Pg. Yancey et al., CELLULAR CHOLESTEROL EFFECT MEDIATED BY CYCLODEXTRINS - DEMONSTRATIONOF KINETIC POOLS AND MECHANISM OF EFFLUX, The Journal of biological chemistry, 271(27), 1996, pp. 16026-16034
In this study, we compared the kinetics of cholesterol efflux from cel
ls with 2-hydroxypropyl-beta-cyclodextrins and with discoidal high den
sity lipoprotein (HDL) particles to probe the mechanisms governing the
remarkably rapid rates of cyclodextrin-mediated efflux. The rate of c
holesterol efflux was enhanced by shaking cells growing in a monolayer
and further enhanced by placing cells in suspension to achieve maxima
l efflux rates, The extent of efflux was dependent on cyclodextrin con
centration, and maximal efflux was observed at concentrations >50 mM.
For several cell types, biexponential kinetics of cellular cholesterol
efflux were observed, indicating the existence of two kinetic pools o
f cholesterol: a fast pool (half-time (t(1/2)) similar to 19-23 s) and
a slow pool with t(1/2) of 15-30 min. Two distinct kinetic pools of c
holesterol were also observed with model membranes (large unilamellar
cholesterol-containing vesicles), implying that the cellular pools are
in the plasma membrane, Cellular cholesterol content was altered by i
ncubating cells with solutions of cyclodextrins complexed with increas
ing levels of cholesterol. The number of kinetic pools was unaffected
by raising the cellular cholesterol content, but the size of the fast
pool increased, After depleting cells of the fast pool of cholesterol,
this pool was completely restored after a 40-min recovery period. The
temperature dependence of cyclodextrin-mediated cholesterol efflux fr
om cells and model membranes was compared; the activation energies wer
e 7 kcal/mol and 2 kcal/mol, respectively. The equivalent activation e
nergy observed with apo-HDL-phospholipid acceptor particles was 20 kca
l/mol, It seems that cyclodextrin molecules are substantially more eff
icient than phospholipid accepters, because cholesterol molecules deso
rbing from a membrane surface can diffuse directly into the hydrophobi
c core of a cyclodextrin molecule without having to desorb completely
into the aqueous phase before being sequestered by the acceptor.