B-100 Particles Research Articles

Effect of transport distance on the size distribution, graphitized structure, surface functional groups and oxidation activity of PM from diesel engine: A comparison of waste cooking oil biodiesel and petroleum diesel

In order to provide a theoretical basis for the development of aftertreatment devices, this paper investigated the exhaust particles of petroleum diesel (B0) and waste cooking oil biodiesel (B100) using EEPS, XPS and TGA. The results showed that the particle number distributions of different particle sizes showed a bimodal distribution. With the increase of transport distance, the peak value of the nuclear mode particles decreases and of accumulation mode particles increases. With the increase of transport distance, the O/C ratio of the particles decreased, but the ratios of sp3/sp2, C–OH and CO content increased, i.e., the farther the transport distance was, the higher the degree of disordered graphitization structure was, and the surface oxygen-containing functional groups increase. The oxidation characteristic temperature decreased with the increase of transport distance, and the activation energy of exhaust particles decreased by more than 10%, i.e., the farther the transport distance, the higher the oxidation activity of the particles. Compared with B0, B100 particles have less PN, more disordered graphitized structure, more C–OH and CO content, and higher oxidation activity. The transport distance had similar effects on the size distribution, graphitized structure, surface functional groups and oxidation activity of B0 and B100 particles.

Lipoprotein (a) and Low-density lipoprotein apolipoprotein B metabolism following apheresis in patients with elevated lipoprotein(a) and coronary artery disease.

Lipoprotein apheresis effectively lowers lipoprotein(a) [Lp(a)] and low-density lipoprotein (LDL) by approximately 60%-70%. The rebound of LDL and Lp(a) particle concentrations following lipoprotein apheresis allows the determination of fractional catabolic rate (FCR) and hence production rate (PR) during non-steady state conditions. We aimed to investigate the kinetics of Lp(a) and LDL apolipoprotein B-100 (apoB) particles in patients with elevated Lp(a) and coronary artery disease undergoing regular apheresis. A cross-sectional study was carried out in 13 patients with elevated Lp(a) concentration (>500mg/L) and coronary artery disease. Lp(a) and LDL-apoB metabolic parameters, including FCR and PR were derived by the fit of a compartment model to the Lp(a) and LDL-apoB concentration data following lipoprotein apheresis. The FCR of Lp(a) was significantly lower than that of LDL-apoB (0.39 [0.31, 0.49] vs 0.57 [0.46, 0.71] pools/day, P=0.03) with no significant differences in the corresponding PR (14.80 [11.34, 19.32] vs 15.73 [11.93, 20.75] mg/kg/day, P=0.80). No significant associations were observed between the FCR and PR of Lp(a) and LDL-apoB. In patients with elevated Lp(a), the fractional catabolism of Lp(a) is slower than that of LDL-apoB particles, implying that different metabolic pathways are involved in the catabolism of these lipoproteins. These findings have implications for new therapies for lowering apolipoprotein(a) and apoB to prevent atherosclerotic cardiovascular disease.

Abstract 11339: Relationship of LPA Single Nucleotide Polymorphisms With Apolipoprotein (a) Isoforms and Biomarkers of Oxidized Lipoproteins in African-Americans, Caucasians and Hispanics: Results From the DALLAS Heart Study

Background: Oxidized phospholipids (OxPL) measured on apolipoprotein B-100 particles (OxPL-apoB) are a measure of the pro-inflammatory activity of Lp(a). Lp(a) in plasma is genetically determined by variability in kringle IV (KIV) repeats and single nucleotide polymorphisms (SNPs) in the LPA gene, is causally related to cardiovascular disease (CVD) and has become a target of therapy. Methods and Results: OxPL-apoB IgG and IgM autoantibodies to oxidized LDL and apoB-immune complexes, KIV repeats of the major apolipoprotein (a) isoform, LPA snps rs3798220, rs10455872 and rs9457951 were measured in 3454 subjects (1798 African-Americans, 1032 Caucasian and 576 Hispanic) subjects enrolled in the Dallas Heart Study (Table). All 3 snps were highly correlated with the number of KIV repeats in African-Americans but only rs3798220 and rs10455872 were correlated with KIV repeats in Caucasians or Hispanics. rs3798220 and rs10455872 demonstrated strong correlations with Lp(a) and OxPL-apoB levels in subjects with low numbers of KIV repeats, whereas rs9457951 showed similar associations only in African-Americans. None of the LPA snps correlated with the other biomarkers of oxidation. Conclusion: Significant racial differences exist among LPA SNPs and their relationship to Lp(a), apolipoprotein (a) isoforms and OxPL-apoB. These findings may provide insights into the clinical variability of Lp(a)-mediated risk in different racial groups and suggests novel avenues of investigation and therapeutic potential in mitigating the CVD risk of Lp(a) and OxPL-apoB.

