Cholesteryl esters hydrolysis

Avart SJ, Bernard DW, Jerome WG et al. (1999) Cholesteryl ester hydrolysis in J774 macrophages occurs in the cytoplasm and lysosomes. J Lipid Res 40 405 14... 

Noel, A-P, R. Dupras, and A-M. Fillion. 1983. The activity of cholesteryl ester hydrolysis in the enzymatic determination of cholesterol Comparison of five enzymes obtained commercially. Clinical Chemistry 129 464-471. 

Wiebe, D. A., and J. T. Bernert. 1984. Influence of incomplete cholesteryl ester hydrolysis on enzymic measurements of cholesterol. Clinical Chemistry 30 352-356. 

Another pancreatic enzyme, cholesterol esterase, is responsible for intraluminal cholesteryl ester hydrolysis. Complete hydrolysis of dietary cholesteryl esters occurs in the lumen before absorption of cholesterol can occur (Shiratori and Goodman, 1965). This enzyme is discussed further below. 

The pancreatic enzyme mainly responsible for retinyl ester hydrolysis appears to be the same enzyme that catalyzes intraluminal cholesteryl ester hydrolysis (Erlanson and Boigstibm, 1968 Lombardo and Guy, 1980). This enzyme has been purified from rat (Calame et al.. 1975) and from porcine (Momsen and Brockman, 1977) pancreas, and from human pancreatic juice (Lombardo et al.. 1979). The enzyme appears to be a relatively nonspecific carboxylic ester hydrolase that can act on a wide variety of esters as substrates. Thus the purified human enzyme hydrolyzes triacetin, tributyrin, p-nitrophenylacetate, and lyso-phosphatidylcholine, as well as esters of cholesterol and of vitamins A, Dj, and E and glycerides solubilized by bile salts. Its molecular weight (approximately 100,000) is greater than that of the rat or pig enzyme, and it can hydrolyze... 

The anatomic sites (subcellularly) and the details of the enzymatic processes involved in the hydrolysis of chylomicron cholesteryl esters newly taken up by the liver have not been fully defined. It is clear that one of the major processes consists of receptor-mediated endocytosis of chylomicron remnants, followed by hydrolysis of cholesteryl esters and other remnant components within lysosomes. In rare genetic diseases characterized by lysosomal acid lipase deficiency (Wol-man s disease and cholesteryl ester storage disease), cholesteryl esters accumulate in liver cells and in other tissues as well [see Assmann and Frederickson (1983) for review and references]. An acid cholesteryl ester hydrolase from rat liver lysosomes has been partially purified and characterized (Brown and Sgoutas, 1980 Van Berkel etal., 1980). Enzymatic activity was found in preparations of both parenchymal and nonparenchymal liver cells (Van Berkel et al., 1980). Hydrolysis of chylomicron cholesteryl esters taken up by isolated rat hepatocytes was inhibited by chloroquine (Florin and Nilsson, 1977), an agent which inhibits the action of acid hydrolases in lysosomes. Finally, there is also evidence that the rate of cholesteryl ester hydrolysis may be limited by the rate at which internalized remnant particles are moved to the presumably lysosomal site of hydrolysis (Nilsson, 1977 Florin and Nilsson, 1977 Cooper and Yu, 1978). 

On the other hand, there are other processes that may participate as well, to some extent, in the initial metabolism and hydrolysis of chylomicron cholesteryl esters in the liver. Liver homogenates and homogenate fractions display cholesteryl ester hydrolase activity at neutral pH, and the enzyme(s) responsible for such activity have been partially purified and characterized (Deykin and Goodman, 1962 Stein et al., 1969 Tuhackova et al., 1980). It is possible that some uptake of cholesteryl esters can occur without uptake of the entire remnant particle [see, e.g., Chajek-Shaul et al. (I981a,b) for such evidence in other tissues]. It is also possible that dissociation of the constituents of the remnant can occur to some extent, permitting cholesteryl ester hydrolysis to take place before remnants are delivered to lysosomes. The extent to which these alternative processes might occur in normal physiology is not known. 

Belkner et al. [32] demonstrated that 15-LOX oxidized preferably LDL cholesterol esters. Even in the presence of free linoleic acid, cholesteryl linoleate continued to be a major LOX substrate. It was also found that the depletion of LDL from a-tocopherol has not prevented the LDL oxidation. This is of a special interest in connection with the role of a-tocopherol in LDL oxidation. As the majority of cholesteryl esters is normally buried in the core of a lipoprotein particle and cannot be directly oxidized by LOX, it has been suggested that LDL oxidation might be initiated by a-tocopheryl radical formed during the oxidation of a-tocopherol [33,34]. Correspondingly, it was concluded that the oxidation of LDL by soybean and recombinant human 15-LOXs may occur by two pathways (a) LDL-free fatty acids are oxidized enzymatically with the formation of a-tocopheryl radical, and (b) the a-tocopheryl-mediated oxidation of cholesteryl esters occurs via a nonenzymatic way. Pro and con proofs related to the prooxidant role of a-tocopherol were considered in Chapter 25 in connection with the study of nonenzymatic lipid oxidation and in Chapter 29 dedicated to antioxidants. It should be stressed that comparison of the possible effects of a-tocopherol and nitric oxide on LDL oxidation does not support importance of a-tocopherol prooxidant activity. It should be mentioned that the above data describing the activity of cholesteryl esters in LDL oxidation are in contradiction with some earlier results. Thus in 1988, Sparrow et al. [35] suggested that the 15-LOX-catalyzed oxidation of LDL is accelerated in the presence of phospholipase A2, i.e., the hydrolysis of cholesterol esters is an important step in LDL oxidation. 

