scholarly journals An alternative procedure for incorporating radiolabelled cholesteryl ester into human plasma lipoproteins in vitro

1985 ◽  
Vol 226 (1) ◽  
pp. 319-322 ◽  
Author(s):  
D C K Roberts ◽  
N E Miller ◽  
S G L Price ◽  
D Crook ◽  
C Cortese ◽  
...  
Keyword(s):  
Cholesteryl Ester ◽  
Human Plasma ◽  
Specific Radioactivity ◽  
Density Lipoprotein ◽  
Plasma Lipoproteins ◽  
Cholesteryl Esters ◽  
Simple Method ◽  
High Specific Radioactivity

A simple method has been developed for labelling human plasma lipoproteins to high specific radioactivity with radioactive cholesteryl esters in vitro. After isolation by preparative ultracentrifugation, the selected lipoprotein was incubated for 30 min at 4 degrees C in human serum (d greater than 1.215) that had been prelabelled with [4-14C]cholesteryl oleate or [1,2-3H]cholesteryl linoleate, and was then re-isolated by ultracentrifugation. All major lipoprotein classes were labelled by the procedure. Specific radioactivities of up to 18 d.p.m. pmol-1 (46 d.p.m. ng-1) were achieved. When radiolabelled high-density lipoprotein was infused intravenously, the radioactive cholesteryl ester behaved in vivo indistinguishably from endogenous cholesteryl esters produced by the lecithin (phosphatidylcholine): cholesterol acyltransferase reaction.

Rapid transformation of [3H]cholesteryl ester m rat high-density lipoprotein: in vivo and in vitro studies

Steroids ◽  
1990 ◽  
Vol 55 (7) ◽  
pp. 308-313
Author(s):  
I.J. Goldberg ◽  
R.S. Rosenfeld ◽  
I. Paul ◽  
L.K. Miller ◽  
M.L. Tiell
Keyword(s):  
High Density Lipoprotein ◽  
Cholesteryl Ester ◽  
Density Lipoprotein ◽  
High Density ◽  
In Vitro Studies ◽  
Rapid Transformation

scholarly journals Exogenous cholesterol transport in rabbit plasma lipoproteins

1973 ◽  
Vol 134 (2) ◽  
pp. 531-537 ◽  
Author(s):  
L. L. Rudel ◽  
J. M. Felts ◽  
M. D. Morris
Keyword(s):  
Free Cholesterol ◽  
Specific Radioactivity ◽  
Plasma Lipoproteins ◽  
Cholesterol Transport ◽  
Low Density ◽  
Cholesteryl Esters ◽  
Exogenous Cholesterol ◽  
Rate Of Increase ◽  
Chylomicron Metabolism

1. The appearance of exogenous cholesterol in free cholesterol and ester cholesterol of plasma chylomicra, very-low-density (VLD), low-density (LD) and high-density (HD) lipoproteins was studied in unanaesthetized rabbits after ingestion of a meal containing 5% fat and 0.08% [3H]cholesterol. 2. The specific radioactivity of ester cholesterol of VLD lipoproteins reached the highest value of any lipoprotein fraction and for each lipoprotein it increased at a faster rate and reached a higher maximum than that of free cholesterol; the maximum in VLD lipoproteins occurred later than in chylomicra. 3. The pattern of appearance of exogenous cholesterol in chylomicra and VLD lipoproteins of plasma was similar to the pattern previously observed in lymph. The specific radioactivity of ester cholesterol in plasma VLD lipoproteins was higher than that in chylomicra in spite of a larger pool size and dilution of cholesteryl esters from VLD lipoproteins produced by the liver. These results support the concept that during absorption the major portion of exogenous cholesterol is transported in VLD lipoproteins as ester cholesterol. 4. The specific radioactivity of ester cholesterol of chylomicra and VLD lipoproteins increased at a faster rate than that of LD and HD lipoproteins. However, the rate of increase and the absolute values of the specific radioactivity in LD and HD lipoproteins were identical. Since cholesteryl esters are thought not to exchange between lipoproteins, this observation supports the hypothesis that a result of VLD lipoprotein and chylomicron metabolism is the formation of LD and HD lipoproteins. 5. Results in vivo showed that the free cholesterol of individual plasma lipoproteins does not equilibrate within a period of 24h.

scholarly journals Reverse cholesterol transport in the isolated perfused rat spleen

