摘要:

Little is known about racial differences in lipoprotein [a] (Lp[a]) concentrations and apolipoprotein[a] (apo[a]) phenotypes. Lp[a] protein concentrations were determined by a double monoclonal antibody enzyme-linked immunosorbent assay method in 4165 Caucasian and African American men and women from four US communities. Apo[a] phenotypes were determined by polyacrylamide gel electrophoresis and immunoblotting on a random subset of these participants (n=690). The distribution of Lp[a] protein levels in Caucasians was highly skewed (mean, 6.9 mg/dL; median, 3.7 mg/dL). In contrast, the distribution in African Americans was less skewed (mean, 13.0 mg/dL; median, 11.6 mg/dL), and Lp[a] protein levels were approximately double those in Caucasians within most apo[a] phenotypes. The previously described inverse relationship between apo[a] size and Lp[a] concentration was generally confirmed in Caucasians, but the B phenotype had lower Lp[a] levels than the SI or S2 phenotype. In African Americans, both the B and SI phenotypes had lower Lp[a] levels than the S2 phenotype. The frequencies of the apo[a] phenotypes in African Americans differed from those in Caucasians (p<.001) and also differed from the frequencies reported in a Sudanese population (p<.002). African Americans had a lower frequency of the S2 phenotype than Caucasians (8% vs 18%;p<.01) and a higher frequency of S3 (36% vs 25%;p<.01). As compared with the data reported in Sudanese, African Americans also had a higher frequency of the S3 phenotype (36% vs 14%; P-c.OOl) and a lower frequency of S4 (29% vs 44%;p<.01). The apo[a] polymorphism explained 35% of the variability in Lp[a] concentrations in Caucasians and 27% of the variability in African Americans. Though frequencies of the apo[a] phenotypes differ between Caucasians and African Americans, they do not explain the differences in Lp[a] levels between the two racial groups.

ISSN:1049-8834    

出版商:OVID    

年代:1993

数据来源: OVID

摘要:

Lipoprotein composition was determined using ultracentrifugation in 20 non&#151;insulin-dependent (NIDDM) diabetic patients on diet only (D), 20 NIDDM patients on diet and sulfonylorea therapy (T), and 20 nondiabetic control subjects (C), all of whom had total plasma cholesterol concentrations <6.5 mmol/L and total plasma triglyceride concentrations <3.0 mmol/L. Although the groups were well matched for age, body mass index, total triglyceride levels, and total cholesterol concentrations, there were significant compositional abnormalities in the low-density lipoprotein (LDL) fractions of diabetic subjects. The LDL total lipid to apolipoprotein B weight ratio (representing the density distributions of LDL particles) was reduced in both diabetic groups: 3.75±0.3, 3.50±0.28, and 3.54±0.22 in C, D, and T groups, respectively (mean±SD;p<.05). This was associated with a significant shift in the hydrated density distributions of LDL in the diabetic groups, with the average peak densities being 1.0320 g/mL (in C), 1.0365 g/mL (in D), and 1.0380 g/mL (in T) (p<.05). The LDL particles were also smaller in the NIDDM patients: 21.1±0.7, 20.4±0.5, and 20.6±0.5 nm in C, D, and T groups, respectively (p<.05). When the NIDDM groups were analyzed together, the LDL peak density was found to correlate with both insulin resistance (measured by a modified Harano technique; r=0-37, /><.O15) and total triglyceride concentrations (r=0.40,P<.0l). The results show that diabetic patients have small, dense LDL particles, which may be related to insulin resistance, and that these occur with minimal elevations of total triglyceride concentrations. These potentially atherogenic changes may contribute to the increased coronary heart disease in diabetic patients with mild hyperlipidemia.

ISSN:1049-8834    

出版商:OVID    

年代:1993

数据来源: OVID

摘要:

