Sterol carrier protein-2 alters high density lipoprotein-mediated cholesterol efflux

Although sterol carrier protein-2 (SCP-2) participates in the uptake and in tracellular trafficking of cholesterol, its effect on "reverse cholesterol transport" has not been explored. As shown herein, SCP-2 expression inhibit ed high density lipoprotein (HDL)-mediated efflux of [H-3]cholesterol and f luorescent 22-(N-(7-nitro-benz-oxa-1,3-diazol-4-yl)amino) -23,24-bisnor-5-c holen-3b-ol (NBD-cholesterol) up to 61 and 157%, respectively. Confocal mic roscopy of living cells allowed kinetic analysis of two intracellular pools of HDL-mediated NBD-cholesterol efflux: the highly fluorescent lipid dropl et pool and the less fluorescent pool outside the lipid droplets, designate d the cytoplasmic compartment, Both the whole cell and the cytoplasmic comp artment exhibited two similar kinetic pools, the half-times of which were c onsistent with protein (t(1/2)(b) near 1 min) and vesicular (t(1/2)(d) = 10 -20 min) mediated sterol transfer. Although SCP-2 expression did not alter cytoplasmic sterol pool sizes, the rapid t(1/2)(b) decreased 36%, while the slower t(1/2)(d) increased 113%. Lipid droplets also exhibited two kinetic pools of NBD-cholesterol efflux but with half-times over 200% shorter than those of the cytoplasmic compartment. The lipid droplet slower effluxing p ool size and t(1/2)(d), were increased 48% and 115%, respectively, in SCP-a -expressing cells. Concomitantly, the level of the lipid droplet-specific a dipose differentiation-related protein decreased 70%. Overall, HDL-mediated sterol efflux from L-cell fibroblasts reflected that of the cytoplasmic ra ther than lipid droplet compartment, SCP-2 differentially modulated sterol efflux from the two cytoplasmic pools. However, net efflux was determined p rimarily by inhibition of the slowly effluxing pool rather than by accelera tion of the rapid protein-mediated pool. Finally, SCP-2 expression also inh ibited sterol efflux from lipid droplets, an effect related to decreased ad ipose differentiation-related protein, a lipid droplet surface protein that binds cholesterol with high affinity.