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D S Leake

Publications and source records attributed to D S Leake.

At least 37 records · Page 2Linked to original sources
PubMed1995

Transition metal ions within human atherosclerotic lesions can catalyse the oxidation of low density lipoprotein by macrophages.

The oxidation of low density lipoprotein (LDL) in the arterial wall may contribute to atherogenesis. The oxidation of LDL by cells usually requires catalytically active transition metal ions. We show here some that gruel samples from human advanced atherosclerotic lesions are capable of catalysing the oxidation of LDL by macrophages as measured by thiobarbituric acid-reactive substances, enhanced electrophoretic mobility and increased macrophage uptake. This catalysis could be inhibited by pretreatment of the gruel with Chelex-100, which binds transition metal ions. The presence of catalytically active transition metal ions in atherosclerotic lesions may help to explain why LDL oxidation occurs at these sites.

Animals↗
PubMed1995

Induction of the antioxidant stress proteins heme oxygenase-1 and MSP23 by stress agents and oxidised LDL in cultured vascular smooth muscle cells.

Enhanced expression of the antioxidant stress proteins heme oxygenase-1 (HO-1) and macrophage stress protein (MSP23) by oxidative stress agents and oxidatively modified low density lipoproteins (LDL) was investigated in cultured porcine aortic smooth muscle cells. Treatment of smooth muscle cells with glucose oxidase, CdCl2 or diethylmaleate resulted in a time-dependent (6-48 h) induction of HO-1 and MSP23 expression. Exposure of cells to 100 micrograms protein/ml highly oxidised LDL increased the expression of HO-1 and MSP23 within 24 h, and the induction was dependent on the degree of LDL oxidation. The induction of HO-1 and MSP23 may thus play an important cytoprotective role against oxidative stress in atherogenesis.

Animals↗
PubMed1995

Oxidation of low density lipoprotein by iron or copper at acidic pH.

Oxidized low density lipoprotein (LDL) may play a significant role in atherosclerosis. We have investigated the effect of pH on the oxidation of LDL by iron or copper. When LDL was oxidized by iron in the presence of cysteine in either Hanks' balanced salt solution (HBSS) or Ham's F-10 medium, an acidic pH greatly decreased the lag period and increased the rate of formation of hydroperoxides and thiobarbituric acid-reactive substances (TBARS), and increased its uptake by macrophages. There was a dose-dependent increase of LDL oxidation at acidic pH in the presence of increasing concentrations of cysteine. When LDL was oxidized by copper in HBSS, an acidic pH increased the lag phase before the rapid formation of conjugated dienes, hydroperoxides, and TBARS, but increased its uptake by macrophages. Similar results were obtained using Ham's F-10 medium. Cysteine (100 microM) inhibited the modification of LDL by copper in HBSS at both pH 7.4 and 5.5 As atherosclerotic lesions may be acidic, these observations may help to explain why LDL oxidation occurs locally at these sites.

Arteriosclerosis↗
PubMed1994

The effects of free radical scavengers on the oxidation of low-density lipoproteins by macrophages.

Oxidised LDL has been implicated in the pathogenesis of atherosclerosis. Macrophages can oxidatively modify low-density lipoprotein (LDL) in vitro. The mechanisms of this oxidation process are presently unclear. In this study, we have investigated the effects of compounds and enzymes widely used to quench or scavenge active oxygen species to try to identify the oxidative species involved in this process. The data obtained suggest that hydrogen peroxide may possibly play a role in LDL oxidation by macrophages, whereas singlet oxygen and hydroxyl radicals may not. The role of superoxide anions was uncertain because copper-zinc superoxide dismutase (Cu/Zn-SOD) and manganese SOD (Mn-SOD), widely used to determine superoxide-dependency in other systems may be unsuitable in this particular system. Cu/Zn-SOD at high concentrations displayed a variability in its effects, sometimes augmenting LDL oxidation and sometimes inhibiting it. In the experiments in which Cu/Zn-SOD augmented LDL oxidation, heat inactivation of the enzyme decreased the augmentation; in the experiments in which Cu/Zn-SOD inhibited LDL oxidation, it retained its inhibitory effect after heat inactivation. Mn-SOD always inhibited modification even after heat inactivation. We have therefore concluded that superoxide involvement in LDL oxidation by macrophages is still uncertain and the uncertainty will remain until a suitable probe is found.

