https://www.cell.com/cell/pdf/0092-8674(92)90362-G.pdf

Severe hypercholesterolemia and atherosclerosis in apolipoprotein E-deficient mice created by homologous recombination in ES cells

Apolipoprotein E (apoE) is a ligand for receptors that clear remnants of chylomicrons and very low density lipoproteins Lack of apoE is, therefore, expected to cause accumulation in plasma of cholesterol-rich remnants whose prolonged circulation should be atherogenic ApoE-deficient mice generated by gene targeting were used to test this hypothesis and to make a mouse model for spontaneous atherosclerosis The mutant mice had five times normal plasma cholesterol, and developed foam cell-rich depositions in their proximal aortas by age 3 months These spontaneous lesions progressed and caused severe occlusion of the coronary artery ostium by 8 months The severe yet viable phenotype of the mutants should make them valuable for investigating genetic and environmental factors that modify the atherogenic process

https://science.sciencemag.org/content/sci/258/5081/468.full.pdf

Spontaneous Hypercholesterolemia and Arterial Lesions in Mice Lacking Apolipoprotein E

High density lipoprotein metabolism.

Lipoprotein receptors and cholesterol homeostasis

The liver plays a key role in lipid metabolism. Depending on species it is, more or less, the hub of fatty acid synthesis and lipid circulation through lipoprotein synthesis. Eventually the accumulation of lipid droplets into the hepatocytes results in hepatic steatosis, which may develop as a consequence of multiple dysfunctions such as alterations in beta-oxidation, very low density lipoprotein secretion, and pathways involved in the synthesis of fatty acids. In addition an increased circulating pool of non-esterified fatty acid may also to be a major determinant in the pathogenesis fatty liver disease. This review also focuses on transcription factors such as sterol-regulatory-element-binding protein-1c and peroxisome proliferator-activated receptor alpha, which promote either hepatic fatty acid synthesis or oxidation.

https://onlinelibrary.wiley.com/doi/epdf/10.1111/j.1439-0396.2007.00752.x

Liver lipid metabolism

The major apoprotein of high density lipoprotein is apolipoprotein A-I (ApoA-I). In addition to being a structural component of this class of lipoproteins, ApoA-I also has a physiologic role as an activator of lecithin-cholesterol acyl transferase, an enzyme important in the metabolism of all lipoproteins.

To measure ApoA-I content in human plasma, to assess its immunologic activity in hyperlipoproteinemia, and to carry out certain structural studies of high density lipoproteins, we have developed a double antibody radioimmunoassay. ApoA-I, isolated by gel filtration, was used to produce monospecific antisera. ApoA-I was iodinated by chloramine-T and the resulting [125I]-ApoA-I was purified by gel filtration. > 85% of [125I]-ApoA-I was precipitated by antibody, and 90% of bound [125I]ApoA-I was displaced by “cold” ApoA-I. Other lipoproteins and apoproteins did not react.

Plasma and high density lipoprotein from normals and subjects with hyperlipoproteinemia displaced counts in parallel with ApoA-I, suggesting that the same antigenic determinants were reacting with antibody on lipid-free and lipid-associated ApoA-I. However, less than 5% of ApoA-I of high density lipoprotein reacted in the assay. Removal of the lipid by extraction increased the reactivity of ApoA-I in high density lipoprotein 15-20-fold; thus more than 95% of the ApoA-I molecules in “intact” high density lipoprotein are unreactive with antibody. Normal and hyperlipoproteinemic plasma and high density lipoproteins isolated from the same subjects continued to display parallelism with ApoA-I standard after lipid extraction, suggesting that ApoA-I of normal and hyperliproteinemic subjects are immunologically identical.

