Biomedical subjects

K A Rye

Publications and source records attributed to K A Rye.

47 records · Page 3Linked to original sources

PubMed1993

Relationship between the size and phospholipid content of low-density lipoproteins.

In studies performed in vivo and in vitro, it has been found that the Stokes' diameter of human low-density lipoproteins (LDL) correlates positively and significantly with the molar ratio of phospholipid/apo B in LDL but not with the LDL molar ratios of either cholesterol/apo B or triacylglycerol/apo B. It has been concluded that the phospholipid content of LDL is an important determinant of LDL size.

Apolipoproteins B↗

PubMed1992

Changes in the size of reconstituted high density lipoproteins during incubation with cholesteryl ester transfer protein: the role of apolipoproteins.

It has been reported previously that the particle size distribution of discoidal, reconstituted HDL (r-HDL) changes dramatically during incubation in vitro with cholesteryl ester transfer protein (CETP). The present study was undertaken in order to determine whether these changes are influenced by the apolipoprotein composition of the r-HDL. Two preparations of r-HDL that contained egg phosphatidylcholine (egg PC) and unesterified cholesterol (UC) but differed in their apolipoprotein composition were used for the study. One preparation contained apolipoprotein (apo) A-I only (A-I w/o A-II r-HDL) while the other contained apoA-I and apoA-II (A-I w A-II r-HDL). The Stokes' radius of the major population of particles in the (A-I w/o A-II) and (A-I w A-II) r-HDL was, respectively, 4.8 and 4.9 nm. When the (A-I w/o A-II) r-HDL were incubated with CETP, most of the particles of radius 4.8 nm were converted to populations of smaller and larger particles. The smaller particles had Stokes' radii of 4.3 and 3.9 nm. The radii of the larger particles ranged from 8.2 to 13.7 nm. When the (A-I w A-II) r-HDL were incubated with CETP larger particles (Stokes' radii = 8.4-11.0 nm) appeared but there was minimal conversion to smaller particles. In addition, a significant proportion of the original (A-I w A-II) r-HDL of radius 4.9 nm was still present at the end of the incubation. These results are consistent with apoA-II inhibiting the conversion of r-HDL to small particles. It is concluded that the apolipoprotein content of r-HDL is an important determinant of the sizes of the particles that are formed during incubation with CETP.

Apolipoproteins↗

PubMed1990

The interaction of cholesteryl ester transfer protein and unesterified fatty acids promotes a reduction in the particle size of high-density lipoproteins.

Purified human cholesteryl ester transfer protein (CETP) has been found, under certain conditions, to promote changes to the particle size distribution of high-density lipoproteins (HDL) which are comparable to those attributed to a putative HDL conversion factor. When preparations of either the conversion factor or CETP are incubated with HDL3 in the presence of very-low-density lipoproteins (VLDL) or low-density lipoproteins (LDL), the HDL3 are converted to very small particles. The possibility that the conversion factor may be identical to CETP was supported by two observations: (1) CETP was found to be the main protein constituent of preparations of the conversion factor and (2) an antibody to CETP not only abolished the cholesteryl ester transfer activity of the conversion factor preparations but also inhibited changes to HDL particle size. In additional studies, the changes to HDL particle size promoted by purified CETP were inhibited by the presence of fatty-acid-free bovine serum albumin; by contrast, albumin had no effect on the cholesteryl ester transfer activity of the CETP. The possibility that albumin may inhibit changes to HDL particle size by removing unesterified fatty acids from either the lipoproteins or CETP was tested by adding exogenous unesterified fatty acids to the incubations. In incubations of HDL with either VLDL or LDL, sodium oleate had no effect on HDL particle size. However, when CETP was also present in the incubation mixtures the capacity of CETP to reduce the particle size of HDL was greatly enhanced by the addition of sodium oleate. It is concluded that the changes in HDL particle size which were previously attributed to an HDL conversion factor can be explained in terms of the interacting effects of CETP and unesterified fatty acids.

Adult↗

PubMed1990

Evidence in vitro that hepatic lipase reduces the concentration of apolipoprotein A-I in rabbit high-density lipoproteins.