Animals↗
PubMed1994

Production of oxidized lipids during modification of low-density lipoprotein by macrophages or copper.

The oxidation of low-density lipoprotein (LDL) is implicated in atherosclerosis. Lipids and oxidized lipids were analysed by gas chromatography and gas chromatography-mass spectrometry in human LDL incubated with mouse peritoneal macrophages (MPM) or copper (II) sulphate in Ham's F-10 medium or medium alone (control). MPM-modification and copper-catalysed oxidation of LDL resulted in the formation of oxysterols, mainly cholest-5-en-3 beta,7 beta-diol (7 beta-OH-CHOL); 7%-19% of the initial cholesterol was converted to 7 beta-OH-CHOL in 24 h. 7 beta-OH-CHOL levels in control LDL were very low. The increase in 7 beta-OH-CHOL in MPM and copper-oxidized LDL was accompanied by decreases in linoleate and arachidonate and increases in the electrophoretic mobility and degradation of LDL protein by 'target' macrophages. The concerted occurrence of these processes and their similarity in both MPM-modification and copper-catalysed oxidation of LDL were suggested by the highly significant cross-correlations. The fall in polyunsaturated fatty acid (PUFA) was accompanied by a directly proportional increase in electrophoretic mobility of the LDL. Production of 7 beta-OH-CHOL and protein degradation by macrophages showed modest elevations during the initial steep fall in PUFA, and showed their greatest increases as the levels of PUFA slowly approached zero. The levels of 7 beta-OH-CHOL and the degradation of LDL by macrophages were directly proportional. The degradation of LDL by macrophages increased rapidly as the electrophoretic mobility of LDL was slowly approaching its maximum level.

Animals↗
PubMed1994

Iron released from transferrin at acidic pH can catalyse the oxidation of low density lipoprotein.

Low density lipoprotein (LDL) oxidation within the arterial wall may contribute to the disease of atherosclerosis. We have investigated the conditions under which transferrin (the major iron-carrying protein in plasma) may release iron ions to catalyse the oxidation of LDL. Transferrin that had been incubated at pH 5.5 released approximately 10% of its bound iron in 24 h, as measured by ultrafiltration and atomic absorption spectroscopy. Furthermore, transferrin co-incubated with LDL and L-cysteine at pH 5.5 resulted in the oxidation of the LDL as measured by thiobarbituric acid-reactive substances and electrophoretic mobility. This effect was observed at transferrin concentrations as low as 40% of its average plasma concentration. The release of iron from transferrin in atherosclerotic lesions due to a localised acidic pH may help to explain why LDL oxidation occurs in these lesions.

Humans↗
PubMed1994

NADPH oxidase is not essential for low density lipoprotein oxidation by human monocyte-derived macrophages.

NADPH oxidase has been reported to be involved in low density lipoprotein (LDL) oxidation by monocytes. We have investigated the ability of monocyte-derived macrophages from four chronic granulomatous disease patients, which lack NADPH oxidase, to oxidise LDL. All the cells oxidised LDL to significantly increase its uptake by mouse macrophages. We conclude therefore that NADPH oxidase is not essential for LDL oxidation by macrophages.

Animals↗
PubMed1994

The oxidation of low density lipoprotein by cells or iron is inhibited by zinc.

We have examined the effect of zinc ions on low density lipoprotein (LDL) oxidation by macrophages, endothelial cells and iron ions in terms of the increased uptake of the LDL by macrophages. Zinc ions inhibited LDL modification by both cell types (which is dependent on the presence of iron ions in the culture medium) and by iron ions alone. As oxidised LDL is believed to be involved in atherogenesis, this raises the possibility that zinc may be an endogenous protective factor against atherosclerosis.

Animals↗
PubMed1994

Ascorbic acid can either increase or decrease low density lipoprotein modification.

