Jj. Klansek et al., COMPARTMENTAL ISOLATION OF CHOLESTEROL PARTICIPATING IN THE CYTOPLASMIC CHOLESTERYL ESTER CYCLE IN CHINESE-HAMSTER OVARY 25-RA CELLS, The Journal of biological chemistry, 271(9), 1996, pp. 4923-4929
Using the Chinese hamster ovary cell line, 25-RA, we have demonstrated
that lipoprotein-derived cholesterol and endogenously synthesized cho
lesterol are selectively differentiated with respect to their cellular
locations, These cells lack sterol-mediated regulation, spontaneously
storing large amounts of esterified cholesterol, which turns over wit
h a half-time of 7.5 h. When [H-3]cholesterol was provided to the cell
s in serum to trace cellular cholesterol, the specific activities of c
ellular free and esterified cholesterol (6238 +/- 273 and 5128 +/- 277
cpm/mu g, respectively) failed to equilibrate, indicating that bulk c
ellular free cholesterol is isolated from that participating in the ch
olesteryl ester cycle. Using [H-3]acetate to trace the fate of endogen
ously synthesized cholesterol, a failure of equilibration was also obs
erved (specific activities of free and esterified cholesterol = 280 +/
- 37 and 458 +/- 8 cpm/mu g, respectively), The lower specific activit
y of the precursor indicates that endogenously synthesized cholesterol
is preferentially esterified, When cells radiolabeled with [H-3]aceta
te were post-incubated in the absence of radiolabel, the specific acti
vity of the esterified cholesterol pool remained significantly higher
than that of the free cholesterol, suggesting that cholesterol derived
from hydrolysis of esterified cholesterol is preferentially re-esteri
fied.