Selective Removal of Low-Density Lipoproteins from Blood by Induced Precipitation with AAS in Vitro(Ⅰ)

1 INTRODUCTION It’s evident that high level of cholesterol in blood is associated with the formation and devel-opment of familial hypercholestrolemia(FH)and atherosclerosis(AS).In general,choles-terol in blood is mainly combined with low-density lipoproteins(LDL),very low-densitylipoproteins(VLDL)and high density lipoproteins(HDL).About 60%-80% cholesterolexists in LDL and VLDL.LDL and VLDL have been recognized as the principal

Overexpression of low-density lipoprotein receptor prevents neurotoxic polarization of astrocytes via inhibiting NLRP3 inflammasome activation in experimental ischemic stroke

Neurotoxic astrocytes are a promising therapeutic target for the attenuation of cerebral ischemia/reperfusion injury.Low-density lipoprotein receptor,a classic cholesterol regulatory receptor,has been found to inhibit NLR family pyrin domain containing protein 3(NLRP3)inflammasome activation in neurons following ischemic stroke and to suppress the activation of microglia and astrocytes in individuals with Alzheimer’s disease.However,little is known about the effects of low-density lipoprotein receptor on astrocytic activation in ischemic stroke.To address this issue in the present study,we examined the mechanisms by which low-density lipoprotein receptor regulates astrocytic polarization in ischemic stroke models.First,we examined low-density lipoprotein receptor expression in astrocytes via immunofluorescence staining and western blotting analysis.We observed significant downregulation of low-density lipoprotein receptor following middle cerebral artery occlusion reperfusion and oxygen-glucose deprivation/reoxygenation.Second,we induced the astrocyte-specific overexpression of low-density lipoprotein receptor using astrocyte-specific adeno-associated virus.Low-density lipoprotein receptor overexpression in astrocytes improved neurological outcomes in middle cerebral artery occlusion mice and reversed neurotoxic astrocytes to create a neuroprotective phenotype.Finally,we found that the overexpression of low-density lipoprotein receptor inhibited NLRP3 inflammasome activation in oxygen-glucose deprivation/reoxygenation injured astrocytes and that the addition of nigericin,an NLRP3 agonist,restored the neurotoxic astrocyte phenotype.These findings suggest that low-density lipoprotein receptor could inhibit the NLRP3-meidiated neurotoxic polarization of astrocytes and that increasing low-density lipoprotein receptor in astrocytes might represent a novel strategy for treating cerebral ischemic stroke.

基于CEEMDAN-QPSO-BLS模型的径流预测研究

准确的径流预测是水资源优化配置和高效利用的前提,是制定防洪减灾决策的基础,然而受到人类活动、环境、气候等因素的影响,径流序列呈现出非线性、非稳态、多尺度变化的特点,这为径流的精准预测增加了难度。为提高径流预测的精准度和可信度,结合自适应噪声完备集合经验模态分解(Complete Ensemble Empirical Mode Decomposition with Adaptive Noise,CEEMDAN)方法,量子粒子群优化算法(Quantum Particle Swarm Optimization,QPSO)、宽度学习系统(Broad Learning System,BLS)模型,提出了一种基于CEEMDAN-QPSO-BLS组合式的径流预测模型。该组合模型首先使用CEEMDAN方法对原始径流信号进行分解,得到若干相对平稳的本征模态分量。其次利用QPSO算法对BLS模型的特征层节点组数、增强层节点组数和组内节点数进行寻优,得到最优的宽度学习网络拓扑结构,进而使用最优的QPSOBLS对多个稳态分量进行预测,并对预测分量进行重构,从而获得更高的预测精度。以黄河流域小浪底水库的日径流值为实验数据,将EMD-QPSO-BLS、QPSO-BLS作为CEEMDAN-QPSO-BLS的对比模型,并采用纳什效率系数(NSE)、均方根误差(RMSE)、平均绝对误差(MAE)和平均绝对百分比误差(MAPE)作为模型预测可信度和精准度的评价指标。实验表明,在预见期4天内,与QPSO-BLS、EMD-QPSO-BLS模型相比,CEEMDAN-QPSO-BLS的预测精准度分别提高了79.87%、19.80%,可信度分别提高了131.2%、10.98%,径流预测精度的提高,可为防洪抗旱保护人民生命财产和可持续发展提供决策支持。

