Nanoscale amphiphilic macromolecules with variable lipophilicity and stereochemistry modulate inhibition of oxidized low-density lipoprotein uptake.

Amphiphilic macromolecules (AMs) based on carbohydrate domains functionalized with poly(ethylene glycol) can inhibit the uptake of oxidized low density lipoprotein (oxLDL) and counteract foam cell formation, a key characteristic of early atherogenesis. To investigate the influence of lipophilicity and stereochemistry on the AMs' physicochemical and biological properties, mucic acid-based AMs bearing four aliphatic chains (2a) and tartaric acid-based AMs bearing two (2b and 2l) and four aliphatic chains (2g and 2k) were synthesized and evaluated. Solution aggregation studies suggested that both the number of hydrophobic arms and the length of the hydrophobic domain impact AM micelle sizes, whereas stereochemistry impacts micelle stability. 2l, the meso analogue of 2b, elicited the highest reported oxLDL uptake inhibition values (89%), highlighting the crucial effect of stereochemistry on biological properties. This study suggests that stereochemistry plays a critical role in modulating oxLDL uptake and must be considered when designing biomaterials for potential cardiovascular therapies.

Coarse grained molecular dynamics of engineered macromolecules for the inhibition of oxidized low-density lipoprotein uptake by macrophage scavenger receptors.

Atherosclerosis is a condition resulting from the accumulation of oxidized low-density lipoproteins (oxLDLs) in arterial walls. Previously developed macromolecules consisting of alkyl chains and polyethylene glycol (PEG) on a mucic acid backbone, termed nanolipoblockers (NLBs) are hypothesized to mitigate the uptake of oxLDL by macrophage scavenger receptors. In this work, we developed a coarse grained model to characterize the interactions between NLBs with a segment of human scavenger receptor A (SR-A), a key receptor domain that regulates cholesterol uptake and foam cell conversion of macrophages, and studied NLB ability to block oxLDL uptake in PBMC macrophages. We focused on four different NLB configurations with variable molecular charge, charge location, and degree of NLB micellization. Kinetic studies showed that three of the four NLBs form micelles within 300 ns and of sizes comparable to literature results. In the presence of SR-A, micelle-forming NLBs interacted with the receptor primarily in an aggregated state rather than as single unimers. The model showed that incorporation of an anionic charge near the NLB mucic acid head resulted in enhanced interaction with the proposed binding pocket of SR-A compared to uncharged NLBs. By contrast, NLBs with an anionic charge located at the PEG tail showed no interaction increase as NLB aggregates were predominately observed to interact away from the oxLDL binding site. Additionally, using two different methods to assess the number of contacts that each NLB type formed with SR-A, we found that the rank order of contacts coincided with our experimental flow cytometry results evaluating the ability of the different NLBs to block the uptake of oxLDL.

Atorvastatin active metabolite inhibits oxidative modification of small dense low-density lipoprotein.

We tested the hypothesis that atorvastatin active metabolite (ATM), on the basis of its distinct structural features and potent antioxidant activity, preferentially inhibits lipid oxidation in human small dense low-density lipoprotein (sdLDL) and other small lipid vesicles. LDL, sdLDL, and various subfractions were isolated from human plasma by sequential ultracentrifugation, treated with ATM, atorvastatin, pravastatin, rosuvastatin, or simvastatin and were subjected to copper-induced oxidation. Lipid oxidation was measured spectrophotometrically as a function of thiobarbituric acid reactive substances formation. Similar analyses were performed in reconstituted lipid vesicles enriched in polyunsaturated fatty acids and prepared at various sizes. ATM was found to inhibit sdLDL oxidation in a dose-dependent manner. The antioxidant effects of ATM in sdLDL were 1.5 and 4.7 times greater (P < 0.001) than those observed in large buoyant LDL and very low-density lipoprotein subfractions, respectively. ATM had similar dose- and size-dependent effects in reconstituted lipid vesicles. None of these effects were reproduced by atorvastatin (parent) or any of the other statins examined in this study. These data suggest that ATM interacts with sdLDL in a specific manner that also confers preferential resistance to oxidative stress. Such interactions may reduce sdLDL atherogenicity and improve clinical outcomes in patients with cardiovascular disease.

Oxidized low-density lipoprotein increases interleukin-8 production in human gingival epithelial cell line Ca9-22.

