Age-related decrease in expression of peroxisome proliferator-activated receptor α and its effects on development of dyslipidemia

Background Ageing is associated with increased incidence of dyslipidemia. To investigate potential molecular mechanisms, the effects of age and fibrate administration on peroxisome proliferator-activated receptor α(PPARα) expression in livers of young and old rats were studied.Methods A total of 16 young (2-month-old) and 16 old rats (24-month-old) were randomly assigned to a control group and fenofibrate group (fenofibrate in a total therapeutic dosage of 0.5% in ratio to each treated rat weight in 14 days). RT-PCR was applied to evaluate hepatic mRNA expression of PPARα and its target genes. Western blotting was used to determine PPARα protein level in liver tissue. Results When compared with 2-month-old rats, the liver tissue from 24-month-old rats showed reduced expression of PPARα mRNA (52%, P<0.05) and protein (109%, P<0.01). Consequently, the mRNA levels of PPAR target genes, LPL, ACO, ACS and CPT-1 were markedly lowered by 19%, 8%, 13% and 9% respectively, and apoCIII increased by 24% in livers from 24-month-old rats, compared with values obtained from 2-month-old rats (P<0.05). Fenofibrate therapy significantly lowered plasma triglyceride and total cholesterol levels in old rats, accompanied with improvement in hepatic expression of genes, including LPL, ACO, ACS, CPT-1 and apoCIII, but no change was found in PPARα expression in livers from either 24-month or 2-month-old rats. Conclusions The decrease in the hepatic PPARα expression is probably directly related to the lipid metabolic disturbances observed in old animals. The beneficial effects of fenofibrate administration in old rats suggests that fibrates may be useful for treating lipid disturbances in old people.     

THE DIFFERENCES OF SERUM CHOLESTEROL AND LIPOPROTEINS IN ANIMALS SUSCEPTIBLE AND NONSUSCEPTIBLE TO ATHEROSCLEROSIS

The differences of serum lipid and lipoprotein (LP,profiles of animals susceptible (rabbit)and nonsusceptible (Beijing duck) to atherosclerosis as well as the distribution of apolipo-protein (apo) A-I and its catabolism in vivo were studied.Eight items,i.e.total cholesterol(TC),high density lipoprotein (HDL) cholesterol,triglyceride,percentage of a-LP and β-LP,β-LP concentration,lecithin cholesterol acyl transferase (LCAT),and agarose and polyacryla-mide gel electrophoresis of both species were assayed before and after feeding a high fat,high cholesterol diet.Results indicate that the exogenous cholesterol consumed by Beijingducks was carried and transported by HDL,while that in rabbits was transported by lowdensity lipoprotein (LDL).The biological half lives of apo A-I in serum and in HDL were41.11±2.4 and 42.8±1.7 h respectively.and its distribution in different organs was in theorder of liver>kidney>spleen>lung>heart>intestine>muscles>aorta.These resultsshow that the liver is the major organ for metabolizing HDL apo A-I.and the kidneyis also important.The results also imply that in Beijing ducks the cholesterol carried byHDL may be catabolized through apo A-I receptors in the liver and kidney.The differencesin cholesterol binding and in lipoprotein and apolipoprotein metabolism of the two speciesprovide important clues for the elucidation of the pathogenesis of atherosclerosis.     

THE EFFECT OF RADIX SALVIAE MILTIORRHIZAE COMPOSITA AND LIGUSTRAZINE ON LIPID PEROXIDATION IN LOW DENSITY LIPOPROTEIN DUE TO COPPER DICHLORIDE

It is well known that lipid peroxide in low density lipoprotein (LDL) plays an important role in atherosclerosis and atherosclerotic diseases. We used oxidized LDL generated by incu-bating LDL from healthy persons with copper dichloride as a model to investigate the anti-lipid-peroxide property of Radix Salviae Miltiorrhizae Composita (RSMC) and ligustrazine. The changes in concentrations of lipid peroxide and lipids in LDL due to Cu were studied, and the effects of RSMC and ligustrazine on the changes were studied. The re-sults in our study indicate that RSMC has a potential role on and-lipid-peroxidation, but it was not found that ligustrazine has similar anti-peroxidation action.     