In freshly prepared low density lipoprotein (LDL), ascorbate inhibited LDL oxidation by macrophages at the higher concentrations tested (60-100 microM). In contrast, with LDL that had been allowed to autoxidise in the refrigerator (3 degrees C) for at least 10 weeks after isolation (mildly oxidised or minimally-modified LDL), ascorbate did not inhibit the modification of LDL in the presence of macrophages. Ascorbate actually modified autoxidised LDL itself in the absence of macrophages to greatly increase its uptake by macrophages. The modification of autoxidised LDL by ascorbate increased the levels of thiobarbituric acid-reactive substances in the medium and was completely inhibited by the antioxidant butylated hydroxytoluene. Thus the effects of ascorbate on unoxidized LDL can be very different to those on mildly oxidised LDL.

Animals↗
PubMed1994

The effect of inhibitors of free radical generating-enzymes on low-density lipoprotein oxidation by macrophages.

Oxidised low-density lipoprotein (LDL) produced by the action of arterial cells, including macrophages, has been implicated in atherosclerosis. We have investigated the effect of inhibitors of various cellular free-radical generating enzymes on macrophage-mediated LDL oxidation. Xanthine oxidase and nitric oxide synthase are not responsible for LDL modification by resident mouse peritoneal macrophages. Eicosatetraynoic acid, a lipoxygenase inhibitor, produced a dose-dependent irreversible inhibition of macrophage modification of LDL, but at concentrations rather close to those toxic to the cells. Diphenyl and diphenylene iodonium, NADPH oxidase and mitochondrial electron transport inhibitors, inhibited macrophage oxidation of LDL, at concentrations that were not obviously toxic. This suggests that NADPH oxidase, or some other flavin nucleotide-dependent process, may be involved in LDL oxidation by macrophages. Wortmannin and thiopropionic acid dilauryl ester did not inhibit LDL oxidation, suggesting that inhibition of NADPH oxidase may not be the means by which the iodonium compounds inhibit LDL oxidation. Macrophages from C3H/HeJ mice, which lack receptors for lipopolysaccharide, modified LDL normally, suggesting that the inadvertent priming of resident macrophages by traces of lipopolysaccharide bound to LDL was not involved in LDL oxidation.

Amino Acid Oxidoreductases↗
PubMed1994

Acidic pH enables caeruloplasmin to catalyse the modification of low-density lipoprotein.

LDL oxidation within the arterial wall may contribute to the disease of atherosclerosis. There is some evidence that elevated plasma levels of copper are associated with an increased risk of coronary artery disease. We have investigated the conditions under which caeruloplasmin (the plasma copper carrier protein) can catalyse the macrophage-mediated modification of LDL. Low concentrations of CuSO4 (< 1 microM) could catalyse the macrophage-mediated modification of LDL. Native caeruloplasmin was unable to catalyse the modification of LDL at pH 7.4, but could do so after preincubation at acidic pH. After preincubation at acidic pH, concentrations of caeruloplasmin as low as 30 micrograms/ml (about one-tenth of the human plasma level) could catalyse significant LDL oxidation when added to macrophages. The activation of copper in caeruloplasmin in atherosclerotic lesions due to a localised acidic pH may help to explain why LDL oxidation occurs in these areas of the body.

Animals↗
PubMed1993

Acidic pH increases the oxidation of LDL by macrophages.

We have investigated the effect of pH on LDL oxidation by macrophages (in the presence of iron ions), using a modification of Hanks' balanced salt solution. Increasing the acidity of the medium greatly increased the oxidation of the LDL by the macrophages as measured by thiobarbituric acid-reactive substances or increased uptake and degradation by a second set of macrophages. The rate of oxidation of LDL by iron ions alone, measured in terms of conjugated dienes, was also increased greatly even at mildly acidic pH. It is quite possible that atherosclerotic lesions have an acidic extracellular pH, particularly in the vicinity of macrophages, and the observation that LDL oxidation by macrophages is increased at acidic pH may therefore help to explain why atherosclerotic lesions are apparently one of the very few sites in the body where LDL oxidation occurs.

Animals↗
PubMed1993

The modification of low density lipoprotein by the flavonoids myricetin and gossypetin.

