Role of PERK/eIF2α/CHOP Endoplasmic Reticulum Stress Pathway in Oxidized Low-density Lipoprotein Mediated Induction of Endothelial Apoptosis

Objective PERK/elF2/CHOP is a major signaling pathway mediating endoplasmic reticulum （ER） stress related with atherosclerosis. Oxidized LDL （ox-LDL） also induces endothelial apoptosis and plays a vital role in the initiation and progression of atherosclerosis. The present study was conducted to explore the regulatory effect of ox-LDL on PERK/elF2a/CHOP signaling pathway in vascular endothelial cells. Methods The effects of ox-LDL on PERK and p-elF2a protein expression of primary human umbilical vein endothelial cells （HUVECs） were investigated by Western blot analysis. PERK gene silencing and selective elF2a phosphatase inhibitor, salubrinal were used to inhibit the process of ox-LDL induced endothelial cell apoptosis, caspase-3 activity, and CHOP mRNA level. Results Ox-LDL treatment significantly increased the expression of PERK, PERK-mediated inactivation of elF2a phosphorylation, and the expression of CHOP, as well as the caspase-3 activity and apoptosis. The effects of ox-LDL were markedly decreased by knocking down PERK with stable transduction of lentiviral shRNA or by selective elF2a phosphatase inhibitor, salubrinal. Conclusion This study provides the first evidence that ox-LDL induces apoptosis in vascular endothelial cells mediated largely via the PERK/elF2a/CHOP ER-stress pathway. It adds new insights into the molecular mechanisms underlying the pathogenesis and progression of atherosclerosis.

Oxidized low-density lipoprotein receptor 1:a novel potential therapeutic target for intracerebral hemorrhage

Oxidized low-density lipoprotein receptor 1(OLR1)is upregulated in neurons and participates in hypertension-induced neuronal apoptosis.OLR1 deletion exerts protective effects on cerebral damage induced by hypertensive-induced stroke.Therefore,OLR1 is likely involved in the progress of intracerebral hemorrhage.In this study,we examined the potential role of OLR1 in intracerebral hemorrhage using a rat model.OLR1 small interfering RNA(10μL;50 pmol/μL)was injected into the right basal ganglia to knock down OLR1.Twenty-four hours later,0.5 U collagenase type VII was injected to induce intracerebral hemorrhage.We found that knockdown of OLR1 attenuated neurological behavior impairment in rats with intracerebral hemorrhage and reduced hematoma,neuron loss,inflammatory reaction,and oxidative stress in rat brain tissue.We also found that silencing of OLR1 suppressed ferroptosis induced by intracerebral hemorrhage and the p38 signaling pathway.Therefore,silencing OLR1 exhibits protective effects against secondary injury of intracerebral hemorrhage.These findings suggest that OLR1 may be a novel potential therapeutic target for intracerebral hemorrhage.

A modified laser-induced choroidal neovascularization animal model with intravitreal oxidized low-density lipoprotein

AIM: To investigate whether intravitreal injection of oxidized low-density lipoprotein(OxLDL) can promote laserinduced choroidal neovascularization(CNV) formation in mice and the mechanism involved, thereby to develop a better animal model.METHODS: C57BL6/J mice were randomized into three groups. Immediately after CNV induction with 532 nm laser photocoagulation, 1.0 μL of OxLDL [100 μg/m L in phosphate-buffered saline(PBS)] was intravitreally injected, whereas PBS and the same volume low-density lipoprotein(LDL;100 μg/m L in PBS) were injected into the vitreous as controls. Angiogenic and inflammatory cytokines were measured by quantitative real-time polymerase chain reaction(q RT-PCR) and Western blotting(WB) after 5 d, and CNV severity was analyzed by choroid flat mount and immunofluorescence staining after 1wk. In vitro, retinal pigment epithelial(RPE) cell line(ARPE19) were treated with OxLDL(LDL as control) for 8 h. Angiogenic and inflammatory cytokine levels were measured. A specific inhibitor of lectin-like oxidized low-density lipoprotein receptor 1(LOX1) was used to evaluate the role of LOX1 in this process.RESULTS: At 7 d after intravitreal injection of 1 μL(100 μg/mL) OxLDL, T15-labeled OxLDL was mainly deposited around the CNV area, and the F4/80-labeled macrophages, the CD31-labeled vascular endothelial cells number and CNV area were increased. Meanwhile, WB and qR T-PCR results showed that vascular endothelial growth factor(VEGF), CC chemokine receptor 2(CCR2), interleukin-6(IL-6), IL-1β, and matrix metalloproteinase 9(MMP9) expressions were increased, which was supported by in vitro experiments in RPE cells. LOX1 inhibitors significantly reduced expressions of inflammatory factors IL-1β and VEGF. CONCLUSION: A modified laser-induced CNV animal model is established with intravitreal injection of 1 μL(100 μg/mL) of OxLDL at 7 d, which at least partially through LOX1. This animal model can be used as a simple model for studying the role of OxLDL in age-related macular degeneration.

