Acid In Cholesterol Esters Research Articles

Hormonal regulation of acid cholesterol ester hydrolase activity: effects of triiodothyronine and 17 alpha-ethynylestradiol.

Cholesterol ester hydrolase activity was measured in isolated rat hepatocytes and adipocytes. Administration of triiodothyronine to rats resulted in a specific and selective increase in lysosomal acid (pH 4.5) cholesterol ester hydrolase activity in hepatocytes. Since the majority of lipoprotein degradation occurs in liver parenchymal cells (hepatocytes), the stimulation of liver (hepatocyte) acid cholesterol ester hydrolase activity by triiodothyronine could contribute to the hypocholesterolemic action of thyroid hormones. Treatment of rats with 17 alpha-ethynylestradiol to increase the hepatic degradation of lipoprotein did not change acid cholesterol ester hydrolase activity in liver, indicating that the thyroid hormone induced stimulation of acid cholesterol ester hydrolase activity in hepatocytes is not a secondary effect owing to the increased hepatic catabolism of low density lipoproteins (LDL). In contrast to the results with hepatocytes, hyperthyroidism did not increase acid cholesterol ester hydrolase activity in rat adipocytes.

Effect of hyperinsulinemia on acid cholesterol ester hydrolase activity in liver, heart and epididymal fat pad preparations form rats and mice

The effect of insulin on lysosomal acid cholesterol ester hydrolase activity was studied inliver, heart and fat pad preparations form rats and mice. Hyperinsulinemia was induced for a period of 6 days in rats by the subcutaneous administration of exogenous insulin by an osmotic minipump. The effect of more chronic endogenous hyperinsulinemia was studied using genetic strains of diabetic ( db/db) ice at 12 weeks of age. Mouse liver and heart preparations were characterized as having an acid pH optimum of 4.5–5 for cholesterol ester hydrolase activity; a smaller but distinct pH optimum could also be observed at pH 7. In contrast, hydrolase activity in mouse fat pad preparatons had only one distinct pH optimum of 6.5. Hyperinsulinemia in rats and mice resulted in a significant decrease in acid cholesterol ester hydrolase activity in heart preparatons, but had no consistent effect on acid hydrolase activity observed in liver and fat pad preparations. This decrease in lysosomal acid cholesterol ester hydrolase activity in cardiac tissue due to hyperinsulinemia cannot be related to any changes in lipoprotein turnover caused by insulin or diabetes.

Fatty acid composition of serum and adipose tissue lipids in males with chronic renal failure. Relations to serum lipoproteins and clinical factors.

We have analysed the fatty acid composition of serum cholesterol esters (CE), phospholipids (PL) and triglycerides (TG) and of adipose tissue (AT) lipids in 72 males and chronic renal failure (CRF). Sixteen patients were treated with protein-poor diet. 16 with hemodialysis. The results have been related to clinical aspects of the disease and to abnormalities in serum lipid and lipoprotein concentrations. The major findings were high relative content of oleic acid and a low relative content of linoleic acid in all lipids in the diet-treated group compared to both controls and to the untreated and dialysed patients. Tendencies in the same directions were seen also in the untreated subjects, significant for oleic and linoleic acid in CE and for oleic acid in PL. These changes did not correlate to serum of lipoprotein lipids, or to serum creatinine concentrations in patients not on diet or dialysis, indicating that other factors are more important in modifying the fatty acid metabolism in CRF. The composition of the protein-poor diet, which was rich in fat but low in linoleic acid in relation to oleic acid, is probably a major factor explaining our findings in the diet-treated group. Since low levels of linoleic acid in plasma and AT lipids have been suggested to be a risk factor for atherosclerosis, more attention should be paid to the content and composition of dietary fat in order to prevent the occurrence of such low levels.

