Secretion of the glucose-regulated selenoprotein SEPS1 from hepatoma cells

SEPS1 (also called selenoprotein S, SelS, Tanis or VIMP) is a selenoprotein, localized predominantly in the ER membrane and also on the cell surface. In this report, we demonstrate that SEPS1 protein is also secreted from hepatoma cells but not from five other types of cells examined. The secretion can be abolished by the ER-Golgi transport inhibitor Brefeldin A and by the protein synthesis inhibitor cycloheximide. Using a sandwich ELISA, SEPS1 was detected in the sera of 65 out of 209 human subjects (31.1%, average=15.7+/-1.1 ng/mL). Fractionation of human serum indicated that SEPS1 was associated with LDL and possibly with VLDL. The function of plasma SEPS1 is unclear but may be related to lipoprotein metabolism.     

Dissociable and nondissociable forms of platelet-activating factor acetylhydrolase in human plasma LDL: implications for LDL oxidative susceptibility

Platelet-activating factor acetylhydrolase (PAF-AH) is transported by lipoproteins in plasma and is thought to possess both anti-inflammatory and anti-oxidative activity. It has been reported that PAF-AH is recovered primarily in small, dense LDL and HDL following ultracentrifugal separation of lipoproteins. In the present studies, we aimed to further define the distribution of PAF-AH among lipoprotein fractions and subfractions, and to determine whether these distributions are affected by the lipoprotein isolation strategy (FPLC versus sequential ultracentrifugation) and LDL particle distribution profile. When lipoproteins were isolated by FPLC, the bulk (～85%) of plasma PAF-AH activity was recovered within LDL-containing fractions, whereas with ultracentrifugation, there was a redistribution to HDL (which contained ～18% of the activity) and the d>1.21 g/ml fraction (which contained ～32%). Notably, re-ultracentrifugation of isolated LDL did not result in any further movement of PAF-AH to higher densities, suggesting the presence of dissociable and nondissociable forms of the enzyme on LDL. Differences were noted in the distribution of PAF-AH activity among LDL subfractions from subjects exhibiting the pattern A (primarily large, buoyant LDL) versus pattern B (primarily small, dense LDL) phenotype. In the latter group, there was a relative depletion of PAF-AH activity in subfractions in the intermediate to dense range (d=1.039–1.047 g/ml) with a corresponding increase in enzyme activity recovered within the d>1.21 g/ml ultracentrifugal fraction. Thus, there appears to be a greater proportion of the dissociable form of PAF-AH in pattern B subjects. In both populations, most of the nondissociable activity was recovered in a minor small, dense LDL subfraction. Based on conjugated dienes as a measure of lipid peroxidation, variations in PAF-AH activity appeared to contribute to variations in oxidative behavior among ultracentrifugally isolated LDL subfractions. The physiologic relevance of PAF-AH dissociability and the minor PAF-AH-enriched oxidation-resistant LDL subpopulation remains to be determined.     

Lipid profiling of FPLC-separated lipoprotein fractions by electrospray ionization tandem mass spectrometry

Glycerophospholipid and sphingolipid species and their bioactive metabolites are important regulators of lipoprotein and cell function. The aim of the study was to develop a method for lipid species profiling of separated lipoprotein classes. Human serum lipoproteins VLDL, LDL, and HDL of 21 healthy fasting blood donors were separated by fast performance liquid chromatography (FPLC) from 50 microl serum. Subsequently, phosphatidylcholine (PC), lysophosphatidylcholine, sphingomyelin (SM), ceramide (CER), phosphatidylethanolamine (PE), PE-based plasmalogen (PE-pl), cholesterol, and cholesteryl ester (CE) content of the separated lipoproteins was quantified by electrospray ionization tandem mass spectrometry (ESI-MS/MS). Analysis of FPLC fractions with PAGE demonstrated that albumin partially coelutes with HDL fractions. However, analysis of an HDL deficient serum (Tangier disease) showed that only lysophosphatidylcholine, but none of the other lipids analyzed, exhibited a significant coelution with the albumin containing fractions. Approximately 60% of lipoprotein CER were found in LDL fractions and 60% of PC, PE, and plasmalogens in HDL fractions. VLDL, LDL, and HDL displayed characteristic lipid class and species pattern. The developed method provides a detailed lipid class and species composition of lipoprotein fractions and may serve as a valuable tool to identify alterations of lipoprotein lipid species profiles in disease with a reasonable experimental effort.     

