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Biomedical subjects

G M Kostner

Publications and source records attributed to G M Kostner.

At least 19 recordsLinked to original sources
PubMed1992

Organization of phosphatidylcholine and sphingomyelin in the surface monolayer of low density lipoprotein and lipoprotein(a) as determined by time-resolved fluorometry.

Fluorescent analogs of phosphatidylcholine (PC) and sphingomyelin (SM) labeled with diphenylhexatrienylpropionic acid (DPH) were prepared and incorporated into the surface layer of human low density lipoprotein (LDL) and lipoprotein(a) (Lp(a)). Fluorescence anisotropy measurements of DPH-PC and DPH-SM in both lipoprotein classes were carried out at different temperatures ranging from 20 to 37 degrees C. DPH-PC as well as DPH-SM were shown to reside in more rigid domains in Lp(a) than in LDL according to higher anisotropy values in Lp(a). In both LDL and Lp(a), DPH-PC experienced a more rigid environment than DPH-SM, suggesting different environments of PC and SM in the surface shell of the lipoproteins. Fluorescence lifetimes of the labeled lipoproteins were determined by phase and modulation fluorometry. We found bimodal Lorentzian distributions for the decay times of DPH-PC and DPH-SM in LDL and Lp(a). Lifetime distribution centers for labeled lipids were very similar except for DPH-PC in Lp(a) which was shifted to longer lifetimes, suggesting a less polar environment of PC in Lp(a) than in LDL. The distributional width of DPH-PC in Lp(a) was broader than in LDL. Accordingly, phosphatidylcholine must be localized in a more homogeneous environment in LDL as compared with Lp(a). On the other hand, no difference in distributional widths was observed for DPH-SM in both lipoproteins, showing that SM organization in Lp(a) is unaffected by apo(a). From the obtained fluorescence data we propose that apoproteins discriminate between the choline phospholipids and preferentially associate with phosphatidylcholine. This effect is enhanced in Lp(a) due to the presence of apolipoprotein(a).

Fluorescence Polarization↗
PubMed1992

High-density lipoprotein antagonizes the inhibitory effects of oxidized low-density lipoprotein and lysolecithin on soluble guanylyl cyclase.

Oxidatively modified LDL (LDLox) reduces the response of soluble guanylyl cyclase to nitrovasodilators. We now demonstrate that this desensitization can be antagonized by HDL. Similar to its protective effect against LDLox, HDL also inhibited the lysolecithin-induced desensitization of soluble guanylyl cyclase. Since the lysolecithin content of LDLox correlated with the amount of lysolecithin necessary to diminish stimulation of soluble guanylyl cyclase, our data support the hypothesis that lysolecithin may be responsible for the inhibitory effect of LDLox on smooth muscle relaxation and provide evidence that the antagonistic effect of HDL against desensitization of soluble guanylyl cyclase by atherogenic compounds could be responsible for the protective role of HDL in atherosclerosis.

Animals↗
PubMed1991

Sulfhydryl-selective fluorescence labeling of lipoprotein(a) reveals evidence for one single disulfide linkage between apoproteins(a) and B-100.

Human lipoprotein(a) and low-density lipoprotein were labeled with two different sulfhydryl-selective fluorescence markers. The hydrophilic fluorophore lucifer yellow iodoacetamide and the apolar compound 6-acryloyl-2-(dimethylamino)naphthalene were used to derivatize free -SH groups in the lipoproteins. Three sulfhydryls could be detected in low-density lipoprotein, whereas only two cysteines were available in lipoprotein(a). One of the three -SH groups in low-density lipoprotein was shown to be located in close proximity to the particle surface. We suggest that this surface-exposed cysteine of apoprotein B-100 serves as a component for the disulfide linkage to apoprotein(a) in lipoprotein(a).

2-Naphthylamine↗
PubMed1991

Lipoprotein(a) mediates high affinity low density lipoprotein association to receptor negative fibroblasts.