The role of oxidized phospholipids, lipoprotein (a) and biomarkers of oxidized lipoproteins in chronically occluded coronary arteries in sudden cardiac death and following successful percutaneous revascularization

OxPL are pro-inflammatory and may mediate atherogenesis, thrombosis and endothelial dysfunction. We studied the histological presence and temporal increases in oxidized phospholipids on apolipoprotein B-100 particles (OxPL/apoB), lipoprotein (a) [Lp(a)] and biomarkers of oxidized lipoproteins in subjects with chronic total coronary occlusions (CTO) with sudden cardiac death (SCD) and following percutaneous coronary intervention (PCI). Eight subjects with SCD and CTO and 33 patients with successful PCI of CTO were included. Blood samples were drawn before PCI, immediately post-PCI, at 6 and 24 h, at 3 days and at 1 week. Plasma levels of OxPL/apoB, Lp(a), IgG and IgM autoantibodies to malondialdehyde (MDA) low-density lipoprotein and apoB-immune complexes were measured in all samples and compared with previous data from 141 patients undergoing PCI of non-CTO vessels. Immunohistochemistry of coronary CTOs revealed OxPL and MDA-like epitopes, particularly in areas of recanalized and organized thrombus and neovascularization. Following PCI, OxPL/apoB and Lp(a) levels, expressed as percent change from baseline levels before PCI, rose gradually and progressively over the next 7 days. In contrast, levels of OxPL/apoB and Lp(a) in non-CTO vessels rose immediately post PCI and then dropped rapidly to baseline within 24 h. CTOs contain immunohistological evidence of OxPL and MDA-like epitopes. Successful PCI of CTOs results in a slower increase in OxPL/apoB and Lp(a) but higher increase in IgM immune complexes compared to non-CTO vessels. Pro-inflammatory oxidation-specific epitopes may impact development of CTOs and affect outcomes following PCI that can be evaluated in larger clinical trials.

Oxidative Modification of Lipoproteins: Mechanisms, Role in Inflammation and Potential Clinical Applications in Cardiovascular Disease

Abnormalities of lipid metabolism often lead to pathologic lipid accumulation in the vessel wall, oxidative and chronic inflammatory sequelae and the formation of atherosclerotic lesions, ultimately leading to clinical events. Oxidation of lipoproteins, and in particular low density lipoprotein (LDL), is a seminal even that mediates many pro-atherogenic and pro-inflammatory pathways. Many in vivo mechanisms exist to oxidize LDL, including transition metals such as divalent iron cations, heme, as well as a number of different enzyme systems, such as lipoxygenases, myeloperoxidase, NADPH oxidases, and nitric oxide synthases. Oxidized LDL is taken up in an unregulated fashion. By macrophages leading to foam cell formation, ultimately generating a potent pro-inflammatory milieu. Minimally modified LDL also induces proinflammatory effects in macrophages, including cytoskeletal rearrangements and macropinocytosis, generation of reactive oxygen species, survival of foam cells, reduced phagocytic capacity toward apoptotic cells, and expression of inflammatory genes, many of these effects mediated through toll-like receptor-4. Using the scientific knowledge gained from understanding these pathways, antibodies binding well-defined oxidation-specific epitopes have been generated and are being used in translational clinical applications. In particular, assays measuring oxidized phospholipids on apolipoprotein B-100 particles (OxPL/apoB) predict the presence and progression of femoral, carotid and coronary artery disease and predict new cardiovascular events independent of established risk factors. Human oxidation-specific antibodies have also been successfully used to image the extent and regression of experimental atherosclerotic lesions using nuclear and magnetic resonance imaging approaches. If validated and translated to humans, this imaging approach may provide a means to non-invasively detect, quantitate and monitor extent of atherosclerosis and potentially image high risk plaques. Further understanding of the role of oxidation of lipoproteins may allow more rational targeted diagnostic and therapeutic modalities in clinical applications.

