PLASMA LIPOPROTEIN PHENOTYPE IN RESPONSE TO CHOLESTERYL ESTER TRANSFER PROTEIN LEVELS IN DYSLIPOPROTEINEMIA

PLASMA LIPOPROTEIN PHENOTYPE IN RESPONSE TO CHOLESTERYL ESTER TRANSFER PROTEIN LEVELS IN DYSLIPOPROTEINEMIA

There are two mamallian systems for reverse cholesterol transport, one of which is dependent on the presence of cholesteryl ester transfer protein (CETP) activity 1 • 2 . The rat is typical of species lacking CETP activity and under these conditions, triglyceride-rich lipoproteins and their remnants transport only that cholesterol which was initially associated with the nascent particles. Rat HDL on the other hand, is rich in apoE and transports cholesterol derived from both splanchnic and peripheral organs, and esterified by LCAT and eventually returns it to the liver, via an apoE receptor. In contrast in the human system, rhe high CETP activity promotes the transfer of 2/3 or more of cholesteryl esters in HDL to the triglyceride-rich lipoproteins and their remnants. These recipient :farticles are then actively removed by hepatic apoE and apoB/E receptors • In individuals with active receptors, this represents the major pathway for reverse cholesterol transport while a minority of HDL cholesterol is cleared directly by an apoE-mediated pathway. Direct selective uptake of HDL cholesteryl esters by the liver has also been suggesteg, but recent evidence indicated that this route is also apoE­ mediated •

e do not know which of the two above pathways is the most efficient in returning cholesterol to the liver and most protective against atherosclerosis, it is thus of interest to consider the different lipoprotein phenotypes that occur under conditions of either low or high CETP activity. In genetic CETP deficiency, the classical phenotype is that of yperalphalipoproteinemia with accumulation of large HDL particles 5 ' • A poE levels have been found to be increased fro about 1.5 to 6 fold above normal in different CETP deficient patients • The lipoprotein profile observed is thus similar to that of the rat. The accumulation of choles teryl esters in HDL suggests that in absence of CETP, maximal transport of cholesteryl esters cannot be maintained. However this apparently inefficient pathway represents a safe form for intravascular storage of cholesterol since HDL cholesterol is not directly taken up by any peripheral receptor. This pathway is apparently of adequate capacity in species with low LDL transport rates.

In contrast, in situations where CETP activity is high, more cholesteryl esters are efficiently transfered to triglyceride-rich lipoproteins and their remnants, which provided that these particles contain sufficient apoE3 or apoE4 are actively cleared by hepatic apoE receptors. Thus the reverse cholesterol transport pathway operating with high CETP activity is also dependent upon apoE for clearance. When the available apoE is not an effective ligand for its receptor, accumulation of -VLDL results in the expression of the type III phenotype.

However no mutation causing an increase in CETP levels and activity has been reported and therefore n·o example of the lipoprotein profile that would prevail under such conditions is available. Increased lipid transfer activity has been measured in certain human dyslipoproteinemia8, notably in familial hypercholesterolemia and in dysbetalipoproteinemia, although decreased lipid transfer activity has also been reported9. Some of these apparent contradictions reflect the composite CETP activity that is measured in plasma. The assay of CETP ac tivi ty is influenced by several factors, notably by the composition and ratio of donor and acceptor lipoproteins and by the presence of putative inhibitors of CETP. It is therefore important to measure the plasma levels of CETP in different types of dyslipoproteinemia where abnormal lipoproteins accumulate for a better understanding of the role of lipid transfer in the development of these lipoprotein phenotypes. Using a radioimmunoassay recently developpediO, we report here the preliminary data on plasma CETP levels in hyperlipoproteinemia.

RESULTS AND DISCUSSION

We have studied hyperlipoproteinemic patients recruted from the Lipid Research Clinic of the Royal Victoria Hospital. This included 17 patients with familial hypercholesterolemia, 18 with familial combined hyperlipoproteinemia, 10 with dysbetalipoproteinemia (type III), 12 with hypertriglyceridemia (typeiV), and 5 with chylomicronemia (type V). The diagnosis was made on the basis of appropriate family history, clinical and biochemical data. Total cholesterol, triglycerides, HDL cholesterol were measured by standard techniques11 and the levels of apoA-I, A-II, B, and E by radioimmunoassays as described earlier10, The control population (79 subjects) was taken from the previously reported groups of normolipemic subjects which were studied under the same conditions and in whom plasma levels of CETP had also been measured10, Plasma levels of CETP were significantly increased in patients with hypercholesterolemia (+26%'),and familial combined hyperlipidemia (+25%'). However CETP levels were unchanged in the patients with moderate hypertriglyceridemia without chylomicronemia (type IV).The highest levels of CETP were observed in patients with type III hyperlipoprteinemia (+68%') and in patients with severe hyperchylomicronemia (+85%'). The present results corroborate the earlier observations of Tall et alB, who showed that CETP activity was increased in type III hyperlipoproteinemia. In hypercholesterolemia as in familial combined hyperlipidemia, we speculate that the modest increase in CETP may be related to increased delivery of cholesterol to the periphery which stimulates the synthesis of CETP by macrophages 12. The higher levels of CETP which are observed in type V and type III may be derived from additional macrophagic intestinal13 and hepatic14,15 synthesis in response to the increased chylomicron and chylomicron remnant levels. Alternatively, assuming that CETP is cleared with these lipoproteins, the high level of CETP seen in types III and V may be related to the decreased catabolism of chylomicrons and their remnants. However the later hypothesis seems improbable since no CETP was found associated with chylomicrons in the plasma of fat-fed subjects10, Calculation of Pearson correlation coefficients between CETP levels and lipoprotein parameters for the total group of hyperlipoproteinemic patients demonstrated the existence of significant relationships between CETP and total cholesterol (r=0.52), CETP and VLDL cholesterol (r=0.63), CETP and triglycerides (r=0.53), and CETP and apoE (r=0.40). The correlation with triglycerides was only observed when hyperchylomicronemic subjects were included. In contrast to earlier observations in normolipemic subjects, CETP levels in hyperlipidemic patients were not correlated with apoA-I. The present observations confirm previous evidence that CETP levels vary in relation with cholesterol. In the cholesterol fed rabbit, the levels of hepatic mRNA are increased together with the plasma levels of CETP16. In response to the diet, apoE plasma levels are also increased16 but hepatic mRNA levels are unchanged 1t, de,nons t rating that although the genes for these two proteins are regulated by dietary cholesterol, there are important differences in dssue specificity for the control of their expression. This is also consistant with observations in human that both CETP and apoE are increased with probucol treatment18 and with dietary cholesterol (McPherson et al, unpublished results).

