Release of Stored Retinol from Adipocytes

Adipose tissue is second only to liver as a retinol depot. Retinol is stored there in part as retinyl esters. In cultured mouse adipocytes, a hormone-sensitive lipase hydrolyzes these esters upon CAMP stimulation, with free retinol being released into the medium.

It is now standard textbook knowledge that vitamin A (retinol) is stored in the liver as retinyl esters and released into the circulation combined with retinol-binding protein (RBP).’ It is not generally known, however, that adipose tissue, already acknowledged as the storage site for I3-carotene, is also an important storage tissue for retinol. About 15-20% of the rat’s store of retinol is in adipocytes; with especially high concentrations in epididymal adipocytes.2 About 60% of this retinol exists in the form of retinyl ester^.^

Recent research from Blaner’s laboratory4 has dem- onstrated that the mechanism of retinol release from adipocytes differs from that from liver cells. Moreover, it was found that the mechanism of uptake of retinol into adipose tissue also differed from uptake into other tis- sues: it appears that adipocytes acquire their retinol not from circulating RBP-retinol but from chylomicron-bound retinyl esters.s

As a starting point, the Blaner group4 tested the hy- pothesis that the release of retinol, stored in adipocytes mostly as retinyl esters, would resemble the release of fatty acids, stored in adipocytes as triglycerides. Fatty acid release from adipocytes is regulated by intracellular CAMP.^

The authors4 first loaded adipocytes with retinol. Mouse BFC- 1 I3 pre-adipocytes were made to differentiate into adipocytes by incubation for 27 days in a medium containing insulin and tri-iodothyronine. On day 22,l pM retinol was added to the medium. After 5 days, the cells contained 425 ng of a mixture of retinol and retinyl esters per lo6 cells, a concentration of the same order of magni- tude as that in freshly isolated rat epididymal fat pads.* The preloaded cells were then exposed to media with or without 1 mM dibutyryl CAMP (dbcAMP) for various times. The cells were found to release significantly more retinol into the media with than without dbcAMP (400 ng/ plate without dbcAMP; 600 ng/plate with dbcAMP), to the extent of 45% of the retinol/retinyl esters present in

This review was prepared by George Wolf, D.Phil., Department of Nutritional Sciences, University of California, Berkeley, CA 94720-31 04. Reprint requests are to be addressed to Nutrition Reviews, 711 Washington Street, Boston, MA 02111, USA.

the cells. Retinyl esters were not released, but the level of the esters declined severely within the cells, in accord with a rise in the level of intracellular retinol. The release of retinol into the media, as well as conversion of intracel- lular retinyl esters to intracellular retinol, was not only time dependent but also dependent on media dbcAMP concentrations between 0.1 and 5.0 mM. Of the three retinyl esters present, most of retinyl linoleate and palmitate were hydrolyzed, whereas retinyl oleate remained largely un- changed. Interestingly, in contrast to adipocytes, primary mouse hepatocytes exposed to dbcAMP showed no in- crease in intracellular or released retinol.

From the experiments described above, the authors concluded that the first step in retinol mobilization from adipocytes must be the hydrolysis of the stored retinyl esters to retinol. Because dbcAMP stimulated the puta- tive hydrolysis, the authors hypothesize that hor- mone-sensitive lipase (HSL) might be the likely enzyme effecting this hydrolysis. HSL is a lipase with broad sub- strate specificity, hydrolyzing triglycerides, and choles- terol esters. It is known6 that hormonal stimulation of HSL is mediated by CAMP. In fact, it was found4 that HSL, partially purified from adipocytes, was able to hydrolyze retinyl esters to retinol. This is in contrast to the situation in liver, where a specific retinyl ester hydrolase performs this function.

To elucidate this property of adipocytes, the Blaner group4 transfected Chinese hamster ovary cells with a cDNA for rat HSL. The resulting cells expressed high lev- els of HSL mRNA and were highly immunoreactive to antirat HSL antibody. Upon homogenization, they showed considerable retinyl ester hydrolase activity, which was time, protein, and substrate dependent. This hydrolytic activity, though, was only about one-tenth the activity of triglyceride (triolein) hydrolysis with the same homoge- nate.

In liver, retinol is released in combination with RBP. Although adipocytes actively synthesize and secrete RBP; the amount secreted by the BFC-1 p adipocytes was found to be small (64k24 ng RBP/106 celldday) and was not increased by incubation with dbcAMP. From the au- thors’ data: it can be calculated that the amount of retinol released by the adipocytes unstimulated by dbcAMP was about 1 00-fold that of the RBP released and about 300- fold that of RBP released upon dbcAMP stimulation. There- fore, adipocytes, in contrast to liver cells, secrete free ret- inol, at least in cell culture. Bovine fetal serum, which is a component of the media in which the adipocytes were incubated, contains RBP. When retinol-loaded adipocytes were incubated in serum-free media, they released no ret- inol, even in the presence of dbcAMP. The authors4 hy-

Nutrition Reviews, Vol. 56, No. I 29

pothesized that the release of retinol may depend on the relative concentration of extracellular RBP, which “draws the newly formed retinol into the media.”

In summary, the CAMP-mediated mobilization of ret- inol from adipose tissue is unlike that from liver and re- sembles the release mechanism of fatty acids from triglyc- eride depots. It is preceded by the hydrolysis of stored retinyl esters by a CAMP-sensitive hormone-dependent lipase. It is possible that different retinol depots respond differently to different hormones.

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Combs GC Jr. The vitamins. New York: Academic Press Inc, 1992; 127-9 Tsutsumi C, Okuno M, Tannous L, et al. Retinoids

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3. Zovich DC, Orologa A, Okuno M, et al. Differentiation-dependent expression of retinoid-binding proteins in BFC-1 R adipocytes. J Biol Chem 1992;267:13884-9 Wei S, Lai K, Patel S, et al. Retinyl ester hydrolysis and retinol efflux from BFC-113 adipocytes. J Biol Chem 1997;272:1459-65 Blaner WS, Obunike JC, Kurlandsky SB, et al. Lipo- protein lipase hydrolysis of retinyl ester. J Biol Chem

Linsheer WG, Vergroesen AJ. Lipids. In: Shils ME, Olson JA, Shike M, eds. Modern nutrition in health and disease, 8th ed. Malvern, PA: Lea and Febiger,

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