Overexpression of Human Lecithin Cholesterol Acyltransferase Leads to Hyperalphalipoproteinemia in Transgenic Mice

Abstract

Lecithin cholesterol acyltransferase (LCAT) is a key enzyme which catalyzes the esterification of free cholesterol present in plasma lipoproteins. In order to evaluate the role of LCAT in HDL metabolism, a 6.2-kilobase (kb) fragment consisting of 0.851 and 1.134 kb of the 5'- and 3'-flanking regions, as well as the entire human LCAT gene, was utilized to develop transgenic mice. Three different transgenic mouse lines overexpressing human LCAT at plasma levels 11-, 14-, and 109-fold higher than non-transgenic mice were established. Northern blot hybridization analysis demonstrated that the injected 6.2-kb fragment contained the necessary DNA sequences to direct tissue specific expression of the human LCAT gene in mouse liver. Compared to age- and sex-matched controls, total cholesterol and HDL cholesterol levels were increased in all 3 transgenic mice lines by 124-218 and 123-194%, respectively, while plasma triglyceride concentrations remained similar to that of control animals. Fast protein liquid chromatography analysis of transgenic mouse plasma revealed marked increases in high density liposportin (HDL)-cholesteryl ester and phospholipid as well as the formation of larger size HDL. Thus, the majority of the increase in transgenic plasma cholesterol concentrations was due to accumulation of cholesteryl ester in HDL consistent with enhanced esterification of free cholesterol in mouse HDL by human LCAT. Plasma concentrations of apoA-I, apoA-II, and apoE were increased in high expressor homozygote mice who also demonstrated an accumulation of an apoE-rich HDL1. Like the mouse enzyme, human LCAT was found to be primarily associated with mouse HDL. Our studies demonstrate a high correlation between plasma LCAT activity and total as well as HDL cholesterol levels establishing that in mice LCAT modulates plasma HDL concentrations. Overexpression of LCAT in mice leads to HDL elevation as well as increased heterogeneity of the HDL lipoprotein particles, indicating that high levels of plasma LCAT activity may be associated with hyperalphalipoproteinemia and enhanced reverse cholesterol transport.

Antioxidative Activity of HDL Particle Subspecies Is Impaired in Hyperalphalipoproteinemia: Relevance of Enzymatic and Physicochemical Properties

“…24 By contrast, decreased LCAT and PAF-AH activities in distinct HDL subfractions in HALP appear to be related to their redistribution between plasma lipoproteins, consistent with earlier data. 25,26 Specific antioxidative activity, PON, PAF-AH, and LCAT activities, and oxidative resistance of HDL subfractions were highest in the densest HDL3c subfraction and tended to decrease with decrement in HDL density. HDL-associated enzymes therefore may not only protect LDL but also protect PON/PAF-AH/LCAT-transporting HDL particles themselves against oxidation.…”

Abnormal lipoprotein metabolism and reversible female infertility in HDL receptor (SR-BI)–deficient mice

“…Consistent with this possibility are earlier studies that showed that (a) other apolipoproteins found in HDL (e.g., apoA-II) can serve as ligands for SR-BI (28); and (b) adenovirus-mediated overexpression of SR-BI in apoA-I KO mice results in the loss of virtually all of the apoA-I-deficient HDL cholesterol (K. Kozarsky and M. Donahee, personal communication). There was a reduction in the amount of HDL-like cholesterol in the SR-BI/apoA-I double -KO relative to SR-BI KO (Figure 1a, open circles) mice, with the greatest reduction seen in the smallest, normal-sized HDL particles (fractions [29][30][31][32][33][34][35][36][37][38]. Therefore, in the context of a SR-BI KO background, the loss of apoA-I resulted in significant changes in the structure of the HDL (e.g., no apoA-I, altered size distribution) and the amount of HDL cholesterol.…”

The ATP binding cassette transporter A1 (ABCA1) modulates the development of aortic atherosclerosis in C57BL/6 and apoE-knockout mice

“…After a 4-h fast, blood was obtained from the retro-orbital plexus of mice anesthetized with methoxyflurane (PitmanMoore, Mundelein, IL), placed into precooled tubes containing EDTA (final concentration 4 mM), and centrifuged at 2500 ϫ g for 20 min at 4°C, and aliquots of plasma were stored at Ϫ70°C. Total cholesterol (TC), triglycerides (TG), phospholipids (PL), and free cholesterol (FC) were assayed as previously described (32). HDL-cholesterol was determined as the cholesterol remaining in the plasma after precipitation of apoB-containing lipoproteins with dextran sulfate (Ciba-Corning, Oberlin, OH).…”