Proportion Of Free Cholesterol Research Articles

Cholesterol and oxysterol metabolism and subcellular distribution in macrophage foam cells: accumulation of oxidized esters in lysosomes

Cholesterol- and cholesteryl ester-rich macrophage foam cells, characteristic of atherosclerotic lesions, are often generated in vitro using oxidized low density lipoprotein (OxLDL). However, relatively little is known of the nature and extent of sterol deposition in these cells or of its relationship to the foam cells formed in atherosclerotic lesions. The purpose of this study was to examine the content and cellular processing of sterols in OxLDL-loaded macrophages, and to compare this with macrophages loaded with acetylated LDL (AcLDL; cholesteryl ester-loaded cells containing no oxidized lipids) or 7-ketocholesterol-enriched acetylated LDL (7KCAcLDL; cholesteryl ester-loaded cells selectively supplemented with 7-ketocholesterol (7KC), the major oxysterol present in OxLDL). Both cholesterol and 7KC and their esters were measured in macrophages after uptake of these modified lipoproteins. Oxysterols comprised up to 50% of total sterol content of OxLDL-loaded cells. Unesterified 7KC and cholesterol partitioned into cell membranes, with no evidence of retention of either free sterol within lysosomes. The cells also contained cytosolic, ACAT-derived, cholesteryl and 7-ketocholesteryl esters. The proportion of free cholesterol and 7KC esterified by ACAT was 10-fold less in OxLDL-loaded cells than in AcLDL or 7KCAcLDL-loaded cells. This poor esterification rate in OxLDL-loaded cells was partly caused by fatty acid limitation. OxLDL-loaded macrophages also contained large (<40–50% total cell sterol content) pools of oxidized esters, containing cholesterol or 7KC esterified to oxidized fatty acids. These were insensitive to ACAT inhibition, very stable and located in lysosomes, indicating resistance to lysosomal esterases. Macrophages loaded with OxLDL do not accumulate free sterols in their lysosomal compartment, but do accumulate lysosomal deposits of OxLDL-derived cholesterol and 7-ketocholesterol esterified to oxidized fatty acids. The presence of similar deposits in lesion foam cells would represent a pool of sterols that is particularly resistant to removal.—Brown, A. J., E. L. Mander, I. C. Gelissen, L. Kritharides, R. T. Dean, and W. Jessup. Cholesterol and oxysterol metabolism and subcellular distribution in macrophage foam cells: accumulation of oxidized esters in lysosomes.

Changes in composition and distribution of LDL subspecies in hypertriglyceridemic and hypercholesterolemic patients during gemfibrozil therapy

The effect of gemfibrozil (GEM) on composition and distribution of LDL subspecies in 10 hypertriglyceridemic (HTG) patients with triglyceride (TG) levels of 300–750 mg/dl and low density lipoprotein cholesterol (LDL-C) < 160 mg/dl, and 8 hypercholesterolemic (HC) patients with LDL-C ≥ 190 mg/dl and TG < 200 mg/dl was investigated. Patients were randomized in a double-blind, crossover design to 12 week periods of placebo and 1200 mg/day GEM with an intervening washout period. All 7 LDL subspecies in the density range of 1.029 to 1.063 g/ml in the HTG group contained more TG ( P < 0.01) and less free cholesterol ( P < 0.05, while the proportion of cholesterol ester was lower only in LDL subspecies 1 to 3 ( P < 0.05) as compared to the HC group. In HTG patients, GEM increased the proportion of cholesterol ester ( P < 0.05) in all subfractions, while the proportion of free cholesterol was increased and TG was decreased in LDL subspecies 1 to 4 only ( P < 0.05). GEM decreased the LDL subspecies score from 4.8 ± 0.7 to 4.2 ± 0.5, and the major LDL subclass density from 1.048 ± 0.006 to 1.043 ± 0.002 g/ml ( P < 0.01) in HTG patients. GEM decreased TG levels to mean of 228 ± 52 mg/dl in HTG patients, but there was no change in LDL phenotype. No significant changes in composition and distribution of LDL subspecies were noted in HC patients with GEM. We conclude that GEM significantly altered the composition and distribution of LDL subspecies with a shift from small dense LDL particles to large, less dense particles in HTG patients.