Myricetin and gossypetin, two hexahydroxylated flavonoids, are capable of modifying low density lipoprotein (LDL) to increase greatly its uptake by macrophages. When human 125I-labelled LDL was incubated with 100-1000 microM myricetin or gossypetin, it was subsequently endocytosed much faster by mouse peritoneal macrophages. This modification did not occur at a concentration of 10 microM. Nine other flavonoids containing up to five hydroxyl substituents did not modify LDL to any great extent at 100 microM. The modification of LDL by 100 microM myricetin was time-dependent and complete by 6 hr. Flavonoids can sometimes act as pro-oxidants but myricetin did not act by oxidizing the LDL, as the LDL lipid hydroperoxide content was not increased by myricetin, nor did it promote the depletion of the endogenous antioxidant alpha-tocopherol in the LDL. High concentrations of myricetin caused the aggregation of LDL particles, as judged by light microscopy, agarose gel electrophoresis, retention by a membrane filter and sedimentability by centrifugation. SDS-PAGE indicated that the apolipoprotein B-100 molecules of LDL particles were covalently crosslinked. The uptake and degradation by macrophages of myricetin-modified 125I-labelled LDL reached saturation at about 10 micrograms protein/mL, suggesting the existence of a high affinity uptake process for the modified LDL. The uptake of myricetin-modified 125I-labelled LDL was not competed for by a large excess of non-labelled native LDL or acetylated LDL. We conclude that myricetin and gossypetin at high concentrations are capable of modifying LDL by a novel non-oxidative mechanism to a form taken up by macrophages by a high affinity process.

Animals↗
PubMed1993

Oxidation of low density lipoprotein by bovine and porcine aortic endothelial cells and porcine endocardial cells in culture.

Oxidation of low density lipoprotein (LDL) in atherosclerotic lesions may be involved in converting macrophages into cholesterol-laden foam cells, a major characteristic of atherosclerotic lesions. It has been reported, and is widely believed, that endothelial cells derived from rabbit, pig and human aortas, but not those derived from bovine aortas, are capable of oxidising LDL in vitro. We have re-investigated this subject and found that during a 48-h incubation period bovine aortic endothelial cells (both in primary culture and in subcultures) were capable of consistently modifying LDL, increasing its uptake and degradation by macrophages by more than 4-fold. Incubation of LDL with bovine aortic endothelial cells for only 24 h, however, produced inconsistent modification of the LDL, whereas mouse peritoneal macrophages consistently modified LDL in 24 h. The modification of LDL by bovine aortic endothelial cells was an oxidative process, as the chain-breaking antioxidants, alpha-tocopherol and probucol, completely or greatly inhibited it. Thus, bovine aortic endothelial cells are capable of oxidising LDL but they are slower at doing so than are certain other types of cells. Nitric oxide generated by activated macrophages has very recently been shown to inhibit their oxidation of LDL. We have therefore investigated whether or not the inhibition of the constitutive nitric oxide synthase of bovine or porcine aortic endothelial cells would increase their rate of oxidation of LDL.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

The effect of EDTA on the oxidation of low density lipoprotein.

Low density lipoprotein (LDL) is routinely isolated and stored in buffers containing ethylene-diaminetetra-acetic acid (EDTA) to inhibit its autoxidation. We have investigated the effect of EDTA on LDL oxidation by both copper ions and macrophages. LDL oxidation by macrophages in Ham's F-10 medium containing 6 microM iron showed a large and concentration-dependent increase when EDTA was added up to about 10 microM. EDTA concentrations above about 10 microM progressively inhibited LDL oxidation as measured by macrophage degradation, thiobarbituric acid-reactive substances and electrophoretic mobility. The oxidation of LDL by 1 microM copper in Ham's F-10 medium, measured by macrophage degradation, also showed a large increase with low concentrations of EDTA (1-3 microM), with higher concentrations (10 microM or above) strongly inhibiting the oxidation. In a simple phosphate buffer, however, EDTA simply inhibited the oxidation of LDL by copper with equimolar amounts of EDTA to copper giving a complete inhibition. The results of this study indicate that when LDL oxidation by cells or by copper in Ham's F-10 medium is investigated, more oxidation may be obtained if the EDTA is not previously removed from the LDL preparation.

Animals↗