Properties of acid and neutral cholesterol ester hydrolases in rat and pigeon aortas

The activity of cholesterol ester hydrolase was measured in subcellular fractions from rat and pigeon aortas using a glycerol-dispersed cholesterol oleate substrate preparation. The specific activity of acid cholesterol ester hydrolase (assayed at pH 5) in adventitia tissue fractions was 40–50 fold greater than in media-intima fractions from rat aorta. Soluble and particulate subcellular fractions from rat aorta (media-intima) were observed to have cholesterol ester hydrolase activity with both an acid (pH 4.5–5) and a neutral (pH 7.5) pH optimum. A comparison of the subcellular distribution of acid cholesterol ester hydrolase with the lysosomal marker enzyme, N-acetylglucosaminidase, suggests that the acid hydrolase activity originated in aortic lysosomes; the neutral cholesterol ester hydrolase was predominantly soluble. Acid and neutral cholesterol ester hydrolases could also be distinguished on the basis of the effects of MgCl 2 and NaCl on hydrolase activity and on rates of thermal denaturation. Both acid and neutral hydrolases from rat aorta (media-intima) were inhibited by chloroquine (half-maximal at 2–4 mM), and both hydrolases were characterized as having the same apparent affinity for the glycerol-dispersed cholesterol oleate substrate. Acid and neutral cholesterol ester hydrolases were also observed in preparations from pigeon aortas. The specific activity for both acid and neutral hydrolases was higher in atherosclerosis-susceptible White Carneau pigeon aortas in comparison to Show Racer pigeon aortas.

Increased synthesis of hexacosanoic acid (C23:0) by cultured skin fibroblasts from patients with adrenoleukodystrophy (ALD) and adrenomyeloneuropathy (AMN).

We studied the metabolism of radioactive stearic acid by cultured skin fibroblasts from patients wtih adrenoleukodystrophy (ALD) and its variant, adrenomyeloneuropathy (AMN), to clarify the mechanism of the increased content of very long chain saturated fatty acids in cholesterol esters and sphingolipids, which are known to be the characteristic biochemical changes in ALD and AMN. A substantial amount of hexacosanoic acid (C26 : 0) wa synthesized from stearic acid by ALD and AMN fibroblasts, whereas only a trace amount of hexacosanoic acid was synthesized by control fibroblasts. This indicates that the primary biochemical defect in ALD and AMN may involved the elongation system of very long chain fatty acids.

Effect of thyroid hormones on acid cholesterol ester hydrolase activity in rat liver, heart and epididymal fat pads

The regulation of acid cholesterol ester hydrolase activity by thyroid hormones was studied in subcellular fractions from rat liver, heart, and epididymal fat pads; hydrolase activity was determined at pH 5 with a glycerol-dispersed cholesterol oleate substrate preparation. Acid cholesterol ester hydrolase activity was decreased in liver preparations from thyroidectomized rats relative to activity in livers from euthyroid control rats. Administration of triidothyronine to either euthyroid or hypothyroid (thyroidectomized) rats resulted in an increase in acid cholesterol ester hydrolase activity in liver preparations. Similar effects of thyroidectomy and the administration of triiodothyronine on acid cholesterol ester hydrolase activity were observed with fat pad preparations. In contrast, no effect of thyroid hormones was observed on acid cholesterol ester hydrolase activity in heart. These results suggest that thyroid hormones may regulate the catabolism of serum lipoproteins, in part, by alterations in lysosomal acid cholesterol ester hydrolase activity in liver and epididymal fat pads.

