scholarly journals Degradation by cultured fibroblasts and macrophages of unmodified and 1,2-cyclohexanedione-modified low-density lipoprotein from normal and homozygous familial hypercholesterolaemic subjects

1982 ◽  
Vol 202 (1) ◽  
pp. 145-152 ◽  
Author(s):  
B L Knight ◽  
A K Soutar
Keyword(s):  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Normal Subjects ◽  
Proteolytic Degradation ◽  
Low Density ◽  
Human Skin Fibroblasts ◽  
Cultured Fibroblasts ◽  
Apparent Affinity ◽  
Monolayer Cultures

Monolayer cultures of human skin fibroblasts and monocyte-derived macrophages were used to examine the effect of cyclohexane-1,2-dione modification on the proteolytic degradation of 125I-labelled low-density lipoprotein (LDL) from normal subjects (NLDL) and homozygous familial hypercholesterolaemic subjects (FHLDL). Normal fibroblasts, pre-incubated in lipoprotein-deficient serum, and macrophages, pre-incubated in whole serum, exhibited both saturable and non-saturable degradation of LDL. In fibroblasts, the saturable receptor-mediated degradation of FHLDL was similar to that of NLDL and was abolished if the lipoproteins were modified with cyclohexanedione. The rate of non-saturable degradation of FHLDL was at least 3-fold higher than that of NLDL and each was decreased by approx. 60% after modification. In macrophages, saturable degradation was decreased but not abolished by modification. The apparent affinity for unmodified LDL was lower than that of the fibroblast receptor and was greater for NLDL than for FHLDL. Non-saturable degradation of FHLDL by macrophages was only slightly higher than that of NLDL. Modification with cyclohexanedione decreased the rate of non-saturable degradation of NLDL by 30%, but increased that of FHLDL by 75%. These experiments show differences between the degradation of 125I-labelled NLDL and FHLDL. They suggest that macrophages can degrade LDL by a saturable process with different properties from that mediated by the fibroblast receptor and that, in vitro, the rate of degradation of the modified LDL is not the same as the rate of non-receptor-mediated degradation of unmodified LDL.

Intralysosomal Accumulation of Colloidal Gold-Low Density Lipoprotein Conjugates in Chloroquine-Treated Fibroblasts

1985 ◽  
Vol 43 ◽  
pp. 546-547 ◽  
Author(s):  
Dean A. Handley ◽  
Cynthia M. Arbeeny ◽  
Larry D. Witte
Keyword(s):  
Colloidal Gold ◽  
Cultured Cells ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Coated Vesicle ◽  
Low Density ◽  
Cholesterol Esters ◽  
Cultured Fibroblasts ◽  
Surface Receptors ◽  
Ldl Particles

Low density lipoproteins (LDL) are the major cholesterol carrying particles in the blood. Using cultured cells, it has been shown that LDL particles interact with specific surface receptors and are internalized via a coated pit-coated vesicle pathway for lysosomal catabolism. This (Pathway has been visualized using LDL labeled to ferritin or colloidal gold. It is now recognized that certain lysomotropic agents, such as chloroquine, inhibit lysosomal enzymes that degrade protein and cholesterol esters. By interrupting cholesterol ester hydrolysis, chloroquine treatment results in lysosomal accumulation of cholesterol esters from internalized LDL. Using LDL conjugated to colloidal gold, we have examined the ultrastructural effects of chloroquine on lipoprotein uptake by normal cultured fibroblasts.

scholarly journals Reductions of copper ion-mediated low-density lipoprotein (LDL) oxidations of trypsin inhibitors, the sweet potato root major proteins, and LDL binding capacities

2020 ◽  
Vol 61 (1) ◽  
Author(s):  
Yeh-Lin Lu ◽  
Chia-Jung Lee ◽  
Shyr-Yi Lin ◽  
Wen-Chi Hou
Keyword(s):  
Sweet Potato ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Trypsin Inhibitors ◽  
Water Soluble ◽  
Affinity Column ◽  
Low Density ◽  
Native Page

