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S Bellosta

Publications and source records attributed to S Bellosta.

At least 37 records · Page 2Linked to original sources
PubMed1998

Oleuropein, the bitter principle of olives, enhances nitric oxide production by mouse macrophages.

The Mediterranean diet, rich in fresh fruits and vegetables, has been associated with a lower incidence of cardiovascular disease and cancer, partly because of its high proportion of bioactive compounds such as vitamins, flavonoids and polyphenols. The major lipid component of such diet is the drupe-derived olive oil, that can be distinguished from other seed oils for the peculiar composition of its non-triglyceride fraction. In fact, several minor components, including polyphenols, grant the oil its particular taste and aroma. Oleuropein, the most abundant among these components, has been shown to be a potent antioxidant endowed with antiinflammatory properties. We investigated the effects of oleuropein on NO release in cell culture and its activity toward nitric oxide synthase (iNOS) expression. The results show that oleuropein dose-dependently enhance nitrite production in LPS-challenged mouse macrophages. This effect was blocked by the iNOS inhibitor L-NAME, indicating increased iNOS activity. Also, Western blot analysis of cell homogenates show that oleuropein increases iNOS expression in such cells. Taken together, our data suggest that, during endotoxin challenge, oleuropein potentiates the macrophage-mediated response, resulting in higher NO production, currently believed to be beneficial for cellular and organismal protection.

Animals↗
PubMed1998

HMG-CoA reductase inhibitors reduce MMP-9 secretion by macrophages.

-Macrophages secrete matrix metalloproteinases (MMPs) that may weaken the fibrous cap of atherosclerotic plaque, predisposing its fissuration. The 92-kDa gelatinase B (MMP-9) has been identified in abdominal aortic aneurysms and in atherosclerotic tissues. Fluvastatin, through the inhibition of the isoprenoid pathway, inhibits major processes of atherogenesis in experimental models (smooth muscle cell migration and proliferation and cholesterol accumulation in macrophages). We studied the effect of fluvastatin on the activity of MMP-9 in mouse and human macrophages in culture. Conditioned media of cells treated for 24 hours with fluvastatin were analyzed by gelatin zymography. In mouse macrophages, fluvastatin (5 to 100 micromol/L) significantly inhibited in a dose-dependent manner MMP-9 activity from 20% to 40% versus control. The drug, at a concentration as low as 5 micromol/L, inhibited MMP-9 activity ( approximately 30%) in human monocyte-derived macrophages as well. Phorbol esters (TPA, 50 ng/mL) stimulated MMP-9 activity by 50%, and fluvastatin inhibited this enhanced activity up to 50% in both mouse and human macrophages. The above results on the secretion of MMP-9 were confirmed by Western blotting and ELISA. The inhibitory effect of fluvastatin was overcome by the simultaneous addition of exogenous mevalonate (100 micromol/L), a precursor of isoprenoids. Fluvastatin's effect was fully reversible, and the drug did not cause any cellular toxicity. The statin did not block directly the in vitro activation of the secreted protease. Similar data were obtained with simvastatin. Altogether, our data indicate an inhibition of MMP-9 secretion by the drug. This effect is mediated by the inhibition of synthesis of mevalonate, a precursor of numerous derivatives essential for several cellular functions.

Animals↗
PubMed1997

Pharmacological control of phagocyte function: inhibition of cholesterol accumulation.