OxPL/apoB, lipoprotein(a) and OxLDL biomarkers and cardiovascular disease in chronic hemodialysis patients

The aim of this study was to assess the correlation between oxidized phospholipids on apolipoprotein B-100 particles (OxPL/ApoB), lipoprotein(a) and low-density lipoprotein (LDL)-producing oxidized phospholipid (OxLDL) biomarkers and cardiovascular disease (CVD) in chronic hemodialysis (HD) patients. In 52 HD patients, we measured OxPL/apoB by antibody E06, IgG/IgM autoantibody titers to copper-oxidized LDL (Cu-OxLDL) and malondialdehyde (MDA)-LDL, IgG/IgM apolipoprotein B-100-immune complexes (IC/apoB), C-reactive protein (CRP) and lipoprotein(a) (Lp(a)). In all patients, the presence of CVD was assessed. OxPL/ApoB, IC IgG/IgM, MDA-LDL Ig G/IgM, Cu-OxLDL IgG/IgM and Lp(a) levels were similar in patients with and without CVD. At multiple logistic regression analysis, the association between age and CVD was independent (OR=1.078; 95% confidence interval, 1.005-1.157; p=0.03). In HD patients, OxPL/apoB, lipoprotein(a) and OxLDL biomarkers are not associated with CVD.

Changes in lipoprotein(a), oxidized phospholipids, and LDL subclasses with a low-fat high-carbohydrate diet

Low-fat diets have been shown to increase plasma concentrations of lipoprotein(a) [Lp(a)], a preferential lipoprotein carrier of oxidized phospholipids (OxPLs) in plasma, as well as small dense LDL particles. We sought to determine whether increases in plasma Lp(a) induced by a low-fat high-carbohydrate (LFHC) diet are related to changes in OxPL and LDL subclasses. We studied 63 healthy subjects after 4 weeks of consuming, in random order, a high-fat low-carbohydrate (HFLC) diet and a LFHC diet. Plasma concentrations of Lp(a) (P < 0.01), OxPL/apolipoprotein (apo)B (P < 0.005), and OxPL-apo(a) (P < 0.05) were significantly higher on the LFHC diet compared with the HFLC diet whereas LDL peak particle size was significantly smaller (P < 0.0001). Diet-induced changes in Lp(a) were strongly correlated with changes in OxPL/apoB (P < 0.0001). The increases in plasma Lp(a) levels after the LFHC diet were also correlated with decreases in medium LDL particles (P < 0.01) and increases in very small LDL particles (P < 0.05). These results demonstrate that induction of increased levels of Lp(a) by an LFHC diet is associated with increases in OxPLs and with changes in LDL subclass distribution that may reflect altered metabolism of Lp(a) particles.

Oxidation-Specific Biomarkers, Lipoprotein(a), and Risk of Fatal and Nonfatal Coronary Events

This study sought to assess whether oxidation-specific biomarkers are associated with an increased risk of coronary artery disease (CAD) events. The relationship of a panel of oxidative biomarkers and lipoprotein(a) [Lp(a)] to CAD risk is not fully determined. A prospective case-control study nested in the EPIC (European Prospective Investigation of Cancer)-Norfolk cohort of 45- to 79-year-old apparently healthy men and women followed for approximately 6 years was designed. Cases consisted of participants in whom fatal or nonfatal CAD developed, matched by sex, age, and enrollment time with controls without CAD. Baseline levels of oxidized phospholipids on apolipoprotein B-100 particles and Lp(a) were measured in 763 cases and 1,397 controls. Their relationship to secretory phospholipase A(2) type IIA mass and activity, myeloperoxidase mass, and lipoprotein-associated phospholipase A(2) activity and association with CAD events were determined. After adjusting for age, smoking, diabetes, low- and high-density lipoprotein cholesterol, and systolic blood pressure, the highest tertiles of oxidized phospholipids on apolipoprotein B-100 particles and Lp(a) were associated with a significantly higher risk of CAD events (odds ratios: 1.67 and 1.64, respectively; p < 0.001) compared with the lowest tertiles. The odds ratio of CAD events associated with the highest tertiles of oxidized phospholipids on apolipoprotein B-100 particles or Lp(a) was significantly potentiated (approximately doubled) by the highest tertiles of secretory phospholipase A(2) activity and mass but less so for myeloperoxidase and lipoprotein-associated phospholipase A(2) activity. The odds ratios for fatal CAD were higher than for the combined end point. After taking into account the Framingham Risk Score, c-index values progressively increased when oxidative biomarkers were added to the model. This EPIC-Norfolk study links pathophysiologically related oxidation-specific biomarkers and Lp(a) with CAD events. Oxidation-specific biomarkers provide cumulative predictive value when added to traditional cardiovascular risk factors.

