METABOLIC ROLE OF HUMAN APOPROTEIN A-IV

METABOLIC ROLE OF HUMAN APOPROTEIN A-IV

INTRODUCTION

Compared with the other human apoproteins, physiological variations and metabolic functions of apo A-IV remain obscure. The apo A-IV has been discovered in rat HDL and has been later found in human plasma, mesenteric lymph and interstitial fluid. Apo A-IV is synthetized by intestine and liver and is catabolized by liver and kidneys. As apo A-IV has been suspected to play a role in triglyceride transport, we undertook the study of its plasma concentration in a population of subjects with various plasma triglyceride levels. Moreover, recent studies brought some evidence for an implication of apo A-IV in the reverse cholesterol transport. Because recent reports suggested that HDL was the major lipoprotein class involved in this metabolic pathway, we investigated the structural and metabolic relation between apo A-IV and HDL.

RQL[ OF APOPROT[IN A-IV IN TRIGLYC[RID[ TRANSPORT

By using an apo A-IV competitive enzyme immunoassay we found a significant positive correlation between apo A-IV and triglyceride concentrations in human sera (1) <Figure1). These data were in good agreement with previous reports suggesting a relation between apo A-IV and triglyceride secretion. An increase of plasma apo A-IV concentration has been found after lipid feeding (2,3) and in circumstances associated with accumulation of remnants of triglyceride-rich lipoproteins, such as chronic renal failure treated by peritoneal dialysis or hemodialysis (4,5). Moreover, a relation between the apo A-IV mRNA synthesis and the triglyceride secretion has been observed in intestine and liver (6,7).

As this correlation between apo A-IV triglycerides was found in non­ chylomicronemic sera from fasting subjects, we can assume that triglycerides were contained mainly in VLDL from hepatic origin and that the liver, in fasted subjects, could'contribute significantly to the plasma apo A-IV pool.

ROLE OF APOPROTEIN A-IV IN REVERSE CHOLESTEROL TRANSPORT

Distribution of apoprotein A-IV in normolipidemic human serum

As apo A-IV has been suspected to play a role in reverse cholesterol transport. its structural relation with HDL was studied. Whereas it is known that in rat most of apo A-IV associates with HDL, in human very little apo A-IV has been found on lipoprotein particles isolated by traditional ultracentrifugation methods. However. the distribution of apo A-IV between lipoprotein free and lipoprotein fractions varies widely and appears to be dependent on the techniques used to fractionate sera. About 20 to 35 Y. of apo A-IV were found associated with HDL after agarose gel permeation chromatography of total plasma (2,3,8), while more than 90 Y.

of human apo A-IV localized in the lipoprotein-free fraction after ultra­ centrifugation of plasma at density 1.21 (2,9).

By using a high performance Superose 12-HR column (Pharmacia), we were able not only to confirm the presence of apo A IV in the HDL fraction but to demonstrate that, in fasting human sera, a majority of apo A-IV eluted with the HDL fraction, mainly within the HDL2 size range (10>. Moreover, the gel filtration procedure allowed us to evidence the potential disruptive effect of ultracentrifugation on the apo A-IV distribution in total serum <Figure 2>. This distribution of apo A-IV could be attributable to the combined effects of high ionic strength and high sheering forces of ultracentrifugation procedure.

Concurrently, the coprecipitation of the most part of apo A-IV after incubation of total serum with anti-apo A-I antibodies indicated that about 70 to 80 Y. of serum apo A-IV were carried by apo A-I containing HDL

<Figure 3). This proportion of apo A-IV coprecipitated with HDL is

considerably lowered (less than 10 %) when HDL particles were dissociated by addition of Tween to the serum dilutions prior to the precipatation step.

These results provided evidence that, in fasted normal human sera. apo A-IV preferably associated with HDL and that this apo A-IV-HDL asso­ ciation is a weak one, easily disrupted. The remodeling of lipoprotein surfaces can modify the equilibrium between free and bound apo A-IV. Particularly, LCAT activity can induce a displacement of apo A-IV from lipoprotein-free fraction to the HDL particles.

Apoprotein A-IV and HDL conversion

Beyond its structural relation with HDL, apo A-IV can also play a dynamic role in the intravascular metabolism of HDL particles, especially in the size redistribution or conversion induced by the Cholesterol [ster Transfer Protein CCKTP> (11>. By using gradient gel electrophoresis, it was shown that the incubation of total lipoproteins in the presence of CKTP at physiological levels induced a general displacement of HDL towards large size particles, a decrease in the HDL3 subpopulation and the appearance of small conversion products with mean diameters of 7.8 and 7.4 nm (Figure 4).

This phenomenon can be significantly altered by apo A-IV. Compared with incubation with CETP alone. incubation with CETP and apo A-IV increased the size redistribution of HDL and particularly favored the formation of the very small sized lipoprotein particles (mean diameter 7.4 nml. Incubation of total lipoproteins and apo A-IV, in the absence of CETP, do not modify the HDL distribution profiles.

DESCRIPTION 0 THE MULTIPOTENTIAL INVOLVEMENT 0 APO A-IV IN THE REVERSE CHOLESTEROL TRANSPORT

The results presented above, together with previously reported data, show that apo A-IV is involved at different levels of HDL metabolism and suggest that this apoprotein could play a major role in cholesterol transport from peripheral tissues to the liver. More precisely, the loca­lization of apo A-IV within HDL. its implication in the formation of very small sized lipoprotein particles, its activating role in plasma choles­terol esterification by LCAT and its potential involvement in the cholesterol movements between intra- and extra-cellular media suggest that this apoprotein could play a major role in the reverse cholesterol transport (Figure 5).