Excessive mural calcium overload — A predominant causal factor in the development of stenosing coronary plaques in humans

Healthy human coronary artery walls contain, over their entire lifetime, more free and total cholesterol than calcium. However, as soon as arteriosclerotic alterations set in, the calcium content increases. Thus in coronary fatty streaks (arteriosclerotic plaques of WHO stage I), calcium was increased 13 times, in stage II plaques 25 times, and in fully developed stage III plaques 80 times above normal on average. The most dramatic calcium incrustation was found in coronary stage III plaques that had produced massive fatal coronary infarction. Here, the proportion of calcium salts (particularly hydroxyapatite) may amount to almost 50% of dry weight. Thus the most excessive accumulation of calcium seems to be correlated with the highest fatality. In contrast, there is no correlation between mural coronary free or total cholesterol content, and plaque severity. Accordingly, stenosing coronary stage III plaques contain less cholesterol than do fatty streaks. Moreover, in coronary stage III plaques the proportion of free cholesterol was 1.37%, and of total cholesterol only 2.34% of the whole mass, certainly not enough for directly causing coronary occlusion. Thus the calcium-rich plaques of human coronary arteries considerably differ from the well-known cholesterol-rich plaques (stage I and II) of human aortae. Our findings justify a new prophylactic approach with suitable calcium antagonists to interfere with deleterious calcium uptake in coronary plaque development.

Lipid composition of isolated epiphyseal cartilage cells, membranes and matrix vesicles

1. 1. Intact cells, cell fragments (membranes) and matrix vesicles were isolated from the proliferating and calcifying layers of epiphyseal cartilage by sequential hyaluronidase and collagenase digestion and differential centrifugation. Lipids were extracted and analyzed for various lipid classes and their fatty acid composition by column, thin-layer, paper and gas-liquid chromatography. 2. 2. On a protein basis the isolated matrix vesicles had more total lipid than either the membrane or cell fractions, the vesicles and membranes being richer in non-polar lipids and containing smaller quantities of phospholipids than whole cells. Expressed as a percentage of the total lipid, the cells were richer in triacylglycerols and lower in free fatty acids than the membrane or vesicle fractions. The proportion of free cholesterol and the cholesterol/phospholipid ratio were nearly twice as high in the matrix vesicles as in the other tissue fractions. Choline and ethanolamine phosphoglycerides progressively declined in the membrane and matrix vesicle fractions, whereas serine phosphoglycerides and sphingomyelin increased. Non-phosphorus-containing polar lipids were present in all fractions, the vesicles being richer in polyhexosyl ceramides, cerebrosides, glycosyldiacylglycerols and certain uncharacterized acidic polar lipids. 3. 3. Fatty acid patterns of the matrix vesicles were distinctive from those of isolated cells, being generally richer in 18 : 0 and 18 : 2, and lower in 16 : 1 and 18 : 1 fatty acids. Monoacyl forms were similarly increased in 16 : 0 and/or 18 : 0, and reduced in 16 : 1,18 : 1 or 20 : 2 fatty acids, depending on the lipid class. The fatty acid composition of diphosphatidylglycerol from cells and matrix vesicles was markedly different, providing evidence that the cardiolipin in the vesicles was not from mitochondrial components. 4. 4. Based on the fact that the matrix vesicles were significantly enriched in free cholesterol, sphingomyelin, glycolipids and serine-phosphoglycerides, it is concluded that they are derived from the plasma membrane of the cell, supporting earlier conclusions based upon morphological and enzymological evidence.

Hypercholesterolaemia in rats

At the last College meeting a possible link between low density lipoproteins, proliferation of arterial cells and the formation of atheroma was proposed. It was suggested that the synthesis of cholesterol in these lipoproteins, which occurs in the liver, is inhibited by dietary cholesterol reaching the liver in remnants of chylomicrons. Variations in proportions of diffusible free cholesterol to fixed cholesteryl esters in chylomicrons were shown to depend upon the amount of dietary fat. This was proposed as a mechanism regulating the synthesis of cholesterol by the liver. Rats, unlike rabbits and monkeys, rarely develop hypercholesterolaemia from dietary experiments alone, unless bile acids or anti-thyroid drugs are fed concurrently with cholesterol. However, in the present feeding experiments of various fats designed to ‘trick' the inhibition of endogenous cholesterol synthesis, a significant increase in serum cholesterol was found in those rats fed cholesterol with coconut oil, rising from the normal level of 58 ± 6 mg/100 ml to 97 ± 9. The proportions of free cholesterol to esters in chylomicrons from thoracic duct lymph from different dietary groups of rats varied and may be an explanation for variations in cholesterol levels in their serum. Whether atheromatous-like lesions develop remains to be seen when the rats die of old age.

Free and Esterified Cholesterol in the Cerebrospinal Fluid of Goats Affected with Experimental Scrapie

SUMMARY The total cholesterol content and the proportions of free and esterified cholesterol have been determined in the cerebrospinal fluid and sera of goats during the onset of experimental scrapie. The total cholesterol in cerebrospinal fluid remained relatively constant but a small rise occurred in clinically affected animals. The proportions of free cholesterol increased in all goats when only slight histological damage to brain tissue was present. The mean value for the total cholesterol was 4·0 mg./l. in the experimental animals and, of this, 48 % was free cholesterol when terminal elevations were ignored. In sera there were periodic fluctuations during 12 months in the total cholesterol content which were associated with rises in the proportions of free cholesterol. The mean value for total cholesterol in serum was 70 mg./ 100 ml. and, of this, free cholesterol on an average was 58 %.