FUCOIDIN INHIBITS OXIDIZED LOW DENSITY LIPOPROTEIN FROM INDUCING HUMAN PERIPHERAL BLOOD MONOCYTE EXPRESSION OF PROINFLAMMATORY CYTOKINES mRNA

Objective To study the significance of scavenger receptor class A(SR A)in mediating human peripheral blood monocyte to uptake oxidized low density lipoprotein(OxLDL) and promoting the atherosclerotic immuno pathological lesion in the local blood vessel. Methods With the Digoxenin labeled Oligonucleotide probes In situ Hybridization, this research investigated the effects of OxLDL on the mRNA expression of proinflammatory cytokines including MCP 1, bFGF, PDGF and IL 10 in the human peripheral blood monocyte and whether fucoidin, a peculiarly inhibitory ligand for SR A, would influence this process. Results Monocyte was significantly increased the mRNA expression of MCP 1, bFGF, PDGF and IL 10 in a dose dependent manner after incubating with OxLDL (10,15,20,25,30?mg·L -1 , respectively)for 24 hours( P < 0.001 ). Fucoidin(50,100,150,200,250?mg·mL -1 , respectively)completely inhibited OxLDL(20?mg·L -1 )from inducing monocyte the mRNA expression of above proinflammatory cytokines( P < 0.001 ). Conclusion OxLDL can stimulate human peripheral blood monocyte to give expression to proinflammatory cytokines mRNA in a dose dependent manner, while a peculiarly inhibitory ligand for SR A fucoidin has an obviously opposed role.

Hepatitis C virus relies on lipoproteins for its life cycle

Hepatitis C virus(HCV) infects over 150 million people worldwide. In most cases, HCV infection becomes chronic causing liver disease ranging from fibrosis to cirrhosis and hepatocellular carcinoma. Viral persistence and pathogenesis are due to the ability of HCV to deregulate specific host processes, mainly lipid metabolism and innate immunity. In particular, HCV exploits the lipoprotein machineries for almost all steps of its life cycle. The aim of this review is to summarize current knowledge concerning the interplay between HCV and lipoprotein metabolism. We discuss the role played by members of lipoproteins in HCV entry, replication and virion production.

Establishment of Real-Time TaqMan-Fluorescence Quantitative RT-PCR Assay for Detection and Quantification of Porcine Lipoprotein Lipase mRNA

Porcine lipoprotein lipase （LPL） cDNA was cloned as the standard for real-time quantifying LPL mRNA and the TaqMan-fluorescence quantitative PCR assay for detection was established. The total RNA extracted from Longissimus dorsi of porcine was reverse-transcribed to cDNA. LPL cDNA was ligated with pGM-T vector and transformed into Escherichia coli TOP10. Plasmid DNA extracted from positive clones was verified by PCR amplification and sequenced. LPL was amplified by real-time fluorescence quantitative PCR from the plasmid DNA. The concentration of DNA template purified was detected by analyzing absorbance in 260 nm and then the combined plasmid was diluted to series as standard for fluorescence quantitative PCR （FQ-PCR）. The method of LPL mRNA real-time PCR was well established, which detected as low as 103 with the linear range 10^3 to 10^10 copies. The standard curves showed high correlations （R2 = 0.9871）. A series of standards for real-time PCR analysis have been constructed successfially, and real-time TaqMan-fluorescence quantitative RT-PCR is reliable to quantitatively evaluate FQ-PCR mRNA in L. dorsi of porcine.

High-density lipoproteins:an emerging target in the prevention of cardiovascular disease

High-density lipoproteins （HDLs） have been well established to protect against the development of atherosclerotic cardiovascular disease. It has become apparent that in addition to the promotion of reverse cholesterol transport, HDLs possess a number of additional functional properties that may contribute to their beneficial influence on the arterial wall. A number of exciting therapeutic strategies have been developed that target HDL and its ability to protect against the development of atherosclerotic plaque. This paper will review how the promotion of the functional properties of HDL inhibits the formation of atherosclerotic plaque and stabilises lesions in patients with established disease.

