Use of Cyclodextrins to Monitor Transbilayer Movement and Differential Lipid Affinities of Cholesterol

In view of the demonstrated cholesterol-binding capabilities of certain cyclodextrins, we have examined whether these agents can also catalyze efficient transfer of cholesterol between lipid vesicles. We here demonstrate that β- and γ-cyclodextrins can dramatically accelerate the rate of cholesterol transfer between lipid vesicles under conditions where a negligible fraction of the sterol is bound to cyclodextrin in steady state. β- and γ-cyclodextrin enhance the rate of transfer of cholesterol between vesicles by a larger factor than they accelerate the transfer of phospholipid, whereas, for α- and methyl- β-cyclodextrin, the opposite is true. Analysis of the kinetics of cyclodextrin-mediated cholesterol transfer between large unilamellar vesicles composed mainly of 1-stearoyl-2-oleoyl phosphatidylcholine (SOPC) or SOPC/cholesterol indicates that transbilayer flip-flop of cholesterol is very rapid (halftime < 1–2 min at 37°C). Using β-cyclodextrin to accelerate cholesterol transfer, we have measured the relative affinities of cholesterol for a variety of different lipid species. Our results show strong variations in cholesterol affinity for phospholipids bearing different degrees of chain unsaturation and lesser, albeit significant, effects of phospholipid headgroup structure on cholesterol-binding affinity. Our findings also confirm previous suggestions that cholesterol interacts with markedly higher affinity with sphingolipids than with common membrane phospholipids.