Lipoprotein(a) (Lp(a)) is an acute phase protein with unknown function. Lp(a) binds to low density lipoprotein (LDL) receptors, as well as to plasminogen (Plg) receptors. Preincubation of normal human skin fibroblasts with Lp(a) or with apo(a) cause a severalfold increase of LDL binding. Plg and kringle-4 of Plg have no effect. LDL receptor-negative fibroblasts respond upon preincubation with apo(a) with high affinity binding of LDL with Kd values that are almost identical with those of LDL binding to the LDL receptor. Incubation of apo(a)-pretreated fibroblasts with anti-apo(a) completely abolishes the increment of LDL binding. The high affinity LDL binding to LDL receptor-negative fibroblasts could be dissociated by approximately 80 and 54% with 5 mg/ml proline and 30 mg/ml NaCl, respectively, but not with dextran sulfate. The Lp(a)- and apo(a)-triggered LDL binding to fibroblasts have no effect on LDL internalization. These findings may reflect a key function in the role as an acute phase protein and may be relevant to the high atherogeneicity of Lp(a).

Apolipoproteins↗
PubMed1991

Oxidation of lipoprotein Lp(a). A comparison with low-density lipoproteins.

Aimed at identifying possible mechanisms of the suggested high atherogenicity of Lp(a), its susceptibility for Cu(II)-induced oxidation was studied and compared with that of LDL. Since the content of antioxidants as well as the fatty acid pattern of a lipoprotein greatly affects its oxidizability, Lp(a) and LDL were characterized first with respect to these substances. Paired samples of low-density lipoproteins (LDL) and Lp(a) were isolated from seven individual donors and compared with each other. This study showed that LDL and Lp(a) are very similar with respect to their fatty acid and antioxidant composition. LDL contains approx. 1132 nmol of total fatty acids/mg lipoprotein and LDL 1466 nmol total fatty acids/mg lipoprotein. Analysis of the fatty acid composition of individual lipid classes (cholesteryl esters, phospholipids and triacylglycerols) revealed also a high similarity in the composition of these lipid classes between the two lipoproteins. A comparison of the antioxidant composition showed that Lp(a) contains less alpha-tocopherol than LDL (1.6 +/- 0.35 nmol/mg vs. 2.1 +/- 0.25 nmol/mg LDL). In copper(II)-induced lipid peroxidation experiments we found a striking difference in the susceptibility of individual lipoprotein classes between all donors. In addition, Lp(a) exhibited a 1.2 to 2.4 longer lag-phase than the corresponding LDL preparation from the same blood donor. Treatment of Lp(a) with neuraminidase resulted in a drastic decrease of the lag-phase of Lp(a). Neuraminidase treatment of LDL on the other hand had no significant effects on its susceptibility to oxidation. Supplementation of neuraminidase-treated Lp(a) with N-acetylneuraminic acid (NANA) at concentrations comparable to the naturally occurring amounts of NANA in the Lp(a) protein moiety led to an increase of the lag-phase yielding values which were comparable to those observed with native Lp(a). These results demonstrate that the fatty acid composition as well as the antioxidant concentrations of Lp(a) and LDL are quite similar; despite this fact, Cu2(+)-mediated oxidation of Lp(a) is retarded in comparison to LDL which might be due to the higher content of NANA in Lp(a).

Adult↗
PubMed1991

Determination of fatty acids in the main lipoprotein classes by capillary gas chromatography: BF3/methanol transesterification of lyophilized samples instead of Folch extraction gives higher yields.

The amount of individual fatty acids contained in the main human lipoproteins VLDL, LDL, lipoprotein (a), HDL2, and HDL3 were determined by two different methods. In Method I, the lipids were first extracted by the classical Folch procedure and then transesterified with BF3/methanol and separated by capillary GC. In Method II the lipoprotein solution was freeze dried prior to transesterification with BF3/methanol. In all lipoproteins except VLDL significantly more fatty acids were found with Method II as compared to Method I. For total fatty acids the increase was up to 17.5%, for polyunsaturated fatty acids up to 24.5%. The total fatty acid content determined by Method II resembled closely the content independently derived from the enzymatically determined lipid composition. The results indicate that in case of lipoproteins quantification of fatty acids should be made with freeze-dried samples rather than with Folch extracts.