1990 ◽  
Vol 268 (2) ◽  
pp. 499-505 ◽  
Author(s):  
M A Mindham ◽  
P A Mayes ◽  
N E Miller
Keyword(s):  
Whole Blood ◽  
Chemical Potential ◽  
Specific Radioactivity ◽  
Density Lipoprotein ◽  
Potential Gradient ◽  
Cholesterol Transport ◽  
Unesterified Cholesterol ◽  
Chemical Potential Gradient ◽  
High Specific Radioactivity

1. A method has been developed which enables the rat spleen to be loaded in vivo with [3H]cholesterol to a high specific radioactivity using cholesterol-labelled erythrocytes. The erythrocytes were shown to be rapidly degraded by the spleen and not released intact during subsequent perfusion. 2. When labelled spleens were perfused with whole blood or serum, lipoproteins in the high-density lipoprotein (HDL) range were shown to be the principal lipoprotein vehicles for the removal of cholesterol, the specific radioactivity of cholesterol being much greater in the HDL fractions than in other lipoproteins, particularly in the d 1.175-1.210 fraction. 3. The formation of [3H]cholesteryl ester was restricted to the major HDL fractions. 4. Experiments utilizing individual HDL fractions added to a basal perfusate indicated that HDL1 (d 1.050-1.085) was of less importance in the removal of cholesterol from the spleen than HDL subfractions of higher density. Also, a decrease in density of the lipoproteins was observed during perfusion, concurrent with uptake of cholesterol, especially in the d 1.085-1.125 subfraction. 5. When [3H]cholesterol-labelled spleens were perfused with whole blood, about half of the radioactivity released was detected in erythrocytes, indicating a rapid exchange or transport of cholesterol. Thus erythrocytes could play an important role in the transfer of unesterified cholesterol when the chemical potential gradient is favourable.

scholarly journals The separation of intracellular serum albumin from rat liver

1971 ◽  
Vol 123 (4) ◽  
pp. 643-648 ◽  
Author(s):  
J. D. Judah ◽  
Marion R. Nicholls
Keyword(s):  
Serum Albumin ◽  
Rat Liver ◽  
Specific Radioactivity ◽  
Exchange Chromatography ◽  
Rat Serum ◽  
Radioactive Impurity ◽  
Simple Method ◽  
High Specific Radioactivity ◽  
Chromatographic Procedure

1. Antibody precipitation of serum albumin from rat liver extracts yields impure preparations of the protein. 2. When rat liver is labelled with l-[1-14C]leucine, antibody precipitation of albumin leads to material that is contaminated with a protein or proteins of very high specific radioactivity. Only 10–25% of the radioactivity of the antibody precipitate is associated with serum albumin. 3. A chromatographic procedure is described that can be used to separate radiochemically pure serum albumin from antibody precipitates obtained from extracts of rat liver. 4. Extracellular albumin secreted by liver slices yields a precipitate with antibody which contains much less radioactive impurity. About 70–90% of the radioactivity is associated with serum albumin. Serum albumin separated by antibody precipitation from rat serum labelled in vivo was not contaminated with the radiochemical impurities associated with intracellular albumin. 5. A simple method is described of obtaining the content of serum albumin in rat liver extracts by the technique of isotope dilution and ion-exchange chromatography.

Use of in vivo and in vitro techniques for the study of the effects of insulin on hepatic triacylglycerol secretion in different insulinaemic states

2000 ◽  
Vol 28 (2) ◽  
pp. 103-109 ◽  
Author(s):  
V. A. Zammit
Keyword(s):  
Cultured Cells ◽  
Physiological State ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Cholesteryl Esters ◽  
In Vitro Techniques ◽  
Specific Targeting ◽  
Hepatic Triacylglycerol

This review illustrates how the use of several in vitro and in vivo techniques was necessary to show that the effect of insulin on hepatic triacylglycerol (TAG) secretion in the rat depends on the prior physiological state of the animal. The effect of insulin was always inhibitory when cultured cells were used, irrespective of the physiological state of the donor rats. By contrast, when perfused livers were used, insulin stimulated TAG secretion by livers isolated from fed, normoinsulinaemic rats, but inhibited it in livers from fasted or streptozotocin diabetic animals. This switch in insulin action was also shown to occur in vivo in experiments that involved the liver-specific targeting of both insulin (delivered within liposomes) and labelled fatty acids (delivered as cholesteryl esters within very-low-density lipoprotein remnants) in awake, unrestrained rats during a euglycaemic clamp. It is concluded that observations obtained with perfused liver preparations are more representative of the actual changes that occur in vivo with respect to the effects of insulin on hepatic TAG secretion.

