Serum activity and hepatic secretion of 1ecithin:cholesterol acyltransferase in experimental hypothyroidism and hypercholesterolemia

Lecithin:cholesterol acyltransferase (LCAT), the major cholesterol esterifying enzyme in plasma, plays an important role in the removal of cholesterol from peripheral tissues. This study in rat focuses upon the effects of hypothyroidism and cholesterol feeding on serum activity and hepatic LCAT secretion. To obviate the effect that inclusion of high concentrations of cholesterol in the rat serum may have on the proteoliposome used in the assay of LCAT, very low and low density lipoproteins (VLDL and LDL) were removed by ultracentrifugation at d 1.063 g/ml. The molar esterification rate in the euthyroid VLDL + LDL-free serum was found to be 0.94 f 0.06 compared to 0.67 f 0.05 in hypothyroid rats and 1.56 0.14 in hypercholesterolemic rats. LCAT secretion by suspension cultures of hepatocytes from hypercholesterolemic rats was found to be significantly depressed when compared to that for euthyroid and hypothyroid animals. Secretion by hepatocytes from hypothyroid rats was depressed for the first 0-4 hr, but rapidly recovered. The depressed 'secretion of LCAT by hepatocytes from hypercholesterolemic rats correlates with the appearance in the media of apoE-rich, discoidal HDL. Discoidal HDL was six times more effective as a substrate for purified human LCAT than HDL from hypercholesterolemic serum, and twice as effective as serum and nascent HDL from euthyroid animals. I It is concluded that the depressed LCAT activity in serum from hypothyroid rats is due to a depressed hepatic secretion of the enzyme and that the elevated serum activity of hypercholesterolemic rats may be related to a defect in LCAT clearance. Finally, the appearance of discoidal HDL in the medium upon culture of hepatocytes from hypercholesterolemic rats appears to be due to an inhibition of LCAT secretion by these cells. Ridgway, N. D., and P. J. Dolphin. Serum activity and hepatic secretion of 1ecithin:cholesterol acyltransferase in experimental hypothyroidism and hypercholesterolemia. J. Lipid Res. 1985. 26: 13001313. Supplementary key words nascent discoidal HDL rat hepatocytes LCAT assay The accumulation of apoproteins B and E can be linked to the appearance of two abnormal lipoproteins; betamigrating VLDL and HDL, (1-3). In addition, hypercholesterolemia also produces fluctuations in the activity of enzymes involved in lipoprotein metabolism in both animal models and clinical situations. The influence of the hypercholesterolemic state on lecithin:cholesterol acyltransferase (LCAT) is of particular interest due to its proposed role in reverse cholesterol transport (4). LCAT activity in the plasma of hypercholesterolemic humans and experimental animals has, in most cases, been shown to be slightly depressed or unchanged (5-8) and the mass of LCAT in the plasma of hypercholesterolemic patients has been reported to be slightly elevated (9). The most notable exception is an observation that the molar esterification rate (MER) is increased in the VLDL + LDL-depleted plasma from cholesterol-fed rabbits (10). Problems exist, however, in the interpretation of LCAT activities assayed by either the endogenous method of Stokke and Norum (11) or the common substrate method of Glomset and Wright (12), particularly in disease states when both enzyme activity and substrate availability may be altered. Rats fed cholesterol require simultaneous induction of hypothyroidism to fully manifest hypercholesterolemia. The reasons seem to lie in reduced clearance of LDL (13) and enhanced intestinal absorption of cholesterol (14). Hypothyroidism and cholesterol feeding have some divergent effects. Most notable of these is the lack of secretion of a cholesteryl ester-rich beta-migrating VLDL by hepatocytes from hypothyroid (HT) rats. Such a lipoproInduction of hypercholesterolemia in the rat by adminAbbreviations: LCAT, 1ecithin:cholesterol acyltransferase; LDL, low istration Of a diet containing 5.0% density lipoprotein; VLDL, very low density lipoprotein; PTU, propylmented with 0.3% of the antithyroid agent propylthiouracil; HT, hypothyroid; HDL, high density lipoprotein; MER, 'Present address: Department of Biochemistry, The University of (mu) to enhance the ~ y p e r c ~ o ~ e s t e r o ~ e m ~ c molar esterification rate; HC, hypercholesterolemia; ET, euthyroid. effect, results in the accumulation of apoB, apoE, and British Columbia, Vancouver, B.C., Canada V6T 1W5. apoA-I and both polar and nonpolar lipids in the serum. *To whom all correspondence and reprint requests should be directed. 1300 Journal of Lipid Research Volume 26, 1985 by gest, on O cber 8, 2017 w w w .j.org D ow nladed fom tein seems to be a de novo synthetic product of hepatocytes from hypothyroid-hypercholesterolemic (HT-HC) rats (15). Although no data are available on the combined effects of hypothyroidism and cholesterol feeding on serum LCAT activity in experimental or clinical conditions, the singular effect of hypothyroidism in both man (16) and rat (17) is to insignificantly depress enzyme levels. It is interesting to note that thyroxine treatment in rat increases enzyme activity (18), while no apparent change is evident in hypothyroid humans. Some indirect evidence is available concerning the hepatic contribution to the serum LCAT pool in hypothyroidism and hypercholesterolemia. It has been demonstrated by us (19) that, while spherical nascent HDL accumulate in suspension cultures of HT and control rat hepatocytes, discoidal HDL reminiscent of the nascent lipoproteins described by Hamilton et al. (20) are secreted by hypercholesterolemic hepatocytes. The perfused livers of cholesterol-fed guinea pigs (21) and African green monkeys (22) also secrete a similar discoidal particle. The appearance of discoidal HDL in these instances may indicate a decreased hepatic secretion or inhibition of endogenously secreted LCAT. High cholesterol to phospholipid ratios in these nascent lipoproteins may also retard their conversion to spherical particles (23). Here we examine the effect of hypothyroidism and cholesterol feeding on serum levels and hepatic secretion of LCAT by isolated rat hepatocytes. LCAT levels were assayed using proteoliposomes described by Chen and Albers (24), but with rat apoA-I substituted for the human apoprotein. We further examined the effectiveness of nascent HDL, secreted by hypercholesterolemic rat hepatocytes, as a substrate for purified human LCAT in light of data concerning hepatic LCAT secretion and lipoprotein composition. MATERIALS AND METHODS