Incubation of rabbit plasma in vitro with hepatic lipase resulted in the hydrolysis of triacylglycerol in high-density lipoproteins (HDL) and a reduction in HDL particle size. These changes were accompanied by a decrease in the concentration of apolipoprotein A-I (apo A-I) in the HDL. The loss of apo A-I was demonstrated independently by ultracentrifugation, size exclusion chromatography and gradient gel-immunoblot analysis. It was unrelated to hydrolysis of HDL phospholipids but did correlate with the reduction in HDL particle size. These studies suggest that the concentration of apo A-I in HDL may be influenced by factors which regulate the metabolism of HDL core lipid constituents.

Animals↗

PubMed1990

Interaction of apolipoprotein A-II with recombinant HDL containing egg phosphatidylcholine, unesterified cholesterol and apolipoprotein A-I.

The preparation of discoidal, recombinant HDL (r-HDL) containing various phospholipids, apolipoproteins and a range of concentrations of unesterified cholesterol has been reported by several investigators. The present study describes the preparation of r-HDL containing both apolipoprotein (apo) A-I and apo A-II. r-HDL with 100:1 (mol:mol) egg PC.apo A-I and 0 (Series I), 5 (Series II) or 10 (Series III) mol% unesterified cholesterol were prepared by the cholate dialysis method. The resulting complexes had a Stokes' radius of 4.7 nm and contained two molecules of apo A-I per particle. When the r-HDL (2.0 mg apo A-I) were supplemented with 1.0 mg of apo A-II, one of the apo A-I molecules was replaced by two molecules of apo A-II. This modification was not accompanied by a loss of phospholipid, nor by major change in particle size. The addition of 2.5 or 4.0 mg of apo A-II resulted in the displacement of both apo A-I molecules from a proportion of the r-HDL and the formation of smaller particles (Stokes' radius 3.9 nm), which contained half the original number of egg PC molecules and three molecules of apo A-II. The amount of apo A-I displaced was dependent on the concentration of unesterified cholesterol in the r-HDL: when 2.5 mg of apo A-II was added to the Series I, II and III r-HDL, 44, 60 and 70%, respectively, of the apo A-I was displaced. Addition of 4.0 mg of apo A-II did not promote further displacement of apo A-I from any of the r-HDL. By contrast, the association of apo A-II with r-HDL was independent of the concentration of unesterified cholesterol and was a linear function of the amount of apo A-II which had been added. It is concluded that (1), the structural integrity of egg PC.unesterified cholesterol.apo A-I r-HDL, which contain two molecules of apo A-I, is not affected when one of the apo A-I molecules is replaced by two molecules of apo A-II; (2), when both apo A-I molecules are replaced by apo A-II, small particles which contain three molecules of apo A-II are formed; and (3), the displacement of apo A-I from r-HDL is facilitated by the presence of unesterified cholesterol in the particles.

Apolipoprotein A-I↗

PubMed1989

Interaction of the high density lipoprotein conversion factor with recombinant discoidal complexes of egg phosphatidylcholine, free cholesterol, and apolipoprotein A-I.

An HDL conversion factor which promotes the conversion of HDL3 to populations of larger and smaller particles has recently been identified in human plasma. In the present report a partially purified preparation of this factor has been used to examine the conversion of 79:0:1, 77:5:1, and 74:10:1 (mol:mol:mol) egg phosphatidylcholine-free cholesterol-apolipoprotein A-I (apoA-I) recombinant discoidal complexes. The study was carried out in order to ascertain whether the conversion process is regulated by the concentration of free cholesterol in the complexes. The complexes comprised one major and two minor populations of particles with respective Stokes' diameters of 96 A, 84 A, and 78 A. The 74:10:1 complexes also contained a population of particles 112 A in diameter. The 79:0:1 and 77:5:1 complexes contained two molecules of apoA-I per particle. The 74:10:1 complexes comprised two classes of particles with two or three molecules of apoA-I. When the 74:10:1 complexes were incubated with the conversion factor, the 96 A and 84 A particles were converted to a population of particles 78 A in diameter that contained two apoA-I molecules. In the case of the 79:0:1 and 77:5:1 complexes, the 96 A particles were converted to 78 A particles but the concentration of 84 A particles did not change. The rate of conversion of 96 A particles to 78 A particles was dependent on the concentration of free cholesterol in the complexes. When the 74:10:1 complexes were incubated for 24 hr with the conversion factor, the 96 A particles were completely converted to particles 78 A in diameter. In the case of the 77:5:1 complexes, complete conversion was achieved by 48 hr. Conversion of the 79:0:1 complexes did not proceed to completion, even when the incubation was extended beyond 48 hr. The rate of conversion of 96 A particles to 78 A particles was also dependent on the concentration of the conversion factor in the incubation mixtures. The previous incubations contained equivalent concentrations of apoA-I and conversion factor. When the concentration of the conversion factor relative to apoA-I was reduced, there was a concomitant decrease in the rate of conversion of 96 A particles to 78 A particles. Conversion was not evident when the concentration of the conversion factor was reduced to one-tenth that of apoA-I.(ABSTRACT TRUNCATED AT 400 WORDS)

