scholarly journals Interaction of plasma lipoproteins with human platelets

Blood ◽  
1984 ◽  
Vol 64 (2) ◽  
pp. 365-374 ◽  
Author(s):  
LK Curtiss ◽  
EF Plow
Keyword(s):  
Binding Site ◽  
Dissociation Constant ◽  
Polyacrylamide Gel Electrophoresis ◽  
Ldl Receptor ◽  
Human Platelets ◽  
Plasma Lipoproteins ◽  
High Density Lipoproteins ◽  
Interaction Sites ◽  
Ldl Particles ◽  
Ldl Binding

Abstract Human plasma low density lipoproteins (LDL) and high density lipoproteins (HDL) were radioiodinated and their interaction with washed human platelets was assessed. Both ligands were bound by the platelet at 37 degrees C, and an apparent equilibrium was attained within two hours. Minimal binding was observed at 22 degrees C or 4 degrees C. The specificity of these interactions was indicated by the observations that: (a) labeled and nonlabeled lipoproteins interacted with the platelet with the same apparent affinity; (b) nonlabeled lipoproteins inhibited binding, whereas unrelated plasma proteins did not; and (c) the platelet-bound ligands exhibited the appropriate apoprotein chain compositions when analyzed by polyacrylamide gel electrophoresis. Binding of HDL and LDL was found to be independent of the state of platelet activation and did not require divalent ions. Binding of HDL to the platelet was saturable, and a class of sites that maximally bound 1,585 +/- 390 HDL particles, with a dissociation constant of 3.1 X 10(-8) mol/L, was identified. Binding of LDL to the platelet was more complex, but evidence for a class of sites that bound 7,075 +/- 4,800 LDL particles, with a dissociation constant of 4 X 10(- 8) mol/L, was found. LDL was a poor inhibitor of 125I-HDL binding to the platelet, whereas HDL was an effective inhibitor of 125I-LDL binding. The capacity of HDL to bind or inhibit LDL binding was not dependent on its apoprotein E content. These results are most readily interpreted in terms of two types of lipoprotein interaction sites on platelets: (1) an HDL binding site that does not bind or interacts poorly with LDL, and (2) an LDL binding site that recognizes or is otherwise altered by HDL. The HDL site may be similar to the HDL receptor expressed by steroidogenic tissues in terms of apoprotein specificity. The LDL site is not the same as the LDL receptor of most extrahepatic cells.

scholarly journals Interaction of plasma lipoproteins with human platelets

Blood ◽  
1984 ◽  
Vol 64 (2) ◽  
pp. 365-374
Author(s):  
LK Curtiss ◽  
EF Plow
Keyword(s):  
Binding Site ◽  
Dissociation Constant ◽  
Polyacrylamide Gel Electrophoresis ◽  
Ldl Receptor ◽  
Human Platelets ◽  
Plasma Lipoproteins ◽  
High Density Lipoproteins ◽  
Interaction Sites ◽  
Ldl Particles ◽  
Ldl Binding

Human plasma low density lipoproteins (LDL) and high density lipoproteins (HDL) were radioiodinated and their interaction with washed human platelets was assessed. Both ligands were bound by the platelet at 37 degrees C, and an apparent equilibrium was attained within two hours. Minimal binding was observed at 22 degrees C or 4 degrees C. The specificity of these interactions was indicated by the observations that: (a) labeled and nonlabeled lipoproteins interacted with the platelet with the same apparent affinity; (b) nonlabeled lipoproteins inhibited binding, whereas unrelated plasma proteins did not; and (c) the platelet-bound ligands exhibited the appropriate apoprotein chain compositions when analyzed by polyacrylamide gel electrophoresis. Binding of HDL and LDL was found to be independent of the state of platelet activation and did not require divalent ions. Binding of HDL to the platelet was saturable, and a class of sites that maximally bound 1,585 +/- 390 HDL particles, with a dissociation constant of 3.1 X 10(-8) mol/L, was identified. Binding of LDL to the platelet was more complex, but evidence for a class of sites that bound 7,075 +/- 4,800 LDL particles, with a dissociation constant of 4 X 10(- 8) mol/L, was found. LDL was a poor inhibitor of 125I-HDL binding to the platelet, whereas HDL was an effective inhibitor of 125I-LDL binding. The capacity of HDL to bind or inhibit LDL binding was not dependent on its apoprotein E content. These results are most readily interpreted in terms of two types of lipoprotein interaction sites on platelets: (1) an HDL binding site that does not bind or interacts poorly with LDL, and (2) an LDL binding site that recognizes or is otherwise altered by HDL. The HDL site may be similar to the HDL receptor expressed by steroidogenic tissues in terms of apoprotein specificity. The LDL site is not the same as the LDL receptor of most extrahepatic cells.

