PPARγ: the dominant regulator among PPARs in dry eye lacrimal gland and diabetic lacrimal gland

AIM: To determine whether lectin-like ox-LDL receptor (LOX-1) regulates adhesion molecules expression and neutrophil infiltration in A. fumigatus keratitis of C57BL/6 mice.METHODS: C57BL/6 mice were pretreated with a neutralizing antibody to LOX-1 (5 μg/5 μL) or control nonspecific IgG (5 μg/5 μL), LOX-1 inhibitor Poly-I (2 μg/5 μL) or PBS by subconjunctival injection. Fungal keratitis mouse models of C57BL/6 mice were established by scraping corneal central epithelium, smearing A. fumigatus on the corneal surface and covering the eye with contact lenses. The corneal response to infection was assessed via clinical score. The mRNA levels of the adhesion molecules ICAM-1, VCAM-1, P-selectin and E-selectin were tested in control and infected corneas by RT-PCR. The protein levels of ICAM-1 were evaluated by immunofluorescence (IF) and Western blot. Neutrophils were extracted from the abdominal cavity of C57BL/6 mice followed by pretreatment using antibody to LOX-1 (10 μg/mL) or control nonspecific IgG (10 μg/mL), the Poly-I (4 μg/mL) or PBS. The cells were then stimulated with A. fumigatus and tested mRNA and protein levels of LFA-1 using RT-PCR and Western blot. IF and myeloperoxidase (MPO) assays were used to assess neutrophil infiltration in mice corneas.RESULTS: Pretreatment of LOX-1 antibody or the Poly-I reduced the degree of inflammation of cornea and decreased the clinical fungal keratitis score compared with pretreatment of IgG or PBS. And these pretreatment also displayed an obvious decline in the mRNA levels of ICAM-1, VCAM-1, P-selectin, E-selectin and LFA-1 expression compared with control groups . Furthermore, pretreated with LOX-1 antibody or Poly-I, the protein levels of ICAM-1 and LFA-1 also decreased compared with control groups. Neutrophil infiltration in the cornea was significantly reduced after pretreatment of LOX-1 antibody or Poly-I compared with control groups by IF and MPO assays. CONCLUSION: These data provide evidence that inhibition of LOX-1 can decrease the expression of adhesion molecules and thus reduce neutrophil infiltration in A. fumigatus infected corneas of C57BL/6 mice.     

Decreased small dense LDL levels in Gilbert's syndrome

Objective

To investigate the role of small dense low density lipoprotein cholesterol (sd-LDL-C) in the mechanism of decreased incidence of cardiovascular disease in Gilbert's syndrome (GS).

Design and methods

sd-LDL-C, ox-LDL, and high sensitive C reactive protein (hs-CRP) levels were investigated in subjects with GS (n = 42) and compared to healthy controls (n = 52).

Results

Age, gender and body mass index (BMI) distributions were similar between the two groups. sd-LDL-C, ox-LDL and hs-CRP levels were lower in GS than the healthy controls (p < 0.001, p < 0.001 and p = 0.001, respectively). Unconjugated bilirubin was negatively correlated with sd-LDL-C, ox-LDL and hs-CRP (r = −0.594, p < 0.001; r = −0.249, p = 0.016 and r = − 0.373, p < 0.001 respectively). In addition, sd-LDL-C was positively correlated with ox-LDL (r = 0.307, p = 0.003).

Conclusions

The findings of this preliminary study suggest that reduced sd-LDL-C, ox-LDL and hs-CRP levels may have a role in preventing atherosclerosis in subjects with GS.     

Cardioprotective effect of β-d-mannuronic acid (M2000) as a novel NSAID on gene expression of oxLDL scavenger receptors in the experimental diabetic model

Context: The investigations have shown that patients with diabetes have the elevated levels of glucose and oxLDL. These two play an important role in increased expression levels of oxLDL scavenger receptors on the surface of macrophages and endothelial cells that leads to deposition of oxLDL and macrophages in vascular walls.

Objective: The present study intends to show the effects of β-d-mannuronic acid (M2000) on the expression profile of ox-LDL scavenger receptors (including SR-A, LOX-1, CD36, and CD68) in an experimental model of diabetes.

