopherols (alpha- and gamma-tocopherol) and carotenoids (alpha- and beta-carotene, zeaxanthin, cryptoxanthin, canthaxanthin, lycopene, and retinol). Plasma antioxidant concentrations were determined by HPLC in 7 patients with familial hypercholesterolemia before and after LDL immunoapheresis treatment. Plasma concentrations of both alpha- and gamma-tocopherol and the different carotenoids were significantly reduced by LDL apheresis. However, when standardized for cholesterol to adjust for cholesterol removal, alpha- and gamma-tocopherol, retinol, and the more polar carotenoids lutein and zeaxanthin increased in response to apheresis treatment, while the more unpolar carotenoids such as beta-carotene and lycopene did not change. These data demonstrate that a single LDL immunoapheresis treatment affects tocopherols and individual carotenoids differently. This may be explained by differences in chemical structure and preferential association with different lipoproteins. These results further imply that tocopherols, lutein, zeaxanthin, and retinol, are associated in part with lipoproteins and other carriers such as retinol-binding protein that are not removed during apheresis treatment.     

The evaluation of altered redox status in plasma and mitochondria of acute and chronic diabetic rats

OBJECTIVES: An increase in plasma oxidative stress and decreased mitochondrial lipid hydroperoxides may contribute to the imbalance in the redox status between intramitochondrial and extramitochondrial milieu in chronic experimental diabetic rats. DESIGN AND METHODS: To determine the effect of hyperglycemia in promoting redox imbalance, we determined lipid hydroperoxides (LHP), protein carbonyl (PCO), total antioxidant activity (ferric reducing/antioxidant power; FRAP) and albumin as markers of redox status of plasma, and mitochondrial lipid hydroperoxide levels as a marker of lipid peroxidation in liver, pancreas and kidney tissue of acute and chronic diabetic male Sprague-Dawley rats and their controls. The levels of the studied markers were determined by colorimetric methods. RESULTS: Plasma and mitochondrial oxidative stress parameter levels of acute diabetic rats were not significantly different from their controls. Plasma LHP and PCO levels of chronic diabetic rats were increased significantly as compared to those of both acute diabetic rats and the controls. Plasma FRAP levels of chronic diabetic animals were decreased significantly as compared to those of the controls. On the other hand, LHP levels in liver, pancreas and kidney mitochondria of chronic diabetic rats were decreased significantly as compared to those of both acute diabetic rats and the controls. We observed a negative correlation between LHP levels in liver mitochondria of chronic diabetic rats, and PCO and fructosamine levels in plasma of chronic diabetic rats were correlated. LHP levels in the pancreatic mitochondria of chronic diabetic rats and plasma oxidative stress parameters of chronic diabetic rats were not significantly correlated. LHP levels in kidney mitochondria of chronic diabetic rats were significantly correlated with serum albumin. There was no correlation between LHP levels in kidney mitochondria and other plasma oxidative stress parameters in chronic diabetic rats. CONCLUSIONS: Our data suggest that redox imbalance between plasma and liver mitochondria might become a major threat to chronic diabetic rats.     

Oxidative stress and abnormal cholesterol metabolism in patients with adult respiratory distress syndrome

Oxidative stress has been implicated in the adult respiratory distress syndrome (ARDS). In this study, we determined the levels of selected antioxidants in the plasma of 25 patients with ongoing ARDS and 16 healthy control subjects. We also examined these plasmas and pulmonary edema fluid of ARDS patients for lipid hydroperoxides. Both ascorbate and ubiquinol-10 concentrations in ARDS plasma were significantly lower than in normal plasma. alpha-Tocopherol concentrations, when standardized to total plasma cholesterol, were not lower in ARDS patients than in normal subjects. A pattern of antioxidant levels virtually identical to that observed in ARDS plasma was obtained after in vitro incubation of healthy plasma with stimulated polymorphonuclear leukocytes: very low ascorbate, decreased ubiquinol-10, and unchanged alpha-tocopherol concentrations. Nanomolar concentrations of lipid hydroperoxides were found in pulmonary edema fluid of ARDS patients, but not in plasma, nor in the plasma of healthy individuals, when a sensitive and selective chemiluminescence assay for hydroperoxides was used. ARDS patients also showed significant decreases in plasma levels of cholesterol esters in conjunction with discoidal high-density lipoprotein profiles, indicating a decrease in lecithin-cholesterol acyltransferase activity. We conclude that ARDS is associated with oxidative stress, possibly exerted by oxidants released from activated phagocytic leukocytes, and major changes in plasma cholesterol metabolism.     

High plasma levels of CD40 are associated with low coenzyme Q and vitamin E content of low-density lipoprotein in healthy men

OBJECTIVE: There is a growing body of evidence to suggest that low-density lipoprotein (LDL) cholesterol, inflammation and oxidative stress are pivotal in the development of cardiovascular disease, but their interconnections are not well known. The objective of this study was to determine whether immunological activation, reflected by the plasma levels of soluble CD40 (sCD40), interleukin (IL)-1beta, tumor necrosis factor-alpha and IL-6 are associated with the antioxidant potential of LDL particles or with common lipid, immunological or thrombotic markers in 51 young healthy men. MATERIAL AND METHODS: We determined the coenzyme Q level from an oxidized LDL fraction, obtaining the concentration for ubiquinone, which indicates total coenzyme Q levels. RESULTS: The plasma level of sCD40 was negatively correlated with LDL ubiquinone (r=-0.45, p=0.001) and E vitamin (r=-0.37, p=0.008) and positively correlated with plasma concentration of plasminogen activator inhibitor-1 (PAI-1, r=0.52, p=0.002) and caspase-1 (r=0.40, p=0.004). No correlation was detected between sCD40 and plasma lipid or C-reactive protein concentrations. As sCD40 was strongly correlated with the content of LDL ubiquinone and vitamin E, their values were compared according to groups formed by sCD40 tertiles. Analysis of variance showed that there were significant differences in LDL ubiquinone (p<0.0001) and vitamin E (p=0.004) concentrations between sCD40 tertiles. CONCLUSIONS: The data indicate that increased activation of the CD40 system is related to low levels of LDL ubiquinone and vitamin E. This suggests that chronic or increased immunological activation may consume the antioxidant potential of LDL particles.