EFFLUX OF NEWLY SYNTHESIZED CHOLESTEROL AND BIOSYNTHETIC STEROL INTERMEDIATES FROM CELLS - DEPENDENCE ON ACCEPTOR TYPE AND ON ENRICHMENT OFCELLS WITH CHOLESTEROL
Wj. Johnson et al., EFFLUX OF NEWLY SYNTHESIZED CHOLESTEROL AND BIOSYNTHETIC STEROL INTERMEDIATES FROM CELLS - DEPENDENCE ON ACCEPTOR TYPE AND ON ENRICHMENT OFCELLS WITH CHOLESTEROL, The Journal of biological chemistry, 270(42), 1995, pp. 25037-25046
Previous studies suggest that during sterol synthesis in cells, choles
terol and precusor sterols are transported to the plasma membrane and
that this transport is stimulated by the binding of high density lipop
rotein (HDL) to its putative cell surface receptor, leading to enhance
d sterol efflux. Little is known about the identities of synthesized s
terols subject to efflux or whether efflux of cholesterol and precurso
r sterols are stimulated equally by HDL. To address these issues, cell
s were incubated with [H-3]acetate or [H-3]mevalonate and sterol accep
ters, and then the labeled sterols in cells and efflux media were anal
yzed by high pressure liquid chromatography methods that resolved chol
esterol and precursor sterols. In non-hepatic cells (Chinese hamster o
vary (CHO), fibroblasts, and smooth muscle), cholesterol and multiple
precursor sterols accumulated. In CHO cells, the major products were c
holesterol and des-mosterol, which together constituted 50% of labeled
nonsaponifiable lipids. When media contained human HDL(3) (1 mg of pr
otein/ml), the molar efflux of synthesized desmosterol was four times
that of cholesterol, and the 8-h efflux of those sterols, each normali
zed to its own production, averaged 48 and 16%, respectively. When med
ia contained egg phosphatidylcholine vesicles (1 mg/ml), the efflux of
these sterols averaged 18 and 2.4%, respectively. Thus, with both acc
epters, desmosterol was the major synthesized sterol released from cel
ls, and its efflux was substantially greater than that of synthesized
cholesterol. High relative efflux of desmosterol (or a desmosterol-lik
e sterol) occurred in all cell types and in both cholesterol-enriched
and unenriched cells. These results demonstrated qualitatively similar
efflux of synthesized sterols in the presence of HDL(3) and phospholi
pid vesicles, arguing against an absolute requirement for accepters th
at interact with the HDL receptor. To probe for possible quantitative
differences in the capabilities of these two accepters, the ratios of
(efflux to HDL(3))/(efflux to phosphatidylcholine vesicles) were calcu
lated for synthesized cholesterol and desmosterol, plasma membrane cho
lesterol, and lysosomal cholesterol. In comparison to plasma membrane
cholesterol, there was little or no HDL selectivity for lysosomal chol
esterol or synthesized desmosterol, whereas there was a 2-3-fold selec
tivity for synthesized cholesterol, suggesting that the ability of HDL
to enhance the efflux of synthesized sterols is a modest quantitative
effect and confined to cholesterol.