A flow-cytometric method with fluorescence-labeled monoclonal antibodies (MABs) against the low density lipoprotein (LDL) receptor (C7A MAB) or 3,3′-dioctadecylindocarbocyanin-iodide (Dil) LDL has been developed that allows the quantification of LDL receptors on leukocytes and the identification of patients with familial hypercholesterolemia (FH) within 48 hours. Leukocytes were isolated from 10 mL anticoagulated blood by density gradient centrifugation. To induce maximal expression of LDL receptors, mononuclear cells were preincubated with either phytoheraagglutinine (PHA) or iipoprotein-deflcient serum (LPDS). LPDS-treated raonocytes provided a more homogeneous cell population with regard to LDL receptor activity than did the PHA-treated lymphocytes; they also provided a greater discrimination between the fluorescence of the receptor probes and cellular autofluorescence. The C7A MAB was able to compete for Dil LDL binding by about 40%. In competition with unlabeled LDL, Dil LDL revealed linear binding, indicating an affinity similar to native LDL. The binding characteristics of Dil LDL were also similar toIZ5I-LDL binding. LDL isolated from familial defective apolipoprotein B-100 was not able to compete for Dil LDL binding on monocytes, whereas native LDL reduced it by about 80%. In monocytes from FH heterozygous patients, the cellular mean fluorescence using either C7A MAB or Dil LDL at 4°C was 30% to 70%; in FH homozygotes, cellular mean fluorescence was less than 20% of that in monocytes from normal individuals. In patients with familial defective apolipoprotein B-100 antibody binding was normal, but one patient's own LDL failed to compete with normal Dil LDL for 4°C binding on U937 test monocytes. Patient monocytes having internalization defects showed normal 4°C Dil LDL binding, but at 20°C cell-associated fluorescence was reduced by about 40%. In our study 384 hypercholesterolemic patients (preselected according to serum cholesterol levels, clinical symptoms, and family history) were analyzed for LDL receptor expression using the C7A MAB-based assay. In 71.8% of the patients with cholesterol levels higher than 300 mg/dL, an LDL receptor deficiency was observed. Apolipoprotein E isofonns and lipoprotein [a] were found to be independent from the LDL receptor status. In some patients with high cholesterol levels but normal LDL receptor expression with the C7A MAB assay, LDL receptor defects could be diagnosed when either reduced binding or internalization of Dil LDL or familial defective apolipoprotein B-100 was detected. We conclude that fluorescence flow cytometry provides an appropriate, easily performed assay system for the differential diagnosis of LDL receptor defects, including LDL receptor deficiencies and internalization defects, and also allows the discovery of ligand defects.

ISSN:1049-8834    

出版商:OVID    

年代:1993

数据来源: OVID

摘要:

Human plasma contains a multivalent, Kunitz-type proteinase inhibitor termed tissue factor pathway inhibitor (TFPI), which specifically inhibits the action of the factor VII(a) -tissue factor complex in coagulation. In the present study, we examined the distribution and anticoagulant activity of TFPI among plasma lipoprotein subspecies separated by isopycnic density gradient ultracentrifugation; this procedure permitted the simultaneous fractionation of the major lipoprotein classes (very-low-density lipoprotein [VLDL], intermediate-density lipoprotein [IDL], low-density lipoprotein [LDL], high-density lipoprotein [HDL] 2 and 3, and very-high-density lipoprotein [VHDL]). Studies of eight nonnolipidemic subjects revealed two major lipoprotein carriers of TFPI activity: dense LDL subspecies (d=1.039 to 1.063 g/mL) and both dense HDL particles and VHDL (rf=1.133 to 1.190 g/mL), representing 33.8% and 35.9%, respectively, of the total Upoprotein-assodated TFPI activity in plasma. TFPI activity was also associated with lipoprotein(a) (Lp[a]), whose density distribution (</=1.044 to 1.100 g/mL) overlapped that of LDL and HDL]; such association was related to Lp(a)'s particle size and phenotype. VLDL, IDL, and LDL, through LDL, (</= 1.019 to 1.039 g/mL), HDL2(<f= 1.063 to 1.100 g/mL), and light subtractions of HDL, (d=1.100 to 1.167 g/mL) conveyed only 1.8%, 10%, and 18.5%, respectively, of lipoprotein-associated TFPI activity. Such anticoagulant activity was dependent on the presence of TFPI protein. The dense subspecies or HDL, (d=1.133 to 1.167 g/mL) with which TFPI was preferentially associated were small, displayed a cholesteryl ester to protein ratio of -02, and were deficient in phospholipid (13.6% to 18.3%). HDL subspecies of d=1.110 to 1.167 g/mL mainly contained the higher relative molecular mass form of TFPI of 41 kD (a form that is known to be covalently associated with apolipoprotein [apo] A-II) and minor bands of the 35- and 52-kD forms. The second major localization of TFPI was within the hydrated density range of small, dense LDL particles (<f= 1.033 to 1.063 g/mL), which in comparison with light LDL (</= 1.019 to 1.033 g/mL) exhibited a markedly lower proportion of triglyceride and enrichment in cholesteryl ester. Analysb of the ratios of the percent mass of cholesteryl ester to free cholesterol in LDL subtractions showed that the increase in cholesteryl ester content was positively correlated with an increase in TFPI activity (r=.86, /" =5.0013); equally, a positive correlation between an increase in the free cholesterol to protein ratio in LDL and an increase in TFPI activity (r=.89, PS .0006) was detected. In contrast to dense apo A-I-containing lipoprotein particles, dense LDL subspecies of d= 1.039 to 1.058 g/mL mainly transported the 35-kD form of TFPI. We conclude that among the spectrum of apo B-containing lipoprotein subspecies, small, dense LDL particles provide the most favorable surface structure for efficient TFPI binding and equally, for the expression of its anticoagulant activity.