Effects of Oxidized Low Density Lipoprotein on Transformation of Valvular Myofibroblasts to Osteoblast-like Phenotype

In order to investigate the roles of Wnt signal pathway in transformation of cardiac valvular myofibroblasts to the osteoblast-like phenotype, the primary cultured porcine aortic valve myofibroblasts were incubated with oxidized low density lipoprotein（ox-LDL, 50 mg/L）, and divided into four groups according to the ox-LDL treatment time： control group, ox-LDL 24-h group, ox-LDL 48-h group, and ox-LDL 72-h group. Wnt signal pathway blocker Dickkopf-1（DDK-1, 100 μg/L） was added in ox-LDL 72-h group. The expression of α-smooth muscle actin（α-SMA）, bone morphogenetic protein 2（BMP2）, alkaline phosphatase（ALP）, and osteogenic transcription factor Cbfa-1 was detected by Western blotting, and that of β-catenin, a key mediator of Wnt signal pathway by immunocytochemical staining method. The Wnt/β-catenin was observed and the transformation of myofibroblasts to the osteoblast-like phenotype was examined. The expression of α-SMA, BMP2, ALP and Cbfa-1 proteins in the control group was weaker than in the ox-LDL-treated groups. In ox-LDL-treated groups, the protein expression of α-SMA, BMP2, ALP, and Cbfa-1 was significantly increased in a time-dependent manner as compared with the control group, and there was significant difference among the three ox-LDL-treated groups（P〈0.05 for all）; β-catenin protein was also up-regulated in the ox-LDL-treated groups in a time-dependent manner as compared with the control group（P〈0.05）, and its transfer from cytoplasm to nucleus and accumulation in the nucleus were increased in the same fashion（P〈0.05）. After addition of DKK-1, the expression of α-SMA, bone-related proteins and β-catenin protein was significantly reduced as compared with ox-LDL 72-h group（P〈0.05）. The Wnt/ β-catenin signaling pathway may play an important role in transformation of valvular myofibroblasts to the osteoblast-like phenotype.

Editorial on hemoglobin A1c, blood pressure, and lowdensity lipoprotein cholesterol goals in diabetics

The American Diabetes Association (ADA) 2013 guidelines state that a reasonable hemoglobin A1c goal for many nonpregnant adults with diabetes is less than 7.0% a hemoglobin A1c level of less than 6.5% may be considered in adults with short duration of diabetes, long life expectancy, and no significant cardiovascular disease if this can be achieved without significant hypoglycemia or other adverse effects of treatment. A hemoglobin A1c level less than 8.0% may be appropriate for patients with a history of severe hypoglycemia, limited life expectancy, advanced macrovascular and microvascular complications, extensive comorbidities, and long-standing diabetes in whom the hemoglobin A1c goal is difficult to attain despite multiple glucoselowering drugs including insulin. The ADA 2013 guidelines recommend that the systolic blood pressure in most diabetics with hypertension should be reduced to less than 140 mmHg. These guidelines also recommend use of an angiotensin-converting enzyme inhibitor or angiotensin receptor blocker in the treatment of hypertension in diabetics unless they are pregnant. Diabetics at high risk for cardiovascular events should have theirserum low-density lipoprotein (LDL) cholesterol lowered to less than 70 mg/dL with statins. Lower-risk diabetics should have their serum LDL cholesterol reduced to less than 100 mg/dL. Combination therapy of a statin with either a fibrate or niacin has not been shown to provide additional cardiovascular benefit above statin therapy alone and is not recommended. Hypertriglyceridemia should be treated with dietary and lifestyle changes. Severe hypertriglyceridemia should be treated with drug therapy to reduce the risk of acute pancreatitis.

