Plasma phospholipid transfer protein activity in patients with low HDL and cardiovascular disease treated with simvastatin and niacin

Plasma phospholipid transfer protein (PLTP) is an important modulator of high-density lipoprotein (HDL) metabolism, regulating its particle size, composition, and mass. In patients with low HDL and cardiovascular disease (CVD), plasma PLTP activity is positively correlated with the concentration of HDL particles containing apo A-I but not apo A-II (Lp(A-1)). We recently completed a study to determine the effect of simvastatin and niacin (S-N) therapy on disease progression/regression in these patients, and found that this therapy selectively increased Lp(A-I). To determine if PLTP was also increased with this drug therapy, we measured the PLTP activity in the plasma of 30 of these patients obtained at baseline and after 12 months of therapy, and compared the changes to a similar group of 31 patients who received placebo for the drugs. No significant increase in PLTP activity was observed in either group of patients. However, changes in apo A-I and A-II between these two time points were correlated with the corresponding change in PLTP activity. The correlation coefficients were r=0.57 (P=0.001) and r=0.43 (P=0.02) for apo A-I, and r=0.54 (P=0.002) and r=0.41 (P=0.02) for apo A-II in the placebo and S-N group, respectively. At baseline, PLTP activity correlated positively with the percent of plasma apo A-I associated with Lp(A-I) (r=0.38, P=0.04) and the amounts of apo A-I in these particles (r=0.43, P=0.02). These relationships persisted in patients who took placebo for 12 months (r=0.46, P=0.009 and r=0.37, P=0.04, respectively), but was attenuated in those treated with S-N. These data indicate that S-N-induced increase in Lp(A-I) was PLTP-independent. It also confirms our previous observation that an interrelationship exists between PLTP and apo-specific HDL particle subclasses in CVD patients with low HDL, and that this relationship is altered by drug intervention.     

Lipoprotein lipase and hepatic lipase: their relationship with HDL subspecies Lp(A-I) and Lp(A-I,A-II)

HDL subspecies Lp(A-I) and Lp(A-I,A-II) have different anti-atherogenic potentials. To determine the role of lipoprotein lipase (LPL) and hepatic lipase (HL) in regulating these particles, we measured these enzyme activities in 28 healthy subjects with well-controlled Type 1 diabetes, and studied their relationship with Lp(A-I) and Lp(A-I,A-II). LPL was positively correlated with the apolipoprotein A-I (apoA-I), cholesterol, and phospholipid mass in total Lp(A-I), and with the apoA-I in large Lp(A-I) (r >or= 0.58, P >or= 0.001). HL was negatively correlated with all the above Lp(A-I) parameters plus Lp(A-I) triglyceride (r >or= -0.53, P or= 0.50, P 相似文献   

Protein transfer between A-I-containing lipoprotein subpopulations: evidence of non-transferable A-I in particles with A-II.

Transfer of apolipoproteins (apo) between the two subpopulations of apo A-I-containing lipoproteins in human plasma: those with A-II [Lp(AI w AII)] and those without [Lp(AI w/o AII)], were studied by observing the transfer of 125I-apo from a radiolabeled subpopulation to an unlabeled subpopulation in vitro. When Lp(AI w AII) was directly radioiodinated, 50.3 +/- 7.4 and 19.5 +/- 7.7% (n = 6) of the total radioactivity was associated with A-I and A-II, respectively. In radioiodinated Lp(AI w/o AII), 71.5 +/- 6.8% (n = 6) of the total radioactivity was A-I-associated. Time-course studies showed that, while some radiolabeled proteins transferred from one population of HDL particles to another within minutes, at least several hours were necessary for transfer to approach equilibrium. Incubation of the subpopulations at equal A-I mass resulted in the transfer of 51.8 +/- 5.0% (n = 4) of total radioactivity from [125I]Lp(AI w/o AII) to Lp(AI w AII) at 37 degrees C in 24 h. The specific activity (S.A.) of A-I in the two subpopulations after incubation was nearly identical. Under similar incubation conditions, only 13.4 +/- 4.6% (n = 4) of total radioactivity was transferred from [125I]Lp(AI w AII) to Lp(AI w/o AII). The S.A. of A-I after incubation was 2-fold higher in particles with A-II than in particles without A-II. These phenomena were also observed with iodinated high-density lipoproteins (HDL) isolated by ultracentrifugation and subsequently subfractionated by immunoaffinity chromatography. However, when Lp(AI w AII) radiolabeled by in vitro exchange with free [125I]A-I was incubated with unlabeled Lp(AI w/o AII), the S.A. of A-I in particles with and without A-II differed by only 18% after incubation. These data are consistent with the following: (1) in both populations of HDL particles, some radiolabeled proteins transferred rapidly (minutes or less), while others transferred slowly (hours); (2) when Lp(AI w AII) and Lp(AI w/o AII) were directly iodinated, all labeled A-I in particles without A-II were transferable, but some labeled AI in particles with A-II were not; (3) when Lp(AI w AII) were labeled by in vitro exchange with [125I]A-I, considerably more labeled A-I were transferable. These observations suggest the presence of non-transferable A-I in Lp(AI w AII).     