Lipid and fatty acid composition of canine lipoproteins

Lipid classes and their fatty acids were studied in the major lipoprotein fractions from canine, in comparison with human, plasma. In dogs, high-density-lipoprotein (HDL), the main carrier of plasma phospholipid (PL), cholesterol ester (CE) and free cholesterol, was the most abundant lipoprotein, followed by low and very-low density lipoproteins (LDL and VLDL). Notably, LDL and VLDL contributed similarly to the total dog plasma triacylglycerol (TG). The PL composition was similar in all three lipoproteins, dominated by phosphatidylcholine (PC). Even though the content and composition of lipids within and among lipoproteins differed markedly between dog and man, the total amount of circulating lipid was similar. All canine lipoproteins were relatively richer than those from humans in long-chain (C20-C22) n-6 and n-3 polyunsaturated fatty acids (PUFA) but had comparable proportions of total saturated and monoenoic fatty acids, with 18:2n-6 being the main PUFA in both mammals. The fatty acid profile of canine and human lipoproteins differed because they had distinct proportions of their major lipids. There were more n-3 and n-6 long-chain PUFA in canine than in human plasma, because dogs had more HDL, their HDL had more PC and CE, and both these lipids were richer in such PUFA.     

脂蛋白质组学研究进展

脂蛋白质组学(lipoproteomics)是一门应用蛋白质组学技术对脂蛋白(lipoprotein)进行全面、系统地分析和鉴定,进而了解脂蛋白的组成和功能以及与相关疾病发生、发展之间关系的新兴学科.近些年来,脂蛋白质组学研究促进了脂蛋白中蛋白质组分的急性期响应、补体激活、免疫响应、炎症响应、蛋白酶抑制等新功能的发现,显示了广阔的应用前景.本文对脂蛋白的功能和分类,以及目前应用于脂蛋白质组学研究的脂蛋白分离方法和蛋白质鉴定方法进行了简述,并综述了高密度脂蛋白、低密度脂蛋白和极低密度脂蛋白的脂蛋白质组学研究,及其在冠状动脉疾病、糖尿病、类风湿性关节炎等疾病研究的最新进展.     

Lipoprotein size is a main determinant for the rate of hydrolysis by exogenous LPL in human plasma

LPL is a key player in plasma triglyceride metabolism. Consequently, LPL is regulated by several proteins during synthesis, folding, secretion, and transport to its site of action at the luminal side of capillaries, as well as during the catalytic reaction. Some proteins are well known, whereas others have been identified but are still not fully understood. We set out to study the effects of the natural variations in the plasma levels of all known LPL regulators on the activity of purified LPL added to samples of fasted plasma taken from 117 individuals. The enzymatic activity was measured at 25°C using isothermal titration calorimetry. This method allows quantification of the ability of an added fixed amount of exogenous LPL to hydrolyze triglyceride-rich lipoproteins in plasma samples by measuring the heat produced. Our results indicate that, under the conditions used, the normal variation in the endogenous levels of apolipoprotein C1, C2, and C3 or the levels of angiopoietin-like proteins 3, 4, and 8 in the fasted plasma samples had no significant effect on the recorded activity of the added LPL. Instead, the key determinant for the LPL activity was a lipid signature strongly correlated to the average size of the VLDL particles. The signature involved not only several lipoprotein and plasma lipid parameters but also apolipoprotein A5 levels. While the measurements cannot fully represent the action of LPL when attached to the capillary wall, our study provides knowledge on the interindividual variation of LPL lipolysis rates in human plasma.     

Can modulators of apolipoproteinB biogenesis serve as an alternate target for cholesterol-lowering drugs?

Understanding the molecular defects underlying cardiovascular disease is necessary for the development of therapeutics. The most common method to lower circulating lipids, which reduces the incidence of cardiovascular disease, is statins, but other drugs are now entering the clinic, some of which have been approved. Nevertheless, patients cannot tolerate some of these therapeutics, the drugs are costly, and/or the treatments are approved for only rare forms of disease. Efforts to find alternative treatments have focused on other factors, such as apolipoproteinB (apoB), which transports cholesterol in the blood stream. The levels of apoB are regulated by endoplasmic reticulum (ER) associated degradation as well as by a post ER degradation pathway in model systems, and we suggest that these events provide novel therapeutic targets. We discuss first how cardiovascular disease arises and how cholesterol is regulated, and then summarize the mechanisms of action of existing treatments for cardiovascular disease. We then review the apoB biosynthetic pathway, focusing on steps that might be amenable to therapeutic interventions.