Binding of thyroxine and 3,5,3'-triiodothyronine to trout plasma lipoproteins

1992 ◽  
Vol 262 (5) ◽  
pp. E712-E720 ◽  
Author(s):  
P. J. Babin
Keyword(s):  
Nutritional Status ◽  
Binding Sites ◽  
Density Lipoprotein ◽  
High Density ◽  
Plasma Lipoproteins ◽  
High Density Lipoproteins ◽  
Minor Role ◽  
Very Low Density Lipoproteins

The plasma vectors of thyroid hormones (TH) in trout have been characterized. Plasma components were separated by density gradient ultracentrifugation after first labeling binding sites with trace levels of radioactive hormones, both in vivo and in vitro. Lipoproteins play only a minor role in humans but are major carriers of thyroxine (T4) and 3,5,3'-triiodothyronine (T3) in trout plasma. More than 67% of T4 and 89% of T3 were bound to lipoproteins (density less than 1.210 g/ml), predominantly to high-density lipoproteins (HDL), regardless of the nutritional status of the animals. The percentage of hormone bound to very-low-density lipoproteins, on the other hand, was proportional to their concentration and thus to nutritional status. T3 and T4 could also bind to vitellogenin, a very-high-density lipoprotein, which could transfer TH to the yolk of oocytes. Homologous ligand displacement indicated that T3 could bind to at least two classes of saturable sites in the plasma. In addition, plasma HDL were the major binding sites with low affinity (1.7 +/- 0.4 x 10(5) M-1) but with high capacity (3.1 +/- 0.3 x 10(-5) M). In conclusion, these results show that lipoproteins are the principal binding sites of TH in trout plasma.

scholarly journals Increased selective uptake in vivo and in vitro of oxidized cholesteryl esters from high-density lipoprotein by rat liver parenchymal cells

1996 ◽  
Vol 319 (2) ◽  
pp. 471-476 ◽  
Author(s):  
Kees FLUITER ◽  
Helene VIETSCH ◽  
Eric A. L. BIESSEN ◽  
Gert M. KOSTNER ◽  
Theo J. C. van BERKEL ◽  
...  
Keyword(s):  
High Density Lipoprotein ◽  
Cholesteryl Ester ◽  
Density Lipoprotein ◽  
High Density ◽  
Biliary Secretion ◽  
Liver Uptake ◽  
Selective Uptake ◽  
Parenchymal Cells

Oxidation of low-density lipoprotein (LDL) leads initially to the formation of LDL-associated cholesteryl ester hydroperoxides (CEOOH). LDL-associated CEOOH can be transferred to high-density lipoprotein (HDL), and HDL-associated CEOOH are rapidly reduced to the corresponding hydroxides (CEOH) by an intrinsic peroxidase-like activity. We have now performed in vivo experiments to quantify the clearance rates and to identify the uptake sites of HDL-associated [3H]Ch18:2-OH in rats. Upon injection into rats, HDL-associated [3H]Ch18:2-OH is removed more rapidly from the circulation than HDL-associated [3H]Ch18:2. Two minutes after administration of [3H]Ch18:2-OH-HDL, 19.6±2.6% (S.E.M.; n = 4) of the label was taken up by the liver as compared with 2.4±0.25% (S.E.M.; n = 4) for [3H]Ch18:2-HDL. Organ distribution studies indicated that only the liver and adrenals exhibited preferential uptake of [3H]Ch18:2-OH as compared with [3H]Ch18:2, with the liver as the major site of uptake. A cell-separation procedure, employed 10 min after injection of [3H]Ch18:2-OH-HDL or [3H]Ch18:2-HDL, demonstrated that within the liver only parenchymal cells take up HDL-CE by the selective uptake pathway. Selective uptake by parenchymal cells of [3H]Ch18:2-OH was 3-fold higher than that of [3H]Ch18:2, while Kupffer and endothelial cell uptake of the lipid tracers reflected HDL holoparticle uptake (as analysed with iodinated versus cholesteryl ester-labelled HDL). The efficient uptake of [3H]Ch18:2-OH by parenchymal cells was coupled to a 3-fold increase in rate of radioactive bile acid secretion from [3H]Ch18:2-OH-HDL as compared with [3H]Ch18:2-HDL. In vitro studies with freshly isolated parenchymal cells showed that the association of [3H]Ch18:2-OH-HDL at 37 °C exceeded [3H]Ch18:2-HDL uptake almost 4-fold. Our results indicate that HDL-associated CEOH are efficiently and selectively removed from the blood circulation by the liver in vivo. The selective liver uptake is specifically exerted by parenchymal cells and coupled to a rapid biliary secretion pathway. The liver uptake and biliary secretion route may allow HDL to function as an efficient protection system against potentially atherogenic CEOOH.

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