About 90% of ApoA-I was in the d 1.063-1.21 fractions of normal plasma, trace quantities were found in the lipoproteins of d 1.21 fraction. Normal plasma levels, assessed in extracted plasmas with a precision of 8%, were 100±35 mg/dl. Levels were normal in small groups of subjects with types II and IV hyperlipoproteinemia and high in pregnancy. However, larger population studies need to be performed to determine the distribution of ApoA-I levels in the various hyperlipoproteinemias.

/pdf/the-structure-of-human-high-density-lipoprotein-and-the-4uk66os4he.pdf

The Structure of Human High Density Lipoprotein and the Levels of Apolipoprotein A-I in Plasma as Determined by Radioimmunoassay

We have developed a double antibody radioimmunoassay (RIA) for human apolipoprotein B (ApoB). The assay measures not only the ApoB content of beta-lipoproteins (low density lipoproteins [LDL]) but also that contained in the other lipoproteins in plasma. Purified lymph and plasma chylomicrons and plasma very low density lipoproteins (VLDL) produced displacement curves in the assay system which paralleled those produced by pure LDL. Thus, the ApoB found in chylomicrons, VLDL, and LDL were immunologically identical. ApoB accounted for about 25 and 35%, respectively, of the total protein of chylomicrons and VLDL by RIA. VLDL and LDL preparations from normal and hyperlipoproteinemic subjects also produced parallel displacement curves, suggesting that the ApoB of normal and hyperlipoproteinemic subjects were immunologically identical. High density lipoproteins and abetalipoproteinemic plasma displaced no counts, nor did the sera of several animal species produce any useful displacement curves in this system. The fasting total plasma ApoB concentration of normal subjects was 83+/-16 mg/dl (mean+/-SD). ApoB levels were high in Type II (162+/-16), and less so in Type IV (112+/-24) and Type V (105+/-17).When plasma ApoB concentration in Type IV patients was graphed against plasma glycerides, two subpopulations, which may represent different genetic or biochemical subgroups, were apparent.ApoB concentration in individuals on constant diet and drug regimen was stable over weeks to months. Greater than 90% of ApoB of normal and Type II subjects was in the d > 1.006 plasma fraction. By contrast, only 50-80% of ApoB was in the d > 1.006 fraction in Types IV and V. Thus, hypertriglyceridemia was associated primarily with a redistribution of ApoB to the lighter density fractions; by contrast, in hypercholesterolemia absolute ApoB concentration was markedly increased.

/pdf/assay-of-total-plasma-apolipoprotein-b-concentration-in-2c5okw7xdu.pdf

Assay of Total Plasma Apolipoprotein B Concentration in Human Subjects

To assess genetic variation of murine lipoprotein profiles, plasma lipoproteins of 11 inbred strains, AKR/J, BALB/cByJ, C3H/HeJ, C57BL/6J, C57BL/6ByJ, C57L/J, DBA/1LacJ, 129/J, NZB/B1NJ, PL/J, and SWR/J, were analyzed by gel-permeation chromatography (fast peptide liquid chromatography) and nondenaturing gradient gel electrophoresis. Vena caval blood was drawn after 18 to 20 hours of fasting. Plasma triglyceride and cholesterol concentrations ranged from 12.9 mg/dL (C57BL/6ByJ) to 66.9 mg/dL (C3H/HeJ) and from 54.8 mg/dL (AKR/J) to 128.5 mg/dL (NZB/B1NJ), respectively. Mouse strain-related heterogeneities of very low-, low-, and high-density lipoprotein (VLDL, LDL, and HDL, respectively) concentrations were documented; VLDL-triglyceride concentrations ranged from 7.5 mg/dL to 38.8 mg/dL, LDL cholesterol from 12.0 mg/dL to 39.6 mg/dL, and HDL cholesterol from 41.3 mg/dL to 92.4 mg/dL. Hyper-VLDL-triglyceridemia was present in C3H/HeJ and SWR/J strains and hyper-LDL-cholesterolemia in NZB/B1NJ, C3H/HeJ, and DBA/1LacJ. VLDL cholesterol/VLDL triglyceride ratios also ranged widely among strains (0.13 to 0.43), with C57BL/6J, C57BL/6ByJ, and C57L/J, the strains particularly susceptible to diet-induced atherosclerosis, having the highest VLDL-lipid ratio. LDL and HDL size heterogeneities were also observed. LDL and HDL diameters ranged between 24.1 nm and 29.4 nm, and between 9.24 nm and 10.32 nm, respectively. Although LDL sizes showed no segregation, HDL sizes fell into two groups. C57L/J and C57BL/6J possessed low HDL-cholesterol concentrations and small-sized HDL. HDL sizes were positively correlated with HDL-cholesterol concentrations (r = .90, P less than .001) and LDL-cholesterol concentrations (r = .85, P less than .001), but LDL sizes did not correlate with lipoprotein concentrations.(ABSTRACT TRUNCATED AT 250 WORDS)