Phagocytes play a major role in several diseases. In particular mononuclear phagocyte-derived foam cells have a prominent role in the development of the atherosclerotic lesions. Macrophages are present in all stages of atherogenesis; they internalize lipoproteins and accumulate cholesterol. Moreover, lipid-filled macrophages, by secreting extracellular matrix-degrading enzymes, may weaken rupture-prone atherosclerotic plaques, thus increasing the probability of precipitating atherosclerotic acute symptoms (i.e., myocardial infarction, angina, etc.). Therefore, control of cellular functions and cholesterol accumulation in macrophages represent pharmacological targets against atherosclerosis. In our laboratory we studied the effect of calcium antagonists on cellular cholesterol esterification in cultured macrophages. We also demonstrated that the HMG-CoA reductase inhibitors (vastatins) fluvastatin and simvastatin prevented cholesterol deposition in cultured human and murine macrophage by inhibiting modified LDL endocytosis. Interestingly, vastatin activity was more pronounced in cholesterol-loaded macrophages (i.e., foam cells) than in normal cells. In conclusion, in vitro pharmacological control of cholesterol accumulation in macrophages may be achieved with some calcium antagonists and vastatins independently of their effects on blood pressure or cholesterolemia.

Animals↗
PubMed1997

Effect of lacidipine on cholesterol esterification: in vivo and in vitro studies.

1. Cholesterol esterification and accumulation in the arterial wall is a hallmark of atherogenesis. Several preclinical studies suggest that calcium antagonists may exert antiatherosclerotic activity by directly affecting atherogenesis in the arterial wall. We investigated the effect of the second generation dihydropyridine calcium antagonist lacidipine on cholesterol metabolism in vivo in the aortic arch of cholesterol fed rabbits, and in vitro in mouse cultured peritoneal macrophages. 2. Treatment of cholesterol-fed rabbits with 1, 3 and 10 mg kg-1 day-1 of lacidipine for two months reduced, in a dose-dependent manner, cholesterol esterification in the aortic arch: 24 +/- 6, 30 +/- 12, and 41 +/- 8% inhibition, respectively (P < 0.001 vs HC control). Concomitantly, drug treatment reduced total cholesterol content of the vessel wall. Lacidipine 3 and 10 mg kg-1 day-1 reduced cholesterolaemia (approximately 20%); no effect was observed at the lowest dose used (1 mg kg-1 day-1). These results suggest that the action of lacidipine on cholesterol esterification in the arterial wall involves, at least in part, a direct effect on cellular cholesterol metabolism. Inhibition of cholesterol esterification in the arterial wall was observed also in a reference group of animals treated with the specific ACAT inhibitor CI-976. 3. To evaluate the action of lacidipine on intracellular cholesterol metabolism we performed in vitro experiments with murine macrophages, the main cell type that accumulates cholesterol in the arterial wall. Lacidipine almost completely inhibited cholesterol esterification in cholesterol loaded macrophages in culture. The effect was observed independently of esterification stimuli and in cell free homogenates. The drug modified intracellular cholesterol distribution, doubling the free- to esterified sterol ratio, but did not influence the cellular rate of cholesteryl ester hydrolysis in the cell. All together these results indicate an inhibitory effect of lacidipine on cholesterol esterification catalyzed by the enzyme ACAT in murine macrophages. 4. We concluded that lacidipine influences cellular cholesterol metabolism. This effect may contribute to the potential antiatherosclerotic activity of this drug.

Animals↗
PubMed1996

Disruption of the acyl-CoA:cholesterol acyltransferase gene in mice: evidence suggesting multiple cholesterol esterification enzymes in mammals.

The microsomal enzyme acyl-CoA:cholesterol acyltransferase (ACAT; EC 2.3.1.26) catalyzes the esterification of cellular cholesterol with fatty acids to form cholesterol esters. ACAT activity is found in many tissues, including macrophages, the adrenal glands, and the liver. In macrophages, ACAT is thought to participate in foam cell formation and thereby to contribute to atherosclerotic lesion development. Disruption of the gene for ACAT (Acact) in mice resulted in decreased cholesterol esterification in ACAT-deficient fibroblasts and adrenal membranes, and markedly reduced cholesterol ester levels in adrenal glands and peritoneal macrophages; the latter finding will be useful in testing the role of ACAT and macrophage foam cell formation in atherosclerosis. In contrast, the livers of ACAT-deficient mice contained substantial amounts of cholesterol esters and exhibited no reduction in cholesterol esterification activity. These tissue-specific reductions in cholesterol esterification provide evidence that in mammals this process involves more than one form of esterification enzyme.