Differential effect of ultracentrifugation on apolipoprotein A-I-containing lipoprotein subpopulations

Two populations of apolipoprotein (apo) A-I-containing lipoprotein particles are found in high density lipoproteins (HDL): those that also contain apo A-II[Lp(A-I w A-II)] and those that do not [Lp(A-I w/o A-II)]. Lp(A-I w/o A-II) comprised two distinct particle sizes with mean hydrates Stokes diameter of 10.5 nm for Lp(A-I w/o A-II)1 and 8.5 nm for Lp(A-I w/o A-II)2. To study the effect of ultracentrifugation on these particles, Lp(A-I w/o A-II) and Lp(A-I w A-II) were isolated from the plasma and the ultracentrifugal HDL (d 1.063-1.21 g/ml fractions) of five normolipidemic and three hyperlipidemic subjects. The size subpopulations of these particles were studied by gradient polyacrylamide gel electrophoresis. Several consistent differences were detected between plasma Lp(A-I w/o A-II) and HDL Lp(A-I w/o A-II). First, in all subjects, the relative proportion of Lp(A-I w/o A-II)1 to Lp(A-I w/o A-II)2 isolated from HDL was reduced. Second, particles larger than Lp(A-I w/o A-II)1 and smaller than Lp(A-I w/o A-II)2 were considerably reduced in HDL. Third, a distinct population of particles with approximate Stokes diameter of 7.1 nm usually absent in plasma was detected in HDL Lp(A-I w/o A-II). Little difference in subpopulation distribution was detected between Lp(A-I w A-II) isolated from the plasma and HDL of the same subject. When plasma Lp(A-I w/o A-II) and Lp(A-I w A-II) were centrifuged, 14% and 4% of A-I were, respectively, recovered in the D greater than 1.21 g/ml fraction. Only 2% A-II was found in this density fraction. These studies show that the Lp(A-I w/o A-II) particles are less stable than Lp(A-I w A-II) particles upon ultracentrifugation. Among the various Lp(A-I w/o A-II) subpopulations, particles larger than Lp(A-I w/o A-II)1 and smaller than Lp(A-I w/o A-II)2 are most labile.     

Association of plasma phospholipid transfer protein activity with IDL and buoyant LDL: impact of gender and adiposity

Current data suggest that phospholipid transfer protein (PLTP) has multiple metabolic functions, however, its physiological significance in humans remains to be clarified. To provide further insight into the role of PLTP in lipoprotein metabolism, plasma PLTP activity was measured, and lipoproteins were analyzed in 134 non-diabetic individuals on a controlled diet. Insulin sensitivity index (Si) and body fat composition were also determined. Plasma PLTP activity was comparable between men (n=56) and women (n=78). However, in women but not in men, plasma PLTP activity was positively correlated with cholesterol, triglyceride, low density lipoprotein (LDL) cholesterol, and apolipoprotein (apo) B (r=0.38-0.45, P< or =0.001), and with body mass index (BMI), subcutaneous and intra-abdominal fat (SCF, IAF) (r=0.27-0.29, P<0.02). Among the different apo B-containing lipoproteins (LpB) in women, PLTP was most highly correlated with intermediate density lipoproteins (IDL) and buoyant LDL (r=0.45-0.46, P<0.001). The correlation with IDL was significant only in women with BMI < or =27.5 kg/m(2) (n=56). In men with BMI < or =27.5 kg/m(2) (n=35), PLTP activity was significantly correlated with buoyant LDL (r=0.40, P<0.02) and high density lipoprotein (HDL) (r=0.43, P<0.01). These data provide evidence for a role of PLTP in LpB metabolism, particularly IDL and buoyant LDL. They also suggest that gender and obesity-related factors can modulate the impact of PLTP on LpB.     