In conclusion CETP levels are increased in conditions where cholesterol transport is increased, perhaps especially when peripheral delivery of cholesterol is increased. There is also evidence that CETP and apoE are coregulated by cholesterol although with different tissue specificities. Finally, elevated CETP is associated with a lipoprotein phenotype of decreased HDL cholesterol and increased cholesteryl esters and apoE in chylomicrons and in their remnants. This suggests that increase CETP activity may be essential for the expression of type III hyperlipoproteinemia in E2 homozygosity.

REFERENCES

1. A.R. Tall, Plasma lipid transfer proteins, J. Lipid Res. 27:361 (1986).

2. Y.C. Ha, and P.J. Barter, Differences in plasma cholesteryl ester transfer activity in sixteen vertebrate species, Comp. Biochem. Physiol. 71B:265 (1982).

3. R.J. Havel, Functional activities of hepatic lipoprotein receptors, Ann. Rev. Physiol. 48:119 (1986).

4. F. Rinniger, and R.C. Pitman, Mechanism of cholesteryl ester transfer protein-mediated uptake of high density lipoprotein cholesteryl esters by HepG2 cells, J. Biol. Chern. 264:6118 (1989).

5. J. Koizumi, H. Mabuchi, A. Yoshimura, Itoh, Y. Sakai, T. Sakai, K. Ueda, serum cholesteryl ester transfer familial hyperalphalipoproteinemia.

I. Michishita, and R. Takeda, ac tivi ty in M. Takeda, H. Deficiency of patients with

6. S. Yamashita, Y. Matsuzawa, M. Okazaki, H. Kako, T. Yasugi, H. Akioka, K, Hircho, and S. Tarui, human apolipoprotein E-rich high density lipoproteins (HDLc) in subjects with deficiency of cholesteryl ester transfer , Atherosclerosis 70:7 (1988).

7. R. \l. Mahley, Apolipoprotein E: cholesterol transport protein with expanding role in cell biology, Science 240:622 (1988).

8. A.R. Tall, E. Granot, R. Broda, J. Tabas, C. Hessler, K. Williams, and M. Denke, Accelerated transfer of cholesteryl esters in dyslipidemic plasma, J. Clin. Invest. 79:1217 (1987).

9. P.E Fielding, C.J. Fielding, R.J. Havel, J. P. Kane, and P. Tun, Cholesterol net transport, esterification and transfer in human hyperlipidemic plasma, J. Clin. Invest. 71:449 (1983).

10. Y.L. Marcel, R. McPherson, M. Hogue, H. Czarnecka, z. Zawadski, P.K. eech, M.E. hitlock, A.R. Tall, and R. . Milne, Distribution and concentration of choles teryl ester transfer protein in plasma of normolipemic subjects, J. Clin. Invest. 85:10 (1990).

11. Lipid Research Clinics Program, l!!.. "Manual of Laboratory operations".

U.S. Government printing office, Bethesda Md, p. 1-81 (1974).

12. J.H. Tollefson, R. Faust, J.J. Albers, and A. Chait, Secretion of a lipid transfer proteinn by human moncyte-deri ved macro phages, ..::L_ Biol. Chern. 260:5887 (1985).

13. R.A. Faust, and J.J. Albers, Regulated vectorial secretion of cholesteryl ester transfer proteinn (LTP-I) by the Calo2 model of human enterocyte epithelium, J. Biol. Chern. 263:8786 (1988).

14. T.L. Swenson, J.S. Simmons, C.B. Hesler, C. Bisgaier, and A.R. Tall, Cholesteryl ester transfer protein is secreted by HepG2 cells and contains asparagine-linked carbohydrates and sialic acid, J. Biol. Chern. 262:16211 (1987).

15. R.A. Faust, and J.J. Albers, Synthesis and secretion of plasma choles teryl ester trans fer protein by human hepa tocarcinoma cell line, HepG2, Arteriosclerosis 7:267 (1987).

16. E.M. Quinet, L.B. Agellon, P.A., Kroon, Y.L. Marcel, Y.C. Lee, M.E. hi tlock, and A.R. Tall, Atherogenic diet increased cholesteryl ester transfer protein mRNA levels in rabbit liver, J. Clin. Invest. 85:357 (19??).

17. Y.S. Chao, T.T. Yamin, G.M. Thompson, and P.A. Kroon, Tissue-specific expression of genes encoding apolipoprotein E in rabbits. J. Biol. Chern. 259:5306 (1984).

18. R. McPherson, M. Hogue, R. . Milne, A.R. Tall, and Y.L. Marcel, Increase in plasma cholesteryl ester transfer protein during Probucol treatment: relationship to changes in HDL compostion, submitted for publication.