STUDIES ON THE ANTI-FATIGUE EFFECT OF THE SAPONINS OF STEMS AND LEAVES OF PANAX GINSENG(SSLG)

SSLG was orally administered to rats dailyfor 6 successive days at a dosage of 100 mg/kgor 200 mg/kg body weight.Swimming experi-ment was then performed,and showed that theswimming time of the SSLG-treated rats wasmuch longer than that of a control group.Further,SSLG significantly raised the level ofblood lipid in rats both at rest and during swim-ming exercise and inhibited the increase ofblood lactic acid as well as the reduction ofthe liver and rectus femoris muscle(RFM)gly-cogens.SSLG also enhanced the amount ofprotein in the liver and RFM in rats at restand during swimming,and the content of RNAin the tissues at rest.However,for swimmingrats of both dosage groups the amounts of RNAand DNA in the liver and RFM show no signi-ficant difference.In vivo,SSLG showed noeffect on the respiratory quotient(RQ)of RFMof rats at rest,though the RQ of SSLG-treatedrats was lowered after 2 hrs of swimming.Invitro,the drug had no effect on RQ.The ex-periments showed the ability of saponins tofacilitate the release of fatty acid from epidi-dymal adipose tissue in rats.It was inferredthat the raising of blood lipid level by SSLGcould be one linkage in its anti-fatigue mechan-ism.     

The new strategy for modulating dyslipidemia: Consideration from updated understanding on high-density lipoprotein

With further researches on blood lipids and atherosclerosis,especially after the conception of"residual cardiovascular risk",high-density lipoprotein cholesterol(HDL-C) becomes a new therapeutic target against atherosclerosis.However,the failure of ILLUMINATE study that was targeted at raising HDL-C level disappointed cardiologists all over the world,which indicates the limitation of HDL-C concentration in representing HDL function.The updated understanding of HDL from"quantity"to"quality"brings a new cut-in point for integrative Chinese and Western medicine in preventing and treating dyslipidemia and atherosclerosis. In addition to highlighting statins in directly intervening low-density lipoprotein cholesterol,we should make full use of the superiority of Chinese medicine in overall regulation and individualized treatment to promote the self-healing capacity of the patients,which further regulates abnormity of both concentration and function of the whole blood lipid family to equilibrium.This new strategy for modulating dyslipidemia with mutual complement of advantages from Western and Chinese medicines will no doubt play an important role in future therapies.     

Protection of salvianolate against atherosclerosis via regulating the inflammation in rats

Inflammation plays an essential role in the pathophysiology of atherosclerosis. Our study was aimed to investigate whether salvianolate, a novel water-soluble phenolic compound of Danshen, alleviates atherosclerosis via regulating the inflammation in rats. High fat diet feeding plus vitamin D3 injection was used to induce atherosclerosis in rats. Salvianolate （60, 120 or 240 mg/kg） or placebo was given to atherosclerotic rats. The plasma lipids, interleukin 6 （IL-6） and C reactive protein （CRP） were measured by ELISA. CD4＋CD25＋Foxp3＋ cells were determined by flow cytometry. Histological changes were examined by hematoxylin and eosin staining. The results showed that the levels of plasma IL-6 and CRP were elevated in the rats fed on high fat diet, and the histological analysis demonstrated the successful establishment of atherosclerosis models. Treatment with salvianolate alleviated the atherosclerotic process and decreased the levels of plasma IL-6 and CRP. Also the number of CD4＋CD25＋Foxp3＋ cells was increased in salvianolate-treated rats. It was concluded that salvianolate could treat atherosclerosis via modulating the inflammation at cytokine and cell levels.     