Characterization of lipoprotein particles isolated by immunoaffinity chromatography. Particles containing A-I and A-II and particles containing A-I but no A-II

Two populations of A-I-containing lipoprotein particles: A-I-containing lipoprotein with A-II (Lp (A-I with A-II], and A-I-containing lipoprotein without A-II (Lp (A-I without A-II] have been isolated from plasma of 10 normolipidemic subjects by immunoaffinity chromatography and characterized. Both types of particles possess alpha-electrophoretic mobility and hydrated density in the range of plasma high-density lipoproteins (HDL). Lp (A-I without A-II) and Lp (A-I with A-II) are heterogeneous in size. Lp (A-I without A-II) comprised two distinct particle sizes with mean apparent molecular weight and Stokes diameter of 3.01 X 10(5), and 10.8 nm for Lp (A-I without A-II)1, and 1.64 X 10(5), and 8.5 nm for Lp (A-I without A-II)2. Lp (A-I with A-II) usually contained particles of at least three distinct molecular sizes with mean apparent molecular weight and Stokes diameter of 2.28 X 10(5) and 9.6 nm for Lp (A-I with A-II)1, 1.80 X 10(5) and 8.9 nm for Lp (A-I with A-II)2, and 1.25 X 10(5) and 8.0 nm for Lp (A-I with A-II)3. Apoproteins C, D, and E, and lecithin:cholesterol acyltransferase (LCAT) were detected in both Lp (A-I without A-II) and Lp (A-I with A-II) with most of the apoprotein D, and E, and LCAT (EC 2.3.1.43) in Lp (A-I with A-II) particles. Lp (A-I without A-II) had a slightly higher lipid/protein ratio than Lp (A-I with A-II). Lp (A-I with A-II) had an A-I/A-II molar ratio of approximately 2:1. The percentage of plasma A-I associated with Lp (A-I without A-II) was highly correlated with the A-I/A-II ratio of plasma (r = 0.96, n = 10). The variation in A-I/A-II ratio of HDL density subfractions therefore reflects different proportions of two discrete types of particles: particles containing A-I and A-II in a nearly constant ratio and particles containing A-II but no A-II. Each type of particle is heterogeneous in size and in apoprotein composition.     

Interaction between high-density lipoprotein subpopulations in apo B-free and abetalipoproteinemic plasma.

Two populations of high-density lipoprotein (HDL) particles exist in human plasma. Both contain apolipoprotein (apo) A-I, but only one contains apo A-II: Lp(AI w AII) and Lp(AI w/o AII). To study the extent of interaction between these particles, apo B-free plasma prepared by the selective removal of apo B-containing lipoproteins (LpB) from the plasma of three normolipidemic (NL) subjects and whole plasma from two patients with abetalipoproteinemia (ABL) were incubated at 37 degrees C for 24 h. Apo B-free plasma samples were used to avoid lipid-exchange between HDL and LpB. Lp(AI w AII) and Lp(AI w/o AII) were isolated from each apo B-free plasma sample before and after incubation and their protein and lipid contents quantified. Before incubation, ABL plasma had reduced levels of Lp(AI w AII) and Lp(AI w/o AII), (40% and 70% of normals, respectively). Compared to the HDL of apo B-free NL plasma, ABL HDL had higher relative contents of free cholesterol, phospholipid and total lipid, and contained more particles with apparent hydrated Stokes diameter in the 9.2-17.0 nm region. These differences were particularly pronounced in particles without apo A-II. Despite their differences, the total cholesterol contents of Lp(AI w AII) increased, while that of Lp(AI w/o AII) decreased in all five plasma samples and the amount of apo A-I in Lp(AI w AII) increased by 6-8 mg/dl in four during the incubation. These compositional changes were accompanied by a relative reduction of particles in the 7.0-8.2 nm Stokes diameter size region and an increase of particles in the 9.2-11.2 nm region. These data are consistent with intravascular modulation between HDL particles with and without apo A-II. The observed increase in apo A-II-associated cholesterol and apo A-I, could involve either the transfer of cholesterol and apo A-I from particles without apo A-II to those with A-II, or the transfer of apo A-II from Lp(AI w AII) to Lp(AI w/o AII). The exact mechanism and direction of the transfer remain to be determined.     

