Abstract-The prevention of atherosclerosis by apolipoprotein E (apoE) is generally attributed to the removal of plasma lipoprotein remnant particles. We developed transgenic apoE-knockout mice expressing apoE specifically in the adrenal gland and found that only 3% of the wild-type plasma level of apoE was sufficient to normalize plasma cholesterol levels in the apoE-deficient mouse. As expected, mice expressing apoE at levels that correct hypercholesterolemia had almost no cholesteryl ester deposition in their aortas. In contrast, their nontransgenic siblings had significant atherosclerosis. Unexpectedly, we found that atherosclerosis was also reduced in 2 transgenic lines expressing too little apoE (Ͻ1% to 2% of wild type) to correct their hypercholesterolemia. Gel exclusion chromatographic profiles of plasma lipoproteins and the size distributions of lipoproteins with density Ͻ1.063 (low density and very low density lipoproteins), as determined by dynamic laser light scattering, were the same in mice expressing Ͻ2 g/mL plasma apoE and their nontransgenic littermates. We conclude that the antiatherogenic action of low levels of plasma apoE is not due to the clearance of remnant lipoproteins. Thus, low levels of apoE provided systemically, but not made in the liver or in macrophages, can block atherogenesis in the vascular wall independently of normalizing the plasma concentration of atherogenic remnant lipoprotein particles. (Arterioscler Thromb Vasc

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Long-term insulin treatment selectively stimulates secretion of the truncated form of apolipoprotein B (apoB), apoB-48, from primary rat hepatocytes in culture. Chronic treatment with insulin at 400 ng/ml causes a 3-fold increase in total apoB secretion, with apoB-48 making up about 75% of that increase. apoB-48 is the protein product generated by translation of full-length apoB mRNA which has been modified by a posttranscriptional editing mechanism. Editing changes codon 2153 in the middle ofthe apoB-100 coding region from CAA, coding for glutamine, to UAA, a translation stop signal. We therefore examined the effect ofinsulin treatment on the ratio of edited to nonedited apoB mRNA in RNA isolated from primary rat hepatocyte cultures. There was a dramatic shift in the ratio of edited versus nonedited forms of apoB mRNA, from about 1:1 in untreated cells to 7:1 in insulintreated cells. Insulin exerted a dose-dependent effect on apoB secretion and apoB mRNA editing over the range of insulin concentrations studied (0.4-400 ng/ml). In contrast, oleic acid, which also increased apoB (B-48 and B-100) secretion, had no significant effect on the ratio of apoB-48 to apoB-100 particles secreted and no effect on the proportion of edited apoB mRNA. Neither insulin nor oleic acid affects total apoB mRNA levels as assayed by Northern blot analysis. These data strongly suggest that insulin stimulates biosynthesis and secretion of apoB-48 in rat hepatocytes by regulating the proportion of edited apoB mRNA.Apolipoprotein B (apoB) is an essential structural component of triglyceride-rich very low density lipoproteins (VLDLs) and chylomicrons. The VLDLs are the metabolic precursors of the low density lipoprotein (LDL), the main carrier of cholesterol in humans. apoB is one of the ligands involved in clearance of LDL through binding to the LDL receptor; thus apoB is central to cholesterol homeostasis in the circulation. apoB is also essential for triglyceride (TG) transport to peripheral tissues (1).There are two forms of apoB, which are coded for by a single gene. The full-length translation product is apoB-100; it is a 512,000-Da polypeptide synthesized in the liver and secreted as an obligatory component of VLDL. The shorter form, apoB-48, is a 250,000-Da polypeptide identical to the amino-terminal 48% of apoB-100 and synthesized in the intestine as an integral part of the chylomicrons. In the rat, apoB-48 is also synthesized in the liver and secreted in VLDL. The mechanism for generating these two forms of apoB is a post-transcriptional RNA modification in which the CAA codon for glutamine-2153 in apoB-100 is changed to UAA, resulting in an in-frame translation stop signal (2). The polypeptide translation product of this edited RNA is apoB-48.As part of studies to define the role of insulin in the pathogenesis of dyslipidemia in the hyperinsulinemic state, we examined the effects oflong-term (5-day) insulin exposure on apoB secretion from primary cultures of rat hepatocytes. Chronic exposure to insulin selectively stimu...

Our previous studies have indicated that lecithin-cholesterol acyltransferase (LCAT) contributes significantly to the apoB lipoprotein cholesteryl ester (CE) pool. Cholesterol esterification rate (CER) in apoA-I(-)(/)(-) apoE(-)(/)(-) mouse plasma was <7% that of C57Bl/6 (B6) mouse plasma, even though apoA-I(-)(/)(-) apoE(-)(/)(-) plasma retained (1)/(3) the amount of B6 LCAT activity. This suggested that lack of LCAT enzyme did not explain the low CER in apoA-I(-)(/)(-) apoE(-)(/)(-) mice and indicated that apoE and apoA-I are the only major activators of LCAT in mouse plasma. Deleting apoE on low-density lipoprotein (LDL) reduced CER (1% free cholesterol (FC) esterified/h) compared to B6 (6% FC esterified/h) and apoA-I(-)(/)(-) (11% FC esterified/h) LDL. Similar sized LDL particles from all four genotypes were isolated by fast protein liquid chromatography (FPLC) after radiolabeling with [(3)H]-free cholesterol (FC). LDLs (1 microg FC) from each genotype were incubated with purified recombinant mouse LCAT; LDL particles from B6 and apoA-I(-)(/)(-) plasma were much better substrates for CE formation (5.7% and 6.3% CE formed/30 min, respectively) than those from apoE(-)(/)(-) and apoE(-)(/)(-) apoA-I(-)(/)(-) plasma (1.2% and 1.1% CE formed/30 min). Western blot analysis showed that the amount of apoA-I on apoE(-)(/)(-) LDLs was higher compared to B6 LDL. Adding apoE to incubations of apoA-I(-)(/)(-) apoE(-)(/)(-) very low density lipoprotein (VLDL) resulted in a 3-fold increase in LCAT CER, whereas addition of apoA-I resulted in a more modest 80% increase. We conclude that apoE is a more significant activator of LCAT than apoA-I on mouse apoB lipoproteins.