Abstract 14697: Novel Assays for Quantification of Lipoprotein-Associated (PCSK9-apoB, PCSK9-Lp(a)) Proprotein Covertase Subtilisin/Kexin Type 9 (PCKS9)

Circulation ◽  
2015 ◽  
Vol 132 (suppl_3) ◽  
Author(s):  
Calvin Yeang ◽  
Yun-Seok Choi ◽  
Sang-Rok Lee ◽  
Monica L Bertoia ◽  
Eric B Rimm ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
Human Plasma ◽  
Polyclonal Antibodies ◽  
Human Monoclonal Antibody ◽  
Ldl Receptor ◽  
Plasma Lipoproteins ◽  
Health Study ◽  
Clinical Samples ◽  
Ldl Particles

Background: PCSK9 is a major regulator of plasma LDL-C. Monoclonal antibodies to PCSK9 lower LDL-C by 45-65% and Lp(a) by 9-38%. The canonical function of PCSK9 is binding of LDL-receptor (LDLR) via its extracellular EGF-A domain, and subsequently mediating LDLR degradation. However, PCSK9 also weakly associates with plasma lipoproteins, with 20-40% of total plasma PCSK9 found on LDL. However, most LDL particles do not contain PCSK9. Whether PCSK9 also associates with other lipoproteins such as Lp(a) are not well described. Methods: Sensitive and quantitative sandwich-based ELISA assays were developed to measure PCSK9 associated plasma lipoproteins in both mouse and human plasma. For human plasma, commercial rabbit polyclonal antibodies binding to the C-terminal region of PCSK9 (Abgent, ThermoFisher) or REGN727 human monoclonal antibody were bound to microtiter well plates. Plasma was added and monoclonal antibodies MB47 and LPA4, binding to apoB-100 and apo(a) respectively, were used to detect PCSK9-apoB-100 and PCSK9-Lp(a) complexes with a chemiluminescent ELISA. For mouse assays, REGN727 was used as the capture antibody as it detects mouse PCSK9 and monoclonal antibody LF3 was used to detect mouse apoB. Results: PCSK9-apoB and PCSK9-Lp(a) complexes could be detected in both human plasma and in various mouse models expressing apo(a) or Lp(a). The signal to noise ratio was ~20 fold in various clinical samples, including in healthy subjects and in patients with cardiovascular disease. In 536 clinical samples from the Health Professional Follow-Up Study, PCSK9-Lp(a) correlated strongly with Lp(a) (r=0.59, p<0.001, age-adjusted) but not other lipid variables. PCSK9-apoB correlated weakly with PCSK9-Lp(a) (r=0.30, p<0.001, age-adjusted) and LDL-C (r=0.22, p<0.001, age-adjusted). These associations were virtually the same in 526 women in the Nurses’ Health Study. Conclusions: Novel ELISAs were generated to quantitate lipoprotein-associated PCSK9 in transgenic mouse and human plasma, including on apoB and Lp(a). Changes in PCSK9-Lp(a) complexes may provide insights into the Lp(a)-lowering effect of PCSK9 antibodies. Whether these assays will predict CVD outcomes waits to be determined in PCSK9 antibody and epidemiological studies.

LIPOPROTEIN BINDING TOHUMAN PLATELETS IS LOCATED AT GPIIb/IIIa COMPLEX

1987 ◽  
Author(s):  
E Koller ◽  
F Koller
Keyword(s):  
Specific Binding ◽  
Human Platelets ◽  
Plasma Lipoproteins ◽  
High Density Lipoproteins ◽  
Molecular Weights ◽  
Disulfide Reduction ◽  
Specific Binding Sites ◽  
First Results ◽  
Lipoprotein Binding ◽  
High Concentrations

Human Platelets possess specific binding sites for low density lipoproteins (LDL) and high density lipoproteins(HDL)(1). Binding of both classes of plasma lipoproteins, though competitive, has been shown by several groups to facilitate platelet activation.Isolated washed platelets occasionally aggregate upon addition of high concentrations of LDL even in the absence of known platelet activators. The proteins responsible for this binding have been visualized by ligand blotting (2). Both types of ligand specifically bind to two glycoproteins with molecular weights of 135 and 115 kD, respectively. The conditions of binding to these two proteins, however, markedly differ from those known for other lipoprotein receptors.Following extensive purification, these two species are still present at concentrations relative to each other that depend markedly on the conditions of purification. The purified, solubilized receptor was tested under various conditions, including in the absence and presence of calcium, after disulfide-reduction, and following chymotrypsin digestion. In parallel experiments, the same preparations were tested with respect to binding of fibrinogen, different lectins, and thealloantibody anti-PlAI . The results strongly support the assumption, that the two protein bands associated with lipoprotein binding are constituents of the GP-IIb/IIIa complex.These first results may have greatimplications for our understanding ofthe mechanism by which lipoproteins facilitate platelet stimulation.