Studies on the activity of acyl-coA:Cholesterol o-acyltransferase and acid cholesterol ester synthetase in rat aortas

Acyl-CoA: cholesterol O-acyltransferase (ACAT) activity was measured in microsomal preparations from rat aorta (intima-media) with [ 14C]oleoyl CoA and endogenous cholesterol as substrates. The specific activity of ACAT in liver and adrenal microsomal preparations was 10–20-times greater than ACAT activity in aortic microsomes; no ACAT activity could be detected in fat pad microsomes. ACAT activity in liver and adrenal microsomes was enhanced by the addition of exogenous cholesterol. In contrast, exogenous cholesterol did not increase ACAT activity in rat aortic microsomes. Levels of endogenous cholesterol and ACAT activity in microsomal preparations from rat aorta were not reduced when circulating plasma cholesterol levels were decreased by the administration of 4-aminopyrazolopyrimidine to rats. Acid cholesterol ester synthetase activity was not detectable in high-speed supernatant fractions from rat aorta; low levels of activity could be measured in rat aorta microsomal preparations but this was less than 10% of ACAT activity. Thus, ACAT would seem to be the principal enzymatic route for the synthesis of cholesterol esters in aorta.

Acid cholesterol ester and glycerol ester hydrolase activities: Evidence for the individuality of these enzymes in human mononuclear leucocytes

Hydrolysis of cholesterol oleate and glycerol trioleate was measured in homogenates of human leucocytes at optimum pH of 4.0 and 5.25, respectively. Both enzyme activities appeared to reside in the 15 000 × g, 20-min fraction of mononuclear leucocytes. Solubilization of cholesterol ester hydrolase activity was strongly dependent on the detergent to protein ratio, showing optimal solubilization at weight ratios of 1.0 in cell homogenates and of 3.0 in the 15 000 × g, 20-min fraction, whereas solubilization of glycerol ester hydrolase was independent to protein ratio over the tested range of 0.3 to 5.8. Using a sequential solubilization procedure, about 60% of the granule proteins as well as 88% of glycerol ester hydrolase activity were solubilized at a detergent to protein ratio of 0.3, whereas cholesterol ester hydrolase activity was solubilized from the remaining membranes at a ratio of about 3.0. Thus, the acid glycerol ester hydrolase and acid cholesterol ester hydrolase were related to different proteins. Since solubilization of cholesterol ester hydrolase required drastic treatment, it is suggested that this enzyme is related to a protein within the lysosomal membrane.

Studies in normal pregnancy: III. Fatty acid composition of serum phosphoglycerides and cholesterol esters

Metabolic and hormonal influences on liver lipid synthesis are reflected in the relative fatty acid composition of serum lecithin. Thus, a successive decrease in essential fatty acids (EFA), i.e. n-6 fatty acids, during pregnancy would indicate an incipient EFA deficiency at the time of delivery. Furthermore, reciprocal changes in linoleic and arachidonic acids during the 1st trimester with an increase in arachidonic acid would suggest an enhancement of the Greenbeg Pathway (Pathway II) in lecithin synthesis presumably caused by estrogen influence. During the 2nd and 3rd trimester, reciprocal changes in lecithin content of palmitic and stearic acids are similar to changes appearing under cholestasis. It is furthermore suggested that variations in the utilization of linoleic acid in cholesterol esters (expressed as cholesterol ester 18:2/lecithin 18:2) might reflect changes in plasma cholesterol esterification by the enzyme lecithin-cholesterol-acyl-transferase (LCAT). Then, in late pregnancy, in relation to a subclinical cholestatic liver engagement, LCAT activity would increase as judged from this ratio as well as the increased linoleic acid in cholesterol esters.