High density lipoproteins and type 2 diabetes:Emerging concepts in their relationship

Patients with type 2 diabetes mellitus(T2DM) frequently exhibit macrovascular complications of atherosclerotic cardiovascular(CV) disease. High density lipoproteins(HDL) are protective against atherosclerosis. Low levels of HDL cholesterol(HDL-C) independently contribute to CV risk. Patients with T2 DM not only exhibit low HDL-C, but also dysfunctional HDL. Furthermore, low concentration of HDL may increase the risk for the development of T2 DM through a decreased β cell survival and secretory function. In this paper, we discuss emerging concepts in the relationship of T2 DM with HDL.

POLYMORPHISMS OF THE HUMAN LIPOPROTEIN LIPASE GENE:POSSIBLE ASSOCIATION WITH LIPID LEVELS IN PATIENTS WITH CORONARY HEART DISEASE IN BEIJING AREA

The polymorphisms(Pvu Ⅱand Hind Ⅲ) on the lipoprotein lipase(LPL) gene locus was investigated in a sample of 100 patients surviving previous myocardial infarction and 100 age matched healthy individuals selected from Han Chinese of Beijing area.In patient group a strong association was found between H+allele of Hind Ⅲ polymorphism and raised TG levels(P<0.01).In control group P-P-genotype was observed to be associated with higher TG levels compared with P+P genotype of Pvu Ⅱ polymorphism(P<0.05).Combination of H+H+ genotype with P-P-genotype showed the highest TG levels among all nine kinds of genotype combinations in patient group(P<0.01).However,comparison of distribution of alleles and genotypes of these polymorphisms between patient group and control group demonstrated no significant difference. Our data suggest that the polymorphisms at the LPL gene,as the linkage markers with an aetiologic mutation at or around LPL gene,may constitute one of the genetic determinants for the population variation in plasma TG levels,as well as for the common dyslipidemia in Chinese population.

Expression of Monocyte Chemoattractant Protein-1 in Monocytes and Effects of Native and Oxidized Very Low Density Lipoproteins

Monocyte chemoattractant protein-1(MCP-1), a potent chemoattractant, is thought to play an important role in migration of monocytes into atherosclerotic lesions. The present study was designed to investigate the capacity of human peripheral blood monocytes to express MCP-1 and effects of native very low density lipoprotein (VLDL) and oxidized VLDL(OX-VLDL) on the expression. The total RNA was extracted from cultured monocytes, which were exposed to VLDL and OX-VLDL, and the media conditioned by monocytes were collected. MCP-1 mRNA expression was examined by Northern blot analysis. MCP-1 protein in conditioned media was determined by using sandwich ELISA. The results showed that monocytes can express MCP-1 after a 24 h incubation at 37℃，and the expression was markedly increased by a exposure to OX-VLDL, whereas the expression was slightly increased when exposed to VLDL. It suggests that the capacity of monocytes to produce MCP-1 that recruits and activates circulating monocytes may be of considerable importance in atherogenesis, and oxidation of VLDL enhances its potential to promote atherogenesis.

C57BL/6小鼠微小三毛滴虫的观察与种系发育分析

为鉴定从某外单位引进的C57BL/6小鼠粪样检测中发现的虫体种类,本试验对虫体进行形态学观察,并进行种系发育分析。取C57BL/6小鼠粪样涂片和瑞士染色后进行显微镜观察,提取粪样总DNA,根据三毛滴虫的16S rRNA进行PCR扩增后测序,运用MEGA11进行种系发育分析。显微镜观察发现,虫体符合微小三毛滴虫的形态学特征,测序发现微小三毛滴虫的基因序列与鼠三毛滴虫高度同源。

Metabolism of high density lipoproteins in liver cancer

Liver plays a vital role in the production and catabolism of plasma lipoproteins. It depends on the integrity of cellular function of liver, which ensures homeostasis of lipid and lipoprotein metabolism. When liver cancer occurs these processes are impaired and high-density lipoproteins are changed.