Reduction in VLDL, but not HDL, in plasma of rats deficient in choline

1990 ◽  
Vol 68 (2) ◽  
pp. 552-558 ◽  
Author(s):  
Zemin Yao ◽  
Dennis E. Vance
Keyword(s):  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
High Density ◽  
Plasma Lipoproteins ◽  
Male Rats ◽  
High Density Lipoproteins ◽  
Choline Deficiency ◽  
Deficient Diet ◽  
Apo B

We have analyzed plasma lipoprotein levels in young male rats fed a choline-deficient diet for 3 days. We confirmed previous studies that choline deficiency promotes 6.5-fold accumulation of triacylgycerol in the liver (23.9 ± 6.0 versus 3.69 ± 0.92 μmol/g liver) and reduction of triacylglycerol concentration in plasma by 60% (0.17 ± 0.04 versus 0.46 ± 0.10μmol/mL plasma). Agarose gel electrophoresis showed that the plasma very low density lipoprotein (VLDL) levels were reduced in choline-deficient rats, but the concentration of plasma high density lipoproteins (HDL) was not affected. Sodium dodecyl sulfate – polyacrylamide gel electrophoresis of fractionated plasma lipoproteins revealed that the concentrations of apolipoproteins (apo) BH, BL, and E in VLDL from choline-deficient rats were 37.1, 11.0, and 37.2% of normal levels, respectively. In contrast, the amount of apo A-I, the major one in HDL, was almost unchanged. Correspondingly, there were decreased lipid (mainly phosphatidylcholine and triacylglycerol) levels in VLDL from choline-deficient rats, but no change in the levels of phosphatidylcholine, cholesterol, and cholesterol ester in HDL. There were similar levels of apo B and E (components of VLDL) in homogenates of livers from normal and choline-deficient rats, as determined by immunoblotting. These results support the hypothesis that choline deficiency causes reduction of VLDL, but not HDL, levels in plasma as a consequence of impaired hepatic VLDL secretion.Key words: choline deficiency, very low and high density lipoproteins, apolipoproteins, rat plasma.

scholarly journals Lipid Disorders and Mutations in the APOB Gene

2004 ◽  
Vol 50 (10) ◽  
pp. 1725-1732 ◽  
Author(s):  
Amanda J Whitfield ◽  
P Hugh R Barrett ◽  
Frank M van Bockxmeer ◽  
John R Burnett
Keyword(s):  
Ldl Receptor ◽  
Lipoprotein Metabolism ◽  
Plasma Lipoproteins ◽  
Premature Coronary Artery Disease ◽  
Missense Mutations ◽  
Apob Gene ◽  
Apo B ◽  
Ldl Particles ◽  
Genetic Features ◽  
Familial Hypobetalipoproteinemia

Abstract Background: Plasma lipoproteins are important determinants of atherosclerosis. Apolipoprotein (apo) B is a large, amphipathic glycoprotein that plays a central role in human lipoprotein metabolism. Two forms of apoB are produced from the APOB gene by a unique posttranscriptional editing process: apoB-48, which is required for chylomicron production in the small intestine, and apoB-100, required for VLDL production in the liver. In addition to being the essential structural component of VLDL, apoB-100 is the ligand for LDL-receptor-mediated endocytosis of LDL particles. Content: The study of monogenic dyslipidemias has revealed important aspects of metabolic pathways. In this review, we discuss the regulation of apoB metabolism and examine how APOB gene defects can lead to both hypo- and hypercholesterolemia. The key clinical, metabolic, and genetic features of familial hypobetalipoproteinemia and familial ligand-defective apoB-100 are described. Summary: Missense mutations in the LDL-receptor-binding domain of apoB cause familial ligand-defective apoB-100, characterized by hypercholesterolemia and premature coronary artery disease. Other mutations in APOB can cause familial hypobetalipoproteinemia, characterized by hypocholesterolemia and resistance to atherosclerosis. These naturally occurring mutations reveal key domains in apoB and demonstrate how monogenic dyslipidemias can provide insight into biologically important mechanisms.