2D-NMR reveals different populations of exposed lysine residues in the apoB-100 protein of electronegative and electropositive fractions of LDL particles

Several potentially atherogenic LDL subfractions present low affinity for the LDL receptor, which result in impaired plasma clearance. Electronegative LDL [LDL(-)] is one of these minor subfractions and the molecular basis for its reduced receptor affinity is not well understood. In the present study, high-resolution 2D-NMR spectroscopy has been employed to characterize the surface-exposed lysine residues of the apolipoprotein (apo)B-100 protein in both LDL(-) and LDL(+) subfractions. LDL(+) showed two populations of lysine residues, similar to those previously described in total LDL. "Normal" Lys have a pk(a) of 10.4 whereas "active" Lys have a pk(a) of 8.8 and have been suggested to be involved in receptor binding. In contrast to LDL(+), the LDL(-) subfraction presented a third type of Lys, named as "intermediate" Lys, with a different microenvironment and higher basicity (pk(a) 10.7). These intermediate Lys cannot be reliably identified by 1D-NMR. Because the abundance of normal Lys is similar in LDL(+) and LDL(-), the intermediate Lys in the apoB-100 molecule of LDL(-) should come from a group of active Lys in LDL(+) particles that have a less basic microenvironment in the LDL(-) particle. These differences between LDL(+) and LDL(-) are indicative of a distinct conformation of apoB-100 that could be related to loss of affinity of LDL(-) for the LDL receptor.

Pitavastatin Reduces Lectin‐Like Oxidized Low‐Density Lipoprotein Receptor‐1 Ligands in Hypercholesterolemic Humans

The aim of this study was to determine the impact of pitavastatin on low-density lipoprotein cholesterol (LDL-C) and lectin-like oxidized LDL receptor-1 (LOX-1) in patients with hypercholesterolemia. Twenty-five hypercholesterolemic patients (8 male, 17 female; age 66 +/- 13, 21-80 years) who had not received anti-dyslipidemic agents and had LDL-C levels of more than 160 mg/dL were examined. Biochemical factors were measured at baseline and after treatment with pitavastatin (2 mg/day) for 6 months. Serum levels of LOX-1 with apolipoprotein B-100 particle ligand and a soluble form of LOX-1 (sLOX-1) were measured by ELISA. All subjects completed the study with no adverse side effects. Total-C (268 +/- 26 vs. 176 +/- 17 mg/dL), LDL-C (182 +/- 21 vs. 96 +/- 14 mg/dL), and LOX-1 ligand (867 +/- 452 vs. 435 +/- 262 ng/mL) were reduced with pitavastatin treatment (P < 0.0001 for each). Significant decreases in triacylglycerols were noted (P < 0.0001), but there were no changes in high-density lipoprotein cholesterol. After 6 months, there were no significant changes in high-sensitivity CRP or soluble LOX-1. At baseline, there were no significant correlations between LOX-1 ligand and either LDL-C or sLOX-1. The decrease in LOX-1 ligand was not correlated with the decrease in LDL-C, but was correlated with the decrease in sLOX-1 (r = 0.47, P < 0.05). In conclusion, pitavastatin therapy had beneficial effects on markers of oxidative stress in hypercholesterolemic subjects. Serum levels of LOX-1 ligand may be a useful biomarker of the pleiotropic effects of statins.

Relation of Oxidative Biomarkers, Vascular Dysfunction, and Progression of Coronary Artery Calcium

The relation between oxidative stress and coronary artery calcium (CAC) progression is currently not well described. The present study evaluated the relation among the biomarkers of oxidative stress, vascular dysfunction, and CAC. Sixty asymptomatic subjects participated in a randomized trial evaluating the effect of aged garlic extract plus supplement versus placebo and underwent measurement of CAC. The postcuff deflation temperature-rebound index of vascular function was assessed using a reactive hyperemia procedure. The content of oxidized phospholipids (OxPL) on apolipoprotein B-100 (apoB) particles detected by antibody E06 (OxPL/apoB), lipoprotein(a), IgG and IgM autoantibodies to malondialdehyde-low-density lipoprotein and apoB-immune complexes were measured at baseline and after 12 months of treatment. CAC progression was defined as an annual increase in CAC >15%. Vascular dysfunction was defined according to the tertiles of temperature-rebound at 1 year of follow-up. From baseline to 12 months, a strong inverse correlation was noted between an increase in CAC scores and increases in temperature-rebound (r(2) = -0.90), OxPL/apoB (r(2) = -0.85), and lipoprotein(a) (r(2) = -0.81) levels (p <0.0001 for all). The improvement in temperature-rebound correlated positively with the increases in OxPL/apoB (r(2) = 0.81, p = 0.0008) and lipoprotein(a) (r(2) = 0.79, p = 0.0001) but inversely with autoantibodies to malondialdehyde-low-density lipoprotein and apoB-immune complexes. The greatest CAC progression was noted with the lowest tertiles of increases in temperature-rebound, OxPL/apoB and lipoprotein(a) and the highest tertiles of increases in IgG and IgM malondialdehyde-low-density lipoprotein. In conclusion, the present results have documented a strong relation among markers of oxidative stress, vascular dysfunction, and progression of coronary atherosclerosis. Increases in OxPL/apoB and lipoprotein(a) correlated strongly with increases in vascular function and predicted a lack of progression of CAC.