Characterization of A-I-containing lipoproteins in subjects with A-I Milano variant

The A-I Milano variant of apolipoprotein A-I (A-IM), by virtue of its Arg-173----Cys substitution, is capable of forming a disulfide bond with the 77-amino-acid apolipoprotein A-II polypeptide (A-IIS) as well as with itself to produce dimers, A-IM/A-IIS and A-IM/A-IM, respectively. A-I-containing lipoproteins (Lp): particles with A-II (Lp(A-I with A-11)) and particles without A-II (Lp(A-I without A-II)) in the plasma of two nonhyperlipidemic A-IM carriers were investigated to determine the effect of A-IM on these lipoproteins. Despite the existence of abnormal apolipoprotein dimers and the unusually low HDL cholesterol (17 and 14 mg/dl), A-I (67 and 75 mg/dl), and A-II (18 and 18 mg/dl) levels in the two carriers, the plasma A-I of the carriers was distributed between Lp(A-I with A-II) and Lp(A-I without A-II) in a proportion comparable to that observed in normals. As expected, A-IM/A-IIS mixed dimer was found in carrier Lp(A-I with A-II). However, A-IM/A-IM dimer was located almost exclusively in carrier Lp(A-I without A-II). Chemical (dimethylsuberimidate) crosslinking of the protein moieties of the major subpopulations of Lp(A-I with A-II) and Lp(A-I without A-II) of normal and A-IM carriers showed that Lp(A-I with A-II), which is located predominantly in the 7.8-9.7 nm interval ((HDL2a + 3a + 3b)gge), had an apparent protein molecular weight equivalent to two molecules of A-I and one to two molecules of A-II per particle. Most of the Lp(A-I without A-II) particles, located predominantly in the size intervals of 9.7-12.9 nm (designated (HDL2b)gge) and 8.2-8.8 nm (HDL3a)gge) had protein moieties exhibiting a molecular weight equivalence predominantly of four and three molecules of A-I, respectively. A small quantity of particles with apparent protein content of two molecules of A-I in the 7.2-8.2 nm interval ((HDL3b + 3c)gge) was also detected. These studies showed that in nonhyperlipidemic A-IM carriers, the occurrence of apolipoprotein dimers had not markedly affected the protein stoichiometry of Lp(A-I with A-II) and Lp(A-I without A-II).     

Characterization of human high-density lipoprotein subclasses LP A-I and LP A-I/A-II and binding to HepG2 cells

Plasma HDL can be classified according to their apolipoprotein content into at least two types of lipoprotein particles: lipoproteins containing both apo A-I and apo A-II (LP A-I/A-II) and lipoproteins with apo A-I but without apo A-II (LP A-I). LP A-I and LP A-I/A-II were isolated by immuno-affinity chromatography. LP A-I has a higher cholesterol content and less protein compared to LP A-I/A-II. The average particle mass of LP A-I is higher (379 kDa) than the average particle weight of LP A-I/A-II (269 kDa). The binding of 125I-LP A-I to HepG2 cells at 4 degrees C, as well as the uptake of [3H]cholesteryl ether-labelled LP A-I by HepG2 cells at 37 degrees C, was significantly higher than the binding and uptake of LP A-I/A-II. It is likely that both binding and uptake are mediated by apo A-I. Our results do not provide evidence in favor of a specific role for apo A-II in the binding and uptake of HDL by HepG2 cells.     

Characterization of apoA-I-containing lipoprotein subpopulations secreted by HepG2 cells

Recent immunoaffinity studies demonstrate two populations of high density lipoprotein (HDL) particles: one contains both apolipoprotein (apo) A-I and A-II [Lp(A-I w A-II)], and the other contains apoA-I but no A-II [Lp(A-I w/o A-II)]. To investigate whether these two populations are derived from different precursors, we applied sequential immunoaffinity chromatography to study the lipoprotein complexes in HepG2 conditioned serum-free medium. The apparent secretion rates of apoA-I, A-II, E, D, A-IV, and lecithin:cholesterol acyltransferase (LCAT) were 4013 +/- 1368, 851 +/- 217, 414 +/- 64, 171 +/- 51, 32 +/- 14, and 2.9 +/- 0.7 ng/mg cell protein per 24 h, respectively (n = 3-5). Anti-A-II removed all apoA-II but only 39 +/- 5% (n = 5) apoA-I from the medium. These HepG2 Lp(A-I w A-II) also contained 31 +/- 1% (n = 5) of the apoD and 82 +/- 2% (n = 3) of the apoE in the medium. The apoE existed both as E and E-A-II complex. Lipoproteins isolated from the apoA-II-free medium by anti-A-I contained, besides apoA-I, 60 +/- 3% of the medium apoD and trace quantities of apoE. The majority of HepG2 apoA-IV (78 +/- 4%) (n = 3) and LCAT (85 +/- 6%) (n = 3) was not associated with either apoA-I or A-II. HepG2 Lp(A-I w A-II) contained relatively more lipids than Lp(A-I w/o A-II) (45 vs. 37%).(ABSTRACT TRUNCATED AT 250 WORDS)     