Apolipoproteins A↗

PubMed1988

Isolation of a high-density-lipoprotein conversion factor from human plasma. A possible role of apolipoprotein A-IV as its activator.

1. A high-density-lipoprotein (HDL) conversion factor was partially purified from human plasma by precipitation with (NH4)2SO4, ultracentrifugation, cation-exchange chromatography, anion-exchange chromatography and chromatography on a column of hydroxyapatite. 2. This factor modulates the particle size of HDL by converting a homogeneous population into new populations of particles, some of which are smaller and others larger than those in the original population. 3. The isolated HDL conversion factor appeared as one major band and at least three minor bands on SDS/polyacrylamide-gel electrophoresis; attempts to purify this factor further resulted in loss of conversion activity. 4. Preparations of the HDL conversion factor were stable after heating to 58 degrees C for 1 h, and were shown not to possess proteolytic activity. 5. The conversion factor was distinct from the known apolipoproteins, none of which had HDL conversion activity. 6. Addition of apolipoprotein A-IV had a dose-dependent potentiating effect on the process promoted by the HDL conversion factor.

Apolipoproteins A↗

PubMed1988

Lipid transfers between reconstituted high density lipoprotein complexes and low density lipoproteins: effects of plasma protein factors.

In this study we examined the transfer of lipids between reconstituted high density lipoprotein discs (r-HDL) and human low density lipoproteins (LDL) in the presence and absence of lecithin:cholesterol acyltransferase (LCAT) or of plasma phospholipid transfer protein (PLTP). We found that spontaneous transfer of phospholipids from r-HDL to LDL occurred by an apparent first order reaction with a half-time of 5.8 to 6.9 hr depending on the phospholipid. During the time of incubation of r-HDL with LDL (from 0 to 25 hr), the phospholipid content of r-HDL decreased more than 30%, the free cholesterol content increased 2.5-fold, and low levels of cholesteryl esters appeared in r-HDL. These compositional changes gave rise to small discoidal particles with a limiting diameter of 77 A and two molecules of apoA-I per particle. When LCAT was included in the reaction mixture, the r-HDL lost even more phospholipid, lost some free cholesterol, and gained cholesteryl esters relative to the apolipoprotein content, due to the enzymatic reaction. The products of the LCAT reaction had a diameter of 93 A and three, rather than two, apoA-I molecules per particle. Inclusion of PLTP into the reaction mixture accelerated the transfer of phospholipids (half-time of 1.7 hr) and the formation of the 77 A product. In addition to these compositional and morphological changes, which may be important in the interconversions of native HDL subspecies, the prolonged incubations revealed some slow reactions, such as the esterification of LDL cholesterol by LCAT, a background formation of cholesteryl esters in r-HDL, and an apparent hydrolysis of cholesteryl esters in LDL in the presence of r-HDL.

Apolipoprotein A-I↗

PubMed1986

Changes in the size and density of human high-density lipoproteins promoted by a plasma-conversion factor.