Blood Proteins↗
PubMed1991

Platelet membrane fluidity in type IIA, type IIB and type IV hyperlipoproteinemia.

Fluorescence spectroscopy, a very sensitive index for measuring the biophysical properties of living cell systems, was used to examine the structural order of intact, resting, gel-filtered platelets from hyperlipidemic subjects (n = 48, 25-70 years) and normolipemic subjects (n = 34, 19-68 years). Fluorescence anisotropy (r[s]), which is inversely related to membrane fluidity, was estimated using 3 different fluorescent dyes, DPH, TMA-DPH, and 6-AS, known to label different regions of biological membranes. Increased membrane fluidity was observed in type IIB (n = 24, 36-62 yrs; r[s] = 0.0692 +/- 0.09) and type IV (n = 10, 33-57 yrs; r[s] = 0.058 +/- 0.006) hyperlipidemics in comparison to type IIA (n = 14, 25-70 yrs; r[s] = 0.086 +/- 0.019) and control subjects (n = 24, 28-68 yrs; r[s] = 0.079 +/- 0.012). The temperature dependency of r[s]-DPH values was significantly different (P less than 0.01) in platelets from type IIB and type IV patients compared to type IIA and control subjects of similar age. A significant positive correlation (P less than 0.005) between membrane fluidity and age was found only in healthy control subjects (n = 34, 19-68 yrs). Despite significant (P less than 0.01) differences in plasma lipid concentrations in hyperlipidemic patients and controls, significant ex vivo relations between membrane fluidity and lipoprotein concentrations, free fatty acid distribution, and increased age were found only in healthy control subjects. Plasma levels of thromboxane as well as serum selenium concentrations did not significantly differ between hypercholesterolemic, hypertriglyceridemic, and control subjects.

Adult↗
PubMed1991

Effects of dietary fish oil supplementation on platelet aggregability and platelet membrane fluidity in normolipemic subjects with and without high plasma Lp(a) concentrations.

The purpose of this study was to compare the relative effect of n-3 fatty acids on plasma lipids and platelet function in normolipemic subjects (n = 8) with plasma Lp(a) levels greater than 30 mg/dl and normolipemic subjects (n = 7) without detectable plasma Lp(a) concentrations. Six weeks of dietary supplementation (3.8 g EPA and 2.9 g DHA/d) significantly reduced (P less than 0.005) plasma TGs in both groups whereas no changes of plasma TC, LDL-C, HDL-C, and Lp(a), respectively, were found. Collagen- or thrombin-stimulated platelet aggregation and collagen- or thrombin-induced TXB2 generation from platelets decreased by approx. 45% in Lp(a)-negative and Lp(a)-positive platelet donors after a 6 week dietary intake. Four more weeks without n-3 supplementation restored the pretreatment values of TGs, platelet aggregability and TXB2 release. The biophysical properties of platelets from normolipemics with and without high plasma Lp(a) concentrations revealed a similar structural order of platelets at 37 degrees C using DPH, TMA-DPH, or 6-AS as fluorescent probes. Also similar temperature-dependent changes in platelet fluidity from 37 degrees C to 17 degrees C were observed in platelet preparations from Lp(a)-positive and Lp(a)-negative subjects. However, no subtle changes in the structural order of platelets due to nutrient intakes were found in all subjects (n = 15, 19-28 yrs) using fluorescence polarization technique. The present data suggest a similar in vitro platelet behaviour from normolipemic subjects with and without high plasma levels of Lp(a) (which is considered a risk for premature atherosclerosis) in contrast to platelet aggregability and platelet fluidity in certain hyperlipidemic stages.