Oxidized low-density lipoprotein stimulates CD206 positive macrophages upregulating CD44 and CD133 expression in colorectal cancer with high-fat diet

BACKGROUND Oxidized low-density lipoprotein(ox-LDL),which is abnormally increased in the serum of colorectal cancer(CRC)patients consuming a high-fat diet(HFD),may be one of the risk factors for the development of CRC.Ox-LDL exerts a regulatory effect on macrophages and may influence CRC through the tumor microenvironment.The role of ox-LDL in CRC remains unclear.AIM To investigate the role of ox-LDL through macrophages in HFD associated CRC.METHODS The expression of ox-LDL and CD206 was detected in colorectal tissues of CRC patients with hyperlipidemia and HFD-fed mice by immunofluorescence.We stimulated the macrophages with 20μg/mL ox-LDL and assessed the expression levels of CD206 and the cytokines by cell fluorescence and quantitative polymerase chain reaction.We further knocked down LOX-1,the surface receptor of ox-LDL,to confirm the function of ox-LDL in macrophages.Then,LoVo cells were co-cultured with the stimulated macrophages to analyze the CD44 and CD133 expression by western blot.RESULTS The expression of ox-LDL and the CD206 was significantly increased in the stroma of colorectal tissues of CRC patients with hyperlipidemia,and also upregulated in the HFD-fed mice.Moreover,an increased level of CD206 and decreased level of inducible nitric oxide synthase were observed in macrophages after ox-LDL continuous stimulation.Such effects were inhibited when the surface receptor LOX-1 was knocked down in macrophages.Ox-LDL could induce CD206+macrophages,which resulted in high expression of CD44 and CD133 in co-cultured LoVo cells.CONCLUSION Ox-LDL stimulates CD206+macrophages to upregulate CD44 and CD133 expression in HFD related CRC.

Effects of Pitavastatin on Lipoprotein Subfractions and Oxidized Low-density Lipoprotein in Patients with Atherosclerosis

It has been demonstrated that pitavastatin can significantly reduce low-density lipoprotein(LDL)cholesterol(LDL-C),but its impact on lipoprotein subfractions and oxidized low-density lipoprotein(oxLDL)has not been determined.The aim of the present study was to investigate the potential effects of pitavastatin on subfractions of LDL and high-density lipoprotein(HDL)as well as oxLDL in untreated patients with coronary atherosclerosis(AS).Thirty-six subjects were enrolled in this study.O f them,18 patients with AS were administered pitavastatin 2 mg/day for 8 weeks and 18 healthy subjects without therapy served as controls.The plasma lipid profile,lipoprotein subfractions and circulating oxLDL were determined at baseline and 8 weeks respectively.The results showed that pitavastatin treatment indeed not only decreased LDL-C,total cholesterol(TC),triglycerides(TG)and apolipoprotein B(ApoB)levels,and increased HDL cholesterol(HDL-C),but also reduced the cholesterol concentration of all of the LDL subfractions and the percentage of intermediate and small LDL subfractions.Meanwhile,pitavastatin could decrease plasma oxLDL levels.Furthermore,a more close correlation was found between oxLDL and LDL-C as well as LDL subfractions after pitavastatin treatment.We concluded that a moderate dose of pitavastatin therapy not only decreases LDL-C and oxLDL concentrations but also improves LDL subfractions in patients with AS.