ISSN:1049-8834    

出版商:OVID    

年代:1993

数据来源: OVID

摘要:

Familial defective apolipoprotein B-100 (FDB) and familial hypercholesterolemia (FH) are the common causes of monogenic primary hypercholesterolemia. An individual of mixed English and Afrikaner descent with both FDB and the FH Afrikaner-1 low-density lipoprotein receptor mutation was identified in our laboratory. Subsequent analysis of her extended family revealed the presence of heterozygotes for either FH Afrikaner-1, FH Afrikaner-2, or FDB as well as five additional double heterozygotes for FH Afrikaner-1 and FDB and one "complex" heterozygote with all three mutations. The hypercholesterolemic and clinical features of the pure FDB subjects were similar to those of the pure FH heterozygotes. The double heterozygotes with both FH and FDB have lipid levels and clinical features that are intermediate in severity between heterozygous and homozygous FH.

ISSN:1049-8834    

出版商:OVID    

年代:1993

数据来源: OVID

摘要:

Prostacyclin analogues have been reported to inhibit the expression of tissue factor procoagulant activity in human monocytes, primarily by elevating intracellular levels of adenoslne 3′,5′-cyclic monophosphate (cAMP). The present studies have investigated whether prostacyclins can also inhibit tissue factor expression in endothelial cells. Iloprost, carbacyclin, and ciprostene had no effect on human umbilical vein endothelial tissue factor activity induced by lipopolysaccharide (LPS), tumor necrosis factor-* (TNF-a), or interleukin-10 (IL-10). Iloprost failed to elevate intracellular levels of cAMP, even when combined with a phosphodiesterase inhibitor. In contrast, forskolin increased endothelial cAMP and inhibited tissue factor expression. Conditioned medium from LPS-challenged monocytic THP-1 cells, which contained both TNF-aand IL-1/3, induced endothelial cell procoagulant activity to levels 20-fold higher than those achieved in response to LPS alone. Iloprost abolished LPS-induced TNF-or secretion by THP-1 cells and inhibited IL-10 secretion by 45%. In keeping with this, iloprost reduced levels of TNF-a andTL-lpmRNA in LPS-challenged cells. Treatment of THP-1 cells with iloprost strongly inhibited the ability of conditioned medium to induce endothelial tissue factor expression, an effect that was mimicked by treating the medium with blocking antibodies to the cytokines. We conclude that although prostacyclin analogues do not directly suppress endothelial tissue factor expression due to their failure to elevate cAMP, they may do so indirectly by inhibiting the amplification produced by monocyte-derived cytokines.

ISSN:1049-8834    

出版商:OVID    

年代:1993

数据来源: OVID

摘要:

High plasma levels of plasminogen activator inhibitor type1 (PAI-1), the principal inhibitor of the fibrinolytic system, have been associated with thrombotic and arterial disease. To study PAI-1 expression in healthy and atherosclerotic human arteries, a detailed analysis was made by light and electron microscopy immunocytochemishy and by in situ hybridization. In healthy arteries PAI-1 was found both at the level of endothelial cells and of smooth muscle cells (SMCs) of the arterial media. In early atherosclerotic lesions PAI-1 was also detected in intimal SMCs and in extracellular areas in association with vitronectin. Immunogold analysis by electron microscopy revealed PAI-1 in vesicular structures in endothelial cells and in SMCs with normal or foam cell characteristics. In advanced atheromatous plaques, PAI-1 mRNA expression in SMCs within the fibrous cap was increased compared with SMCs located in the adjacent media or in normal arterial tissue. PAI-1 mRNA was also detected in macrophages located at the periphety of the necrotic core. The increased synthesis of PAI-1 by cellular components of the atherosclerotic plaque and the extracellular accumulation of PAI-1 may contribnte to the thrombotic complications associated with plaque rupture and possibly play a role in the accumulation of extracellular matrix deposits.