By its combined effects with C[TP, apo A-IV can promote the formation of very small sized HDL particles. Such particles are susceptible to mi­ grate easily in the interstitial space where they could participate to the cell cholesterol efflux <11). Moreover, as apo A-IV is relatively stable in aqueous solution, the free apoprotein itself could also participate directly to the peripheral cholesterol uptake. The particles issued from interstitial space, enriched with free cholesterol and phospholipids can then interact with plasma LCAT. The small HDL, enriched with unesterified cholesterol <UC> and phospholipids <PL>, are known to be good substrates

for this enzyme <12) which can be furthermore activated by apo A-IV <13, 14). These uptake and esterification of cholesterol, which constitute a key step in the Reverse Cholesterol Transport, lead to the formation of large sized HDL2-like particles enriched in esterified cholesterol. In vitro studies on cell cultures have indicated that apo A-IV could specifi­ cally interact with rat hepatocytes <15,16). As we observed that apo A-IV

in human serum localized mainly in large sized HDL particles, it can be postulated that apo A-IV could facilitate specific uptake of HDL by the liver.

In conclusion, by promoting efflux of peripheral cholesterol, by activating cholesterol esterification in plasma, and by facilitating cholesterol uptake by the liver, apo A-IV could play a central role in the Reverse Cholesterol Transport.

REFERENCES

1. L. Lagrost, P. Gambert, S. Meunier, P.Morgado, J. Degres, P. d'Athis, and C. Lallemant, Correlation between apolipoprotein A-IV and tri­ glyceride concentrations in human sera, J. Lipid Res., 30:701 <1989).

2. P. H. Green, R.P. Glickman, J.W. Riley, and [. Quinet, Human apolipoprotein A-IV : intestinal origin and distribution in plasma, J. Clin. Invest., 65:911 <1980).

3. C. L. Bisgaier, 0. P. Sachdev, I. L. Megna, and R. M. Glickman, Distri­ bution of apolipoprotein A-IV in human plasma, J. Lipid Res., 26:11 (1985).

4. P. J. Nestel, N. H. Fidge, and M. H. Tan, Increased lipoprotein-remnant formation in chronic renal failure, N. [ngl. J. Med., 307:329 <1982).

5. M. Seishira, andY. Muto, An increased apo A-IV serum concentration of patients with chronic renal failure on hemodialysis, Clin. Chim. Acta, 167:303 <1987).

6. N. A. [lshourbagy, M. S. Boguski, W. S. L. Liao, L. S. Jefferson, J. I. Gordon, and J. M. Taylor, [xpression of rat apolipoprotein A-IV and A-I genes : mRNA induction during development and in response to glucocorticoids and insulin, Proc. Natl. Acad. Sci. USA, 82:8242 (1985).

7. M. Pessah, C. Salvat, S. R. Wang, and R. Infante, In vitro synthesis of apo A-IV and apo C by liver and intestinal mRNAs from lean and obese Zucker rats. Biochem. Biophys. Res. Commun, 142:78 <1987).

8. M. Rosseneu, G. Michiels, W. De Keersgieter, J. Bury, J. P. De Slypere, H. Dieplinger, and G. Utermann, Quantification of human apolipoprotein A-IV by"sandwich"-type enzyme-linked immunosorbent assay, Clin. Chern., 34:739 <1988).

9. G. Utermann, and U. Beisiegel, Apolipoprotein A-IV : a protein occur­ ring in human mesenteric lymph chylomicron and free in plasma. Isolation and quantification, [ur. J. Biochem., 99:333 <1979).

10. L. Lagrost, P. Gambert. M. Boquillon, and C. Lallemant. Evidence for high density lipoprotein as the major apolipoprotein A-IV containing fraction in normal human serum, J. Lipid Res., 30:1525 <1989).

11. P. Gambert, L. Lagrost, A. Athias, S. Bastiras, and C. Lallemant, Role of apolipoprotein A-IV in the interconversion of HDL subclasses, Advances in Experimental Medicine and Biology, 243:263 <1988).

12. P. J. Barter, G. J. Hopkins, and L. Gorjatschko, Lipoprotein substrates for plasma cholesterol esterification. Influence of particle size and composition of the high-density lipoprotein subfraction 3, Atherosclerosis, 58:97 <1985).

13. A. Steinmetz, and G. Utermann, Activation of lecithin : cholesterol acyltransferase by human apolipoprotein A-IV, J. Biol. Chern., 260:2258 (1985).

14. C. H. Chen, and J. J. Albers, Activation of lecithin : cholesterol

acyltransferase by apolipoprotein E-2, E-3, and A-IV isolated from

plasma, Biochim. Biophys. Acta, 836:279 (1985)

15. E. Dvorin, N. L. Gorder, D. M. Benson, and A.M. Gotto, Jr., Apolipoprotein A-IV. A determinant for binding and uptake of high density lipoproteins by rat hepatocytes. J. Biol. Chern .. 261:15714 (1986).

16. Y. B. Mitchel. V. A. Rifici. and H. A. Eder. Characterization of the specific binding of rat apolipoprotein E-deficiency HDL to rat hepatic plasma membranes, Biochim. Biophys. Acta, 917:324 <1987).