title: Use of cyclodextrin for AFM monitoring of model raft formation.
abstract: The lipid rafts membrane microdomains, enriched in sphingolipids and cholesterol, are implicated in numerous functions of biological membranes. Using atomic force microscopy, we have examined the effects of cholesterol-loaded methyl-beta-cyclodextrin (MbetaCD-Chl) addition to liquid disordered (l(d))-gel phase separated dioleoylphosphatidylcholine (DOPC)/sphingomyelin (SM) and 1-palmitoyl-2-oleoyl phosphatidylcholine (POPC)/SM supported bilayers. We observed that incubation with MbetaCD-Chl led to the disappearance of domains with the formation of a homogeneously flat bilayer, most likely in the liquid-ordered (l(o)) state. However, intermediate stages differed with the passage through the coexistence of l(o)-l(d) phases for DOPC/SM samples and of l(o)-gel phases for POPC/SM bilayers. Thus, gel phase SM domains surrounded by a l(o) matrix rich in cholesterol and POPC could be observed just before reaching the uniform l(o) state. This suggests that raft formation in biological membranes could occur not only via liquid-liquid but also via gel-liquid immiscibility. The data also demonstrate that MbetaCD-Chl as well as the unloaded cyclodextrin MbetaCD make holes and preferentially extract SM in supported bilayers. This strongly suggests that interpretation of MbetaCD and MbetaCD-Chl effects on cell membranes only in terms of cholesterol movements have to be treated with caution.
authors.affiliation: Centre de Biochimie Structurale, CNRS UMR 5048-Université Montpellier I, INSERM UMR554, 34090 Montpellier Cedex, France.
authors.forename: Marie-Cécile
authors.initials: MC
authors.lastname: Giocondi
authors.forename: Pierre Emmanuel
authors.initials: PE
authors.lastname: Milhiet
authors.forename: Patrice
authors.initials: P
authors.lastname: Dosset
authors.forename: Christian
authors.initials: C
authors.lastname: Le Grimellec
mesh_terms: D002784:Cholesterol / Q000737:chemistry*; D003505:Cyclodextrins / Q000737:chemistry*; D008051:Lipid Bilayers / Q000737:chemistry*; D046911:Macromolecular Substances; D008560:Membrane Fluidity; D021962:Membrane Microdomains / Q000737:chemistry* / Q000000981:diagnostic imaging*; D018625:Microscopy, Atomic Force / Q000379:methods*; D008968:Molecular Conformation; D010539:Permeability; D044367:Phase Transition; D010713:Phosphatidylcholines / Q000737:chemistry; D013109:Sphingomyelins / Q000737:chemistry; D014463:Ultrasonography
chemical_list: D003505:Cyclodextrins; D008051:Lipid Bilayers; D046911:Macromolecular Substances; D010713:Phosphatidylcholines; D013109:Sphingomyelins; D002784:Cholesterol; C017251:1,2-oleoylphosphatidylcholine; C028694:1-palmitoyl-2-oleoylphosphatidylcholine