Incubation of human high-density lipoprotein subfraction-3 (HDL3) with rabbit lipoprotein-depleted plasma resulted in marked changes in the density and size of the HDL. After 24 h of incubation at 37 degrees C, the original HDL3 were converted into populations of larger (less dense) and smaller (more dense) particles. The degree of conversion increased with increasing concentrations of lipoprotein-depleted plasma and increasing incubation time. Furthermore, lecithin:cholesterol acyltransferase, lipoprotein lipase and lipid-transfer protein were shown not to be involved in the process. It was therefore proposed that a separate factor, the HDL-conversion factor, was responsible for the observed changes. Conversion-factor activity was assessed in the lipoprotein-depleted plasma of several species and found to be greater in rabbits and rats than in pigs and human subjects. It was also established that the conversion factor was able to be precipitated from rabbit lipoprotein-depleted plasma between 40 and 50% saturation of (NH4)2SO4. This information was used to partially purify the factor from human plasma. The proteins of human plasma which precipitated between 35 and 55% saturation of (NH4)2SO4 were recovered and subjected to ultracentrifugation to isolate the fraction of density 1.21-1.25 g/ml. This fraction, which was rich in HDL-conversion activity, was further purified by cation-exchange chromatography. In conclusion, a factor which promotes the conversion of HDL to populations of larger and smaller particles has been found to exist at various levels of activity in the plasma of several species. Partial purification of the factor from human plasma has been achieved.

Animals↗

PubMed1984

Evidence of the existence of a high-density lipoprotein transformation factor in pig and rabbit plasma.

Experiments have been conducted to elucidate the incubation conditions necessary for the transformation of pig high-density lipoproteins (HDL) from particles resembling the HDL3 subfraction of human plasma into particles resembling human HDL2. All incubations were performed at 37 degrees C for 24 h in the presence of 0.002 M p-chloromercuriphenylsulphonic acid, a chemical inhibitor of lecithin-cholesterol acyltransferase. There was no transformation when pig HDL (1.09 less than d less than 1.21 g/ml) was incubated either in isolation or in the presence of bovine serum albumin. In addition, there was no transformation when the 1.25 g/ml infranatant of pig plasma was added to the incubation. In the presence of the 1.25 g/ml infranatant of rabbit plasma, by contrast, a significant proportion of the pig HDL was transformed into HDL2. The possibility that this result may have been the consequence of the lipid-transfer activity in rabbit plasma was excluded by the observation that a partially purified preparation of rabbit lipid-transfer protein was ineffective in promoting the HDL transformation. Experiments were also performed with the 1.09 g/ml infranatant of pig plasma. This fraction contained HDL and the plasma proteins but no other lipoprotein. In contrast to the absence of change in incubations of a mixture containing isolated pig HDL (1.09 less than d less than 1.21 g/ml) and the 1.25 g/ml infranatant of pig plasma, during incubation of the 1.09 g/ml infranatant there was an obvious formation of HDL2. It has been concluded that there exists in plasma an HDL-transformation factor which is distinct from both lecithin-cholesterol acyltransferase and the lipid-transfer protein. This factor was detectable in the 1.25 g/ml infranatant of rabbit plasma and in the 1.09 g/ml infranatant of pig plasma. As the factor was not detected in isolated pig HDL or in the 1.25 g/ml infranatant of pig plasma, the 1.21-1.25 g/ml fraction of plasma may be a useful starting point for further investigation of this factor.

Animals↗

PubMed1984

A transient increase in diacylglycerols is associated with the action of vasopressin on hepatocytes.

Vasopressin induced a transient increase of 50% in the total concentration of diacylglycerols (determined by g.l.c.) in isolated hepatocytes. The increase was maximal at 0.25 min, and the concentration of diacylglycerols in cells treated with vasopressin had returned to the basal value by 4 min. No change in the concentration of diacylglycerols was observed after the treatment of cells with glucagon. The dependency of this effect on the concentration of vasopressin was similar to that of the effect of the hormone on 45Ca2+ efflux measured at 0.1 mM extracellular Ca2+. Vasopressin increased the proportion of arachidonic acid and stearic acid and decreased the proportion of oleic acid present in the diacylglycerols. In hepatocytes prelabelled with [14C]arachidonic acid, vasopressin increased the amount of [14C]diacylglycerol. The effects of vasopressin on the total concentration of diacylglycerols and [14C]diacylglycerol were mimicked by an exogenous phospholipid phosphodiesterase (phospholipase C) from Clostridium perfringens. The results are consistent with the conclusion that the transient increase in diacylglycerols induced by vasopressin is caused by the rapid hydrolysis of both the phosphoinositides and one or more other phospholipids.

Animals↗