geranylgeranyl-pyrophosphate and Atherosclerosis

It has been proposed that high-density lipoprotein (HDL) loses its cardioprotective ability through oxidative modifications by reactive oxygen species (ROS) and promote atherogenesis. However, the pro-atherogenic pathways undergone by oxidized HDL remain poorly understood. Since monocytes play a crucial role in atherogenesis, this study was aimed to investigate the influence of both native and oxidized HDL (oxHDL) on monocytes-macrophages functions relevant to atherogenesis. HDL particles were isolated from human blood samples by ultracentrifugation and subjected to in vitro oxidation with CuSO(4). The extent of oxidation was quantitated by measurement of lipid peroxides. Human peripheral blood mononuclear cells were isolated and cultured under standard conditions. Cells were treated with native and oxHDL at varying concentrations for different time intervals and used for several analyses. Intracellular ROS production was assessed based on ROS-mediated DCFH fluorescence of the cells. The release of TNF-α and matrix metalloproteinases (MMPs) was quantitated using ELISA kit and gelatine zymography, respectively. Treatment of cells with oxidized HDL enhanced the production of ROS in a concentration-dependent way, while native HDL had no such effect. Further, the release of TNF-α, MMP-9, and MMP-2 was found to be remarkably higher in cells incubated with oxHDL than that of native HDL. Results demonstrate that oxidative modification of HDL induces pro-inflammatory response and oxidative stress in human monocytes-macrophages.

Topics: Atherosclerosis; Cell Death; Cell Survival; Cells, Cultured; Humans; Inflammation; Inflammation Mediators; Lipid Metabolism; Lipoproteins, HDL; Liver X Receptors; Macrophages; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Monocytes; Orphan Nuclear Receptors; Oxidation-Reduction; Oxidative Stress; Polyisoprenyl Phosphates; Reactive Oxygen Species; Tumor Necrosis Factor-alpha

Panax notoginseng (Burk.) F.H. Chen has been used as a health product and natural remedy in traditional medicine for cardiovascular diseases for more than 1000 years in Asia, including China, Japan, and Korea. Panax notoginseng saponins (PNS) are the major effective ingredients extracted from Panax notoginseng.. The purpose of this study was to investigate whether Panax notoginseng saponins (PNS) attenuated atherosclerosis by inducing liver X receptor alpha (LXRα) expression and to elucidate the mechanisms responsible for the effects.. The AS rats were treated once daily with PNS (100 mg/kg, i.p.), and pathological changes in the aorta were observed using Sudan IV staining. The expression of LXRα in the aortic wall was measured by Western blot analysis. THP-1 macrophages were cultured with PNS in the presence or absence of geranylgeranyl pyrophosphate ammonium salt (GGPP), an LXRα antagonist. The expression of LXRα and its target genes ATP-binding cassette A1 and G1 (ABCA1, ABCG1) were determined by qRT-PCR. The transcriptional activation of the LXRα gene promoter was analyzed by a reporter assay. The NF-κB DNA binding activity and the expression of interleukin (IL)-6, monocyte chemotactic protein-1 (MCP-1) was evaluated respectively by Trans-AM NF-κB ELISA and ELISA in THP-1 macrophages that were stimulated with LPS after treatment with PNS and GGPP.. PNS treatment alleviated the typical pathological changes associated with atherosclerosis in rats. The expression of LXRα was increased in rat aortas after treatment with PNS. In vitro, PNS increased LXRα mRNA levels in THP-1 macrophages. The reporter assays showed that PNS enhanced transcriptional activation of the LXRα gene promoter and led to the upregulation of ABCA1 and ABCG1 expression. This upregulation could be reversed by treatment with GGPP. Additionally, PNS inhibited NF-κB DNA binding activity and reduced secretion of IL-6 and MCP-1 in LPS-stimulated THP-1 macrophages. These effects could be reversed by GGPP.. The results indicated that the PNS-mediated attenuation of AS may, at least partly, due to LXRα uprergulation. The mechanisms of action included enhancement transcriptional activation of the LXRα gene promoter by PNS and subsequent upregulation of ABCA1 and ABCG1 and inhibition of NF-κB DNA binding activity.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Aorta; Atherosclerosis; ATP Binding Cassette Transporter 1; ATP Binding Cassette Transporter, Subfamily G, Member 1; ATP-Binding Cassette Transporters; Cell Line; Humans; Inflammation; Lipid Metabolism; Liver X Receptors; Macrophages; Male; NF-kappa B; Orphan Nuclear Receptors; Panax notoginseng; Phytotherapy; Polyisoprenyl Phosphates; Rats; Rats, Wistar; RNA, Messenger; Saponins