Low-density lipoprotein receptor-related protein 2(LRP2)is required for lipid export in the midgut of the migratory locust,Locusta migratoria

Low-density lipoprotein receptor-related protein 2(LRP2)is a multifunctional endocytic receptor expressed in epithelial cells.In mammals,it acts as an endocytic receptor that mediates the cellular uptake of cholesterol-containing apolipoproteins to maintain lipid homeostasis.However,little is known about the role of LRP2 in lipid homeostasis in insects.In the present study,we investigated the function of LRP2 in the migratory locust Locusta migratoria(LmLRP2).The mRNA of LmLRP2 is widely distributed in various tissues,including integument,wing pads,foregut,midgut,hindgut,Malpighian tubules and fat body,and the amounts of LmLRP2 transcripts decreased gradually in the early stages and then increased in the late stages before ecdysis during the nymphal developmental stage.Fluorescence immunohistochemistry revealed that the LmLRP2 protein is mainly located in cellular membranes of the midgut and hindgut.Using RNAi to silence LmLRP2 caused molting defects in nymphs(more than 60%),and the neutral lipid was found to accumulate in the midgut and surface of the integument,but not in the fat body,of dsLmLRP2-treated nymphs.The results of a lipidomics analysis showed that the main components of lipids(diglyceride and triglyceride)were significantly increased in the midgut,but decreased in the fat body and hemolymph.Furthermore,the content of total triglyceride was significantly increased in the midgut,but markedly decreased in the fat body and hemolymph in dsLmLRP2-injected nymphs.Our results indicate that LmLRP2 is located in the cellular membranes of midgut cells,and is required for lipid export from the midgut to the hemolymphand fat body in locusts.

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.

Lectin-like oxidized low-density lipoprotein receptor-1:protein,ligands,expression and pathophvsiological significance

Objective To review the recent research progress in lectin-like oxidized low-density lipoprotein receptor-1 （LOX-1） including its protein, ligands, expression and pathophysiological significance. Data sources Information included in this article was identified by searching of PUBMED （1997-2006） online resources using the key term LOX-1. Study selection Mainly original milestone articles and critical reviews written by major pioneer investigators of the field were selected. Results The key issues related to the LOX-1 protein as well as ligands for LOX-1. Factors regulating the expression of LOX-1 were summarized. The pathophysiological functions of LOX-1 in several diseases were discussed. Conclusions Identification of LOX-1 and a definition of its biological role in pathophysiologic states provide deeper insight into the pathogenesis of some cardiovascular diseases especially in atherosclerosis and provide a potential selective therapeutic approach. LOX-1 is unlocking and drugs targeting LOX-1 might be a promising direction to explore.

Inhibitory Effects of Simvastatin on Oxidized Low-Density Lipoprotein-lnduced Endoplasmic Reticulum Stress and Apoptosis in Vascular Endothelial Cells