Genetic heterogeneity of lipoproteins in inbred strains of mice: analysis by gel-permeation chromatography.

Smaller very low density lipoprotein (VLDL) remnants interact more readily with tissues than do larger "intact" VLDL. This may be related to changes in the availability of VLDL apoproteins on the surface of the lipoproteins. To test this hypothesis VLDL were incubated at 37 degrees C with bovine milk lipase (LPL), and the abilities of LPL-treated VLDL preparations to compete with (125)I-low density lipoproteins (LDL) for interaction with cultured normal human fibroblasts were measured. At the same time, the immunologic activities of these preparations were also tested by double antibody radioimmunoassay. Triglyceride (TG) contents of VLDL fell by 30-90% during incubation with LPL and, on zonal ultracentrifugation, VLDL of faster Svedberg unit of flotation (S(f1.063)) rates (>150) were gradually converted to smaller VLDL with lower S(f) rates (21-60). LPL-treated VLDL competed two to five times more effectively with (125)I-LDL for binding to cellular receptors than did control VLDL. Control VLDL incubated with heat-inactivated LPL at 37 degrees C, or with active LPL at 4 degrees C had unaltered cell reactivities and TG contents compared with VLDL incubated without any enzyme. The direct uptake and degradation of LPL-treated VLDL was also assessed by using VLDL (125)I-labeled in apoprotein (Apo)B. LPL-treated VLDL-(125)I-ApoB were taken up and degraded by fibroblast at greater rates than were control VLDL-(125)I-ApoB. Thus, hydrolysis of VLDL lipids was accompanied by an increased ability of VLDL to interact with fibroblasts. The immunoreactivity of ApoB in the same VLDL preparations, expressed as the "apparent ApoB contents" of LPL-treated VLDL, increased by 10-50% (P < 0.02) in those assays that contained anti-LDL antisera, but the ApoB of control VLDL remained constant. However, assays that contained antisera directed against ApoB isolated from VLDL did not distinguish between LPL-treated and control VLDL. Thus, VLDL lipid hydrolysis was accompanied by changes in the immunoreactivity of VLDL-ApoB, which probably reflect changes in the disposition of ApoB on the surface of VLDL. The altered disposition of ApoB on VLDL "remnants" may be related to their enhanced interaction with cells.

/pdf/lipolysis-produces-changes-in-the-immunoreactivity-and-cell-4fk6kg8p23.pdf

Barbara Pfleger

Papers

The Structure of Human High Density Lipoprotein and the Levels of Apolipoprotein A-I in Plasma as Determined by Radioimmunoassay

Assay of Total Plasma Apolipoprotein B Concentration in Human Subjects

Genetic heterogeneity of lipoproteins in inbred strains of mice: analysis by gel-permeation chromatography.

Lipolysis Produces Changes in the Immunoreactivity and Cell Reactivity of Very Low Density Lipoproteins

Lipolysis of LDL with phospholipase A2 alters the expression of selected apoB-100 epitopes and the interaction of LDL with cells