Major Subfractions Research Articles

An Automatic Fast High Resolution HDL Subfractioning Assay Using Microfluidic Electrophoretic System

Objectives: Studies in recent decade demonstrated the importance of HDL subfractionation in precision medicine. It is notable that not all HDL subfractions are “good” lipoprotein. The lack of a reliable technique that can precisely identify each particle of HDL subfractions remains the major obstacle in clinical application. Based on the principle of microfluidic electrophoresis, we have developed an automatic system (MFE) that meets the need of clinical applications. The objectives of this paper is to test the system for its application, precision, and also the normal range in healthy population using one of major subfraction, HDL-2b, as a target. Methods: The MFE was compared with traditional non-denaturing polyacrylamide gradient gel electrophoresis (GGE). Intra-chip, inter-chip, inter-instrument, inter-lab, and inter-operator precisions were all tested by using the sample in various conditions. An instruction was provided with the instrument and HDL subfractioning kit. Data were analyzed with SPSS for correlation of variation, regressions, etc. Results: The new system exhibited a linear correlation with the traditional GGE system. The intra-chip %CVs, inter-chip %CVs, inter-instrument %CVs of HDL-2b were less than 10%, which was the set acceptance criteria. With 243 samples from healthy individuals demonstrated that the mean value of HDL-2b% is about 25% in both male and female. Conclusions: Our data demonstrated that the MFE system is a fast, automatic, and reliable system that can obtain the percentage of HDL-2b with high precision

Pro‑differentiating compounds for human intervertebral disc cells are present in Violina pumpkin leaf extracts.

Intervertebral disc (IVD) degeneration (IDD) is closely associated with inflammation, oxidative stress and loss of the discogenic phenotype, which current therapies are unable to reverse. In the present study, the effects of acetone extract from Violina pumpkin (Cucurbitamoschata) leaves on degenerated IVD cells were investigated. IVD cells were isolated from the degenerated disc tissue of patients undergoing spinal surgery and were exposed to acetone extract and three major thin layer chromatography subfractions. The results revealed that, in particular, the cells benefited from exposure to subfraction Fr7, which consisted almost entirely of p‑Coumaric acid. Western blot and immunocytochemical analysis showed that Fr7 induced a significant increase in discogenic transcription factors (SOX9 and tricho‑rhino‑phalangeal syndrome typeI protein, zinc finger protein), extracellular matrix components (aggrecan, collagen typeII), cellular homeostasis and stress response regulators, such as FOXO3a, nuclear factor erythroid2‑related factor 2, superoxide dismutase 2 and sirtuin 1. Two important markers related to the presence and activity of stem cells, migratory capacity and OCT4 expression, were assessed by scratch assay and western blotting, respectively, and were significantly increased in Fr7‑treated cells. Moreover, Fr7 counteracted H2O2‑triggered cell damage, preventing increases in the pro‑inflammatory and anti‑chondrogenic microRNA (miR), miR‑221. These findings strengthen the hypothesis that adequate stimuli can support resident cells to repopulate the degenerated IVD and restart the anabolic machinery. Taken together, these data contribute to the discovery of molecules potentially effective in slowing the progression of IDD, a disease for which there is currently no effective treatment. Moreover, the use of part of a plant, the pumpkin leaves, which is usually considered a waste product in the Western world, indicated that it contains substances with potential beneficial effects on human health.

Prioritizing the Role of Major Lipoproteins and Subfractions as Risk Factors for Peripheral Artery Disease.