Adult↗
PubMed1991

Oxidized low-density lipoprotein antagonizes the activation of purified soluble guanylate cyclase by endothelium-derived relaxing factor but does not interfere with its biosynthesis.

Oxidized low-density lipoprotein (LDLox) is a molecule with strong atherogenic properties. In a concentration dependent fashion, LDLox antagonized the activation of purified soluble guanylate cyclase by endothelium-derived relaxing factor (EDRF), which was produced in vitro by incubation of a partially purified EDRF-forming enzyme in the presence of L-arginine, Ca2+ and NADPH. The inhibitory effect of LDLox was potentiated by preincubation of the soluble guanylate cyclase with LDLox, but not when the EDRF-forming enzyme was pretreated with LDLox. As LDLox did not diminish the calmodulin-dependent conversion of L-arginine into L-citrulline by the EDRF-forming enzyme it would appear that EDRF-biosynthesis was not affected by LDLox. It is suggested that the impaired relaxant response of atherosclerotic blood vessels to endothelium-dependent vasodilators was not due to a reduced formation of EDRF but due to a diminished responsiveness of soluble guanylate cyclase.

Animals↗
PubMed1991

Lipoprotein (a) concentrations as risk indicators for atherosclerosis.

The plasma concentration of different lipoproteins were measured in 102 control children, in 42 children with a parent suffering from coronary heart disease (CHD), and in 50 children with a parent with cerebrovascular disease (CVD). Significant differences between controls and children in the other two groups were found for apolipoprotein A I, apolipoprotein B, and high density lipoprotein-cholesterol. Children of parents with CHD differed from controls in total cholesterol and apolipoprotein A II concentrations. A highly significant difference furthermore was found in lipoprotein (a) concentrations from children of parents with CHD in comparison with controls, but not between children of parents with CVD and controls. The difference in lipoprotein (a) concentrations (children of parents with CHD compared with controls) were only noticed in children above the age of 10 years. This could be explained by the observed rise of lipoprotein (a) between age 2 and 13 years, which was much more pronounced in the group with parents who had CHD. Plasma glycosaminoglycan concentrations were also measured in the three groups. They were significantly higher in children of parents with CHD and CVD compared with controls; they also varied with age.

Adolescent↗
PubMed1991

Lipoprotein metabolism and atherogenesis: implications for therapy.

The causes of atherosclerosis are numerous, but disturbances in lipid and lipoprotein (LP) metabolism undoubtedly play a key role. Although there exist multiple forms of genetic and secondary hyperlipoproteinemias linked with premature vascular diseases there are only a few LP that need to be considered: low-density LP, beta-very-low-density LP, chylomicron remnants and LP(a). In addition, low HDL levels have been found to represent an independent risk factor. Prolonged residence times of these LP lead to chemical modification and interaction with platelets, smooth muscle cells, endothelial cells and macrophages. Atherogenesis is thus a concerted action. Knowledge of the metabolic pathways of most of these LP is necessary in order to be able to specifically influence hyperlipoproteinemia with dietary measures or, ultimately, with lipid-lowering drugs.

Arteriosclerosis↗
PubMed1990

Activation of soluble guanylate cyclase by nitrovasodilators is inhibited by oxidized low-density lipoprotein.

In the presence of oxidized low-density lipoprotein the stimulatory effects of nitric oxide, sodium nitroprusside and S-nitrosoglutathione on soluble guanylate cyclase partially purified from bovine platelets were diminished in a concentration-dependent manner with IC50 values around 100 micrograms total cholesterol/ml. This inhibitory effect was potentiated about 10-fold when the enzyme was pre-incubated with the lipoprotein for 10 minutes at 37 degrees C which indicates a direct interaction of the lipoprotein with the guanylate cyclase. As oxidized low-density lipoprotein is present in the wall of atherosclerotic arteries, we suggest that the impaired response of atherosclerotic blood vessels to vasodilators may be due to a diminished activation of smooth muscle guanylate cyclase.