Materials and methods: Eighteen Sprague-Dawley rats were randomly divided into three 6-member groups of the healthy control, diabetic control, and treated rats by M2000. Diabetes was induced in rats by intraperitoneal (IP) administration of 60?mg/kg streptozotocin. The treated rats were given daily intraperitoneal injections of M2000 with a dose of 25?mg/kg for 28 days and at the end of the 28th day, their aortas were removed. The qRT-PCR technique was then used to evaluate the expression levels of the proposed gene.

Results: The gene expression levels of the SR-A, LOX-1, CD36, and CD68 significantly declined in the diabetic group that received M2000 compared with untreated diabetic rats.

Conclusions: The M2000, as a novel NSAID is able to modify by lowering the gene expression levels of SR-A, LOX-1, CD36, and CD68 in treated rats compared to the untreated diabetic group, which may play an important role in preventing the complications that could lead to a cardioprotective efficacy.     

Atorvastatin reduces platelet-oxidized-LDL receptor expression in hypercholesterolaemic patients

BACKGROUND: Oxidized-LDL (ox-LDL) are proatherogenic and platelet-activating molecules. Atorvastatin reduces platelet activity before cholesterol-lowering action. CD36 and lectin-like oxidized-LDL receptor-1 (LOX-1) are specific ox-LDL receptors expressed also in platelets. This study was planned to address whether the possible rapid effect of atorvastatin on platelets could be related to modulation of ox-LDL receptors. MATERIALS AND METHODS: Forty-eight hypercholesterolaemic subjects requiring statin treatment (atorvastatin 20 mg day(-1)) after an ineffective diet regimen were evaluated for complete lipid-profile (chromogenic); P-selectin (P-sel), CD36 and LOX-1 expression (cytofluorimetric detection); circulating and platelet-associated ox-LDL (ox- and Pox-LDL, ELISA); and intracellular citrullin recovery (iCit, HPLC) at baseline and 3, 6 and 9 days after inclusion in the study. Moreover, we studied 48 normal controls matched for sex and age. RESULTS: Platelet activity expressed by P-sel (in resting and thrombin-activated cells), CD36 and LOX-1 were increased in hypercholesterolaemic subjects (all P < 0.01). Atorvastatin induced a reduction of CD36 at 6 days (P < 0.05); and P-sel in resting (P < 0.001) and activated cells (P < 0.001) and LOX-1 were reduced at 9 days (all P < 0.001) in association with decreased Pox-LDL (P < 0.001) and increased iCit (P < 0.01). All data were obtained before a significant reduction of LDL and ox-LDL was achieved (P = 0.109 and 0.113). DISCUSSION: Present data suggest that platelet deactivation by atorvastatin is related to CD36 and LOX-1 expression reduction before significant LDL changes. Moreover, the modulation of LOX-1 can be considered a self-relevant antiatherothrombotic action of atoravastin owing to the important role of this receptor in the ox-LDL-mediated vascular damage.     

老年代谢综合征患者血清氧化低密度脂蛋白水平的变化及检测意义

目的探讨老年代谢综合征患者血清氧化低密度脂蛋白（ox-LDL）水平的变化及其相关因素分析。方法老年代谢综合征组169例,健康对照组89例,分别测量血压、身高、体重、腰围、臀围,检测空腹血糖、胰岛素、血脂及ox-LDL水平,计算腰臀比、体重指数、HOMA-IR。结果代谢综合征组的体重指数、腰臀比、空腹血糖及胰岛素、HOMA-IR、甘油三酯、胆固醇、ox-LDL较健康对照组升高,代谢综合征组HDL水平较健康对照组降低（P〈0.05）。ox-LDL水平与腰臀比、体重指数、空腹血糖、空腹胰岛素、HOMA-IR、甘油三酯、LDL水平呈正相关,与HDL水平呈负相关（P〈0.001）。代谢综合征合并糖尿病、冠心病、脑梗塞时其ox-LDL水平较无合并症时增高（P〈0.05）。结论ox-LDL检测对评估代谢综合征患者发生动脉粥样硬化的危险度具有重要意义。     