Adrenal Glands↗
PubMed1995

Stable expression and secretion of apolipoproteins E3 and E4 in mouse neuroblastoma cells produces differential effects on neurite outgrowth.

Previously, we demonstrated in cultured dorsal root ganglion neurons that, in the presence of beta-migrating very low density lipoproteins (beta-VLDL), apolipoprotein (apo) E4, but not apoE3, suppresses neurite outgrowth. In the current studies, murine neuroblastoma cells (Neuro-2a) were stably transfected with human apoE3 or apoE4 cDNA, and the effect on neurite outgrowth was examined. The stably transfected cells secreted nanogram quantities of apoE (44-89 ng/mg of cell protein in 48 h). In the absence of lipoproteins, neurite outgrowth was similar in the apoE3- and apoE4-secreting cells. The apoE4-secreting cells, when incubated with beta-VLDL, VLDL, cerebrospinal fluid lipoproteins (d < 1.21 g/ml), or with triglyceride/phospholipid (2.7:1 (w/w)) emulsions, showed a reduction in the number of neurites/cell, a decrease in neurite branching, and an inhibition of neurite extension, whereas in the apoE3-secreting cells in the presence of a lipid source, neurite extension was increased. Uptake of beta-VLDL occurred to a similar extent in both the apoE3- and apoE4-secreting cells. With low density lipoproteins or with dimyristoylphosphatidylcholine emulsions, either alone or complexed with cholesterol, no differential effect on neurite outgrowth was observed. A slight differential effect was observed with apoE-containing high density lipoproteins. The differential effect of apoE3 and apoE4 in the presence of beta-VLDL was blocked by incubation of the cells with heparinase and chlorate, with lactoferrin, or with receptor-associated protein, all of which prevent the uptake of lipoproteins by the low density lipoprotein receptor-related protein (LRP). The data suggest that the secreted and/or cell surface-bound apoE interact with the lipoproteins and facilitate their internalization via the heparan sulfate proteoglycan-LRP pathway. The mechanism by which apoE3 and apoE4 exert differential effects on neurite outgrowth remains speculative. However, the data suggest that apoE4, which has been shown to be associated with late onset familial and sporadic Alzheimer's disease, may inhibit neuronal remodeling and contribute to the progression of the disease.

Alzheimer Disease↗
PubMed1995

The inhibitory effect of apolipoprotein E4 on neurite outgrowth is associated with microtubule depolymerization.

Evidence is presented for the differential effects of two isoforms of apolipoprotein (apo) E, apoE3 and apoE4, on neurite outgrowth and on the cytoskeleton of neuronal cells (Neuro-2a) in culture. In the presence of a lipid source, apoE3 enhances and apoE4 inhibits neurite outgrowth. Immunocytochemical studies demonstrate that there is a higher concentration of apoE3 than apoE4 in both the cell bodies and neurites. Cells treated with apoE4 showed fewer microtubules and a greatly reduced ratio of polymerized to monomeric tubulin than did cells treated with apoE3. The effect of apoE4 on depolymerization of microtubules was shown by biochemical, immunocytochemical, and ultrastructural studies. The depolymerization of microtubules and the inhibition of neurite outgrowth associated with apoE4 suggest a mechanism whereby apoE4, which has been linked to the pathogenesis of Alzheimer's disease, may prevent normal neuronal remodeling from occurring later in life, when this neurodegenerative disorder develops.