Acid acylhydrolase deficiency in I-cell disease and pseudo-hurler polydystrophy

Fibroblasts from patients with I-cell disease and pseudo-Hurler poly-dystrophy are characterized by multiple lysosomal enzyme deficiencies; only acid β- d-galactosidase and α- d-neuraminidase deficiency have been described in liver. Acid cholesterol ester hydrolase and acid triacylglycerol hydrolase have not previously been studied in either tissue. Acid cholesterol ester hydrolase activity in five lines of I-cell fibroblasts was 18 ± 5% (S.D.) of normal. Acid triacylglycerol hydrolase was 17 ± 3% of normal. Equivalence in deficiency of the two activities was also seen in fibro-blasts from patients with pseudo-Hurler poly dystrophy, Wolman's disease and cholesterol ester storage disease, suggesting that both activities reside in one enzyme. Although the apparent K m of cholesterol ester hydrolase (substrate emulsified with Triton X-100) was moderately elevated in two of three I-cell lines examined, deficiency was more a function of diminished catalytic capacity in all cases. In autopsied liver from three I-cell patients acid cholesterol ester hydrolase was 1–11% and acid triacylglycerol was 8–22% of normal; triacylglycerol hydrolase was less deficient in each case. In the liver of another I-cell patient, KZ, cholesterol ester hydrolase was 25–60% and triacylglycerol hydrolase was 86–194% of normal. Both activities in KZ fibroblasts were 17–20% of normal, indicating that the deficiency in fibroblasts does not necessarily denote triacylglycerol deficiency in liver. The apparent K m of liver cholesterol ester hydrolase was moderately elevated in two of the four samples.

Lecithin : cholesterol acyl transfer (LACT) and fatty acid composition of lecithin and cholesterol esters in young male myocardial infarction survivors

Lecithin : cholesterol acyl transfer (LCAT) and relative fatty acid composition of serum lecithin and cholesterol esters were studied in 20 young male survivors of myocardial infarction (MI). Comparisons were made with controls matched for serum cholesterol. There was no difference in LCAT rate between MI patients and controls. The relative content of arachidonic acid in cholesterol esters was higher in MI patients. The fatty acid composition of lecithin and cholesterol esters suggests an equal transfer of linoleic and oleic acids from lecithin to cholesterol. Furthermore negative correlations were found between LCAT and linoleic acid content of lecithin ( r = −0.43, P < 0.01) and cholesterol esters ( r = −0.45, P < 0.01). This inverse relationship does not seem to be linked to substrate specificity, but rather to be mediated by influences in common on serum lipid content and turnover.

Hepatic cholesterol ester hydrolase in human liver disease

Human liver contains an acid cholesterol ester hydrolase (CEH) of presumed lysosomal origin, but its significance is unknown. We developed a modified CEH radioassay suitable for needle biopsy specimens and measured hepatic activity of this enzyme in 69 patients undergoing percutaneous liver biopsy. Histologically normal livers hydrolyzed 5.80 ± 0.78 sem μmoles of cholesterol ester per hr per g of liver protein (n, 10). Values were similar in alcoholic liver disease (n, 17), obstructive jaundice (n, 9), and miscellaneous hepatic disorders (n, 21). In contrast, mean hepatic CEH activity was more than 3-fold elevated in 12 patients with acute hepatitis, 21.05 ± 2.45 sem μmoles per hr per g of protein (P < 0.01). In 2 patients studied serially, CEH returned to normal as hepatitis resolved. CEH activity in all patients paralleled SGOT levels (r, 0.84; P < 0.01). There was no correlation with serum levels of free or esterified cholesterol nor with serum activity of lecithin-cholesterol acyltransferase, the enzyme responsible for cholesterol esterification in plasma. These studies confirm the presence of CEH activity in human liver and show markedly increased activity in acute hepatitis. The pathogenesis and clinical significance of altered hepatic CEH activity in liver disease require further study.

Metabolic studies of adrenoleukodystrophy.