Background：Oxidized low-density lipoprotein （ox-LDL）-induced oxidative stress and endothelial apoptosis are essential for atherosclerosis. Our previous study has shown that ox-LDL-induced apoptosis is mediated by the protein kinase RNA-like endoplasmic reticulum kinase （PERK）/eukaryotic translation initiation factor 2α-subunit （eIF2α）/CCAAT/enhancer-binding protein homologous protein （CHOP） endoplasmic reticulum （ER） stress pathway in endothelial cells. Statins are cholesterol-lowering drugs that exert pleiotropic effects including suppression of oxidative stress. This study aimed to explore the roles of simvastatin on ox-LDL-induced ER stress and apoptosis in endothelial cells.Methods：Human umbilical vein endothelial cells （HUVECs） were treated with simvastatin （0.1, 0.5, or 2.5 μmol/L） or DEVD-CHO （selective inhibitor of caspase-3, 100 μmol/L） for 1 h before the addition of ox-LDL （100 μg/ml） and then incubated for 24 h, and untreated cells were used as a control group. Apoptosis, expression of PERK, phosphorylation of eIF2α, CHOP mRNA level, and caspase-3 activity were measured. Comparisons among multiple groups were performed with one-way analysis of variance （ANOVA） followed by post hoc pairwise comparisons using Tukey’s tests. A value of P 〈 0.05 was considered statistically significant.Results：Exposure of HUVECs to ox-LDL resulted in a significant increase in apoptosis （31.9% vs. 4.9%, P 〈 0.05）. Simvastatin （0.1, 0.5, and 2.5 μmol/L） led to a suppression of ox-LDL-induced apoptosis （28.0%, 24.7%, and 13.8%, F = 15.039, all P 〈 0.05, compared with control group）. Ox-LDL significantly increased the expression of PERK （499.5%, P 〈 0.05） and phosphorylation of eIF2α （451.6%, P 〈 0.05）, if both of which in the control groups were considered as 100%. Simvastatin treatment （0.1, 0.5, and 2.5 μmol/L） blunted ox-LDL-induced expression of PERK （407.8%, 339.1%, and 187.5%, F = 10.121, all P 〈 0.05, compared with control group） and phosphorylation of eIF2α （407.8%, 339.1%, 187.5%, F = 11.430, all P 〈 0.05, compared with control group）. In contrast, DEVD-CHO treatment had no significant effect on ox-LDL-induced expression of PERK （486.4%） and phosphorylation of eIF2α （418.8%）. Exposure of HUVECs to ox-LDL also markedly induced caspase-3 activity together with increased CHOP mRNA level; these effects were inhibited by simvastatin treatment.Conclusions：This study suggested that simvastatin could inhibit ox-LDL-induced ER stress and apoptosis in vascular endothelial cells.

Aqueous Extracts of Tribulus terrestris Protects Against Oxidized Low-Density Lipoprotein-Induced Endothelial Dysfunction

Objective： To investigate the role of aqueous extracts of Tribulus terrestris （TT） against oxidized low-density lipoprotein （ox-LDL）-induced human umbilical vein endothelial cells （HUVECs） dysfunction in vitro. Methods： HUVECs were pre-incubated for 60 min with TT （30 and 3 μg/mL respectively） or 105 mol/L valsartan （as positive controls） and then the injured endothelium model was established by applying 100 μg/mL ox-LDL for 24 h. Cell viability of HUVECs was observed by real-time cell electronic sensing assay and apoptosis rate by Annexin V/PI staining. The cell migration assay was performed with a transwell insert system. Cytoskeleton remodeling was observed by immunofluorescence assay. The content of endothelial nitric oxide synthase （eNOS） was measured by enzyme-linked immunosorbent assay. Intracellular reactive oxygen species （ROS） generation was assessed by immunofluorescence and flow cytometer. Key genes associated with the metabolism of ox-LDL were chosen for quantitative real-time polymerase chain reaction to explore the possible mechanism of TT against oxidized LDL-induced endothelial dysfunction. Results： TT suppressed ox-LDL-induced HUVEC proliferation and apoptosis rates significantly （41.1% and 43.5% after treatment for 3 and 38 h, respectively; P〈0.05）. It also prolonged the HUVEC survival time and postponed the cell＇s decaying stage （from the 69th h to over 100 h）. According to the immunofluorescence and transwell insert system assay, TT improved the endothelial cytoskeletal network, and vinculin expression and increased cell migration. Additionally, TT regulated of the synthesis of endothelial nitric oxide synthase and generation of intracellular reactive oxygen species （P〈0.05）. Both 30 and 3μg/mL TT demonstrated similar efficacy to valsartan. TT normalized the increased mRNA expression of PI3Kα and Socs3. It also decreased mRNA expression of Aktl, AMPKα, JAK2, LepR and STAT3 induced by ox-LDL. The most notable changes were JAK2, LepR, PI3Kα, Socs3 and STAT3. Conclusions： TT demonstrated potential lowering lipid benefits, anti-hypertension and endothelial protective effects. It also suggested that the JAK2/STAT3 and/or PI3K/AKT pathway might be a very important pathway which was involved in the pharmacological mechanism of TT as the vascular protective agent.