Abstract Background The root major proteins of sweet potato trypsin inhibitors (SPTIs) or named sporamin, estimated for 60 to 80% water-soluble proteins, exhibited many biological activities. The human low-density lipoprotein (LDL) showed to form in vivo complex with endogenous oxidized alpha-1-antitrypsin. Little is known concerning the interactions between SPTIs and LDL in vitro. Results The thiobarbituric-acid-reactive-substance (TBARS) assays were used to monitor 0.1 mM Cu2+-mediated low-density lipoprotein (LDL) oxidations during 24-h reactions with or without SPTIs additions. The protein stains in native PAGE gels were used to identify the bindings between native or reduced forms of SPTIs or soybean TIs and LDL, or oxidized LDL (oxLDL). It was found that the SPTIs additions showed to reduce LDL oxidations in the first 6-h and then gradually decreased the capacities of anti-LDL oxidations. The protein stains in native PAGE gels showed more intense LDL bands in the presence of SPTIs, and 0.5-h and 1-h reached the highest one. The SPTIs also bound to the oxLDL, and low pH condition (pH 2.0) might break the interactions revealed by HPLC. The LDL or oxLDL adsorbed onto self-prepared SPTIs-affinity column and some components were eluted by 0.2 M KCl (pH 2.0). The native or reduced SPTIs or soybean TIs showed different binding capacities toward LDL and oxLDL in vitro. Conclusion The SPTIs might be useful in developing functional foods as antioxidant and nutrient supplements, and the physiological roles of SPTIs-LDL and SPTIs-oxLDL complex in vivo will investigate further using animal models.

scholarly journals Development of Capture Assays for Different Modifications of Human Low-Density Lipoprotein

2005 ◽  
Vol 12 (1) ◽  
pp. 68-75 ◽  
Author(s):  
Gabriel Virella ◽  
M. Brooks Derrick ◽  
Virginia Pate ◽  
Charlyne Chassereau ◽  
Suzanne R. Thorpe ◽  
...  
Keyword(s):  
Oxidized Ldl ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Gas Chromatography Mass Spectrometry ◽  
Low Density ◽  
Modified Ldl ◽  
Capture Assay ◽  
Carboxymethyl Lysine

ABSTRACT Antibodies to malondialdehyde (MDA)-modified low-density lipoprotein (LDL), copper-oxidized LDL (oxLDL), N ε(carboxymethyl) lysine (CML)-modified LDL, and advanced glycosylation end product (AGE)-modified LDL were obtained by immunization of rabbits with in vitro-modified human LDL preparations. After absorption of apolipoprotein B (ApoB) antibodies, we obtained antibodies specific for each modified lipoprotein with unique patterns of reactivity. MDA-LDL antibodies reacted strongly with MDA-LDL and also with oxLDL. CML-LDL antibodies reacted strongly with CML-LDL and also AGE-LDL. oxLDL antibodies reacted with oxLDL but not with MDA-LDL, and AGE-LDL antibodies reacted with AGE-LDL but not with CML-LDL. Capture assays were set with each antiserum, and we tested their ability to capture ApoB-containing lipoproteins isolated from precipitated immune complexes (IC) and from the supernatants remaining after IC precipitation (free lipoproteins). All antibodies captured lipoproteins contained in IC more effectively than free lipoproteins. Analysis of lipoproteins in IC by gas chromatography-mass spectrometry showed that they contained MDA-LDL and CML-LDL in significantly higher concentrations than free lipoproteins. A significant correlation (r = 0.706, P < 0.019) was obtained between the MDA concentrations determined by chemical analysis and by the capture assay of lipoproteins present in IC. In conclusion, we have developed capture assays for different LDL modifications in human ApoB/E lipoprotein-rich fractions isolated from precipitated IC. This approach obviates the interference of IC in previously reported modified LDL assays and allows determination of the degree of modification of LDL with greater accuracy.

Sign in / Sign up

Export Citation Format

Share Document