scholarly journals A transient amphipathic helix in the prodomain of PCSK9 facilitates binding to low-density lipoprotein particles

2020 ◽  
Vol 295 (8) ◽  
pp. 2285-2298
Author(s):  
Samantha K. Sarkar ◽  
Alexander C. Y. Foo ◽  
Angela Matyas ◽  
Ikhuosho Asikhia ◽  
Tanja Kosenko ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Low Density Lipoprotein ◽  
Ldl Receptor ◽  
Density Lipoprotein ◽  
Low Density ◽  
Loss Of Function ◽  
Ldl Particles ◽  
Ldl Binding ◽  
Α Helix

Proprotein convertase subtilisin/kexin type-9 (PCSK9) is a ligand of low-density lipoprotein (LDL) receptor (LDLR) that promotes LDLR degradation in late endosomes/lysosomes. In human plasma, 30–40% of PCSK9 is bound to LDL particles; however, the physiological significance of this interaction remains unknown. LDL binding in vitro requires a disordered N-terminal region in PCSK9's prodomain. Here, we report that peptides corresponding to a predicted amphipathic α-helix in the prodomain N terminus adopt helical structure in a membrane-mimetic environment. This effect was greatly enhanced by an R46L substitution representing an atheroprotective PCSK9 loss-of-function mutation. A helix-disrupting proline substitution within the putative α-helical motif in full-length PCSK9 lowered LDL binding affinity >5-fold. Modeling studies suggested that the transient α-helix aligns multiple polar residues to interact with positively charged residues in the C-terminal domain. Gain-of-function PCSK9 mutations associated with familial hypercholesterolemia (FH) and clustered at the predicted interdomain interface (R469W, R496W, and F515L) inhibited LDL binding, which was completely abolished in the case of the R496W variant. These findings shed light on allosteric conformational changes in PCSK9 required for high-affinity binding to LDL particles. Moreover, the initial identification of FH-associated mutations that diminish PCSK9's ability to bind LDL reported here supports the notion that PCSK9-LDL association in the circulation inhibits PCSK9 activity.

scholarly journals Native and modified low-density-lipoprotein interaction with human platelets in normal and homozygous familial-hypercholesterolaemic subjects

1984 ◽  
Vol 224 (1) ◽  
pp. 13-20 ◽  
Author(s):  
A Shmulewitz ◽  
J G Brook ◽  
M Aviram
Keyword(s):  
Platelet Aggregation ◽  
Binding Site ◽  
Opposite Effect ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Normal Subjects ◽  
Human Platelets ◽  
Low Density ◽  
Platelet Activity ◽  
Ldl Binding

The binding of low-density lipoproteins (LDL) as well as LDL modified by cyclohexanedione (CHD-LDL) to gel-filtered platelets (GFP) and its effect on platelet function were studied in normal and in homozygous familial hypercholesterolaemic (HFH) subjects. Only normal-derived LDL could significantly compete with normal 125I-labelled LDL for binding to normal platelets. When GFP from normal subjects were incubated with normal LDL at concentrations of 25-200 micrograms of protein/ml, platelet aggregation in the presence of thrombin (0.5 i.u./ml) was increased by 65-186%. CHD-LDL, at similar concentrations, caused the opposite effect and decreased platelet aggregation by 26-47%. Both LDL and CHD-LDL (100 micrograms/ml) from HFH patients, when incubated with normal GFP, caused a significant reduction in platelet aggregation (33 and 50% respectively). When HFH-derived platelets were used, both patient LDL and CHD-LDL (but not the normal lipoprotein) could markedly compete with the patient 125I-labelled LDL for binding to the platelets. LDL and CHD-LDL (100 micrograms/ml) from normal subjects decreased aggregation of HFH-platelets by 52 and 85% respectively, while corresponding concentrations of LDL derived from HFH subjects (HFH-LDL) and CHD-LDL derived from HFH subjects (CHD-HFH-LDL) increased platelet aggregation by 165 and 65% respectively. The present results support the following conclusions: platelet activation by LDL in normal subjects is through the arginine-rich apoprotein-binding site; more than one binding site for LDL exists on platelets; under certain circumstances, LDL binding can cause a reduction in platelet activity; specificity for LDL binding to the platelets resides in different regions of the lipoprotein in HFH and in normal subjects. We have thus suggested a model for LDL-platelet interaction in normal and in HFH subjects.

scholarly journals Properties of the factor Xa binding site on human platelets.