Lipoprotein-related traits have been consistently identified as risk factors for atherosclerotic cardiovascular disease, largely on the basis of studies of coronary artery disease (CAD). The relative contributions of specific lipoproteins to the risk of peripheral artery disease (PAD) have not been well defined. We leveraged large-scale genetic association data to investigate the effects of circulating lipoprotein-related traits on PAD risk. Genome-wide association study summary statistics for circulating lipoprotein-related traits were used in the mendelian randomization bayesian model averaging framework to prioritize the most likely causal major lipoprotein and subfraction risk factors for PAD and CAD. Mendelian randomization was used to estimate the effect of apolipoprotein B (ApoB) lowering on PAD risk using gene regions proxying lipid-lowering drug targets. Genes relevant to prioritized lipoprotein subfractions were identified with transcriptome-wide association studies. ApoB was identified as the most likely causal lipoprotein-related risk factor for both PAD (marginal inclusion probability, 0.86; P=0.003) and CAD (marginal inclusion probability, 0.92; P=0.005). Genetic proxies for ApoB-lowering medications were associated with reduced risk of both PAD (odds ratio,0.87 per 1-SD decrease in ApoB [95% CI, 0.84-0.91]; P=9×10-10) and CAD (odds ratio,0.66 [95% CI, 0.63-0.69]; P=4×10-73), with a stronger predicted effect of ApoB lowering on CAD (ratio of effects, 3.09 [95% CI, 2.29-4.60]; P<1×10-6). Extra-small very-low-density lipoprotein particle concentration was identified as the most likely subfraction associated with PAD risk (marginal inclusion probability, 0.91; P=2.3×10-4), whereas large low-density lipoprotein particle concentration was the most likely subfraction associated with CAD risk (marginal inclusion probability, 0.95; P=0.011). Genes associated with extra-small very-low-density lipoprotein particle and large low-density lipoprotein particle concentration included canonical ApoB pathway components, although gene-specific effects were variable. Lipoprotein(a) was associated with increased risk of PAD independently of ApoB (odds ratio, 1.04 [95% CI, 1.03-1.04]; P=1.0×10-33). ApoB was prioritized as the major lipoprotein fraction causally responsible for both PAD and CAD risk. However, ApoB-lowering drug targets and ApoB-containing lipoprotein subfractions had diverse associations with atherosclerotic cardiovascular disease, and distinct subfraction-associated genes suggest possible differences in the role of lipoproteins in the pathogenesis of PAD and CAD.

Anti-mycobacterial and immunomodulatory activity of n-hexane fraction and spathulenol from Ocotea notata leaves

Abstract Ocotea notata (Lauraceae) is popularly known as white-cinnamon. Ocotea species have several medicinal uses, especially for treating chest pain, rheumatism and wounds. The present study aimed to analyze the chemical composition of O. notata n-hexane fraction, in addition to its anti-mycobacterial and immunomodulatory activities. The n-hexane fraction was analyzed by GC-MS and was chromatographed to afford 15 subfractions (SF1-15), where SF5 was identified, by GC-MS and NMR, as the sesquiterpene spathulenol. The n-hexane fraction was the most potent in inhibiting nitric oxide (NO) and tumor necrosis factor-alpha (TNF-α) production on LPS-stimulated macrophages (IC50 8.3 ± 0.9 and 5.9 ±1.0 μg/mL, respectively). SF4, a major subfraction, that presents a spathulenol analogous as a constituent, also inhibited NO and TNF-α production. Spathulenol only modulated NO production (IC50 45.6 ± 1.4 μg/mL). The n-hexane fraction, SF4, and spathulenol revealed antimycobacterial activity against Mycobacterium bovis BCG, M. tuberculosis H37Rv, and M299 strains. Spathulenol inhibited the growth of Mtb H37Rv with MIC50 36.9 ± 1.5 μg/mL (167.5 ± 6.8 μM), and Mtb M299 with MIC5042.1 ± 0.5 μg/mL (191.0 ± 2.2 μM). This is the first report describing the isolation of spathulenol from O. notata leaves and its anti-mycobacterial activity.

Effects of TM6SF2 E167K on hepatic lipid and very low-density lipoprotein metabolism in humans.