The I4399M variant of apolipoprotein(a) is associated with increased oxidized phospholipids on apolipoprotein B-100 particles

Objectives The LPA I4399M (rs3798220) single nucleotide polymorphism (SNP) is associated with increased plasma levels of Lp(a) and advanced coronary artery disease (CAD). We hypothesized that carriers of the Met allele of the I4399M SNP would also have elevated levels of oxidized phospholipids (OxPL) on apoB (OxPL/apoB) particles. Methods and results Plasma levels of Lp(a) and OxPL/apoB were measured in non-carriers (TT genotype, n = 116) and carriers (CT/CC genotype, n = 58) of the I4399M SNP. Carriers had significantly higher Lp(a) levels [median (interquartile range, IQR)] [43 (9–57) mg/dl vs. 5.5 (2–20) mg/dl, p < 0.0001] and smaller apolipoprotein(a) isoforms than non-carriers (number of kringle IV repeats: 18(17–20) vs. 22(18–25), p = 0.002). Median (IQR) OxPL/apoB levels were significantly higher in carriers than non-carriers [8019 (6254–31,785) relative light units (RLU) vs. 2168 (1303–5869), p < 0.0001]. Patients with small apolipoprotein(a) isoforms had the highest OxPL/apoB levels. The number of kringle IV repeats was inversely related to Lp(a) ( r = −0.43, p < 0.001) and OxPL/apoB ( r = −0.36, p < 0.0001) levels. Conclusion The CT and CC genotypes of the I4399M SNP in the LPA gene are associated with elevated OxPL/apoB levels, which primarily represent OxPL on Lp(a). The concomitant increase of OxPL/apoB levels in the setting of small apolipoprotein(a) isoforms may potentiate the atherogenic effect on CAD of elevated Lp(a) levels in carriers of the I4399M SNP.

Genome-wide association study of plasma lipoprotein(a) levels identifies multiple genes on chromosome 6q

Plasma lipoprotein(a) (Lp[a]) level is an independent risk factor of cardiovascular disease that is under strong genetic control. We conducted a genome-wide association study of plasma Lp(a) in 386 members of a founder population that adheres to a communal lifestyle, proscribes cigarette smoking, and prepares and eats meals communally. We identified associations with 77 single nucleotide polymorphisms (SNPs) spanning 12.5 Mb on chromosome 6q26-q27 that met criteria for genome-wide significance (P <or= 1.3 x 10(-7)) and were within or flanking nine genes, including LPA. We show that variation in at least six genes in addition to LPA are significantly associated with Lp(a) levels independent of each other and of the kringle IV repeat polymorphism in the LPA gene. One novel SNP in intron 37 of the LPA gene was also associated with Lp(a) levels and carotid artery disease number in unrelated Caucasians (P = 7.3 x 10(-12) and 0.024, respectively), also independent of kringle IV number. This study suggests a complex genetic architecture of Lp(a) levels that may involve multiple loci on chromosome 6q26-q27.

Relationship of oxidized phospholipids on apolipoprotein B-100 particles to race/ethnicity, apolipoprotein(a) isoform size, and cardiovascular risk factors: results from the Dallas Heart Study.