Actins↗
PubMed1995

Macrophage-specific expression of human apolipoprotein E reduces atherosclerosis in hypercholesterolemic apolipoprotein E-null mice.

apoE deficiency causes hyperlipidemia and premature atherosclerosis. To determine if macrophage-specific expression of apoE would decrease the extent of atherosclerosis, we expressed human apoE in macrophages of apoE-null mice (apoE-/-) and assessed the effect on lipid accumulation in cells of the arterial wall. Macrophage-specific expression of human apoE in normal mice was obtained by use of the visna virus LTR. These animals were bred with apoE-/- mice to produce animals hemizygous for expression of human apoE in macrophages in the absence of murine apoE (apoE-/-,hTgE+/0). Low levels of human apoE mRNA were present in liver and spleen and high levels in lung and peritoneal macrophages. Human apoE was secreted by peritoneal macrophages and was detected in Kupffer cells of the liver. Human apoE in the plasma of apoE-/-,hTgE+/0 mice (n = 30) was inversely correlated (P < 0.005) with the plasma cholesterol concentration. After 15 wk on a normal chow diet, atherosclerosis was assessed in apoE-/-,hTgE+/0 animals and in apoE-/-,hTgE0/0 littermates matched for plasma cholesterol level (approximately 450 mg/dl) and lipoprotein profile. There was significantly less atherosclerosis in both the aortic sinus and in the proximal aorta (P < 0.0001) in the animals expressing the human apoE transgene. In apo-E-/-,hTgE+/0 animals, which had detectable atherosclerotic lesions, human apoE was detected in the secretory apparatus of macrophage-derived foam cells in the arterial wall. The data demonstrate that expression of apoE by macrophages is antiatherogenic even in the presence of high levels of atherogenic lipoproteins. The data suggest that apoE prevents atherosclerosis by promoting cholesterol efflux from cells of the arterial wall.

Animals↗
PubMed1994

Differential effects of apolipoproteins E3 and E4 on neuronal growth in vitro.

Apolipoprotein E4 (apoE4), one of the three common isoforms of apoE, has been implicated in Alzheimer's disease. The effects of apoE on neuronal growth were determined in cultures of dorsal root ganglion neurons. In the presence of beta-migrating very low density lipoproteins (beta-VLDL), apoE3 increased neurite outgrowth, whereas apoE4 decreased outgrowth. The effects of apoE3 or apoE4 in the presence of beta-VLDL were prevented by incubation with a monoclonal antibody to apoE or by reductive methylation of apoE, both of which block the ability of apoE to interact with lipoprotein receptors. The data suggest that receptor-mediated binding or internalization (or both) of apoE-enriched beta-VLDL leads to isoform-specific differences in interactions with cellular proteins that affect neurite outgrowth.

Animals↗
PubMed1993

27-Hydroxycholesterol modulation of low density lipoprotein metabolism in cultured human hepatic and extrahepatic cells.

27-Hydroxycholesterol**, 25-hydroxycholesterol and cholesterol suppressed LDL uptake and degradation in human extrahepatic and hepatic cell lines in a concentration-dependent manner. Cholesterol was the least potent, and the inhibitory effect of oxysterols was more pronounced in skin fibroblasts and in endothelial cell line EAhy 926 than in hepatoma HepG2 cells. Shorter incubations were required for oxysterols to achieve 50% inhibition of LDL uptake and degradation in all cultured cells. The inhibition of LDL catabolism in extrahepatic cells by 27-hydroxycholesterol occurred at concentrations close to those observed in human plasma (0.2-0.6 microM). The results support a possible role of 27-hydroxycholesterol, a physiological oxysterol, in the regulation of cellular cholesterol homeostasis in non-hepatic tissues.

Cells, Cultured↗
PubMed1993

Requirement for mevalonate in acetylated LDL induction of cholesterol esterification in macrophages.