Nonalcoholic fatty liver disease (NAFLD) is characterized by hepatic lipid accumulation. The transmembrane 6 superfamily member 2 (TM6SF2) E167K genetic variant associates with NAFLD and with reduced plasma triglyceride levels in humans. However, the molecular mechanisms underlying these associations remain unclear. We hypothesized that TM6SF2 E167K affects hepatic very low-density lipoprotein (VLDL) secretion and studied the kinetics of apolipoprotein B100 (apoB100) and triglyceride metabolism in VLDL in homozygous subjects. In 10 homozygote TM6SF2 E167K carriers and 10 matched controls, we employed stable-isotope tracer and compartmental modeling techniques to determine apoB100 and triglyceride kinetics in the 2 major VLDL subfractions: large triglyceride-rich VLDL1 and smaller, less triglyceride-rich VLDL2. VLDL1-apoB100 production was markedly reduced in homozygote TM6SF2 E167K carriers compared with controls. Likewise, VLDL1-triglyceride production was 35% lower in the TM6SF2 E167K carriers. In contrast, the direct production rates for VLDL2-apoB100 and triglyceride were not different between carriers and controls. In conclusion, the TM6SF2 E167K genetic variant was linked to a specific reduction in hepatic secretion of large triglyceride-rich VLDL1. The impaired secretion of VLDL1 explains the reduced plasma triglyceride concentration and provides a basis for understanding the lower risk of cardiovascular disease associated with the TM6SF2 E167K genetic variant.

Antiatherogenic properties of high-density lipoproteins from arterial plasma are attenuated as compared to their counterparts of venous origin

Background and aimsHigh-density lipoprotein (HDL) particles play atheroprotective roles by their ability to efflux cholesterol from foam cells and to protect low-density lipoproteins (LDLs) from oxidative damage in the arterial intima. We hypothesized that antioxidative properties of HDLs can be attenuated in the oxygen-rich prooxidative arterial environment, contributing to the development of atherosclerosis. To evaluate this hypothesis, we compared antioxidative activity of HDLs from arterial and venous plasmas. Methods and resultsArterial and venous blood samples were simultaneously obtained from 16 patients (age 68 ± 10 years; 75% males) presenting with ischemic or valvular heart disease. Major HDL subfractions and total HDLs were isolated by density gradient ultracentrifugation and their chemical composition and the capacity to protect LDLs from in vitro oxidation were evaluated. HDL-cholesterol, triglycerides and apolipoprotein (apo) B-100 levels were slightly but significantly reduced by −4 to −8% (p < 0.01) in the arterial vs. venous samples. Total mass of HDL subpopulations was similar and HDL subpopulations did not reveal marked compositional differences between the arterial and venous circulation. Potent antioxidative activity of the small, dense HDL3c subpopulation was significantly reduced in the particles of arterial origin vs. their counterparts from venous plasma (increase of +21% in the propagation rate of LDL oxidation, p < 0.05). Interestingly, antioxidative properties of venous HDLs were enhanced in statin-treated patients relative to untreated subjects. ConclusionAntioxidative properties of small, dense HDLs from arterial plasma are attenuated as compared to the particles of venous origin, consistent with the development of atherosclerosis in the arterial wall.

Abstract 17162: Ambient Ultra Fine Particle Exposure Impairs Gut Vascular Barrier via Notch Signaling

Introduction: Ultrafine particles (UFP, d &lt; 0.1 μm), a major sub-fraction of particulate matter (PM 2.5 , d &lt; 2.5μm) in air pollutant, have been reported to increase cardiovascular morbidity. UFP promote endothelial dysfunction/permeability associated with Notch inhibition and degradation of tight junction protein. Gut vascular barrier (GVB) is a distinct protective barrier that inhibits translocations of antigens across the gut lumen, preventing infections by oral digestions. Here, we hypothesized UFP impair GVB by attenuating Notch signaling and disrupting tight junction protein. Method: The transgenic Tg( flk1:mCherry ) zebrafish embryos were immobilized in neutralized tricaine solution and mounted in 2% low melting agarose to perform micro-gavage at 5 days post fertilizations (5dpf). A mA mixture of 10 kD FITC conjugated dextran and phenol red dye wasere micro-gavaged on the intestinal bulb with or without UFP at 25 μg/mL. Anterior trunk and posterior cardinal vein (PCV) post micro-gavage were imaged under a confocal microscope. Notch signaling related genes including the Notch ligand, Dll4, and the target, HES1, and the level of tight junction protein genes zonula occludens-1 (ZO-1) and Occludin (OCLN) mRNA expression following different courses of exposure to UFP were assessed in Human Aortic Endothelial Cells (HAEC) with quantitative RT-PCR., Result: In the control group, the majority of FITC-dextran remained inside the gastrointestinal system. On the other hand, UFP exposure developed a significant accumulation of FITC-dextran in the intersomatic spaces (ISS) between the dorsal aorta (DA) and the PCV. This finding supports evidence of UFP exposure disrupted both the epithelial boundary and the endothelial vascular layer of the gut. In addition, Notch signaling related gene (HES1, DLL4) and the tight junction proteins genes (ZO-1, OCLN) expressions in HAEC were downregulated in concentration and time-course dependent manner by the exposure to UFP. Conclusion: The UFP exposure affected GVB homeostasis and increased endothelial permeability. This finding supports the link between ambient air pollutant exposure to the impairment in the gastrointestinal system.