Protective Effect of Propyl Gallate Against Oxidized Low-Density Lipoprotein-Induced Injury of Endothelial Cells

Objective： To evaluate the protective effect of propyl gallate （PG）, an alkyl ester of gallic acid which is an active ingredient of Radix Paeoniae, against oxidized low-density lipoprotein （ox-LDL）-induced apoptosis and death in endothelial cells （ECs） and to find out its preliminary mechanism. Methods： The cultured endothelial cells were divided into normal, model （ox-LDL）, control （fetal bovine serum）, PG high dose （20 p,g/mL）, PG middle dose （10 μg/mL）, and PG low dose （5 μg/mL） groups, each derived from three different pools of umbilical cords. The model of injured human umbilical vein endothelial cells （HUVECs） was induced by ox-LDL. The 3-（4,5-dimethylthiazol-2-yl）-2,5-diphenyl-2H-tetrazolium bromide （MTT） assay, Hoechst 33258 staining, flow cytometry and measurement of nitrogen monoxidum （NO） release were used to evaluate the protective effect of PG against ox-LDL-induced apoptosis and death in HUVECs. To find out the mechanism of this protective effect, the expression of endothelial nitric oxide synthase （eNOS） mRNA, eNOS protein expression, immunofluorescence of intracellular reactive oxygen species （ROS） and activities of malondialdehyde （MDA）, superoxidedismutase （SOD） and glutathione peroxidase （GPx） were observed. Results： PG significantly reduced ox-LDL-induced apoptosis and cell death. The percentage of cells death and apoptosis was significantly higher in the ox-LDL group than that in the control group （P〈0.05）. Compared with the control group, the cells death and apoptosis of PG group was no different （P〉0.05）. As compared with the ox-LDL group, results of the PG high dose group showed that cell viability was significantly increased （P〈0.05）, the level of NO release, expression of eNOS mRNA, densitometric value of eNOS protein expression, as well as the activities of SOD and GPx were all significantly higher （all P〈0.05）. Conclusion： PG could potentially serve as a novel endothelial protective agent against ox-LDL-induced injury of endothelial cell.

Metformin improved oxidized low-density lipoprotein-impaired mitochondrial function and increased glucose uptake involving Akt-AS160 pathway in raw264.7 macrophages

Background:Macrophage accumulation in the vascular wall is a hallmark of atherosclerosis.Studies showed that shifting of oxidized lipids-induced inflammatory macrophages towards an anti-inflammatory phenotype by promoting oxidative metabolism attenuated atherosclerosis progression.Therefore,this study aimed to investigate whether metformin,which has ameliorated atherosclerosis in animal models and clinical trials,modulated oxidized low-density lipoprotein (Ox-LDL) induced inflammatory status in macrophages by regulating cellular oxidative metabolism.Methods:Murine raw264.7 macrophages were incubated with Ox-LDL (50 μg/mL) in the presence or absence of metformin (15 μmol/L) for 24 h.Real-time polymerase chain reaction was used to quantify the transcription of classically activated (M1) proinflammatory and alternatively activated (M2) anti-inflammatory markers and mitochondrial DNA copy numbers.Cellular reactive oxygen species (ROS) production and mitochondrial membrane potential were detected by immunofluorescence.Cellular adenosine triphosphate (ATP) synthesis,glucose uptake,and lactic acid production were measured by commercial kit and normalized to cellular lysates.Western blotting analysis was performed to detect the expression of mitochondrial fusion/fission related proteins,enzymes mediating lipid metabolism and signaling pathway of glucose transport.Differences between groups were analyzed using one-way analysis of variance.Results:Metformin improved Ox-LDL-impaired anti-inflammatory phenotype in raw264.7 macrophages as shown by up-regulated transcription of anti-inflammatory markers including interleukin 10 (0.76 ± 0.04 vs.0.94 ± 0.01,P =0.003) and Resistin-like molecule alpha (0.67 ± 0.08 vs.1.78 ± 0.34,P =0.030).Conversely,Ox-LDL-diminished phosphorylation of Akt was up regulated by metformin treatment (0.47 ± 0.05 vs.1.02 ± 0.08,P =0.040),associated with an improvement of mitochondrial function,characterized by decreased ROS generation (2.50 ± 0.07 vs.2.15 ± 0.04,P =0.040),increased lipid oxidation,and elevated cellular ATP production (0.026 ± 0.001 vs.0.035 ± 0.003,P =0.020).Moreover,metformin-mediated Akt activation increased Akt substrate of 160 kDa (AS160) phosphorylation (0.51 ± 0.04 vs.1.03 ± 0.03,P =0.0041),promoted membrane translocation of glucose transporter 1,and increased glucose influx into the cells (4.78 ± 0.04 vs.5.47 ± 0.01,P < 0.001).Contusion:This study suggested that targeting macrophage metabolism with new or existing drugs had therapeutic potential for the prevention and treatment of diabetes-accelerated atherosclerosis.

