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J H Shand

Publications and source records attributed to J H Shand.

At least 37 records · Page 2Linked to original sources
PubMed1993

Inhibition of neutral cholesteryl ester hydrolase by a naturally occurring cytosolic protein in macrophages.

The soluble fraction from a number of macrophage cell lines has been shown to contain a protein(s) that inhibits neutral cholesteryl ester hydrolase. The inhibition is dependent on the concentration of soluble protein used, and the efficiency of the inhibitor can be influenced by the inclusion of oleate and an inhibitor of ACAT. It is suggested that the presence of this material indicates that the macrophage contains a means of negatively controlling the activity of the hydrolytic phase of the cholesterol/cholesteryl ester cycle.

Animals↗
PubMed1993

The esterification of cholesterol in the yolk sac membrane of the chick embryo.

The uptake of lipid from the yolk by the yolk sac membrane of the chick embryo is accompanied by the rapid esterification of a large proportion of the yolk cholesterol. This could arise from enhanced acyl-CoA:cholesterol acyltransferase (ACAT) activity and/or inhibition of cholesteryl ester hydrolase (CEH) activity. The activity of ACAT was therefore measured in microsomes obtained from yolk sac membranes at various stages of development. A high level of activity (up to 929 pmol of cholesteryl oleate formed per min per mg protein) was found during the second half of this period. Supplementation with exogenous cholesterol stimulated ACAT activity in microsomes obtained from the tissue at the earlier, but not at the later, stages of development suggesting that the enzyme became saturated with microsomal cholesterol as development proceeded. Correlating with this, the concentration of cholesterol in the microsomes increased 4-fold between 9 and 20 d of development. The activity of CEH was very low in the microsomes and could not be detected in the cytosolic fraction. The activity of a protein, which has been shown to function as an inhibitor of CEH, was found to be present at all stages of development. The high activity of ACAT, together with the low activity of CEH and an active CEH inhibitor protein is a combination well suited to promote an essentially unidirectional conversion of cholesterol to cholesteryl ester. This process may be a major determinant of the rate of lipid transfer from the yolk to the embryo.

Animals↗
PubMed1992

Characterization of a cytosolic protein in rat liver inhibiting neutral cholesteryl ester hydrolase.

Neutral cholesteryl ester hydrolase activity (EC 3.1.1.13) present in microsomes isolated from lactating rat mammary glands was found to be inhibited by a factor (or factors) occurring in the cytosolic fraction of male rat liver. The inhibitor was heat-labile, non-dialyzable, destroyed by proteolysis, and was stable following preparation of an acetone/diethyl ether powder of the cytosolic fraction. The protein also inhibited the activity of hormone-sensitive lipase (HSL) (from bovine adipose tissue) and esterase from Candida cylindracea, but seemed to be more active against the neutral hydrolase found in rat liver microsomes. For the mammary gland microsomal cholesteryl ester hydrolase, the extent of the inhibitory effect was dependent on the concentration of the cytosolic protein, 50% inhibition being achieved by about 100 micrograms of cytosolic protein, and on the method of initiating the enzyme assay. Kinetic analysis indicated that, under circumstances where the reaction was initiated by the addition of substrate, the inhibition was characterized as "uncompetitive." When an inhibitor/substrate complex was allowed to form in the absence of enzyme, an element of "competitive" inhibition was introduced into the reaction. Food withdrawal reduced the activity of the inhibitor in liver by 56%, but activity was fully restored by short-term re-feeding. In contrast, feeding a diet high in fat led to a 34% increase in activity. The present findings suggest that the inhibitory factor(s) may be involved in the regulation of the hydrolysis of cholesteryl esters in the liver and also in other cell types.

Animals↗
PubMed1992

The inhibition of neutral cholesteryl ester hydrolase by a cytosolic protein factor in female rat liver: the influence of varying hormonal and nutritional conditions on the inhibitory activity.

A cytosolic protein, that is inhibitory to neutral cholesteryl ester hydrolase, has been investigated in the livers of female rats using microsomes isolated from the mammary gland of lactating rats as an enzyme source. To facilitate comparisons, inhibitory activity is expressed in terms of the amount (micrograms) of cytosolic protein required to reduce esterase activity by 50% and is compared to the hepatic content of both cholesterol and cholesteryl esters. The experiments revealed a sexual difference in the level of inhibitory activity, with the livers of both suckling and mature male animals containing less of the material than the corresponding females. Alterations in the physiological status of the females, such as pregnancy and lactation, led to a decrease in the activity of the protein. This was reversed by blocking lactation with a combination of an antiserum to rat growth hormone and the anti-prolactin drug, bromocriptine, but not by premature weaning of the animals. Food withdrawal for 24 hr also had the effect of increasing inhibitory activity. In general the cholesteryl ester content of the livers correlated with the level of inhibitory activity. Thus the activity of the cytosolic inhibitor of neutral cholesteryl ester hydrolase responded to changes in both the hormonal and the nutritional status of the female animal. It is suggested that the presence of the greater cholesteryl ester hydrolase inhibitory activity in the female liver may help to explain the lower risk of coronary heart disease in premenopausal females by facilitating increased hepatic storage of the sterol in the form of the ester.