HMG-CoA reductase inhibitors simvastatin, fluvastatin and fluvastatin enantiomers (0.1 to 5 microM) were utilized to block both mevalonate formation and cholesterol esterification in mouse peritoneal macrophages in the presence of a large excess of cholesterol supplied by acetylated LDL. Supplementation of cultures with mevalonate fully reversed, in a dose-dependent manner, the inhibitory effect of the drugs on cholesterol esterification. Mevalonate alone, in the range of the tested concentrations, did not affect cholesterol esterification in the absence of the HMG-CoA reductase inhibitors, indicating that its effect was linked to the restoration of the endogenous pool depleted by the pharmacological block of HMG-CoA reductase. The inhibitory effect of fluvastatin was also prevented by the non-sterol mevalonate isoprenoid derivative geranylgeraniol. Evaluation of fluvastatin enantiomers demonstrated the stereospecificity of drug action with most of the effect associated to the antipode with the highest inhibitory activity of HMG-CoA reductase. We conclude that mevalonate or a mevalonate product(s), possibly a non-sterol derivative(s), are required in cholesterol esterification induced by acetylated LDL in macrophages.

Acetylation↗
PubMed1993

Effect of trapidil derivative AR 12456 on intracellular cholesterol homeostasis in human hepatoma cell line Hep G2.

The effect of trapidil derivative AR12456 on intracellular cholesterol metabolism was investigated in human hepatoma cell line HepG2. AR12456 enhanced the uptake and degradation of 125I-LDL in a dose-dependent manner. The drug inhibited cholesterol synthesis and esterification without affecting cellular cholesterol content and bile acid synthesis; cholesterol efflux was slightly increased. These results show that the inhibition of cholesterol synthesis together with the enhanced expression of LDL receptors may partially explain the hypocholesterolemic activity of compound AR12456.

Anticholesteremic Agents↗
PubMed1991

Cholesterol stimulation of HDL binding to human endothelial cells EAhy 926 and skin fibroblasts: evidence for a mechanism independent of cellular metabolism.

The properties of the HDL binding site on the permanent human cell line EAhy 926 were studied. This cell line presents with highly differentiated functions of vascular endothelium. EAhy 926 cells possess HDL3 saturable binding sites with a Kd of about 20 micrograms/ml, which were up-regulated by cholesterol and were pronase- and EDTA-insensitive. Furthermore, HDL3 promoted cholesterol efflux from EAhy 926 cells in a dose-dependent manner. Thus, the HDL-binding site in EAhy 926 cells is similar to that present in fibroblasts, smooth muscle cells and endothelial cells. Up-regulation of HDL binding by cholesterol did not require de novo synthesis of HDL 'receptor' protein, as shown by the lack of effect of cycloheximide and alpha-amanitin and also occurred in fixed, non-living cells. Similar results were obtained using human skin fibroblasts. From these data we conclude that: (a) EAhy 926 cells are a good model for studying the HDL interaction with endothelial cells; (b) a mechanism independent of cellular metabolism is involved in the cholesterol-mediated up-regulation of HDL binding sites in EAhy 926 cells and human skin fibroblasts.

Binding, Competitive↗
PubMed1991

Influence of trapidil and derivatives on cholesterol synthesis and esterification in cultured cells.

The effect of trapidil (Rocornal) and its derivatives AR 12456 and AR 12463 on endogenous cholesterol synthesis and on cholesterol esterification rate was studied in human skin fibroblasts (HSF), in human hepatoma cell line Hep G2 and in primary culture of peritoneal macrophages from mouse (PMM). The cholesterol esterification rate was not influenced by the drugs in the tested cell lines. The incorporation of [14C]acetate into cholesterol in HSF was inhibited by AR 12463 and AR 12456, but not by trapidil. The inhibitory potency of AR 12456 in HSF was enhanced after preincubation of the drug with Hep G2 and removal of the medium to HSF, suggesting that the formed metabolite(s) are more potent inhibitors than the parent substance. The metabolite(s) formed seem(s) to influence the first steps in the endogenous formation of cholesterol, because the incorporation of [14C]mevalonate into cholesterol was not significantly inhibited. These findings suggest that the demonstrated inhibition of the endogenous cholesterol synthesis by AR 12456, especially after transformation into a probably more active substance(s), together with the recently described enhanced expression of LDL receptors in Hep G2 cells may partially explain the hypocholesterolaemic activity of AR 12456.