Parathyroid hormone-related protein (PTHrP) is overexpressed in atherosclerotic plaques by unknown mechanisms. We have examined here the putative mechanism(s) responsible for this overexpression in the atherosclerotic lesion and its potential modulation by simvastatin, both in vitro and in vivo.. Atherosclerosis was induced in rabbits by femoral endothelial dessication and atherogenic diet. After 2 weeks, animals were randomized to receive either 5mg/(kgd) simvastatin (n=7) or no treatment (n=6) during 4 additional weeks. An increase in PTHrP immunostaining was observed in atherosclerotic lesions of hyperlipidemic rabbits, which was significantly reduced by simvastatin. However, PTH/PTHrP type 1 receptor staining was similar in both groups. In cultured vascular smooth muscle cells (VSMCs), atherogenic concentrations of native LDL (0.125-0.5mg/mL) increased PTHrP expression. This effect was prevented by preincubation with simvastatin (1micromol/L) and was reversed by mevalonate, geranylgeranylpyrophosphate and, to a lesser extent, by farnesylpyrophosphate. Moreover, in transfection studies, we showed that RhoA appears to participate in the mechanism whereby LDL induces PTHrP in VSMC. Finally, native LDL-induced VSMC growth and this mitogenic effect was blocked by PTHrP silencing.. LDL might be responsible for PTHrP overexpression in atherosclerotic plaques of hyperlipidemic rabbits. The inhibition of this effect by simvastatin provides further insights into the mechanisms of action of statins.

Topics: Animals; Atherosclerosis; Cells, Cultured; Disease Models, Animal; Femoral Artery; Humans; Hydroxymethylglutaryl-CoA Reductase Inhibitors; Lipoproteins, LDL; Macrophages; Male; Mevalonic Acid; Muscle Cells; Muscle, Smooth, Vascular; Parathyroid Hormone-Related Protein; Polyisoprenyl Phosphates; Rabbits; Receptor, Parathyroid Hormone, Type 1; RNA, Small Interfering; Simvastatin; Tunica Intima; Up-Regulation

Resistin is a novel cysteine-rich protein that plays a role in the development of insulin resistance and atherosclerosis in rodents, while its role in humans is unclear. C-reactive protein (CRP) is an important risk predictor for coronary heart disease, and it can also modify the expression of genes involved in atherogenesis. Statins have been demonstrated to possess lipid lowering effects as well as pleiotropic properties. We hypothesize that CRP may result in overexpression of resistin, and statin may decrease CRP-induced resistin expression in cultured human peripheral blood monocytes (PBMC).. The aim of the present study, therefore, was to assess the effects of both CRP on resistin expression and simvastatin on CRP-induced of resistin expression in cultured human PBMC.. Human PBMC were isolated from the whole blood of healthy volunteers by density gradient centrifugation. First, cells were incubated with varying concentrations of CRP (0, 5, 10, 25 and 50 microg/ml) for 24h for assessing the dose-dependent effects on resistin expression. Second, 25 microg/ml of CRP was used to time-dependent evaluation on resistin expression (0, 3, 6, 12 and 24h). Moreover, cells were pretreated with simvastatin at concentrations from 0.1 to 1 microM for 2h, and then co-incubated with 25 microg/ml CRP for 24h for evaluating effect of statin on resistin production subjected to CRP. Finally, in additional experiments, monocytes were incubated with 1 microM simvastatin in the absence or presence of 100 microM mevalonate or 10 microM geranylgeranyl-pyrophosphate (GGPP) or 10 microM farnesylpyrophosphate (FPP) for 2h, then co-incubated with CRP for 24h for examining whether effects of statin on CRP-induced resistin expression was independent of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors.. The results showed that CRP induced both mRNA expression and protein secretion of resistin in a dose- and time-dependent manner. Co-incubation with simvastatin significantly inhibited CRP-induced up-regulation of mRNA and protein expression of resistin. Treatment with mevalonate, GGPP, but not FPP, reversed the inhibition of resistin expression caused by simvastatin, suggesting that simvastatin regulated resistin expression in culture human PBMC through the mevalonate-GGPP signal pathway.. In the present study, the data showed that CRP could significantly increase resistin expression in cultured human PBMC, and this effect was inhibited by simvastatin, suggesting that CRP and resistin might be involved in the pathogenesis of atherosclerosis, and statin therapy might be beneficial for atherosclerotic disease by modifying CRP-induced resistin overexpression in PBMC.

Topics: Animals; Atherosclerosis; C-Reactive Protein; Cells, Cultured; Coronary Disease; Diterpenes; Dose-Response Relationship, Drug; Humans; Hydroxymethylglutaryl-CoA Reductase Inhibitors; Insulin Resistance; Mevalonic Acid; Monocytes; Polyisoprenyl Phosphates; Resistin; Risk Factors; Rodentia; Signal Transduction; Simvastatin; Time Factors; Up-Regulation