Functional Lecithin:Cholesterol Acyltransferase Deficiency and High Density Lipoprotein Deficiency in Transgenic Mice Overexpressing Human Apolipoprotein A-II

Abstract

The concentration of high density lipoproteins (HDL) is inversely related to the risk of atherosclerosis. The two major protein components of HDL are apolipoprotein (apo) A-I and apoA-II. To study the role of apoA-II in lipoprotein metabolism and atherosclerosis, we have developed three lines of C57BL/6 transgenic mice expressing human apoA-II (lines 25.3, 21.5, and 11.1). Northern blot experiments showed that human apoA-II mRNA was present only in the liver of transgenic mice. SDS-polyacrylamide gel electrophoresis and Western blot analysis demonstrated a 17.4-kDa human apoA-II in the HDL fraction of the plasma of transgenic mice. After 3 months on a regular chow, the plasma concentrations of human apoA-II were 21 +/- 4 mg/dl in the 25.3 line, 51 +/- 6 mg/dl in the 21.5 line, and 74 +/- 4 mg/dl in the 11.1 line. The concentration of cholesterol in plasma was significantly lower in transgenic mice than in control mice because of a decrease in HDL cholesterol that was greatest in the line that expressed the most apoA-II (23 mg/dl in the 11.1 line versus 63 mg/dl in control mice). There was also a reduction in the plasma concentration of mouse apoA-I (32 +/- 2, 56 +/- 9, 91 +/- 7, and 111 +/- 2 mg/dl for lines 11.1, 21.5, 25.3, and control mice, respectively) that was inversely correlated with the amount of human apoA-II expressed. Additional changes in plasma lipid/lipoprotein profile noted in line 11.1 that expressed the highest level of human apoA-II include elevated triglyceride, increased proportion of total plasma, and HDL free cholesterol and a marked (>10-fold) reduction in mouse apoA-II. Total endogenous plasma lecithin:cholesterol acyltransferase (LCAT) activity was reduced to a level directly correlated with the degree of increased plasma human apoA-II in the transgenic lines. LCAT activity toward exogenous substrate was, however, only slightly decreased. The biochemical changes in the 11.1 line, which is markedly deficient in plasma apoA-I, an activator for LCAT, are reminiscent of those in patients with partial LCAT deficiency. Feeding the transgenic mice a high fat, high cholesterol diet maintained the mouse apoA-I concentration at a normal level (69 +/- 14 mg/dl in line 11.1 compared with 71 +/- 6 mg/dl in nontransgenic controls) and prevented the appearance of HDL deficiency. All this happened in the presence of a persistently high plasma human apoA-II (96 +/- 14 mg/dl). Paradoxical HDL elevation by high fat diets has been observed in humans and is reproduced in human apoA-II overexpressing transgenic mice but not in control mice. Finally, HDL size and morphology varied substantially in the three transgenic lines, indicating the importance of apoA-II concentration in the modulation of HDL formation. The LCAT and HDL deficiencies observed in this study indicate that apoA-II plays a dynamic role in the regulation of plasma HDL metabolism.

Overexpression of Human Apolipoprotein A-II in Mice Induces Hypertriglyceridemia Due to Defective Very Low Density Lipoprotein Hydrolysis

“…1 illustrates Northern blot analysis of mRNA prepared from several tissues. Expression of human apoA-II was restricted to liver, as expected (10,12). Despite similar relative amounts of human apoA-II mRNA to 18 S (2.6 and 1.9 in lines ␦ and , respectively), the human apoA-II concentration in plasma was 56 and 116 mg/dl in mice ␦ and , respectively, suggesting that the higher plasma human apoA-II in line resulted from post-transcriptional regulation or decreased clearance.…”

“…Indeed, nondenaturing gradient gel electrophoresis showed that HDL of our transgenic mice comprised two populations, both smaller than control mouse HDL (Fig. 3A), as reported for other human apoA-II-expressing transgenic mouse lines (10,12). Mouse apoA-I accounted for 80% of total apolipoproteins in control HDL and drastically decreased down to 12 and 1.5% in HDL from ␦ and mice, respectively (Fig.…”

“…A careful survey of the literature shows slight increases in plasma TG proportionately to apoA-II expression in other transgenic lines that also exhibited increased VLDL and LDL levels, even after an overnight fast (12,15). However, apoA-II has never been detected in the VLDL fraction of these transgenic mice.…”

“…When human apoA-II was expressed at normal levels, overall lipoprotein metabolism was not markedly modified, except for the appearance of a smaller HDL population containing solely human apoA-II (10). At greater expression levels of human (12) or murine (15) apoA-II, plasma HDL was increased in mice overexpressing murine apoA-II (14, 15) but decreased in mice overexpressing human apoA-II (12,16). Surprisingly, mouse apoA-II overexpression resulted in increased atherosclerotic lesions under standard chow (14), whereas human apoA-II overexpression increased fatty streaks only after long term feeding with an atherogenic diet (13).…”

“…Surprisingly, mouse apoA-II overexpression resulted in increased atherosclerotic lesions under standard chow (14), whereas human apoA-II overexpression increased fatty streaks only after long term feeding with an atherogenic diet (13). Two of the above studies reported mildly increased plasma triglyceride (TG), which was associated with the VLDL fraction when mice expressed high levels of human (12) or murine (15) apoA-II. However, comparisons among the various studies are rendered difficult by the different nutritional status of transgenic mice, namely whether they were fasted overnight (10 -15) or fed ad libitum (16).…”