Animal Nutritional Physiological Phenomena↗
PubMed1991

Acyl-CoA: cholesterol acyltransferase activity in the rat mammary gland: variation during pregnancy and lactation.

Cholesterol esterification by acyl-CoA:cholesterol acyltransferase (ACAT; EC 2.3.1.26) has been studied in microsomes isolated from the mammary glands of rats in late pregnancy, in early and mid-lactation, and following premature weaning. The mammary glands were freeze-clamped and the microsomes prepared in the presence of phosphatase inhibitors to preserve the phosphorylation status of the enzyme. Optimal conditions were established for the assay of enzyme activity in the presence of endogenous cholesterol. Supplementation of the microsomes with exogenous cholesterol as a dispersion in Triton WR-1339 was shown to lead to an increase in enzyme activity. Incubation of microsomes with MgATP led to an increase in ACAT activity which could be reversed by treatment of the microsomes with a phosphoprotein phosphatase preparation from rat liver. The results suggested that ACAT activity in the mammary gland was activated by phosphorylation in a similar way to that observed for the hepatic enzyme. The mammary glands from pregnant animals contained a higher level of ACAT activity than did the glands of the lactating animals and this correlated with the higher cholesteryl ester content of the pregnant glands. The highest level of ACAT activity was found in the weaned animals but the cholesteryl ester content of the microsomes was lower than expected. The influence of progesterone levels and changes in feeding patterns during gestation were considered as factors in these variations in ACAT activity.

Animals↗
PubMed1991

Cholesterol ester hydrolase activity in mammary tissue of the lactating rat.

Cholesterol ester hydrolase activity has been studied in mammary glands of rats. Subcellular fractionation of the glands obtained in mid-lactation indicated that around 80% of the recovered activity was associated with particulate fractions. Two distinct cholesterol ester hydrolase activities were identified, one with an optimum pH of 7.5-9.0 and the second (approximately 5% of the total activity) with a more acidic pH optimum. Although the neutral cholesterol ester hydrolase had some properties in common with the lipoprotein lipase in mammary tissue, it was shown to be a separate entity by several criteria. Its activity could be increased following treatment with Mg-ATP and cAMP-dependent protein kinase, suggesting identity with the hormone sensitive lipase of adipose tissue. The cholesterol ester hydrolase activity in mammary glands just after parturition was greater than in glands obtained either from late-pregnant or midlactating animals. The subcellular distribution of the neutral cholesterol ester hydrolase suggested that it may have a different function to the neutral cholesterol ester hydrolase of adrenals and other tissues. Nevertheless the fact that the activity of the enzyme can be modulated by cAMP-dependent protein kinase suggests the possibility that hormonal control of this enzyme may be involved in the regulation of cholesterol metabolism in the mammary gland.

Adenosine Triphosphate↗
PubMed1991

Effects of an antiserum to rat growth hormone and bromocriptine on cholesterol-metabolizing enzymes in the lactating rat mammary gland.

Rats in mid-lactation were treated, for 2 days, with anti-rat GH serum (anti-rGH) and/or bromocriptine before microsomes were prepared from the freeze-clamped mammary glands. The effects of these anti-hormone treatments on the concentrations of microsomal cholesterol and cholesterol esters and on the activities of acyl-CoA:cholesterol acyltransferase (ACAT), neutral cholesterol ester hydrolase and 3-hydroxy-3-methylglutaryl-CoA reductase (HMG-CoA reductase) were measured. HMG-CoA reductase was determined in microsomes prepared in both the presence and absence of phosphatase inhibitors to determine the expressed and total activities respectively. Anti-rGH reduced HMG-CoA reductase and increased microsomal cholesterol and cholesterol esters. Bromocriptine reduced HMG-CoA reductase but increased all of the other parameters. The results indicate that the initial stage in the stimulation of milk secretion involves a decrease in the activity of ACAT and that the phosphorylation level of HMG-CoA reductase is modulated by both prolactin and GH acting in opposition.

Animals↗
PubMed1987

Measurement of 14CO2 evolution during tissue oxidation in vitro; an alternative to the Kontes well.

1. An alternative method to the use of the disposable Kontes well for trapping 14CO2 produced in the course of biological oxidations is described. 2. A polyethylene miniature scintillation vial was used to contain the hyamine hydroxide-impregnated filter paper wick. 3. The two methods are compared in their abilities to trap 14CO2 produced directly by acidification of sodium [14C]bicarbonate and during beta-oxidation of 1[14C] palmitic acid. 4. The miniature vial and Kontes well methods showed similar efficiencies in the trapping of 14CO2 (97% and 95%, respectively, on average) the radioactivity of which was determined in the miniature vial using 5 ml only of scintillation fluid compared with a minimum of 10 ml required by the standard scintillation vial used to accommodate the Kontes well. 5. The technical advantages of the suggested miniature vial system, during both incubation and counting stages, are discussed.

Animals↗