Danshen protects endothelial progenitor cells from oxidized low-density lipoprotein induced impairment

In this study,we examined the protective effects of Danshen both on endothelial progenitor cells(EPCs) in patients with hypercholesterolemia and on in-vitro EPCs of healthy volunteers.In the clinical study,we randomly divided 24 subjects with hypercholesterolemia into two groups(the control group and the Danshen-treated group).At the end of two weeks of treatment,the EPC cellular functions of both groups were tested.The results indicated that,compared to the control group,EPCs in the Danshen-treated group showed significantly better cellular functions,which was manifested in the cloning number,the proliferation capacity,the number of EPC adhesions,and cell migration.In the subsequent in-vitro experiments,EPCs were treated with vehicle,oxidized low-density lipoprotein(Ox-LDL,100 μg/ml),or Ox-LDL(100 μg/ml) plus different concentrations of Danshen(Danshensu 2,10,or 50 μg/ml,respectively) for 24 h.The results showed that Danshen treatments can prevent the detrimental effects of Ox-LDL on EPC cellular functions measured by proliferation capacity(0.24±0.08,0.37±0.11,0.30±0.04 vs.0.13±0.02,P<0.05,P<0.01,and P<0.01,respectively),and adhesion ability(63.00±11.60,70.00±10.80,85.50±11.41 vs.40.50±6.85,all P<0.01).Compared to the group treated with Ox-LDL alone,Danshen treatment significantly decreased the lipid peroxidation end product malondialdehyde(MDA) [(4.34±0.54),(3.98±0.47),(3.46±0.31) vs.(5.57±0.64) nmol/ml,all P<0.01],increased the production of superoxide dismutase(SOD) [(29.74±0.71),(31.09±0.83),(30.41±0.65) vs.(14.76±3.99) U/ml,all P<0.01],and lowered the expression of interleukin-6(IL-6) [(24.62±7.69),(27.04±3.14),(33.38±18.86) vs.(230.67±33.53) pg/ml,all P<0.01] and tumor necrosis factor-α(TNF-α) [(41.72±6.10),(17.02±6.82),(3.73±2.26) vs.(228.71±41.53) pg/ml,all P<0.01] in Ox-LDL treated EPCs.These results suggest that Danshen may exert a protective effect through its antioxidant and anti-inflammatory features.

Association between moderately oxidized low-density lipoprotein and high-density lipoprotein particle subclass distribution in hemodialyzed and post-renal transplant patients

Disturbances in the metabolism of lipoprotein profiles and oxidative stress in hemodialyzed(HD) and post-renal transplant(Tx) patients are proatherogenic,but elevated concentrations of plasma high-density lipoprotein(HDL) reduce the risk of cardiovascular disease.We investigated the concentrations of lipid,lipoprotein,HDL particle,oxidized low-density lipoprotein(ox-LDL) and anti-ox-LDL,and paraoxonase-1(PON-1) activity in HD(n=33) and Tx(n=71) patients who were non-smokers without active inflammatory disease,liver disease,diabetes,or malignancy.HD patients had moderate hypertriglyceridemia,normocholesterolemia,low HDL-C,apolipoprotein A-I(apoA-I) and HDL particle concentrations as well as PON-1 activity,and increased ox-LDL and anti-ox-LDL levels.Tx patients had hypertriglyceridemia,hypercholesterolemia,moderately decreased HDL-C and HDL particle concentrations and PON-1 activity,and moderately increased ox-LDL and anti-ox-LDL levels as compared to the reference,but ox-LDL and anti-ox-LDL levels and PON-1 activity were more disturbed in HD patients.However,in both patient groups,lipid and lipoprotein ratios(total cholesterol(TC)/HDL-C,LDL-C/HDL-C,triglyceride(TG)/HDL-C,HDL-C/non-HDL-C,apoA-I/apoB,HDL-C/apoA-I,TG/HDL) were atherogenic.The Spearman's rank coefficient test showed that the con-centration of ox-LDL correlated positively with HDL particle level(R=0.363,P=0.004),and negatively with TC(R=?0.306,P=0.012),LDL-C(R=?0.283,P=0.020),and non-HDL-C(R=?0.263,P=0.030) levels in Tx patients.Mul-tiple stepwise forward regression analysis in Tx patients demonstrated that ox-LDL concentration,as an independent variable,was associated significantly positively with HDL particle level.The results indicated that ox-LDL and de-creased PON-1 activity in Tx patients may give rise to more mildly-oxidized HDLs,which are less stable,easily un-dergo metabolic remodeling,generate a greater number of smaller pre-β-HDL particles,and thus accelerate reverse cholesterol transport,which may be beneficial for Tx patients.Further studies are necessary to confirm this.