Two series of metabolic studies were prompted by the previous finding that the brain and adrenal tissues of patients with adrenoleukodystrophy, an X-linked genetic disorder, contain unusually long-chain (C22--C32) fatty acids in cholesterol esters and gangliosides. Postmortem brain tissues from three patients were assayed for activities of the three distinct cholesterol ester hydrolases, using [4-14C]cholesterol oleate, lignocerate and cerotate as the substrates. No deficiency of the crude mitochrondrial (pH 4.2), the microsomal (pH 6.0), or the myelin-localized cholesterol ester hydrolases was detected, although the activities of the myelin-localized cholesterol ester hydrolase against cholesteryl lignocerate and cerotate were too low for reliable assays. The activities of the microsomal and myelin-localized hydrolases were actually higher in adrenoleukodystrophy than in controls. Uptake and exclusion by cultured fibroblasts of [1-14C]stearic, [1-14C]lignoceric and [1-14C)cerotic acids were also examined. All fatty acids were avidly taken up by the fibroblasts. Stearic acid was excluded from the cells much more rapidly than lignoceric or cerotic acid. No difference was observed in the uptake and exclusion of fatty acids between the controls and adrenoleukodystrophy, except that cells from some cases of adrenoleukodystrophy consistently took up the very long chain fatty acids at greater rates than the control cells. Neither did the distribution of the label among individual lipids reveal differences between the controls and adrenoleukodystrophy, although there were interesting and dramatic differences in the metabolism of lignoceric acid and cerotic acid. Cerotic acid appeared largely inert with 90--95% remaining intact over eight days, while lignoceric acid was mostly incorporated into complex lipids. This series of studies did not uncover the fundamental genetic defect underlying adrenoleukodystrophy.

Adrenomyeloneuropathy

Histopathologic features were studied in postmortem examination of two men with adrenomyeloneuropathy, and biochemical studies were performed on one of these individuals. The histopathologic picture of one case included dying-back features in the nervous system and lamellar cytoplasmic inclusions in the brain, adrenal gland, and testis similar to those in adrenoleukodystrophy. Biochemical studies of the cerebral white matter of this individual revealed increased amounts of long-chain saturated fatty acids in cholesterol esters, an abnormality characteristic of adrenoleukodystrophy. Despite differences in clinical presentation and neuropathology, adrenomyeloneuropathy probably represents a distinct variant of adrenoleukodystrophy.

Uptake and degradation of cholesterol ester-labelled rat plasma lipoproteins in purified rat hepatocytes and nonparenchymal liver cells

1. 1. A new method for isolation and purification of rat liver hepatocytes and nonparenchymal cells by differential centrifugation is described. 2. 2. Cholesterol ester-labelled lipoproteins (prepared by the action of lecithin: cholesterol acyltransferase) intravenously injected were taken up by hepatocytes and nonparenchymal cells. 3. 3. Hepatocytes and nonparenchymal cells in suspension were able to take up and hydrolyse the cholesterol ester portion of lipoproteins. 4. 4. Uptake of cholesterol ester labelled whole rat plasma and high density lipoproteins (HDL) increased with increasing concentrations until a distinct saturation level was reached in hepatocytes. In nonparenchymal cells there was no saturation of lipoprotein uptake. 5. 5. Concanavalin A inhibited cholesterol ester-labelled lipoprotein uptake in hepatocytes, indicating that the uptake at least partially depends on carbohydrate sites on the cell surface. The uptake in nonparenchymal cells was unaffected of concanavalin A. 6. 6. The specific activity of the acid cholesterol ester hydrolase was the same in homogenates from hepatocytes and nonparenchymal cells while acyl-CoA: cholesterol acyltransferase was found almost exclusively in hepatocytes.

Ovipositional behavior ofBracon mellitor Say, a parasitoid of the boll weevil (Anthonomus grandis Boheman) III. Isolation and identification of natural releasers of ovipositor probing

The chemical cues in the frass of larvae of the boll weevil,Anthonomus grandis Boheman, that elicit an ovipositional response in females ofBracon mellitor Say were isolated by a combination of column, thin-layer, and gas-liquid chromatography. Derivatization, analytical tests, and mass spectrometry were used to identify the biologically active materials as long-chain fatty acid esters of cholesterol. Bioassays with natural and synthetic cholesteryl esters were used to confirm activity. The activity of the cholesteryl esters was also confirmed by using females that had not been previously exposed to hosts.

Stability of fatty acids in hyperlipoproteinemic plasma during long-term storage.