Small dense HDLs display potent vasorelaxing activity, reflecting their elevated content of sphingosine-1-phosphate

The functional heterogeneity of HDL is attributed to its diverse bioactive components. We evaluated whether the vasodilatory effects of HDL differed across HDL subpopulations, reflecting their distinct molecular composition. The capacity of five major HDL subfractions to counteract the inhibitory effects of oxidized LDL on acetylcholine-induced vasodilation was tested in a rabbit aortic rings model. NO production, an essential pathway in endothelium-dependent vasorelaxation, was studied in simian vacuolating virus 40-transformed murine endothelial cells (SVECs). Small dense HDL3 subfractions displayed potent vasorelaxing activity (up to +31% vs. baseline, P < 0.05); in contrast, large light HDL2 did not induce aortic-ring relaxation when compared on a total protein basis. HDL3 particles were enriched with sphingosine-1-phosphate (S1P) (up to 3-fold vs. HDL2), with the highest content in HDL3b and -3c that concomitantly revealed the strongest vasorelaxing properties. NO generation was enhanced by HDL3c in SVECs (1.5-fold, P < 0.01), a phenomenon that was blocked by the S1P receptor antagonist, VPC 23019. S1P-enriched reconstituted HDL (rHDL) was a 1.8-fold (P < 0.01) more potent vasorelaxant than control rHDL in aortic rings. Small dense HDL3 particles displayed potent protective effects against oxidative stress-associated endothelium dysfunction, potentially reflecting their elevated content of S1P that might facilitate interaction with S1P receptors and ensuing NO generation.

Comparison of flocculant aids as pretreatment reagent for membrane filtration process by fingerprint analysis of organic matters in secondary effluent

Coagulation/flocculation is becoming a conventional and beneficial pretreatment for membrane filtration of wastewater, but how to choose coagulant/flocculant aid and optimal dosage is still a key issue in practice. Two cationic flocculant aids, polyacrylamide (PAM) and Kuriverter EP-301 (EP), were compared for the removal of organic matters from secondary effluents of municipal wastewater plant using FeCl3 as coagulant. EP achieved a better performance than PAM in terms of membrane filtration factor (MFF) and removal of suspended solids, turbidity and dissolved organic carbon. Optimal dosage for PAM was 30 mg Fe/L FeCl3 and 1 mg/L PAM, at which MFF was 2.13, while when 30 mg Fe/L FeCl3 and 1 mg/L EP was injected, MFF was 1.47. Optimal dosage for EP was 50 mg Fe/L FeCl3 and 2 mg/L EP, at which MFF was as low as 1.04. Fingerprint analysis method was first used to analyze the removal characteristics of dissolved organic matters fraction by PAM and EP flocculation. It was found that EP could remove more hydrophobic fractions (HO) than PAM. Moreover, EP could remove part of hydrophilic fractions (HI), while PAM had no effect on HI fractions. Hydrophobic acids (HOA) with MW > 5,000 Da were the major subfraction removed largely by coagulation/flocculation and EP could remove more of it than PAM. Larger MW (>107 Da) and three-dimensional molecular structure made EP more effective based on polymer bridging mechanism and charge neutralization.