1978 ◽  
Vol 253 (19) ◽  
pp. 6908-6916 ◽  
Author(s):  
J.P. Miletich ◽  
C.M. Jackson ◽  
P.W. Majerus
Keyword(s):  
Binding Site ◽  
Factor Xa ◽  
Human Platelets

Selective uptake of cholesteryl esters from various classes of lipoproteins by HepG2 cells

1999 ◽  
Vol 77 (2) ◽  
pp. 157-163 ◽  
Author(s):  
Louise Brissette ◽  
Marie-Claude Charest ◽  
Louise Falstrault ◽  
Julie Lafond ◽  
David Rhainds ◽  
...  
Keyword(s):  
Total Lipid ◽  
Hepg2 Cells ◽  
Ldl Receptor ◽  
Inverse Correlation ◽  
Low Density Lipoproteins ◽  
High Density Lipoproteins ◽  
Low Density ◽  
Cholesteryl Esters ◽  
Very Low Density Lipoproteins ◽  
Selective Uptake

Selective uptake of cholesteryl esters (CE) from lipoproteins by cells has been extensively studied with high density lipoproteins (HDL). It is only recently that such a mechanism has been attributed to intermediate and low density lipoproteins (IDL and LDL). Here, we compare the association of proteins and CE from very low density lipoproteins (VLDL), IDL, LDL and HDL3 to HepG2 cells. These lipoproteins were either labelled in proteins with 125I or in CE with 3H-cholesteryl oleate. We show that, at any lipoprotein concentration, protein association to the cells is significantly smaller for IDL, LDL, and HDL3 than CE association, but not for VLDL. At a concentration of 20 µg lipoprotein/mL, these associations reveal CE-selective uptake in the order of 2-, 4-, and 11-fold for IDL, LDL, and HDL3, respectively. These studies reveal that LDL and HDL3 are good selective donors of CE to HepG2 cells, while IDL is a poor donor and VLDL is not a donor. A significant inverse correlation (r2 = 0.973) was found between the total lipid/protein ratios of the four classes of lipoproteins and the extent of CE-selective uptake by HepG2 cells. The fate of 3H-CE of the two best CE donors (LDL and HDL3) was followed in HepG2 cells after 3 h of incubation. Cells were shown to hydrolyze approximately 25% of the 3H-CE of both lipoproteins. However, when the cells were treated with 100 µM of chloroquine, a lysosomotropic agent, 85 and 40% of 3H-CE hydrolysis was lost for LDL and HDL3, respectively. The fate of LDL and HDL3-CE in HepG2 cells deficient in LDL-receptor was found to be the same, indicating that the portion of CE hydrolysis sensitive to chloroquine is not significantly linked to LDL-receptor activity. Thus, in HepG2 cells, the magnitude of CE-selective uptake is inversely correlated with the total lipid/protein ratios of the lipoproteins and CE-selective uptake from the two best CE donors (LDL and HDL3) appears to follow different pathways.Key words: lipoprotein, receptor, HepG2 cell, selective uptake, lipid, cholesterol, binding.

Kaempferol ameliorates symptoms of metabolic syndrome by improving blood lipid profile and glucose tolerance

2021 ◽  
Author(s):  
Ayasa Ochiai ◽  
Mahmoud Ben Othman ◽  
Kazuichi Sakamoto
Keyword(s):  
Metabolic Syndrome ◽  
Glucose Tolerance ◽  
Molecular Mechanisms ◽  
Ldl Receptor ◽  
Blood Lipid ◽  
Apolipoprotein A1 ◽  
High Density Lipoproteins ◽  
Blood Lipid Profile ◽  
Beneficial Effects ◽  
Apoa1 Gene

Abstract Kaempferol (KPF) is a dietary polyphenol reported to have various beneficial effects on human health. However, its molecular mechanisms in regulating lipid and glucose metabolism are not fully understood. This study examined the effects of KPF on obesity, dyslipidemia, and diabetes in Tsumura, Suzuki, Obese Diabetes (TSOD) mice. The six-week administration of KPF decreased fat weight, serum total cholesterol, and low-density lipoproteins (LDLs); increased high-density lipoproteins (HDLs); and improved glucose tolerance. Additionally, KPF increased LDL receptor (LDLR) and apolipoprotein A1 (ApoA1) gene expression and decreased serum resistin levels. These findings suggest that the decrease in LDL and the increase in HDL caused by KPF may be due to increases in hepatic LDLR and ApoA1 expression, respectively. Furthermore, it is possible that the improvement in glucose tolerance by KPF may occur via resistin reduction. These mechanisms may be parts of complex mechanism by which KPF improves metabolic syndrome.

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