Influence of Oxidized Low Density Lipoprotein on the Proliferation of Human Artery Smooth Muscle Cells in vitro

The effects of oxidized low density lipoprotein (ox-LDL) on the proliferation of cultured human vascular smooth muscle cells (vSMC) were investigated in vitro. By using NaBr density gra-dient centrifugation, LDL was isolated and purified from human plasma. Ox-LDL was produced from LDL by being incubated with CuSO4. ox-LDL was then added to the culture medium at different concentrations (35, 60, 85, 110, 135 and 160 μg/mL) for 7 days. The influence of ox-LDL on vSMC proliferation was observed in growth curve, mitosis index, and in situ determination of apoptosis. The data were analyzed with SPSS 10.0 software. The results showed that the ox-LDL produced in vitro had a good purity and optimal oxidative degree, which was similar to the intrinsic ox-LDL in athero-sclerotic plaque. ox-LDL at a concentration of 35 μg/mL demonstrated the strongest proliferation in-ducement, and at a concentration of 135 μg/mL, ox-LDL could inhibit the growth of vSMC. ox-LDL at concentrations of 35 and 50 μg/mL presented powerful mitotic trigger, and with the increase of ox-LDL concentration, the mitotic index of vSMC was decreased gradually. ox-LDL at higher con-centrations promoted more apoptotic vSMCs. ox-LDL at lower concentrations triggered proliferation of vSMCs, and at higher concentrations induced apoptosis in vSMCs. ox-LDL played a promotional role in the pathogenesis and development of atherosclerosis by affecting vSMC proliferation and apoptosis.     

Effects of andrographolide on the activation of mitogen activated protein kinases and nuclear factor-κB in mouse peritoneal macrophage-derived foam cells

Objective:To observe the effect of andrographolide on the activation of mitogen-activated protein kinases(MAPKs) and expression of nuclear factor-κB(NF-k B) in macrophage foam cells.Methods:The mouse peritoneal macrophages were cultured in the media in the presence of oxidized low-density lipoprotein (ox-LDL),ox-LDL+andrographolide,or neither(control).The phosphorylation of MAPK molecules(p38MAPK, JNK,ERK1/2) and the expressions of NK-κB p65 were examined by Western blot.Results:As compared with cells in the control group,the expressions of phospho-p38 and NF-κB p65 were increased in the cells cultured with either ox-LDL or ox-LDL+andrographolide(P<0.01),but attenuated significantly in the presence of ox-LDL+ andrographolide when compared with ox-LDL(P<0.05).The phospho-JNK increased in the presence of either ox-LDL or ox-LDL+andrographolide when compared with control cells(P<0.01),but no significant difference existed between ox-LDL and ox-LDL+andrographolide(P>0.05).The expression of phospho-ERK1/2 was increased in the presence of ox-LDL compared with the control cells(P<0.01),but no significant differences existed between the cells cultured in the presence of ox-LDL+andrographolide and the control medium(P>0.05). Conclusions:Andrographolide could inhibit the activation of ERK1/2,p38MAPK and NK-k B induced by ox-LDL in macrophage foam cells,which might be one of its mechanisms in preventing atherosclerosis.     

ROLE OF ANTISENSE AND DECOY OLIGONUCLEOTIDE OF NF-KB IN VESSEL STENOSIS AND NEOINTIMA FORMATION IN BALLOON-INJURED RAT ARTERY