Purification, identification and structural modelling of DPP-IV inhibiting peptides from barbel protein hydrolysate.

Inhibition of DPP-IV may improve glycemic control in diabetics by preventing the rapid breakdown and there by prolonging the physiological action of incretin hormones. Barbel muscle protein hydrolysate (BMPH) was noted to exhibit DPP-IV inhibitory activity, with an IC50 value of 1.94mg/mL. It was fractionated into five major fractions (FI-FV) by size exclusion chromatography using a Superdex peptide. The FIII fraction was noted to display the highest inhibitory activity, with an IC50 value of 1.23mg/mL, and was, therefore, further fractionated by RP-HPLC. Four major peptide sub-fractions were selected. The results revealed that the SF4 sub-fraction showed the highest DPP-IV inhibitory activity, with an IC50 value of 0.21mg/mL. This sub-fraction was submitted to RP-HPLC, ESI-MS, and ESI-MS/MS analyses. The findings indicated that SF4 consisted of two peptides (IC50=96μg/mL), namely PP1 and PP2, whose structures were identified as Trp-Ser-Gly (330Da) and Phe-Ser-Asp (349Da), respectively. This is the first report of these sequences from barbel proteins. The structural modelling through docking simulations results with DPP-IV showed that the Trp-Ser-Gly peptide bound to DPP-IV with high affinity. Overall, the results suggested that BMPH can be considered as a promising natural source of DPP-IV inhibitory peptides.

Structural and Functional Study of d-Glucuronyl C5-epimerase

Heparan sulfate (HS) is a glycosaminoglycan present on the cell surface and in the extracellular matrix, which interacts with diverse signal molecules and is essential for many physiological processes including embryonic development, cell growth, inflammation, and blood coagulation. D-glucuronyl C5-epimerase (Glce) is a crucial enzyme in HS synthesis, converting D-glucuronic acid to L-iduronic acid to increase HS flexibility. This modification of HS is important for protein ligand recognition. We have determined the crystal structures of Glce in apo-form (unliganded) and in complex with heparin hexasaccharide (product of Glce following O-sulfation), both in a stable dimer conformation. A Glce dimer contains two catalytic sites, each at a positively charged cleft in C-terminal α-helical domains binding one negatively charged hexasaccharide. Based on the structural and mutagenesis studies, three tyrosine residues, Tyr(468), Tyr(528), and Tyr(546), in the active site were found to be crucial for the enzymatic activity. The complex structure also reveals the mechanism of product inhibition (i.e. 2-O- and 6-O-sulfation of HS keeps the C5 carbon of L-iduronic acid away from the active-site tyrosine residues). Our structural and functional data advance understanding of the key modification in HS biosynthesis.

Association of high-density lipoprotein subclasses and incident coronary heart disease: The Jackson Heart and Framingham Offspring Cohort Studies.

We aimed to clarify the associations of high-density lipoprotein cholesterol (HDL-C) subclasses with incident coronary heart disease (CHD) in two large primary prevention cohorts. We measured cholesterol at baseline from the two major HDL subfractions (larger, more buoyant HDL2 and smaller, denser HDL3) separated by density gradient ultracentrifugation in 4114 (mean age 53.8 years; 64% female) African American participants from the Jackson Heart Study and 818 (mean age 57.3 years, 52% female) predominantly Caucasian participants from the Framingham Offspring Cohort Study. Multivariable adjusted hazard ratios (HRs) for HDL-C and its subclasses were derived from Cox proportional hazards regression models to estimate associations with incident CHD events including myocardial infarction, CHD death, and revascularization. Analyses were performed for each cohort separately and as a combined population. In models adjusted for cardiovascular risk factors for the combined population, HDL3-C (HR 0.76 per SD increase; 95% confidence interval (CI), 0.62-0.94; p = 0.01), rather than HDL2-C (HR 0.88 per SD; 95% CI, 0.72-1.09; p = 0.24) drove the inverse association of HDL-C (HR 0.79 per SD; 95% CI, 0.64-0.98; p = 0.03) with CHD. Similar associations were seen in multivariable analyses within each cohort including after adjusting for apolipoprotein A1 in the Jackson Heart Study. Smaller, denser HDL3-C levels are primarily responsible for the inverse association between HDL-C and incident CHD in this diverse group of primary prevention subjects. These findings have important implications ranging from considerations of HDL biology to interpretations of clinical trials utilizing HDL-C therapeutics.