PLTP activity in premenopausal women. Relationship with lipoprotein lipase, HDL, LDL, body fat, and insulin resistance

Plasma phospholipid transfer protein (PLTP) is thought to play a major role in the facilitated transfer of phospholipids between lipoproteins and in the modulation of high density lipoprotein (HDL) particle size and composition. However, little has been reported concerning the relationships of PLTP with plasma lipoprotein parameters, lipolytic enzymes, body fat distribution, insulin, and glucose in normolipidemic individuals, particularly females. In the present study, 50 normolipidemic healthy premenopausal females were investigated. The relationships between the plasma PLTP activity and selected variables were assessed. PLTP activity was significantly and positively correlated with low density lipoprotein (LDL) cholesterol (r(s) = 0.53), apoB (r(s) = 0.44), glucose (r(s) = 0.40), HDL cholesterol (r(s) = 0.38), HDL(3) cholesterol (r(s) = 0.37), lipoprotein lipase activity (r(s) = 0.36), insulin (r(s) = 0.33), subcutaneous abdominal fat (r(s) = 0.36), intra-abdominal fat (r(s) = 0.29), and body mass index (r(s) = 0.29). HDL(2) cholesterol, triglyceride, and hepatic lipase were not significantly related to PLTP activity. As HDL(2) can be decreased by hepatic lipase and hepatic lipase is increased in obesity with increasing intra-abdominal fat, the participants were divided into sub-groups of non-obese (n = 35) and obese (n = 15) individuals and the correlation of PLTP with HDL(2) cholesterol was re-examined. In the non-obese subjects, HDL(2) cholesterol was found to be significantly and positively related to PLTP activity (r(s) = 0.44). Adjustment of the HDL(2) values for the effect of hepatic lipase activity resulted in a significant positive correlation between PLTP and HDL(2) (r(s) = 0.41), indicating that the strength of the relationship between PLTP activity and HDL(2) can be reduced by the opposing effect of hepatic lipase on HDL(2) concentrations. We conclude that PLTP-facilitated lipid transfer activity is related to HDL and LDL metabolism, as well as lipoprotein lipase activity, adiposity, and insulin resistance.     

GPI-specific phospholipase D associates with an apoA-I- and apoA-IV-containing complex

Glycosylphosphatidylinositol-specific phospholipase D (GPI-PLD) is abundant in serum and associates with high density lipoproteins (HDL). We have characterized the distribution of GPI-PLD among lipoproteins in human plasma. Apolipoprotein (apo)-specific lipoproteins containing apoB (Lp[B]), apoA-I and A-II (Lp[A-I, A-II]), or apoA-I only (Lp[A-I]) were isolated using dextran sulfate and immunoaffinity chromatography. In six human plasma samples with HDL cholesterol ranging from 39 to 129 mg/dl, 79 +/- 14% (mean +/- SD) of the total plasma GPI-PLD activity was associated with Lp[A-I], 9 +/- 12% with Lp[A-I, A-II], and 1 +/- 1% with Lp[B]; and 11 +/- 10% was present in plasma devoid of these lipoproteins. Further characterization of the GPI-PLD-containing lipoproteins by gel-filtration chromatography and nondenaturing polyacrylamide and agarose gel electrophoresis revealed that these apoA-I-containing particles/complexes were small (8 nm) and migrated with pre-beta particles on agarose electrophoresis. Immunoprecipitation of GPI-PLD with a monoclonal antibody to GPI-PLD co-precipitated apoA-I and apoA-IV but little or no apoA-II, apoC-II, apoC-III, apoD, or apoE. In vitro, apoA-I but not apoA-IV or bovine serum albumin interacted directly with GPI-PLD, but did not stimulate GPI-PLD-mediated cleavage of a cell surface GPI-anchored protein. Thus, the majority of plasma GPI-PLD appears to be specifically associated with a small, discrete, and minor fraction of lipoproteins containing apoA-I and apoA-IV. -- Deeg, M. A., E. L. Bierman, and M. C. Cheung. GPI-specific phospholipase D associates with an apoA-I- and apoA-IV-containing complex. J. Lipid Res. 2001. 42: 442--451.     