Objective To examine antisense and decoy oligonucleotides of nuclear factor kappa B in vivo effects on intima proliferation and balloon-injured monocytes chemotactic protein-1 （ MCP-1 ） and extraceUular signal regulated kinase-2 （ERK2） κexpression in the carotid artery of rats. Methods Sprague-Dawley rats underwent balloon-dilation injury of the left carotid artery. Rats are divided into 7 groups （ n = 18 ） and each group includes6 time points （6 h, 1, 3, 5, 7, 14 d） （n =3）. Uninjured right carotid artery of the same rat was used as controls. Results In model group, sense group and scramble group, vessel intima area , media area and intima/media ratio increased after 5 d and reached the maximum after 7 d. The effect of antisense plus decoy group on intimal hyperplasia was more obvious than that of antisense group and decoy group alone. MCP-1 mRNA expression was increased expression continuously at 3, 5 and 7 d and decreased at 14 d. Compared with model group, sense group and scramble group, antisense group, decoy group and antisense plus decoy group had lowered MCP-1 mRNA expression in each time point （ P 〈 0. 05 ）. NF-KB p65 was dispersed positive stain 6 h after injury and increased after 1 d and peaked at 7 d, but the protein expression was weak at 14 d. ERK2 protein synthesis increased at I d and reached the peak at 7 d, while protein expression after 14 d was similar to that at 7 d. Treatment of antisense group, decoy group and antisense plus decoy group inhibited protein synthesis more significantly than those of model group, sense group and scramble group（ P 〈 0. 05）. Conclusion NF-KB modulates genes expression and protein synthesis of MCP-1 and ERK2. Celluar proliferation in vessel wall was dynamically changed after balloon angioplasty injury. Antisense and decoy oligonucleotide of NF-KB by local lipofectaraine transfer inhibit NF-KB activating gene modulation and neointimal hyperplasia.     

EFFECT OF OXIDIZED-LDL ON NF-κB NUCLEAR TRANSLOCATION IN AORTIC SMOOTH MUSCLE CELLS ORIGINATED FROM RATS OF DIFFERENT AGES

Objective To investigate the molecular mechanism of atherosclerosis that related to age.Methods Immunohistochemistry staining and Western blot were adopted to determine the nuclear translocation of nuclear factor-kappa B (NF-Κb) and expression of platelet-derived growth factor B (PDGF-B) in smooth muscle cells (SMCs) co-cultured with low density lipoprotein (LDL), oxidized LDL (ox-LDL), and ox-LDL high density lipoprotein (HDL) originated from rats of 2 and 10 months old respectively. Fat stain was used to identify the lipid intake in SMCs.Results The optimal stimulation time ofox-LDL to SMCs was 12 hours. NF-Κb intensity increased in most nuclei of SMCs that originated from rats of either 2 or 10 months old co-cultured with ox-LDL. The intensity of NF-Κb and the amount of intracellular lipid taken in SMCs were more obvious in cells from 10-month-old rats than from the younger ones.Change of PDGF-B expression in SMCs was not remarkable in each group of rats.Conclusions The 10-month-old rats are more susceptive to ox-LDL than 2-month-old rats in activating nuclear translocation of NF-Κb. Maybe this is one of the important reasons contributing to the difference between the older and younger rats on the initiation and development of atherosclerosis lesion. Expression of PDGF-B is not associated with the activity of nuclear translocation of NF-Κb.     

氧化低密度脂蛋白对人血管平滑肌细胞生长状况的影响

目的：探讨氧化低密度脂蛋白（oxidized low density lipoprotein ox-LDL）对培养的人血管平滑肌细胞（smooth muscle cell,SMC）生长状况的影响。方法：利用密度梯度速离心分离纯化人血浆LDL，体外氧化生成ox-LDL,不同浓度的ox-LDL作用于培养的人SMC不同时间，从不同的角度观察其对SMCT生长状况的影响。结果：ox-LDL对SMC的作用具有时间浓度依赖性，低浓度的ox-LDL促SMC增生及表型转换，高浓度时则诱导SMC发生凋亡，甚至死亡。结论：ox-LDL可能在人动脉粥榇硬化的发生发展过程中起重要作用。     

Effects of oxidized low density lipoprotein on the growth of human artery smooth muscle cells

Atherosclerosis(AS)isacommonandbasicpathologicalchangeofmanycardiovasculardiseases.1Amongtheriskfactorscontributingtohuman AS,oxidizedlowdensitylipoprotein(ox LDL)is consideredtostimulateASdirectlyandplaysanovelrole inthepathogenesisanddevelopmentofAS.2,3Despite proliferation,migrationandphenotypealterationof vascularsmoothmusclecells(vSMCs)arecritical changesinAS,4,5itisstillpoorlyunderstoodwhether ox LDLparticipatesinthepathologicalprocessof vSMCs.6WeisolatedandpurifiedhumanLDL,pro…     