Elevated levels of oxidized phospholipids (OxPLs) on apolipoprotein B-100 particles (OxPL/apoB) are associated with cardiovascular disease and predict new cardiovascular events. Elevated lipoprotein (a) [Lp(a)] levels are a risk factor for cardiovascular disease in whites and also in blacks if they carry small apolipoprotein(a) [apo(a)] isoforms. The relationship of OxPL/apoB levels to race/ethnicity, cardiovascular risk factors, and apo(a) isoforms is not established. OxPL/apoB levels were measured in 3481 subjects (1831 black, 1047 white, and 603 Hispanic subjects) in the Dallas Heart Study and correlated with age, sex, cardiovascular risk factors, and Lp(a) and apo(a) isoforms. Significant differences in OxPL/apoB levels were noted among racial/ethnic subgroups, with blacks having the highest levels compared with whites and Hispanics (P<0.001 for each comparison). OxPL/apoB levels generally did not correlate with age, sex, or risk factors. In the overall cohort, OxPL/apoB levels strongly correlated with Lp(a) (r=0.85, P<0.001), with the shape of the relationship demonstrating a "reverse L" shape for log-transformed values. The highest correlation was present in blacks, followed by whites and Hispanics; was dependent on apo(a) isoform size; and became progressively weaker with larger isoforms. The size of the major apo(a) isoform (number of kringle type IV repeats) was negatively associated with OxPL/apoB (r=-0.49, P<0.001) and Lp(a) (r=-0.61, P<0.001) regardless of racial/ethnic group. After adjustment for apo(a) isoform size, the relationship between OxPL/apoB and Lp(a) remained significant (r=0.67, P<0.001). OxPL/apoB levels vary according to race/ethnicity, are largely independent of cardiovascular risk factors, and are inversely associated with apo(a) isoform size. The association of OxPL with small apo(a) isoforms, in which a similar relationship is present among all racial/ethnic subgroups despite differences in Lp(a) levels, may be a key determinant of cardiovascular risk.

Abstract 4998: Elevated Oxidized Phospholipids on Apolipoprotein B-100 Particles Predict 6-Year Cardiovascular Events in the Epic-Norfolk Study: Potentiation of Risk with Lipoprotein-Associated (Lp-PLA 2 ) and Soluble Phospholipase A2 (sPLA 2 ) Activity

Objective: We aimed to assess whether oxidized phospholipids on apoB-100 particles (OxPL/apoB) predict future coronary heart disease (CHD) in the prospective EPIC-Norfolk cohort of apparently healthy men and women aged 45–79 years followed up 1993–2007. Methods and Results: Cases who developed CHD during follow-up were matched to controls who remained free of cardiovascular disease and who matched to cases by sex, age and enrollment time. OxPL/apoB levels were measured with antibody E06 in 905 cases and 1809 controls. Subjects in the highest quartile of OxPL/apoB at entry in the study had significantly higher risk of future CHD events (OR 1.72, 95%CI 1.32–2.24, p&lt;0.0001) compared to those in the lowest quartile. This relationship was present in both men and women and was also observed after adjustment for sex, age, diabetes, smoking, blood pressure, LDL-C, HDL-C and Framingham Risk Score (FRS). The risk of future CHD events was significantly potentiated by elevated activity levels of sPLA 2 and Lp-PLA 2 . Subjects in the highest tertiles of both OxPL/apoB and LpPLA2 (OR 2.22 [1.51–3.27]) or OxPL/apoB and sPLA 2 (OR 4.34 [2.84 – 6.64]) had significantly higher risk of future CHD than subjects in the lowest tertiles (p&lt;0.0001 for both). Area under receiver operating curves revealed significantly increased values by adding OxPL/apoB and sPLA 2 to traditional risk factors and the FRS. Strong inverse associations were noted between the OxPL/apoB and VLDL and IDL size, CETP mass and RANK Ligand. Conclusion: Elevated OxPL/apoB levels are a strong, independent predictor of CHD events. The predictive values is further enhanced by elevated activity levels of sPLA 2 and Lp-PLA 2 , which mediate breakdown of OxPL present on lipoproteins and within atherosclerotic lesions. This study links several pathophysiologically related oxidative biomarkers in the prediction of CHD events and suggests novel approaches to predicting CHD risk in apparently health men and women.

A novel function of lipoprotein [a] as a preferential carrier of oxidized phospholipids in human plasma