The effects of fucoidans from Laminaria japonica on AAPH mediated oxidation of human low-density lipoprotein

Five fucoidan fractions from Laminaria japonica with different sulfate content and molecular weight were prepared by anion-exchange chromatography and mild acid hydrolysis. Their antioxidant effects on azo radicals 2-2＇-Azobis （2-amidinopropane） dihydrochloride （AAPH） induced oxidation of human low-density lipoprotein （LDL） were evaluated by monitoring cholesteryl ester hydroperoxides （CHL-OOH） formation kinetics through reverse-phase high performance liquid chromatography （RP-HPLC） analysis. Fucoidan F - C with a low molecular mass of 2 000 ～8 000 and a sulfate content of 24.3% had much stronger protective antioxidant effect than other four fucoidan fractions on the hydrophilic radical AAPH induced LDL oxidation. However, the highly sulfated fucoidan fraction L - B with a molecular mass of 20 000 was completely ineffective in protecting LDL from the AAPH induced oxidation. The results suggested that both molecular mass and sulfate content of fucoidan played very important roles in their effects on the AAPH induced LDL oxidation.

C1q/TNF-related protein 5 promotes atherogenesis by enhancing transcytosis and oxidative modification of low-density lipoprotein through increasing 12/15-lipoxygenase

Objective Increased transcytosis of low-density lipoprotein (LDL)across the endothelium and oxidation of LDL deposited within the subendothelial space are crucial early events in atherogenesis. C1q/TNF-related protein (CTRP) 5 is a novel secreted glycoprotein and its biological functions are largely undefined.

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 gradient 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 160μg/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 atherosclerotic plaque, ox-LDL at a concentration of 35 μg/mL demonstrated the strongest proliferation inducement, 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 concentrations 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.

Association between Low-density Lipoprotein Receptor-related Protein 5 Polymorphisms and Type 2 Diabetes Mellitus in Han Chinese:a Case-control Study

Objective To investigate the association between low-density lipoprotein receptor-related protein 5 （LRPS） variants （rs12363572 and rs4930588） and type 2 diabetes mellitus （T2DM） in Han Chinese. Methods A total of 1842 T2DM cases （507 newly diagnosed cases and 1335 previously diagnosed cases） and 7777 controls were included in this case-control study. PCR-RFLP was conducted to detect the genotype of the two single nucleotide polymorphisms （SNPs）. Odds ratios （ORs） and 95% confidence intervals （95% CIs） were calculated to describe the strength of the association by logistic regression. Results In the study subjects, neither rs12363572 nor rs4930588 was significantly associated with T2DM, even after adjusting for relevant covariates. When stratified by body mass index （BMI）, the two SNPs were also not associated with T2DM. Among the 3 common haplotypes, only haplotype ~ was associated with reduced risk of T2DM （OR 0.820, 95% CI 0.732-0.919）. In addition, rs12363572 was associated with BMI （P〈0.001） and rs4930588 was associated with triglyceride levels （P=0.043） in 507 newly diagnosed T2DM cases but not in healthy controls. Conclusion No LRP5 variant was found to be associated with T2DM in Han Chinese, but haplotype TT was found to be associated with T2DM.

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.