Apolipoprotein A-II/A-I ratio is a key determinant in vivo of HDL concentration and formation of pre-beta HDL containing apolipoprotein A-II

Overexpression of human apolipoprotein A-II (apo A-II) in mice induced postprandial hypertriglyceridemia and marked reduction in plasma HDL concentration and particle size [Boisfer et al. (1999) J. Biol. Chem. 274, 11564-11572]. We presently compared lipoprotein metabolism in three transgenic lines displaying plasma concentrations of human apo A-II ranging from normal to 4 times higher, under ad libitum feeding and after an overnight fast. Fasting dramatically decreased VLDL and lowered circulating human apo A-II in transgenic mice; conversely, plasma HDL levels increased in all genotypes. The apo A-I content of HDL was inversely related to the expression of human apo A-II, probably reflecting displacement of apo A-I by an excess of apo A-II. Thus, the molar ratios of apo A-II/A-I in HDL were significantly higher in fed as compared with fasted animals of the same transgenic line, while endogenous LCAT activity concomitantly decreased. The number and size of HDL particles decreased in direct proportion to the level of human apo A-II expression. Apo A-II was abundantly present in all HDL particles, in contrast to apo A-I mainly present in large ones. Two novel findings were the presence of pre-beta migrating HDL transporting only human apo A-II in the higher-expressing mice and the increase of plasma HDL concentrations by fasting in control and transgenic mice. These findings highlight the reciprocal modifications of VLDL and HDL induced by the feeding-fasting transition and the key role of the molar ratio of apo A-II/A-I as a determinant of HDL particle metabolism and pre-beta HDL formation.     

Apolipoproteins A-I,A-II and E are independently distributed among intracellular and newly secreted HDL of human hepatoma cells

Whereas hepatocytes secrete the major human plasma high density lipoproteins (HDL)-protein, apo A-I, as lipid-free and lipidated species, the biogenic itineraries of apo A-II and apo E are unknown. Human plasma and HepG2 cell-derived apo A-II and apo E occur as monomers, homodimers and heterodimers. Dimerization of apo A-II, which is more lipophilic than apo A-I, is catalyzed by lipid surfaces. Thus, we hypothesized that lipidation of intracellular and secreted apo A-II exceeds that of apo A-I, and once lipidated, apo A-II dimerizes. Fractionation of HepG2 cell lysate and media by size exclusion chromatography showed that intracellular apo A-II and apo E are fully lipidated and occur on nascent HDL and VLDL respectively, while only 45% of intracellular apo A-I is lipidated. Secreted apo A-II and apo E occur on small HDL and on LDL and large HDL respectively. HDL particles containing both apo A-II and apo A-I form only after secretion from both HepG2 and Huh7 hepatoma cells. Apo A-II dimerizes intracellularly while intracellular apo E is monomeric but after secretion associates with HDL and subsequently dimerizes. Thus, HDL apolipoproteins A-I, A-II and E have distinct intracellular and post-secretory pathways of hepatic lipidation and dimerization in the process of HDL formation. These early forms of HDL are expected to follow different apolipoprotein-specific pathways through plasma remodeling and reverse cholesterol transport.     

Distribution and localization of lecithin:cholesterol acyltransferase and cholesteryl ester transfer activity in A-I-containing lipoproteins