Animals↗
PubMed1990

A comparative study of the localization of plasminogen and apolipoprotein(a) in human carcinomas.

Recently, it was reported that there is a significant elevation of total plasma apolipoprotein(a) level in cancer patients. Because of their structural homology and immunological cross-reactivity, localization of the plasminogen and the apolipoprotein(a) was comparatively studied in cancerous tissues of breast (N = 4) and colon (N = 3) by immunofluorescence. The following results were obtained: 1) Tumor cells isolated or in nodules were strongly stained in all cancerous samples using antiserum against Pg, absorbed or not with Lp(a). 2) Tumor cells were lightly stained in two breast carcinomas, using anti-Lp(a) serum. All other carcinomas were negative. The staining was abolished when anti-Lp(a) serum was absorbed with Pg. 3) Blood vessels were strongly stained using antiserum against Lp(a) even when it was absorbed with Pg. Anti-Pg serum decorated only a few capillaries. These results show that the two proteins have different localizations: Lp(a) is seen exclusively in the vascular system. The component present at the surface of tumor cells is plasminogen (or plasmin) but not Lp(a).

Adenocarcinoma↗
PubMed1990

Immunochemical determination of lipoprotein Lp(a): comparison of Laurell electrophoresis and ELISA.

The lipoprotein Lp(a) concentrations in serum from 520 persons (317 men and 203 women) were measured by two different Laurell electrophoresis assays (one in house and one commercial) and by an ELISA technique, using polyclonal antibodies from two different animal species. The following results were obtained. 1. The two Laurell techniques gave similar results. The mean values of all three methods were comparable, whereas median values obtained by ELISA were markedly lower (0.85, 0.97 and 0.057 g/l for the two Laurell assays and the ELISA, respectively). 2. All three methods correlated very well with a correlation coefficient of greater than 0.95. 3. Women had significantly higher values than men (p less than 0.01). It was concluded that Lp(a) serum concentrations in the 'pathological range' could be measured with high enough precision by Laurell electrophoresis and by ELISA using polyclonal antibodies from different animal species.

Electrophoresis↗
PubMed1990

Is the atherogenicity of Lp(a) caused by its reactivity with proteoglycans?

Apo B-containing lipoproteins from human plasma were studied for their ability to form complexes with glycosaminoglycans (GAG) and proteoglycans (PG) in the presence of Ca++ and Mg++ ions. We studied low density lipoproteins (LDL), Lp(a) as well as Lpa-, a particle generated from Lp(a) by removing the specific antigen (apo-a). The strongest reactivity with all apo B-containing lipoproteins was found with PG, followed by GAG isolated from human aorta, and by chondroitin-6-sulphate. Lp(a), on the other hand, formed complexes with the highest glycan:lipoprotein ratio using all three complexing agents. Treatment of Lp(a) with neuraminidase did not change the reactivity. The reactivity of Lpa-, on the other hand, was between that of Lp(a) and LDL. Lipoprotein glycan complexes were incubated with mouse peritoneal macrophages. This caused cholesterol ester accumulation and foam cell formation. The amount of cholesterol ester formed correlated highly significantly with the reactivity of a given lipoprotein with different glycans. Sera obtained from patients suffering from myocardial infarction (MI) were incubated with LDL and Lp(a) and the reisolated lipoproteins were also incubated with macrophages. Lipoproteins reisolated from MI plasma caused foam cell formation with MPM to a greater extent than reisolated material incubated with normal plasma. The highest cholesteryl ester accumulation was found when Lp(a) reisolated from MI plasma was incubated with MPM. These results may at least partly explain the higher atherogenicity of Lp(a) in comparison with LDL.

Animals↗
PubMed1990

Postprandial hyperlipemia and platelet eicosanoid metabolism.

Platelet aggregations stimulated with different concentrations of collagen or ADP were reduced during postprandial hyperlipemia in normolipemic subjects. The concomitant formation of platelet AA metabolites (TXB2, 12-HHT, 12-HETE), however, was not significantly altered.

Adult↗