ISSN:1049-8834    

出版商:OVID    

年代:1993

数据来源: OVID

摘要:

Lipoprotein(a) (Lp[a]) is a lipoprotein of high atherogenicity with unknown function. Although it binds in vitro to the low-density lipoprotein (LDL) receptor, it is not clear whether this mechanism also operates in vivo. We studied the interaction of Lp(a) and of apoprotein(a) (apo[a]) with hepatoma cells (HepG2 and Hep3B) with the following results. (1) HepG2 cells exhibited saturable high-affinity binding of LDLs, whereas the majority of Lp(a) binding was of low affinity and nonsaturable. Prelncubatlon of HepG2 cells _ with LDL markedly reduced cholesterol biosynthesis, but Lpfa) had a much lower effect (2) When HepG2 cells were preincubated for 48 to 72 hours with Lp(a) or apo(a), '"I-LDL binding was increased by a factor of >2. During this time, up to =>1/igof apo(a) per 1 milligram cell protein was found to be cell associated in an undegraded form. Monoclonal antibodies against the LDL receptor did not prevent the increase in LDL binding stimulated by apo(a). (3) Coincubation with LDL caused a significant increase of Lp(a) degradation by HepG2 cells that was probably caused by an increase of Lp(a) uptake in a "hitchhiking"- like process.

ISSN:1049-8834    

出版商:OVID    

年代:1993

数据来源: OVID

摘要:

Familial combined hyperlipidemia (FCHL) may be genetically and metabolically more heterogeneous than previously thought A consistent feature is an increase in circulating very-low-density lipoprotein (VLDL) apolipoprotein (apo) B, which could be due to either an increase in apoB production or a decrease in its catabolism. Therefore, we directly measured VLDL apoB production in the postabsorptive state in seven FCHL subjects (four male, three female) and seven normal control subjects (three male, four female) by using L-[l-uC]leucine as an endogenous label. Mean age and body mass index did not differ significantly between the two groups. The mean total cholesterol levels were 4.7±0.8 and 8.8±1.6 mmol/L (±SD,p<.01) and the mean triglyceride levels were 0.84±0.14 and 330±1.10 mmol/L (±SD,p<.01) in the control and FCHL groups, respectively. Although the fractional production rate of VLDL apoB was 38% lower in the FCHL group than in the control subjects (0.11 ±0.03 versus 0.18±0.02 pool/h; mean±SD,p<.01), its absolute production rate was 2.7 times greater (534±193 /ig/kg per hour in FCHL versus 196±71 /tg/kg per hour in control subjects; mean±SD,p<.01). There was a linear relation (r=0.8,P=.O3)between triglyceride levels and the VLDL apoB production rate in FCHL, the slope of which indicated a similar VLDL trigtycerlde-to-apoB ratio in the FCHL and control groups. We conclude that FCHL is a metabolically coherent disorder and that the increase in circulating apoB and triglyceride levels in FCHL is due to secretion of an increased number of VLDL particles, each containing, on average, a normal amount of triglyceride and one molecule of apoB. We hypothesize that FCHL is caused by a defective gene or genes, which enhance intrahepatlc lipid availability for lipoprotein assembly and increase apoB secretion by decreasing the amount of a constitutively synthesized pool of apoB that undergoes Intrahepatic degradation.

ISSN:1049-8834    

出版商:OVID    

年代:1993

数据来源: OVID

摘要:

Two glycoforms of recombinant human thrombomodulin (TM; TMD1-105 and TMD1-75), an endothelial cell membrane protein, were tested for their ability to alter thrombin-induced activation of cultured human umbilical vein endothelial cells (HUVECs). After stimulation with 10 nmol/L thrombin, HUVEC generation of inositol-l,4,5-trisphosphate (IP}), a potent Ca$-mobilizing second messenger, was dosedependently blocked by TMD1-105. Both TMD1-105 (ICM= 10 nmol/L) and TMD1-75 (ICjo=100 nmol/L) blocked the enhanced prostacyclin synthesis by HUVEC monolayers treated with 10 nmol/L thrombin. HUVEC monolayer permeability to Evans blue dye-labeled albumin increased from 0.125 ±0.06 /iL/min in control experiments to 0J80±0.09 jtL/min after treatment with 100 nmol/L thrombin (p<.05). Incubation or HUVECs with TMD1-105 alone (600 nmol/L) had no effect (0.114±0.04fiL/min)on basal permeability. In contrast, incubation of 100 nmol/L thrombin with 600 nmol/L TMD1-105 reduced this increase in HUVEC permeability to almost control levels (0.142±0.06 /tL/min). These results demonstrate that recombinant human TM, a potent in vitro anticoagulant, also functions as an antagonist of thrombin receptor-mediated HUVEC activation. In addition to its anticoagulant functions, the highaffinity endothelial cell receptor TM may play a role in modulating endothelial cell activation by thrombin.

ISSN:1049-8834    

出版商:OVID    

年代:1993

数据来源: OVID