BACKGROUND AND OBJECTIVE: Recent epidemiological studies have shown a correlation between periodontitis and hyperlipidemia. We have found high levels of oxidized low-density lipoprotein (OxLDL) in the gingival crevicular fluid of dental patients. In the present study, we tried to examine the possible role of OxLDL in periodontal inflammation in vitro. MATERIAL AND METHODS: Cells of the human gingival epithelial cell line Ca9-22 were cultured in media containing OxLDL, and the amounts of interleukin-8 (IL-8) and prostaglandin E(2) (PGE(2)) produced were measured using ELISAs. RESULTS: Production of IL-8 by Ca9-22 cells was significantly increased when the cells were treated with OxLDL, but not with native LDL or acetylated LDL. Production of PGE(2) by Ca9-22 cells was enhanced by co-incubation with OxLDL and interleukin-1 beta (IL-1 beta). Scavenger receptor inhibitors, fucoidan and dextran sulfate, inhibited the OxLDL-induced IL-8 and PGE(2) production in the presence of IL-1 beta. The p(38) MAPK inhibitors SB203580 and SB202190 and the ERK inhibitor PD98059 inhibited the OxLDL-induced IL-8 production. Among oxidized lipids and chemically modified LDL, 7-ketocholesterol enhanced IL-8 production. CONCLUSION: This is the first report to show that OxLDL enhances IL-8 production in epithelial cells.

Potentiation of beta-amyloid polymerisation by low-density lipoprotein enhances the peptide's vasoactivity.

Alzheimer's disease (AD) is characterised by the accumulation of insoluble beta-amyloid (A beta) fibrils in the brain. Factors that promote A beta fibrillogenesis may influence the pathogenesis of AD and represent targets for therapeutic intervention. Some A beta deposited in AD may originate in the circulation and plasma factors could promote A beta deposition, particularly in the cerebrovasculature. We investigated the effects of plasma low-density lipoprotein (LDL), in both its native and oxidised forms, on A beta(1-40) fibrillogenesis and vasoactivity. LDL enhanced A beta fibrillogenesis in a process dependent on LDL concentration and the oxidative state of the lipoprotein, as indicated by measurements of thiobarbituric acid reactive substances (TBARS) and conjugated dienes. LDL's actions were inhibited by the iA beta 5 peptide, suggesting that LDL-induced A beta polymerisation involved beta-pleated sheet formation. Potentiated A beta polymerisation was reflected by enhanced A beta-mediated vascular responses. Human endothelial cells exposed to fibrillar A beta generated with LDL, especially oxidised LDL, exhibited decreased 20S proteasome activity. Rat aortic ring constriction induced by noradrenaline was enhanced by A beta fibrils generated with LDL, with oxidised LDL producing the more marked effects. Should plasma lipoproteins prove to play a role in cerebral A beta deposition their modification with statins or antioxidants may offer therapeutic benefit.

Green tea polyphenols inhibit human vascular smooth muscle cell proliferation stimulated by native low-density lipoprotein.

This study investigated whether human vascular smooth muscle cell proliferation induced by native low-density lipoprotein (LDL) is affected by green tea catechins. Furthermore, the effects of native LDL on extracellular signal-regulated kinase (ERK) 1/2 activity were determined. Cell proliferation stimulated by native LDL was concentration-dependently inhibited by epigallocatechin, epigallocatechin-3-gallate, green tea polyphenon, and the nonspecific antioxidant N-acetylcysteine (P<0.05). Combined treatment of green tea polyphenon and N-acetylcysteine markedly potentiated the effect of each drug on vascular smooth muscle cell proliferation. ERK1/2 activity was only partly inhibited by green tea catechins alone or in combination with N-acetylcysteine (P<0.05). These data suggest that green tea constituents inhibit proliferation of human vascular smooth muscle cells exposed to high levels of native LDL. Green tea constituents and antioxidants may exert vascular protection by inhibiting human vascular smooth muscle cell growth associated with hypercholesterolemia.

A novel gene silencer, pyrrole-imidazole polyamide targeting human lectin-like oxidized low-density lipoprotein receptor-1 gene improves endothelial cell function.