This is a broad class of enzymes that catalyze the hydrolysis of esters some of these enzymes are quite specific. See also Cholesteryl Ester Hydrolase specific esterase... 

CHOLESTERYL ESTER HYDROLASE ESTER HYDROLYSIS MECHANISMS ETA (7,)... 

Chylomicrons transport dietary triacylglycerol and cholesteryl ester from the intestine to other tissues in the body. Very-low-density lipoprotein functions in a manner similar to the transport of endogenously made lipid from the liver to other tissues. These two types of triacylglycerol-rich particles are initially degraded by the action of lipoprotein lipase, an extracellular enzyme that is most active within the capillaries of adipose tissue, cardiac and skeletal muscle, and the lactating mammary gland. Lipoprotein lipase catalyzes the hydrolysis of triacylglycerols (see fig. 18.3). The enzyme is specifically activated by apoprotein C-II, which... 

Camarota, L. M., Chapman, J. M., Hui, D. Y., and Howies, P. N. (2004) Carboxyl ester lipase coffactionates with scavenger receptor BI in hepatocyte lipid rafts and enhances selective uptake and hydrolysis of cholesteryl esters from HDL3. J. Biol. Chem. 279, 27599-27606. 

In the next step, the cholesteryl ester entities copolymerized in the shell, were split by carbonate ester hydrolysis. The hydrolysis was carried out in sodium hydroxide in methanol. Thereby, an analogue of cholesterol, the target molecule for later recognition, was removed from the copolymer network. The particles were now ready for non-covalent binding of cholesterol. To quantify the binding behavior of colloidal MIPs, they were mixed with a cholesterol containing solution, separated from the liquid and the cholesterol concentration in the supernatant was quantified by HPLC. 

LDL binds specifically to lipoprotein receptors on the cell surface. The resulting complexes become clustered in regions of the plasma membrane called coated pits. Endocytosis follows (see Fig. 13-3). The clathrin coat dissociates from the endocytic vesicles, which may recycle the receptors to the plasma membrane or fuse with lysosomes. The lysosomal proteases and lipases then catalyze the hydrolysis of the LDL-receptor complexes the protein is degraded completely to amino acids, and cholesteryl esters are hydrolyzed to free cholesterol and fatty acid. New LDL receptors are synthesized on the endoplasmic reticulum (ER) membrane and are subsequently reintroduced into the plasma membrane. The cholesterol is incorporated in small amounts into the endoplasmic reticulum membrane or may be stored after esterification as cholesteryl ester in the cytosol this occurs if the supply of cholesterol exceeds its utilization in membranes. Normally, only very small amounts of cholesteryl ester reside inside cells, and the majority of the free cholesterol is in the plasma membrane. 

Surface materials from IDL, including some phospholipids, free cholesterol, and apolipoproteins, are transferred to HDL, or form HDL de novo in the circulation. Cholesteryl esters are transferred from HDL to LDL. The net result of the coupled lipolysis and the cholesteryl esters exchange reaction is the replacement of much of the triglyceride core of the original VLDL with cholesteryl esters. IDL undergoes a further hydrolysis in which most of the remaining triglycerides are removed and all apolipoproteins except B-lOO are transferred to other hpoproteins. This process ends with ultimate formation of LDL. 

A number of substituted benzenesulfonic acid esters and p-chlorophenoxy-isobutyric acid esters produced hypolipidemic activity.35 Among a series of p-alkoxybenzoic acids, enhanced hypotriglyceridemic and hypocholesterol-emic activity was observed with oximino- (14) and chloro- (15) substituents.98 Hypocholesterolemia was observed after the administration of 16 to rats.97 Tetronic 701, a polymeric surfactant, also lowered serum cholesterol the tetrabenzoate of Tetronic 701 is of particular interest, since it produced comparatively less growth depression.98 Two linoleamide derivatives, (-)N-fa-phenyl-B-(p-tolyl)ethylllinoleamide (PTLA) and N-(a-methyl-benzoyl)linoleamide, suppressed serum and liver cholesterol levels in rats and inhibited cholesterol absorption b interfering with the hydrolysis of cholesteryl esters in the intestine.99 101... 