血凝素样氧化型低密度脂蛋白受体1在人脐静脉内皮细胞表达CD40中的作用

目的研究氧化低密度脂蛋白对人脐静脉内皮细胞表达CD40中血凝素样氧化型低密度脂蛋白受体1的作用。方法在氧化低密度脂蛋白作用人脐静脉内皮细胞前预先用血凝素样氧化型低密度脂蛋白受体1阻断剂角叉(菜)胶(κ-carrageenan)和多聚肌苷酸(PIA)作用1h,应用亲合组织化学技术检测血凝素样氧化型低密度脂蛋白受体1的表达,流式细胞技术检测细胞表面CD40的表达。结果预先加入血凝素样氧化型低密度脂蛋白受体1阻断剂PIA和角叉(菜)胶的CD40的表达低于氧化低密度脂蛋白组(P<0.05)。结论在人脐静脉内皮细胞的炎症反应中,氧化低密度脂蛋白通过血凝素样氧化型低密度脂蛋白受体1途径激活了CD40的表达。     

Quercetin regulates oxidized LDL induced inflammatory changes in human PBMCs by modulating the TLR-NF-κB signaling pathway

Toll-like receptors (TLRs) have been shown to play a pivotal role in both innate and adaptive immune responses. TLR family is the essential recognition and signaling component of mammalian host defense. Both genetic and biochemical data support a common signaling pathway that finally leads to the activation of NF-κB and induction of the cytokines and co-stimulatory molecules required for the activation of the adaptive immune response. The present study was designed to examine the involvement of TLR2 and TLR4 in the oxidized LDL induced inflammation in human PBMCs and the effect of flavonoid quercetin on TLR-NF-κB signaling mechanism. LDL was isolated from human plasma and oxidation of LDL was done by incubating with 10 μM CuSO₄ overnight at 37 °C. The isolated human PBMCs in culture were used as the model system. 50 μg/ml ox-LDL treatment significantly up regulated TLR2 and TLR4 expression in isol human PBMCs after 24 h of culture and this was down regulated by quercetin at 25 μM concentration. ox-LDL caused a significant activation of NF-κB as evidenced by the detection of enhanced p65 subunit in nuclear extracts. Supplementation of quercetin significantly modulates the NF-κB p65 nuclear translocation. The cytokine IL-6 production was significantly increased in ox-LDL treated group and was decreased by quercetin treatment. Quercetin mediated reduction of TLR2 and TLR4 expression and the inhibition of nuclear translocation of NF-κB p65 in turn decreased the inflammatory enzymes like 5-LOX and COX and also decreased the mRNA expression of inducible enzymes like COX-2 and iNOS. Quercetin inhibited the ox-LDL induced TLR2 and TLR4 expression at mRNA level and modulated the TLR-NF-κB signaling pathway thereby inhibited the cytokine production and down regulated the activity of inflammatory enzymes thus have protective effect against the ox-LDL induced inflammation in PBMCs.     

心肾综合征中硫酸吲哚酚对巨噬细胞吞噬氧气型低密度脂蛋白的诱导作用的研究

目的  探讨硫酸吲哚酚（IS）对RAW264.7巨噬细胞吞噬氧气型低密度脂蛋白（ox-LDL）的诱导作用及其作用机制。方法  0、10、50、100、200和400μmol/L IS处理RAW264.7细胞24 h，采用CCK-8法测定RAW264.7细胞存活率，流式细胞术测定RAW264.7细胞的Dil标记氧化型低密度脂蛋白（Dil-ox-LDL）吞噬量。200μmol/L IS处理RAW264.7细胞0、12、24、36和48 h，测定RAW264.7细胞存活率和Dil-ox-LDL吞噬量。IS组RAW264.7细胞用200μmol/L IS处理24 h，UO126+IS组RAW264.7细胞以5μmol/L UO126预处理2 h后再200μmol/L IS处理24 h，测定非处理组、IS组和UO126+IS组RAW264.7细胞存活率和Dil-ox-LDL吞噬量，并采用蛋白质印迹法（Western blot）检测在IS刺激15 min时的ERK1/1蛋白表达情况。结果  IS对RAW264.7细胞存活率的浓度效应和时间效应检测中，不同组间比较差异无统计学意义（P >0.05）。Dil-ox-LDL吞噬量与IS浸润浓度及处理时间均呈正相关（P <0.01）。IS组的Dil-ox-LDL吞噬量和磷酸化ERK1/2蛋白表达量均明显高于非处理组（P <0.01）。UO126+IS组的Dil-ox-LDL吞噬量略低于非处理组，但差异无统计学意义（P >0.05）；磷酸化ERK1/2蛋白表达量大幅低于非处理组，差异有统计学意义（P <0.01）。UO126+IS组同IS组比较，Dil-ox-LDL吞噬量和磷酸化ERK1/2蛋白表达量均呈现大幅降低（P <0.01）。  结论  IS可在体外直接诱导RAW264.7巨噬细胞吞噬ox-LDL，该效应经由活跃MAPK信号转导通路实现，且随IS浓度升高和浸润时间延长而增强。