Two types of A-I-containing lipoproteins are found in human high density lipoproteins (HDL): particles with A-II (Lp(A-I with A-II] and particles without A-II (Lp(A-I without A-II]. We have studied the distribution of lecithin:cholesterol acyltransferase (LCAT) and cholesteryl ester transfer (CET) activities in these particles. Lp(A-I with A-II) and Lp(A-I without A-II) particles were isolated from ten normolipidemic subjects by anti-A-I and anti-A-II immunosorbents. Most plasma LCAT mass (70 +/- 15%), LCAT (69 +/- 16%), and CET (81 +/- 15%) activities were detected in Lp(A-I without A-II). Some LCAT (mass: 16 +/- 7%, activity: 17 +/- 8%) and CET activities (7 +/- 8%) were detected in Lp(A-I with A-II). To determine the size subspecies that contain LCAT and CET activities, isolated Lp(A-I with A-II) and Lp(A-I without A-II) particles of six subjects were further fractionated by gel filtration column chromatography. In Lp(A-I without A-II), most LCAT and CET activities were associated with different size particles, with the majority of the LCAT and CET activities located in particles with hydrated Stokes diameters of 11.6 +/- 0.4 nm and 10.0 +/- 0.6 nm, respectively. In Lp(A-I with A-II), most of the LCAT and CET activities were located in particles similar in size: 11.1 +/- 0.4 nm and 10.6 +/- 0.3 nm, respectively. Ultracentrifugation of A-I-containing lipoproteins resulted in dissociation of both LCAT and CET activities from the particles. Furthermore, essentially all CET and LCAT activities were recovered in the non-B-containing plasma obtained by anti-LDL immunoaffinity chromatography. This report, therefore, provides direct evidence for the association of LCAT and CET protein with A-I-containing lipoproteins. Our conclusions pertain to fasting normolipidemic subjects and may not be applicable to hyperlipidemic or nonfasting subjects.     

Apolipoprotein E genotypes and metabolic risk factors for coronary heart disease in middle-aged women

Apo E genotypes and plasma metabolic risk factors (total cholesterol, triglycerides, HDL and LDL cholesterol, total/HDL cholesterol ratio, lipoprotein Lp (a), apolipoprotein A-I, A-II, apo B, and apo E) were determined in 134 healthy middle-aged (X +/- SD 49.62 +/- 4.83) women. The aim of this study was to investigate metabolic risk markers according to various apo E genotypes, and to evaluate a possible risk for coronary heart disease. The results revealed that the frequencies of apo E3/3 are the most frequent (46%), followed by E4/4 (2%), E3/4 (14%), E2/3 (14%), and E2/4 (2%) in the middle-aged women. Higher mean triglycerides, LDL-C and apo B levels were found with apo E3/4, and lower mean levels of HDL-C i.e. apo A-I than in other analyzed genotypes. Greater mean of total/HDL ratio and lower levels of apo A-II were seen with E2/4. Serum lipoprotein Lp (a) concentration was higher in women with genotypes E3/3. Apo E concentration was the lowest with genotypes E4/4, i.e. the highest with E2/3. Serum total cholesterol tended to be higher in women with genotypes E4/4. Genotype E3/4 is connected with the highest concentrations of (X +/- SD) triglycerides (1.74 +/- 0.78), LDL (4.28 +/- 1.88), apo B (1.03 +/- 0.32) and with the lowest concentrations of HDL cholesterol (1.11 +/- 0.21) in the relation to the other analyzed genotypes. This group of women could possibly represent high risk women for CHD. Genotype E3/3 is associated with the highest concentration of independent genetic risk marker for CHD, lipoprotein Lp (a) (0.19 +/- 0.27). The genotype E4/4 has the highest concentration of total cholesterol (5.93 +/- 1.01), and has to be taken in account for risk evaluation in women. High level of apo E (0.11 +/- 0.05) and low level of apo A-I (1.80 +/- 0.44) were associated with E2/3 genotypes. The significance of E3/4 with the high total/HDL ratio (5.52 +/- 2.21) and low apo A-II (0.53 +/- 0.09) is important indicator, because total/HDL cholesterol ratio represents independent Established Risk Factor (ERF) for CHD. Apolipoprotein E genotypes as genetic markers and investigation of serum metabolic risk markers appear to be important in view for further evaluation of high risk women for CHD in our population.     