Abstract 627: A Deleterious Mutation in SCARB1 Significantly Alters High-Density Lipoprotein Level, Composition and Biological Function in Humans

The membrane protein Scavenger Receptor Class B, Type 1 (SR-BI), coded by the gene SCARB1, facilitates the selective uptake of cholesteryl esters from HDL in the liver. Its impact on human lipoprotein metabolism however is not fully understood. Our laboratory has recently identified two homozygotes and 16 heterozygotes for a coding mutation in SCARB1 (c.1127C&gt;T, p.Pro376Leu). We assessed the hypothesis that this SCARB1 mutation significantly increases HDL-C and alters HDL composition and biological function. Plasma from subjects homozygous (n=2) or heterozygous (n=8) for the SR-BI Pro376Leu mutation was compared with that of normolipidemic controls. Five major HDL subfractions were isolated from EDTA-treated plasma using isopycnic density gradient ultracentrifugation and their chemical composition was assayed using commercial enzymatic assay kits. Quantification of &gt;160 molecular species of HDL phospho- and sphingolipids was accomplished via a novel LC-ESI/MS/MS based analysis. Cholesterol efflux capacity, measured from THP-1 macrophages, was determined in total HDL. We identified a &gt;2-fold increase in HDL-C in homozygotes and a &gt;1.5-fold increase in heterozygotes compared to controls, owing mainly to an increase in large, light HDL2 particles. The ApoA-I/ApoA-II ratio was increased in a gene-dose dependent fashion. HDL isolated from heterozygotes mediated 13.5% less efflux when compared to particles from control subjects. Analysis of the HDL phosphosphingolipidome revealed a 23% decrease in phosphatidylinositol content between carriers and controls. A gene-dose dependent trend towards diminishment of phosphatidylserine and increased ceramide was noticeable. In conclusion, the SR-BI Pro376Leu significantly increases HDL levels, but potentially reduces the capacity of HDL to mediate cellular cholesterol efflux. This may be due in part to a decrease in minor, negatively charged phospholipid subclasses including PI and PS, which have previously been shown to play a functional role in stimulating cellular cholesterol efflux.

Distinct Wnt-driven primitive streak-like populations reflect in vivo lineage precursors

During gastrulation, epiblast cells are pluripotent and their fate is thought to be constrained principally by their position. Cell fate is progressively restricted by localised signalling cues from areas including the primitive streak. However, it is unknown whether this restriction accompanies, at the individual cell level, a reduction in potency. Investigation of these early transition events in vitro is possible via the use of epiblast stem cells (EpiSCs), self-renewing pluripotent cell lines equivalent to the postimplantation epiblast. Strikingly, mouse EpiSCs express gastrulation stage regional markers in self-renewing conditions. Here, we examined the differentiation potential of cells expressing such lineage markers. We show that undifferentiated EpiSC cultures contain a major subfraction of cells with reversible early primitive streak characteristics, which is mutually exclusive to a neural-like fraction. Using in vitro differentiation assays and embryo grafting we demonstrate that primitive streak-like EpiSCs are biased towards mesoderm and endoderm fates while retaining pluripotency. The acquisition of primitive streak characteristics by self-renewing EpiSCs is mediated by endogenous Wnt signalling. Elevation of Wnt activity promotes restriction towards primitive streak-associated lineages with mesendodermal and neuromesodermal characteristics. Collectively, our data suggest that EpiSC pluripotency encompasses a range of reversible lineage-biased states reflecting the birth of pioneer lineage precursors from a pool of uncommitted EpiSCs similar to the earliest cell fate restriction events taking place in the gastrula stage epiblast.