     

ApoAⅠ促进THP-1源性泡沫细胞ApoE分泌的研究

目的 通过建立动脉粥样硬化损伤部位细胞模型,观察ApoAⅠ对THP-1源性泡沫细胞 ApoE分泌的影响,探讨ApoAⅠ与ApoE之间是否存在相互关系以及这种关系对动脉粥样硬化的影响.方法 采用聚乙二醇-6000(PEG-6000)沉淀法制备人血浆HDL,再利用凝胶过滤柱层析法分离ApoAⅠ;佛波酯(PMA)诱导THP-1单核细胞分化为巨噬细胞,氧化型低密度脂蛋白(oxidized low density lipoprotein,ox-LDL)使分化后THP-1细胞荷脂,建立动脉粥样硬化细胞模型;ELISA检测不同浓度ApoAⅠ(0,5,10,15,25 μg/ml)及不同孵育时间(0,3,6,12,24 h),THP-1源性泡沫细胞ApoE的分泌;RT-PCR检测细胞ApoE mRNA的表达情况.结果 ApoAⅠ增加THP-1源性泡沫细胞ApoE的分泌量;且ApoE蛋白质的分泌随着ApoAⅠ孵育时间增加而增加,在24 h ApoE的分泌量达最大;在剂量试验中,ApoE的分泌量随着ApoAⅠ剂量的增加有增加趋势,ApoAⅠ浓度为25 μmol/L时,ApoE分泌量最大;而ApoE基因的表达不受ApoAⅠ的影响.结论 一定浓度的ApoAⅠ促进THP-1源性泡沫细胞ApoE的分泌,且具有时间及剂量依赖性.而在基因水平上,ApoAⅠ对ApoE mRNA表达没有影响.     

不同生活环境藏族脂代谢的特点及与PON1、ox-LDL的关系

目的：探讨高原地区不同生活环境下藏族人群血脂代谢特点及与对氧磷酶1（PON1）及氧化低密度脂蛋白（ox-LDL）变化的关系。方法：对青海省玉树县结古镇（城镇地区,平均海拔3 700m）和隆宝镇（纯牧业地区,平均海拔4 800m）176例藏族人群进行脂代谢的特点及与PON1、ox-LDL的关系进行研究分析。结果：1玉树地区高脂血症患病率为46.0%（81/176例）,其中结古镇组高脂血症的患病率为60.2%（71/118例）,显著高于隆宝镇组17.2%（10/58例）;结古镇组高TC血症、高TG血症、高LDL血症的患病率分别为44.9%、25.4%和47.5%,显著高于隆宝镇组19.0%、5.2%和17.2%）,P〈0.01;2相关分析显示PON1与TC、TG呈负相关,ox-LDL与海拔及Hb呈正相关,P〈0.01;3Logistic回归分析显示,影响脂质代谢的主要因素为年龄、住址（结古镇和隆宝镇）、Hb及腹围。结论：不同海拔不同生活环境藏族血脂水平与PON1及ox-LDL浓度不同,血脂增高是血浆PON1降低原因之一,海拔的差异性、城镇化生活环境、高龄及血红蛋白可能是高原地区藏族PON1及ox-LDL水平不同的一个影响因素。