The pleiotropic vasoprotective functions of high density lipoproteins(HDL)

The pleiotropic functions of circulating high density lipoprotein（HDL） on peripheral vascular health are well established. HDL plays a pivotal role in reverse cholesterol transport and is also known to suppress inflammation,endothelial activation and apoptosis in peripheral vessels. Although not expressed in the central nervous system, HDL has nevertheless emerged as a potential resilience factor for dementia in multiple epidemiological studies. Animal model data specifically support a role for HDL in attenuating the accumulation of P-amyloid within cerebral vessels concomitant with reduced neuroinflammation and improved cognitive performance. As the vascular contributions to dementia are increasingly appreciated, this review seeks to summarize recent literature focused on the vasoprotective properties of HDL that may extend to cerebral vessels, discuss potential roles of HDL in dementia relative to brainderived lipoproteins, identify gaps in current knowledge, and highlight new opportunities for research and discovery.

OXIDIZED HIGH-DENSITY LIPOPROTEIN PROMOTES MATURATION AND MIGRATION OF BONE MARROW DERIVED DENDRITIC CELLS FROM C57BL/6J MICE

Objective To explore the influence of oxidized high-density lipoprotein （oxHDL） on the maturation and migration of bone marrow-derived dendritic cells （BMDCs） from C57BL/6J mice. Methods The C57BL/6J mice bone marrow cell suspension was prepared and purified. Recombinant granulocyte-macrophage colony-stimulating factor （rmGM-CSF） and recombinant interleukin-4 （rmlL-4） were used to promote monocytes to differentiate and suppress lymphocytes. Then 50μg/mL oxHDL was added to stimulate BMDCs, using 50μg/mL high-density lipoprotein （HDL） as homologous protein control, PBS as negative control, and 1 μg/mL lipopolysaccharide （LPS） as positive control. The CD86 and MHCII expression rates were detected with fluorescence-activated cell sorting （FACS）. Liquid scintillation counting （LSC） was used in mixed lymphocyte reactions （MLRs） to reflect the ability of BMDCs in stimulating the proliferation of homologous T cells, Levels of cytokines IL-12 and IL-10 were detected by ELISA. The cell migration was evaluated with the transwell system. Results Compared with PBS group, the expressions of CD86 and MHCII, counts per minute of MLRs, secretion of IL-12 and IL-10, and number of migrated cells in oxHDL group and LPS group significantly increased （all P 〈 0.05）, while the increment was less in oxHDL group than LPS group. The number of migrated cells in oxHDL group was about twice of that in HDL group. Conclusion OxHDL may promote the maturation and migration of BMDCs in vitro.

Effect of low-density lipoproteins, spermatozoa concentration and glycerol on functional and motility parameters of bull spermatozoa during storage at 4℃

An extender has been developed with low-density lipoproteins （LDLs） that eliminates the microbial risks associated with the use of whole egg yolk. The objective of this study was to assess the effects of substituting egg yolk with LDLs for use as an extender in sperm preservation at 4 ℃, as well as on spermatozoa motility, plasma membrane and acrosome integrity, at two different concentrations （80×10^6 and 240× 10^6 sperm per ml） for 8 days and to evaluate glycerol toxicity in both extenders. A total of 12 ejaculates were collected from three bulls. Spermatozoa motility was examined using computer-assisted semen analysis. Plasma membrane integrity was determined using the hypo-osmotic swelling test and acrosome integrity with the fluorescein isothiocyanate-Pisum sativum agglutinin test. The semen was subsequently divided into four aliquots and diluted with Tris-egg yolk-glycerol （TEG）, Tris-egg yolk without glycerol （TE）, LDL with glycerol （LDL＋） and LDL without glycerol （LDL-）, at 80×10^6 and 240 ×10^6sperm per ml. This study showed that the LDL＋ and LDL- extenders were more effective at preserving spermatozoa motility, plasma membrane integrity and acrosome integrity than TEG and TE （P〈0.05） during 8 days of incubation. After 3 days of incubation, a toxicity of glycerol was observed in TEG, whereas no significant difference was observed between LDL＋ and LDL-. We can therefore conclude that the LDL extender can be used to refrigerate semen at 4 ~C instead of TEG and TE at 80×10^6and 240×10^6 sperm per ml for elite bulls. This finding can be used to define a policy for the storage of high-quality bull semen.