Decreased PLTP mass but elevated PLTP activity linked to insulin resistance in HTG: effects of bezafibrate therapy

Hypertriglyceridemia (HTG) is associated with insulin resistance, increased cholesteryl ester transfer (CET), and low HDL cholesterol. Phospholipid transfer protein (PLTP) may be involved in these relationships. Associations between CET, lipids, insulin resistance, CETP and PLTP activities, and PLTP mass were investigated in 18 HTG patients and 20 controls. Effects of 6 weeks of bezafibrate treatment were studied in HTG patients. HTG patients had higher serum triglycerides, insulin resistance, free fatty acid (FFA), and CET, lower levels of HDL cholesterol (-44%) and PLTP mass (-54%), and higher CETP (+20%) and PLTP activity (+48%) than controls. Bezafibrate reduced triglycerides, CET (-37%), insulin resistance (-53%), FFA (-48%), CETP activity (-12%), PLTP activity (-8%), and increased HDL cholesterol (+27%), whereas PLTP mass remained unchanged. Regression analysis showed a positive contribution of PLTP mass (P = 0.001) but not of PLTP activity to HDL cholesterol, whereas insulin resistance positively contributed to PLTP activity (P < 0.01). Bezafibrate-induced change in CET and HDL cholesterol correlated with changes in CETP activity and FFAs, but not with change in PLTP activity. Bezafibrate-induced change in PLTP activity correlated with change in FFAs (r = 0.455, P = 0.058). We propose that elevated PLTP activity in HTG is related to insulin resistance and not to increased PLTP mass. Bezafibrate-induced diminished insulin resistance is associated with a reduction of CET and PLTP activity.     

Plasma lecithin:cholesterol acyltransferase activity modifies the inverse relationship of C-reactive protein with HDL cholesterol in nondiabetic men

Lecithin:cholesterol acyltransferase (LCAT) is instrumental in high-density lipoprotein (HDL) maturation, but high LCAT levels do not predict low cardiovascular risk. LCAT may affect antioxidative or anti-inflammatory properties of HDL. We determined the relationship of plasma high-sensitivity C-reactive protein (CRP) with LCAT activity and evaluated whether LCAT activity modifies the decreasing effect of HDL cholesterol (HDL-C) on CRP, as an estimate of its anti-inflammatory properties. Plasma HDL-C, apolipoprotein (apo) A-I and LCAT activity (exogenous substrate method) were measured in 260 nondiabetic men without cardiovascular disease. CRP was correlated inversely with HDL-C and apo A-I, and positively with LCAT activity (P < 0.01 to 0.001). Multivariate regression analysis demonstrated that age- and smoking-adjusted plasma CRP levels were associated negatively with HDL-C (β = − 0.224, P < 0.001) and positively with LCAT activity (β = 0.119, P = 0.034), as well as with the interaction between HDL-C and LCAT activity (β = 0.123, P = 0.026). There was also an interaction between apo A-I and LCAT activity on CRP (β = 0.159, P = 0.005). These relationships remained similar after adjustment for apo B-containing lipoproteins. In conclusion, the inverse relationship of HDL-C with CRP is attenuated by LCAT activity at higher HDL-C levels. It is hypothesized that LCAT could mitigate HDL's anti-inflammatory or antioxidative properties at higher HDL-C concentrations.     

Active plasma phospholipid transfer protein is associated with apoA-I- but not apoE-containing lipoproteins

Plasma phospholipid transfer protein (PLTP) is a multifaceted protein with diverse biological functions. It has been shown to exist in both active and inactive forms. To determine the nature of lipoproteins associated with active PLTP, plasma samples were adsorbed with anti-A-I, anti-A-II, or anti-E immunoadsorbent, and PLTP activity was measured in the resulting plasma devoid of apolipoprotein A-I (apoA-I), apoA-II, or apoE. Anti-A-I and anti-A-II immunoadsorbents removed 98 +/- 1% (n = 8) and 38 +/- 25% (n = 7) of plasma PLTP activity, respectively. In contrast, only 1 +/- 5% of plasma PLTP activity was removed by anti-E immunoadsorbent (n = 7). Dextran sulfate (DS) cellulose did not bind apoA-I, but it removed 83 +/- 5% (n = 4) of the PLTP activity in plasma. In size-exclusion chromatography, PLTP activity removed by anti-A-I or anti-A-II immunoadsorbent was associated primarily with particles of a size corresponding to HDL, whereas a substantial portion of the PLTP activity dissociated from DS cellulose was found in particles larger or smaller than HDL. These data show the following: 1) active plasma PLTP is associated primarily with apoA-I- but not apoE-containing lipoproteins; 2) active PLTP is present in HDL particles with and without apoA-II, and its distribution between these two HDL subpopulations varies widely among individuals; and 3) DS cellulose can remove active PLTP from apoA-I-containing lipoproteins, and this process creates new active PLTP-containing particles in vitro.     

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.