English

Securinega virosa is a commonly used medicinal plant in African traditional medicine in the management of epilepsy. In an attempt to isolate and characterize the bioactive principles responsible for the anticonvulsant property, the crude methanol root bark extract of the plant was exhaustively partitioned into petroleum ether, chloroform, ethyl acetate and n-butanol. The anticonvulsant potential of the n-butanol fraction (75, 150 and 300 mg/kg) was evaluated using maximal electroshock (MES) test in chicks, strychnine, picrotoxin, 4-aminopyridine and pentylenetetrazole-induced seizures in mice. The fraction did not protect the chicks against tonic-hind limb extension due to MES. Similarly, the fraction did not afford significant protection against seizures induced by strychnine, aminopyridine and picrotoxin contrary to the observations of protections in the crude extract, against pentylenetetrazole-induced seizure, the fraction afforded 66.67% protection and significantly (P&lt;0.01) delayed the onset of seizure in unprotected animals. Column chromatographic separation of the n-butanol fraction yielded three major sub-fractions which were subjected to anticonvulsant screening using pentylenetetrazol (PTZ)-induced seizure. The anticonvulsant potential of the n-butanol fraction was retained in the more polar sub-fractions. The findings of this study suggest that the n-butanol fraction Securinega virosa root bark contains bioactive principle(s) that possess anticonvulsant activities which may be beneficial against absence seizure. This lends further credence to the ethnomedicinal claim of the use of the root of S. virosa in the management of epilepsy. &nbsp; Key words: Securinega virosa, epilepsy, seizure, maximal electroshock, pentyleneterazole, picrotoxin, 4-aminopyridine, strychnine.

Abstract 37: Association between High-Density Lipoprotein Cholesterol Subfractions and Carotid Intima-Media Thickness

Background Recent trials have challenged the protective effect of HDL in atherosclerosis, implicating the need to shift the focus from total HDL concentration to its function and major subfractions. The objective of this cross-sectional study was to investigate the relation between HDL2 and HDL3 subfractions and carotid intima-media thickness (IMT), an early indicator of atherosclerosis and surrogate marker of ischemic stroke. Methods In NOMAS, a prospective population-based study of the incidence and risk factors of stroke, we evaluated 1041 participants (mean age 68±9 years; 38% men; 62% Hispanic, 19% black, 17% white) with available data on HDL subfractions and carotid IMT assessed by a high-resolution 2D carotid ultrasound. Carotid IMT was a composite measure of the mean IMT in all carotid artery segments. The cross-sectional association between HDL-subfractions and IMT was analyzed by multiple linear regression. Results We mean±SD of total HDL was 46.15±14.25 mg/dl (median = 44.00), HDL2 14.59±8.29 mg/dl (median = 13.00), HDL3 31.06±8.32 mg/dl (median = 32.00). The mean IMT was 0.90±0.08 mm. After controlling for demographics, LDL, triglycerides, and cholesterol-lowering medication, a strong association was observed between total HDL (per SD, beta=-0.008, p= 0.01), HDL2 (per SD, beta=-0.006, p=0.03) and HDL3 (per SD, beta=-0.007 p=0.01) in relation to IMT. A significant interaction was observed between diabetes (vs. non-diabetes) and HDL2 in relation to IMT (p=0.01; -0.018mm). Conclusion The observed strong independent inverse associations between HDL, HDL2 and HDL3 and carotid IMT. HDL2 was only significantly associated with IMT among individuals with diabetes. These observations show novel evidence for a potential role of HDL2 and HDL3 subfractions, providing the opportunity to develop novel HDL-targeted therapies for the prevention of atherosclerosis.