The initial steps of reverse cholesterol transport involve export of cholesterol from peripheral cells to plasma lipoproteins for subsequent delivery to the liver. In vivo, HDL or its apolipoproteins act as acceptors of cholesterol from peripheral cells, carrying it to the liver for degradation. When cholesterol acceptors such as HDL are present, cholesterol efflux from macrophages is accelerated, which prevents foam cell formation. To produce this efflux, neutral cholesteryl ester hydrolase catalyzes intracellular hydrolysis of cholesteryl esters into free cholesterol in the lysosome (Avart et al. 1999). 

Chao FF, Blanchette-Mackie EI, et al. (1992) Hydrolysis of cholesteryl ester in low density hpoprotein converts this hpoprotein to a hposome. I Biol Chem 267 4992-4998... 

After partial hydrolysis in the gut, dietary fatty acids, monoacylglycerols, phospholipids, and cholesterol are absorbed into the mucosal enterocytes lining the small intestine (Chapter 12). Once within the cell, the lipids are reesterified and form a lipid droplet within the lumen of the smooth endoplasmic reticulum. These droplets consists of triacylglycerol and small amounts of cholesteryl esters and are stabilized by a surface film of phospholipid. At the junction of the smooth and the rough endoplasmic reticulum, the droplet acquires apoproteins B-48, A-I, A-II, and A-IV, which are produced in the lumen of the rough endoplasmic reticulum in the same way as other proteins bound for export. The lipoprotein particle is then transported to the Golgi stacks where further processing yields chylomicrons, which are secreted into the lymph and then enter the blood circulation at the thoracic duct. 

Essential non-steroidal isoprenoids, such as dolichol, prenylated proteins, heme A, and isopentenyl adenosine-containing tRNAs, are also synthesized by this pathway. In extrahepatic tissues, most cellular cholesterol is derived from de novo synthesis [3], whereas hepatocytes obtain most of their cholesterol via the receptor-mediated uptake of plasma lipoproteins, such as low-density lipoprotein (LDL). LDL is bound and internalized by the LDL receptor and delivered to lysosomes via the endocytic pathway, where hydrolysis of the core cholesteryl esters (CE) occurs (Chapter 20). The cholesterol that is released is transported throughout the cell. Normal mammalian cells tightly regulate cholesterol synthesis and LDL uptake to maintain cellular cholesterol levels within narrow limits and supply sufficient isoprenoids to satisfy metabolic requirements of the cell. Regulation of cholesterol biosynthetic enzymes takes place at the level of gene transcription, mRNA stability, translation, enzyme phosphorylation, and enzyme degradation. Cellular cholesterol levels are also modulated by a cycle of cholesterol esterification mediated by acyl-CoA cholesterol acyltransferase (ACAT) and hydrolysis of the CE, by cholesterol metabolism to bile acids and oxysterols, and by cholesterol efflux. 

The LDL receptor is a key component in the feedback-regulated maintenance of cholesterol homeostasis [1]. In fact, as an active interface between extracellular and intracellular cholesterol pools, it is itself subject to regulation at the cellular level (Fig. 2). LDL-derived cholesterol (generated by hydrolysis of LDL-bome cholesteryl esters) and its intracellularly generated oxidized derivatives mediate a complex series of feedback control mechanisms that protect the cell from over-accumulation of cholesterol. First, (oxy)sterols suppress the activities of key enzymes that determine the rate of cellular cholesterol biosynthesis. Second, the cholesterol activates the cytoplasmic enzyme acyl-CoA cholesterol acyltransferase, which allows the cells to store excess cholesterol in re-esterified form. Third, the synthesis of new LDL receptors is suppressed, preventing further cellular entry of LDL and thus cholesterol overloading. The coordinated regulation of LDL receptors and cholesterol synthetic enzymes relies on the sterol-modulated proteolysis of a membrane-bound transcription factor, SREBP, as described in Chapter 14. 

Other oxidation products include epoxides that can arise from hydroperoxide rearranganent [25-27], but are also formed by enzymic processes [28,29], Products of lipid oxidation may be quantified after formation from simple lipids, for example, oxidation of cholesterol or PUFA. These lipids may be components of more complex lipids like cholesteryl esters [30] or PUFA-containing phospholipids [31]. An analysis of the oxidized products can be performed on the intact lipid or after the individual Upid components are separated by hydrolysis. 

In many instances, oxidized products are considerably more stable when formed in complex lipids, like phospholipids or cholesteryl esters, and alkaline hydrolysis is required to release the oxidized lipid for analysis. Separation and quantitation of mono-hydroxyicosanoids by HPLC has been regularly used for the last 30 years. The procedures have become routine and there are now published procedures for automating the analysis [60]. A recent review by Yin et al. [36] describes in detail how to quantify both mono-hydroxyeicosatetraenoates and F2-isoprostanes. Similar procedures can be used to separate and identify products derived from (5Z,8Z,llZ,14Z,17Z)-eicosa-5,8,11,14,17-pentenoic acid, (7Z,10Z,13Z,16Z,19Z)-docosa-7,10,13,16,19-pentaenoic acid, and (4Z,7Z,10Z,13Z,16Z,19Z)-docosa-4,7,10,13,16,19-hexaenoic acid. 

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