Oxidized phospholipids (OxPLs) on apolipoprotein B-100 (apoB-100) particles are strongly associated with lipoprotein [a] (Lp[a]). In this study, we evaluated whether Lp[a] is preferentially the carrier of OxPL in human plasma. The content of OxPL on apoB-100 particles was measured with monoclonal antibody E06, which recognizes the phosphocholine (PC) headgroup of oxidized but not native phospholipids. To assess whether OxPLs were preferentially bound by Lp[a] as opposed to other lipoproteins, immunoprecipitation and ultracentrifugation experiments, in vitro transfer studies, and chemiluminescent ELISAs were performed. Immunoprecipitation of Lp[a] from human plasma with an apolipoprotein [a] (apo[a])-specific antibody demonstrated that more than 85% of E06 reactivity (i.e., OxPL) coimmunoprecipitated with Lp[a]. Ultracentrifugation experiments showed that nearly all OxPLs were found in fractions containing apo[a], as opposed to other apolipoproteins. In vitro transfer studies showed that oxidized LDL preferentially donates OxPLs to Lp[a], as opposed to LDL, in a time- and temperature-dependent manner, even in aqueous buffer. Approximately 50% of E06 immunoreactivity could be extracted from isolated Lp[a] following exposure of plasma to various lipid solvents. These data demonstrate that Lp[a] is the preferential carrier of PC-containing OxPL in human plasma. This unique property of Lp[a] suggests novel insights into its physiological function and mechanisms of atherogenicity.

The role of oxidized phospholipids in mediating lipoprotein(a) atherogenicity

To review emerging data on the relationship between lipoprotein(a) and oxidized phospholipids. We have recently proposed that a unique physiological role of lipoprotein(a) may be to bind and transport proinflammatory oxidized phospholipids and that this interaction may mediate a common biological influence on cardiovascular disease. In a large series of clinical studies performed to date, a very strong correlation was found between plasma levels of lipoprotein(a) and the content of oxidized phospholipids on apolipoprotein B-100 particles (OxPL/apoB), measured by monoclonal antibody E06, which binds the phosphocholine head group of oxidized phospholipids but not native phospholipids. The correlation of OxPL/apoB to lipoprotein(a) is very strong in individuals with small apolipoprotein(a) isoforms (r = approximately 0.95) and modest in individuals with large isoforms (r = approximately 0.60). In-vitro studies have demonstrated that the vast majority of oxidized phospholipids detected by E06 are bound to lipoprotein(a) in human plasma. A similarly strong association with oxidized phospholipids was also documented in transgenic mice overexpressing lipoprotein(a), even in mice not fed atherogenic diets or with overt atherosclerosis. A better understanding of the ability of human lipoprotein(a) to bind oxidized phospholipids may allow clinically important insights into the role of oxidized phospholipids and lipoprotein(a) in human atherogenesis and cardiovascular disease and may provide novel diagnostic tools and therapeutic interventions aimed at measuring and treating elevated levels of OxPL/apoB and lipoprotein(a).

Antisense Oligonucleotide Directed to Human Apolipoprotein B-100 Reduces Lipoprotein(a) Levels and Oxidized Phospholipids on Human Apolipoprotein B-100 Particles in Lipoprotein(a) Transgenic Mice

Lipoprotein (a) [Lp(a)] is a genetic cardiovascular risk factor that preferentially binds oxidized phospholipids (OxPL) in plasma. There is a lack of therapeutic agents that reduce plasma Lp(a) levels. Transgenic mice overexpressing human apolipoprotein B-100 (h-apoB-100 [h-apoB mice]) or h-apoB-100 plus human apo(a) to generate genuine Lp(a) particles [Lp(a) mice] were treated with the antisense oligonucleotide mipomersen directed to h-apoB-100 mRNA or control antisense oligonucleotide for 11 weeks by intraperitoneal injection. Mice were then followed up for an additional 10 weeks off therapy. Lp(a) levels [apo(a) bound to apoB-100] and apo(a) levels ["free" apo(a) plus apo(a) bound to apoB-100] were measured by chemiluminescent enzyme-linked immunoassay and commercial assays, respectively. The content of OxPL on h-apoB-100 particles (OxPL/h-apoB) was measured by capturing h-apoB-100 in microtiter wells and detecting OxPL by antibody E06. As expected, mipomersen significantly reduced plasma h-apoB-100 levels in both groups of mice. In Lp(a) mice, mipomersen significantly reduced Lp(a) levels by approximately 75% compared with baseline (P<0.0001) but had no effect on apo(a) levels or hepatic apo(a) mRNA expression. OxPL/h-apoB levels were much higher at baseline in Lp(a) mice compared with h-ApoB mice (P<0.0001) but decreased in a time-dependent fashion with mipomersen. There was no effect of the control antisense oligonucleotide on lipoprotein levels or oxidative parameters. Mipomersen significantly reduced Lp(a) and OxPL/apoB levels in Lp(a) mice. The present study demonstrates that h-apoB-100 is a limiting factor in Lp(a) particle synthesis in this Lp(a) transgenic model. If applicable to humans, mipomersen may represent a novel therapeutic approach to reducing Lp(a) levels and their associated OxPL.