外周血ox-LDL浓度与血脂状态关系的研究

目的:观察人外周血氧化型低密度脂蛋白(ox -LDL)浓度与血脂状态的关系。方法:采取病人外周血,酶联免疫吸附试验(ELISA)方法检测血清ox LDL浓度及血总胆固醇(TC)、甘油三酯(TG)、低密度脂蛋白蛋固醇(LDL)、脂蛋白(a) [LP(a) ]的浓度,对检测结果进行相关性分析。结果:ox LDL浓度与TC、LDL的相关系数分别为0 .5 1 6、0 .5 0 4 ,具有统计学意义。外周血ox LDL浓度与TG、LP(a)之间未发现相关性。结论:人外周血ox -LDL浓度与血TC、LDL存在正相关关系。     

坎地沙坦对动脉粥样硬化大鼠血清氧化低密度脂蛋白的影响

目的:探讨坎地沙坦对动脉粥样硬化大鼠血清中氧化低密度脂蛋白(ox-LDL)水平的影响。方法:正常大鼠饲喂高脂饲料建立动脉粥样硬化(AS)大鼠模型。将实验大鼠随机分为正常组(A组)、模型组(B组)、坎地沙坦低(1 mg/kg.d,C组)、中(5 mg/kg.d,D组)、高(10mg/kg.d,E组)剂量组。12周后腹主静脉取血酶联免疫夹心法测定血清中ox-LDL水平;取同部位腹主动脉光镜下观察形态学改变。结果:与B组相比C、D、E组均可以显著降低血清ox-LDL水平(P<0.01)无剂量依赖性;B组与A组相比主动脉出现动脉粥样硬化的形态学改变,与B组相比C、D、E组动脉粥样硬化的形态学改变有所减轻。结论:坎地沙坦可降低动脉粥样硬化大鼠血清中的ox-LDL水平,从而抑制动脉粥样硬化的发生、发展。     

锌指蛋白A20对抑制ox-LDL诱导的平滑肌细胞LOX-1、TLR-4表达的作用

目的 探讨锌指蛋白A20对氧化性低密度脂蛋白（ox-LDL）诱导的血管平滑肌细胞血凝素样氧化低密度脂蛋白受体（LOX-1）和toll样受体-4（TLR-4）表达的作用.方法 体外培养SD大鼠主动脉血管平滑肌细胞（VSMC）,简单随机化分成4组：A组（对照组）;B组（OX-LDL组）;C组（A20组）;D组（ox-LDL＋A20组）.收集以上各组细胞,用RT-PCR检测LOX-1mRNA和TLR-4 mRNA的表达;提取核蛋白,并用western blot的方法检测NF-κ B核易位的量.结果 ox-LDL刺激大鼠主动脉平滑肌细胞后LOX-1mRNA和TLR-4 mRNA的表达明显增加及核因子κ B（NF-κ B）的活化均明显增加.转染含A20基因质粒的平滑肌细胞TLR-4 mRNA的表达达到正常水平,LOX-1 mRNA的表达及NF-κ B的核移位的量甚至降低到正常水平以下.结论 ox-LDL诱导的平滑肌细胞LOX-1mRNA和TLR-4 mRNA表达增加,可能由此激活TLR-4/NF-κB信号系统,启动动脉壁的炎症反应,触发动脉粥样硬化发生.A20明显抑制了ox-LDL对平滑肌细胞的上述诱导作用,可能由此阻断了ox-LDL引发的不断级联扩大的正反馈效应.阻止了动脉粥样硬化的发展.     