Aliquots of hyperlipoproteinemic plasma were stored at -20 degrees C in a nitrogen atmosphere with added disodium ethylenediaminetetraacetate for up to one year, to determine the stability of fatty acids in the four major lipid classes under these conditions. Assays were performed at zero time and at 2, 5, and 12 months. No uniform fatty acid change was found. Minor statistically significant changes were found in total triglyceride fatty acids and in some of the individual fatty acids in cholesterol esters, phospholipid, and free fatty acids. These minor changes could not be accounted for by plasma lipid concentrations, lipolysis, or fatty acid peroxidation. Evidently, plasma can be stored in this manner for as long as a year without substantial change in fatty acids in any of the four major lipid classes.

Fatty Acid Biosynthesis in Mouse Brain Microsomal Fractions

In demyelinating conditions cholesterol is believed to be esterified with fatty acids released by phospholipase action on the phosphatidylcholine of the myelin (Ramsey, 1973). This supposition is supported by Eto & Suzuki (1971), who demonstrated a strikingly similar fatty acid composition in the /?-linked fatty acids of phosphatidylcholine to those of cholesterol esters in brain undergoing demyelination. Demyelination is probably an inexact description of at least one of the congenital myelin disorders that we have studied. Thus, as a result of an electron-microscopic study (W. F. Blakemore, personal communication), it has been suggested that in congenital tremor type AIV, myelination ceases when oligodendrocytes undergo degenerative changes and that myelin breaks down at an early stage of development. Additionally it has been demonstrated that in this condition cerebrosides contain an abnormal range of fatty acids and also that linked with the increase of the C18:1 fraction of the cholesterol ester fatty acids there is a decrease in this particular fatty acid fraction in whole lipid extracts of spinal cord (Patterson et al., 1973). Therefore the present observations showing that cholesterol esters possess an almost universally similar fatty acid pattern in many unrelated pathological states, including congenital tremor type AIV, lends further support to the observations of Eto & Suzuki (1971) that the fatty acids of cholesterol esters must presumably originate in myelin lipid fractions other than cerebroside, probably phospholipids.

CONCENTRATIONS AND FATTY ACID COMPOSITION OF SERUM LIPID FRACTIONS FOLLOWING ACTH IN RABBITS

Abstract In a group of rabbits injection of ACTH over 5 days caused a rise in cholesterol, phospholipids and triglycerides (TG) in serum. Concomitant changes in fatty acid composition included an increase in linoleic acid in cholesterol esters and phospholipids, a lowering of oleic acid in all three fractions, and an increase in palmitic acid in TG. The changes in fatty acid composition may be accounted for by increased lipolysis of adipose tissue effected by ACTH with an increased influx to the liver of free fatty acids (FFA), rich in palmitic and linoleic acid, and relatively poor in oleic acid. Furthermore, inhibition by ACTH of fatty acid synthesis may have favoured the incorporation of linoleic acid at the expense of fatty acids synthetized in the liver. It is concluded that the changes in lipid concentrations and fatty acid composition after injecting ACTH in rabbits are quite different from those observed following serious illness in man. This may indicate that the degradation of low density lipoproteins, with lowering of cholesterol and phospholipids following serious illness in man, is not caused by ACTH or corticosteroids.

Interchangeability of fatty acids between phospholipids and esterified cholesterol in the plasma.

Experiments were performed to investigate incorporation of linoleic acid in lecithins probably synthesized from dietary linoleic acid within the intestinal wall and fatty acids in cholesterolesters, with plasma and liver homogenates pre-pared from albino rats.As a result, evidence has been obtained to demonstrate that linoleic acid con-tained in the α, β-dilinoleylphosphoryl choline molecule is incorporated into chole-sterol palmitate, and the palmitic acid in this cholesterolester into α, β-dilinoleyl-phosphoryl choline, respectively. These findings seem to suggest mutual inter-changeability of these fatty acids between the two forms of fat.