The Influence of Pravastatin and Atorvastatin on Markers of Oxidative Stress in Hypercholesterolemic Humans

The aim of this study was to determine the effects of pravastatin and atorvastatin on markers of oxidative stress in plasma. Hydroxymethylglutaryl coenzyme A reductase inhibitors reduce low-density lipoprotein cholesterol (LDL-C) and cardiovascular risk, but their effects on circulating biomarkers of oxidative stress are not well-defined. Hypercholesterolemic subjects (n = 120, ages 21 to 80 years with LDL-C 130 to 220 mg/dl) were randomized in a double-blind, parallel design to pravastatin 40 mg/day (prava40), atorvastatin 10 mg/day (atorva10), atorvastatin 80 mg/day (atorva80), or placebo. At baseline and 16 weeks, urinary isoprostanes (8, 12-iso-iPF(2 alpha)-VI isoform), plasma lipoprotein-associated phospholipase A2 (Lp-PLA2), Mercodia oxidized LDL (OxLDL) with antibody 4E6, oxidized phospholipids/apolipoprotein B-100 particle (OxPL/apoB) with antibody E06, immunoglobulin (Ig)G/IgM autoantibodies to malondialdehyde (MDA)-LDL, and apolipoprotein B (apoB)-immune complexes (IC) were measured. After 16 weeks, there were no significant changes in urinary 8, 12-iso-iPF(2 alpha)-VI. The Lp-PLA2 and OxLDL were reduced in statin-treated groups, but after adjusting for apoB, only prava40 led to a reduction in Lp-PLA2 (-15%, p = 0.008) and atorva10 to a decrease in OxLDL (-12.9%, p = 0.01). The OxPL/apoB increased 25.8% (p < 0.01) with prava40 and 20.2% (p < 0.05) with atorva80. There were no changes in MDA-LDL autoantibodies, but significant decreases in IC were noted. This study suggests that statin therapy results in variable effects on oxidative stress markers in hypercholesterolemic subjects. Future outcome studies should collectively assess various oxidative markers to define clinical utility.

Oxidized Low-Density Lipoprotein Biomarkers in Patients with End-Stage Renal Failure: Acute Effects of Hemodialysis

Objective: To assess the effect of end-stage renal failure on oxidized low-density lipoprotein (OxLDL) biomarkers and the acute effects of hemodialysis. Oxidized phospholipids (OxPL) on apolipoprotein B-100 (apoB) particles (OxPL/apoB) have been associated with cardiovascular disease and new cardiovascular events. Patients with end-stage renal failure have increased oxidative stress and are at significantly increased risk of cardiovascular disease. Methods and Results: Fifty-two stable patients with end-stage renal failure undergoing chronic hemodialysis were included in the study. Pre and post hemodialysis blood samples were obtained for measurement of OxLDL biomarkers: oxidized phospholipids (OxPL) on apolipoprotein B-100 (apoB) particles (OxPL/apoB) measured by antibody E06, IgG and IgM autoantibody titers to copper-oxidized LDL (Cu-OxLDL) and malondialdehyde (MDA)-LDL, IgG and IgM apolipoprotein B-100-immune complexes (IC/apoB). Traditional laboratory variables as well as C-reactive protein (CRP) and lipoprotein(a) [Lp(a)] were also measured. For the baseline variables, the distribution of OxPL/apoB and Lp(a) were skewed to lower values, and a strong correlation was noted between OxPL/apoB and Lp(a) (r = 0.94, p < 0.0001). No major associations were noted between OxLDL biomarkers and age, gender or dialytic age. There were also no correlations between OxLDL biomarkers and traditional risk factors, CRP, body mass index, serum creatinine, hypertension or intravenous iron therapy. Following dialysis, there as a significant reduction in OxPL/apoB (–7.5%, p = 0.048) and triglyceride levels (–10.8%, p = 0.005). All other OxLDL biomarkers, CRP, total cholesterol, LDL-C, HDL-C and apoB-100 increased significantly (range 6.3–26.9%, p value range 0.005 to <0.0001). Total protein plasma levels increased 8.8% (p = 0.014 compared to predialysis) following dialysis, consistent with a hemoconcentration effect of hemodialysis. Conclusion: In end-stage renal failure patients undergoing hemodialysis, a reduction in OxPL/apoB levels was noted, despite the hemoconcentrating and strong pro-oxidant milieu of hemodialysis. Studies in larger populations of end-stage renal failure patients are needed to assess whether these findings predict future clinical outcomes.