The effect of Omega-3 fatty acids on the atherogenic lipoprotein phenotype in patients with nephrotic range proteinuria

Patients with nephrotic range proteinuria are known to have an increased risk of cardiovascular disease partly due to possessing the atherogenic lipoprotein phenotype. The aim of this study was to examine the effect of high dose omega-3 fatty acids on atherogenic triglyceride rich lipoproteins in patients with nephrotic range proteinuria, comparing their effect on lipoprotein profiles in age and sex matched controls. 17 patients with nephrotic range proteinuria and 17 age and sex matched controls were studied. Fasting lipids and lipoproteins were measured before and after 8 weeks treatment with 4 g daily of omega-3 fatty acids (Omacor®). In patients with proteinuria treatment reduced plasma triglyceride by a mean of 0.45 mmol/l (95%CI 0.16 - 0.74, p = 0.005) and plasma very low density lipoprotein cholesterol by a mean of 0.38 (95%CI 0.01 - 0.75, p = 0.04). LDL III concentration fell from 178.8 mg/dl (61.6 - 231.0) to 96.1 mg/dl (49.3 - 204.5), p = 0.05. In patients treatment altered the LDL profile so that LDLIII which was the major subfraction present at baseline was reduced from 49.9% to 29.8% (p = 0.01). Remnant lipoproteins (RLP) also fell with a mean reduction of 3.5 mg/dl in RLP-Cholesterol (95%CI 0.1 - 6.9, p = 0.05) and 12.4 mg/dl in RLP-triglyceride (95%CI 2.6 - 22.2, p = 0.03). There was however a 0.6 mmol/l rise in LDL-C (p = 0.06) in the patients. Treatment did not alter HDL-C. In patients with nephrotic range proteinuria, omega-3 fatty acids reduced triglyceride rich lipoproteins, LDL III and remnant lipoproteins. A tendency to an increase in LDL-C was observed but this was offset by an alteration in the distribution of the LDL profile towards lighter, larger LDL particles. We propose that treatment with omega-3 fatty acids in conjunction with a statin may be the ideal therapy in these patients.

Synchronous fluorimetric characterization of heavy intermediates of coal direct liquefaction

In order to understand the molecular structure of coal and the mechanism of coal direct liquefaction, the heavy intermediates of coal liquefaction such as asphaltene (AS) and preasphaltene (PA) were separated into different sub-fractions by column chromatography, respectively. The distributions of fused aromatic nucleuses (FAN) of sub-fractions were determined by synchronous fluorescence combined with the fitting technique of multiple peaks of spectrum. The results indicated that the PA and AS obtained from liquefactions of three Chinese coals were separated into 4 and 2–4 sub-fractions, respectively. The content of three rings FAN was the highest in the major sub-fractions of AS. The major sub-fractions of PA contained not only more single ring aromatic nucleus but also larger FAN than those of AS. Meanwhile, there were more complex aggregations in PA compared to AS.

Application of Polydimethylsiloxane/Glass Microchips for Fast Electrophoretic Separation of Serum High-density Lipoprotein Subclasses

When isolated on the basis of density by ultracentrifugation, high-density lipoprotein (HDL) can be separated into two major subfractions: HDL2 and HDL3. The inhibition of oxidative modification of low-density lipoprotein (LDL) by HDL2 is believed to play

Molecular Characteristics and Immunomodulatory Activities of Water-Soluble Sulfated Polysaccharides from Ulva pertusa

Sulfated polysaccharides isolated from Ulva pertusa and fractionated using anion-exchange chromatography were investigated to determine their molecular characteristics and bioactivities. The crude and fractionated polysaccharides (F(1), F(2), and F(3)) were mainly composed of carbohydrates (59.9-65.9%), sulfates (11.6-15.3%), and uronic acid (7.30-16.4%) with small amounts of proteins (1.40-4.80%). Rhamnose (62.5-80.7%) was the major monosaccharide unit of these polysaccharides, with different levels of glucose (13.5-27.4%) and xylose (2.74-11.5%). The polysaccharides contained one or two major subfractions with weight-average molecular mass ranging from 51.1×10(3) to 1,690×10(3) g/mol. The relatively low in vitro anticancer activity of the polysaccharides (22.3-42.4%) suggested that they had little cytotoxicity against the cancer cell line used (AGS). On the other hand, the polysaccharides significantly stimulated Raw 264.7 cells, inducing considerable amounts of nitric oxide and various cytokines production, which suggested that they could be strong immunostimulators.