Acetates↗
PubMed1991

Calcium antagonists and cholesteryl ester metabolism in macrophages.

Experimental data indicate that calcium antagonists modify cellular lipid metabolism in the arterial wall as part of their antiatherosclerotic action observed in animal models. In the present study, we investigated the effect of verapamil, nifedipine, and lacidipine (a new dihydropyridine derivative) on cholesteryl ester metabolism in cultured mouse peritoneal macrophages (MPMs). Cholesteryl esters are formed in the cell via acyl-CoA:cholesterol acyltransferase (ACAT), which senses free cholesterol supplied by lysosomal hydrolysis of lipoprotein cholesterol ester. Verapamil inhibited up to 99% the ability of acetyl-low-density lipoprotein (acLDL) to stimulate cholesterol esterification in macrophages, but was less effective in 25-hydroxycholesterol-stimulated MPMs and in cholesterol-loaded cells after acLDL removal. Cells incubated with [3H]cholesterol ester-acLDL and verapamil showed a reduction in the cholesterol/cholesteryl ester ratio. In the same experimental conditions, nifedipine displays minor or no effects on cholesterol esterification and in the cholesterol/cholesteryl ester ratio. On the contrary, the nifedipine-like lacidipine was active in inhibiting cholesterol esterification in macrophages elicited by acLDL. Our data indicate that calcium antagonists of different structure, even within the same class, may have various effects on cholesterol esterification in macrophages in culture.

Animals↗
PubMed1991

Increased affinity of LDL for their receptors after acipimox treatment in hypertriglyceridemia.

The regulation of cellular LDL metabolism by hypertriglyceridemic LDL was tested before and after treatment with acipimox, a nicotinic acid derivative, in 11 type IV hyperlipidemic patients. Large, less dense LDL particles were found in plasma after acipimox treatment. HDL subfractions were only slightly modified, with an increase of dense, cholesteryl ester-enriched and triglyceride-poor HDL3 particles. The LDL (B, E) receptor activity in human skin fibroblasts of LDL isolated before and after treatment was also evaluated. Hypertriglyceridemic LDL proved rather inefficient in regulating receptor activity, with a close to 30% lower capacity than normal LDL to inhibit receptor-mediated uptake and degradation of 125I-LDL. This abnormality was fully corrected after acipimox. The reported findings indicate that acipimox treatment in type IV patients can normalize the defective interaction of hypertriglyceridemic LDL with the LDL (B, E) receptor.

Adult↗
PubMed1990

Lipoprotein changes and increased affinity of LDL for their receptors after acipimox treatment in hypertriglyceridemia.

Modifications in plasma low and high density lipoprotein (LDL and HDL) subfraction distribution, as well as the regulation of cellular LDL metabolism by hypertriglyceridemic LDL were tested before and after treatment with acipimox, a nicotinic acid derivative, in 11 type IV hyperlipidemic patients. Large, less dense LDL particles were found in plasma after acipimox treatment, reflecting compositional changes, characterized by a 25.4% increase in cholesteryl ester content and by a 46.2% reduction of triglycerides in LDL. HDL subfractions were only slightly modified, with an increase of dense, cholesteryl ester-enriched and triglyceride poor HDL3 particles. The LDL (B,E) receptor activity in humans skin fibroblasts of LDL isolated before and after treatment was also evaluated. Hypertriglyceridemic LDL proved rather inefficient in regulating receptor activity with a close to 30% reduction vs. normal LDL in the capacity to inhibit receptor-mediated uptake and degradation of 125I-LDL. Such abnormality was fully corrected after acipimox. The reported findings indicate that acipimox treatment in type IV patients, in spite of a relatively modest plasma triglyceride reduction, can markedly modify LDL distribution and composition, normalizing the defective interaction of hypertriglyceridemic LDL with the LDL (B,E) receptor.

Humans↗