Lipids, Lipoproteins and Apolipoproteins Abnormalities inPatients undergoing Dialysis

Twenty hemodialysis (HD) patients and 20 patients on continuous am-bulatory peritoneal dialysis (CAPD) were investigated for lipids, lipoproteins andapolipoproteins abnormalities. HD patients had elevated serum triglyceride, de-creased high-density lipoprotein cholesterol (HDL-C ) and apolipoprotein A-I(Apo A-I ), whi1e CAPD patients had elevated total cho1esterol, triglyceride,low-density lipoprotein cholesterol (LDL-C), Apolipoprotein B (Apo B), Apo B/Apo A-Iratio, and decreased HDL-C, Apo A-I. Because of the molecular sievingeffects of peritoneum, CAPD have a negative effect on these abnormalities. CAPDpatients might be at greater risk of developing coronary artery disease than HD patients who are also at increased riskas compared with normals.

Carbamylated lipoproteins in diabetes

Diabetic dyslipidemia is characterized by quantitative and qualitative abnormalities in lipoproteins.In addition to glycation and oxidation,carbamylation is also a post-translational modification affecting lipoproteins in diabetes.Patients with type 2 diabetes(T2D)exhibit higher levels of carbamylated low-density lipoproteins(cLDL)and high-density lipoproteins(cHDL).Accumulating evidence suggests that cLDL plays a role in atherosclerosis in diabetes.cLDL levels have been shown to predict cardiovascular events and all-cause mortality.cLDL facilitates immune cell recruitment in the vascular wall,promotes accumulation of lipids in macrophages,and contributes to endothelial dysf-unction,endothelial nitric oxide-synthase(eNOS)inactivation and endothelial repair defects.Lastly,cLDL induces thrombus formation and platelet aggregation.On the other hand,recent data have demonstrated that cHDL serum level is independently associated with all-cause and cardiovascular-related mortality in T2D patients.This relationship may be causative since the atheroprotective properties of HDL are altered after carbamylation.Thus,cHDL loses the ability to remove cholesterol from macrophages,to inhibit monocyte adhesion and recruitment,to induce eNOS activation and to inhibit apoptosis.Taken together,it seems very likely that the abnormalities in the biological functions of LDL and HDL after carbamylation contribute to atherosclerosis and to the elevated cardiovascular risk in diabetes.

ApoB-containing lipoproteins promote infectivity of chlamydial species in human hepatoma cell line

AIM:To evaluate the direct binding of two main chlamydial biovars(C.trachomatis and C.pneumoniae) to plasma lipoproteins and its effect on chlamydial infection rate in human hepatoma cell line(HepG2 cells). METHODS:Murine plasma lipoproteins were fractionated and isolated using fast-performance liquid chromatography(FPLC),spotted on nitrocellulose membrane and incubated with chlamydial suspensions. Direct binding of chlamydial particles to lipoprotein fractions has been studied using lipopolysaccharide-specific antibodies in immuno-dot blot binding assay and immunoprecipitation analysis.Immunostaining protocol as well as flow cytometry analysis have been employed to study the infectivity rate of chlamydial species in HepG2 cells. RESULTS:Elementary bodies of both C.trachomatis and C.pneumoniae bind ApoB-containing fractions of plasma lipoproteins.That binding becomes stronger when heat-denatured FPLC fractions are used, suggesting a primary role of apolipoproteins in interaction between chlamydial particle and lipoprotein. Both chlamydial biovars efficiently propagate in human hepatoma cell line-HepG2 cells even in serum free conditions forming late-stage inclusion bodies and releasing extracellular elementary bodies.Preincubation of C.trachomatis and C.pneumoniae with native ApoB-containing lipoproteins enhances the rate of chlamydial infection in HepG2 cells.CONCLUSION:A productive infection caused by C. trachomatis and C.pneumoniae may take place in human-derived hepatocytes revealing hepatic cells as possible target in chlamydial infection.Obtained results may suggest the participation of lipoprotein receptors in the mechanism of attachment and/or entry of chlamydial particles into target cells.