Pyrrole-imidazole polyamide can be combined in antiparallel side-by-side dimeric complexes along the minor groove of DNA in a sequence-specific manner. Pyrrole-imidazole polyamides are effective inhibitors of transcription factors as well as viral repressors and transactivators. Recently, lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) was reported to be a major factor contributing to the pathogenesis of coronary atherosclerosis. In this study, we designed a pyrrole-imidazole polyamide specific for the LOX-1 gene and evaluated its effect on LOX-1 gene transcription. A pyrrole-imidazole polyamide was designed to target the AP-1 binding site of the LOX-1 gene and synthesized by solid phase methods. This pyrrole-imidazole polyamide significantly inhibited LOX-1 promoter activity in HEK293 cells, determined by the luciferase assay. LOX-1 mRNA expression was also inhibited by the pyrrole-imidazole polyamide at a concentration of 10-9 mol/l in human umbilical vein endothelial cells (HUVEC), determined by the real-time PCR method. HUVEC were treated by pyrrole-imidazole polyamide targeting the LOX-1 gene, and apoptosis was assessed using Hoechst stain, terminal deoxy nucleotidyl transferase-mediated UTP end labeling method, and dye-uptake bioassay. Treatment of HUVEC for 72 h with LOX-1 targeted pyrrole-imidazole polyamide decreased apoptosis induced by angiotensin II and oxidized low-density lipoprotein (ox-LDL) loading in all assays. This novel therapeutic agent, pyrrole-imidazole polyamide, could specifically inhibit LOX-1 gene expression by reducing the promoter activity of the gene. Pyrrole-imidazole polyamide seems to be a powerful promising new agent that can be used to explore therapies based on inhibition of transcription. Molecular recognition of DNA by small molecules could provide insight into the development of new human medicines.

Engineered polymeric nanoparticles for receptor-targeted blockage of oxidized low density lipoprotein uptake and atherogenesis in macrophages.

Strategies to prevent the uptake of modified low density lipoproteins (LDLs) by immune cells, a major trigger of inflammation and atherogenesis, are challenged by complex interfacial factors governing LDL receptor-mediated uptake. We examine a new approach based on a family of "nanoblockers", which are designed to examine the role of size, charge presentation, and architecture on inhibition of highly oxidized LDL (hoxLDL) uptake in macrophages. The nanoblockers are macromolecules containing mucic acid, lauryl chloride, and poly(ethylene glycol) that self-assemble into 15-20 nm nanoparticles. We report that the micellar configuration of the macromolecules and the combined display of anionic (carboxylate) groups in the hydrophobic region of the nanoblockers caused the most effective inhibition in the uptake of hoxLDL by IC21 macrophages. The nanoblockers primarily targeted SR-A and CD36, the major scavenger receptors and modulated the "atherogenic" phenotype of cells in terms of the degree of cytokine secretion, accumulation of cholesterol, and "foam cell" formation. These studies highlight the promise of synthetically engineered nanoblockers against oxidized LDL uptake.

Dietary fibre, diabetes, and hyperlipidaemia. Progress and prospects.

When diabetes and cardiovascular disease were first classed as possible fibre-deficiency diseases, laboratory and clinical evidence was lacking. Subsequent studies indicated that the gums and viscous types of fibre (e.g., guar and pectin) are more effective than other fibres in slowing carbohydrate absorption and hence in reducing the postprandial rise in blood glucose and serum insulin. This effect has longer term metabolic consequences. In addition, gums and viscous fibres reduce serum cholesterol, possibly by mechanisms other than simply increasing bile-salt loss. If these potential therapeutic effects of fibre are to be exploited, palatable formulations must be developed. The effect of fibre in whole foods should also be determined.

Inhibitory effect of flavonoids on low-density lipoprotein peroxidation catalyzed by mammalian 15-lipoxygenase.

Lipoxygenase-dependent low-density lipoprotein (LDL) oxidation is believed to be involved in atherogenesis. Inhibition of lipoxygenase-induced lipid peroxidation might, therefore, be an important mode to suppress the development of atherosclerosis. Because dietary antioxidants inhibit LDL oxidation in vitro and their intake is inversely associated with coronary heart diseases, we compared the inhibitory effect of three typical flavonoids-quercetin, epicatechin, and flavone-with alpha-tocopherol and ascorbic acid against human LDL oxidation catalyzed by mammalian 15-lipoxygenase. The oxidative modification of LDL was monitored by measurement of cholesteryl ester hydroperoxide (CE-OOH) formation and consumption of antioxidants by using HLPC. Quercetin and epicatechin were the strongest inhibitors of LDL oxidation catalyzed by 15-lipoxygenase; ascorbic acid was an effective inhibitor in the first 3 h of oxidation; and fivefold alpha-tocopherol-enriched LDL showed a partial inhibition of CE-OOH formation only after 4-6 h of incubation. Flavone had no effect. Quercetin, ascorbic acid, and alpha-tocopherol were consumed in the first 3 h of incubation. Consumption of LDL alpha-tocopherol was partially inhibited by ascorbic acid and quercetin, whereas epicatechin and flavone were without effect. These results emphasize the inhibitory effect of the flavonoids quercetin and epicatechin on 15-lipoxygenase-mediated LDL lipid peroxidation. At similar concentrations, they are stronger antioxidants than ascorbic acid, alpha-tocopherol, and flavone.