内脂素对THP-1源性泡沫细胞形成中过氧化物酶体增殖物激活受体γ表达的作用

目的:观察内脂素(visfatin)诱导人类单核细胞白血病细胞株THP-1源性泡沫细胞形成中过氧化物酶体增殖物激活受体γ(PPARγ)的表达作用。方法:给予不同浓度的visfatin处理THP-1源性巨噬细胞,并加入低密度脂蛋白(LDL)孵育24 h后,运用油红O染色观察细胞内脂滴的变化,运用逆转录聚合酶链反应和Western-blot检测PPARγ的mRNA和蛋白表达水平。结果:高浓度的visfatin刺激负荷LDL的THP-1源性巨噬细胞24 h后,细胞质内的脂滴明显增多。visfatin呈浓度依赖性的下调负荷LDL的THP-1源性巨噬细胞中PPARγmRNA和蛋白表达(P<0.05)。结论:visfatin对THP-1源性泡沫细胞形成中PPARγ表达水平的下调,这可能在动脉粥样硬化的发生发展起重要作用。     

褪黑素对ox-LDL所致巨噬细胞TNF-α释放及核因子κB活性的影响

目的探讨褪黑素对氧化型低密度脂蛋白（ox-LDL）所致的巨噬细胞肿瘤坏死因子α（TNF-α）释放及核因子κB活性的影响。方法以ox-LDL、ox-LDL加不同浓度的褪黑素处理RAW264.7小鼠巨噬细胞,采用酶联免疫吸附测定细胞上清液中TNF-α的浓度,免疫印迹法检测p65/核因子κB的核移位,凝胶电泳迁移率分析核因子κB与DNA的结合活性。结果褪黑素能够显著减少ox-LDL所致的巨噬细胞TNF-α的释放,且p65/核因子κB的核移位减少,核因子κB与DNA的结合活性降低。结论褪黑素减少ox-LDL所致的巨噬细胞TNF-α的释放与其调节核因子κB的活性有关。     

血凝素样氧化低密度脂蛋白受体与内皮细胞p38丝裂素活化蛋白激酶关系的研究

目的探讨血凝素样氧化低密度脂蛋白受体1(LOX-1)在氧化低密度脂蛋白(ox-LDL)活化人脐静脉内皮细胞p38丝裂素活化蛋白激酶(p38MAPK)信号通路中的作用。方法培养人脐静脉内皮细胞(HUVEC),用不同浓度的ox-LDL孵育,通过W estern印迹检测LOX-1、p38MAPK蛋白水平,逆转录聚合酶链法观察LOX-1 mRNA表达,并以LOX-1特异阻断性抗体(JTX92)预处理内皮细胞,检测p38MAPK蛋白水平。四氮唑蓝法观察ox-LDL对细胞活力的影响。结果ox-LDL引起内皮细胞形态结构的改变,而且抑制内皮细胞的生长(P<0.05);ox-LDL呈浓度依赖性地上调LOX-1蛋白和mRNA表达(均P<0.05),并呈浓度依赖性激活p38MAPK通路,不同浓度ox-LDL(25μg/m l,50μg/m l,100μg/m l)组p-p38MAPK蛋白质表达水平均明显高于对照组(48±7,79±15,113±14 vs 24±5,P<0.01);JTX92+ox-LDL(100μg/m l)组p-p38MAPK蛋白水平明显低于ox-LDL(100μg/m l)组,(58±13 vs 113±14,P<0.01)。结论ox-LDL通过LOX-1途径激活p38MAPK信号通路,LOX-1上调是ox-LDL致动脉粥样硬化的重要环节。     

雌激素对去势高脂血症兔血脂及氧化型低密度脂蛋白mRNA表达作用的实验研究

目的　观察雌激素对去势高脂血症兔血脂、氧化型低密度脂蛋白 (ox LDL)水平及ox LDLmRNA表达的影响。方法　采用纯种新西兰雌兔去势术 +0 .5 %胆固醇饲料 +静脉注射牛血清白蛋白造模 ,模型动物连续使用雌激素 12周后检测相关指标。结果　与模型组相比 ,雌激素组ox LDL含量及ox LDLmRNA的表达均明显降低 (P值均 <0 .0 5 )。结论　雌激素可改善去势高脂血症模型兔血脂水平 ,调控ox LDLmRNA的表达 ,减少LDL的氧化修饰 ,保护内皮细胞功能 ,防治绝经后动脉粥样硬化。