Quantitative contributions of cholesterol and the individual classes of phospholipids and their degree of fatty acyl (un)saturation to membrane fluidity measured by fluorescence polarization

Wj, van Blitterswijk; Bw, van der Meer; Hilkmann, Henk

doi:10.1021/bi00380a038

Cited by 200 publications

(100 citation statements)

References 42 publications

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“…This possibly indicates a saturation of particular membrane insertion sites. Once in the membrane, C 6 -SM might not be randomly dispersed but, like natural long-chain sphingolipids, show a tendency to selfaggregate in the plane of the membrane (van Blitterswijk et al, 1987(van Blitterswijk et al, , 2003Holopainen et al, 1998). It is therefore appealing to speculate that these physical properties allow C 6 -SM to form their own 'artificial' rafts, which, however, remain detergent soluble (as found here).…”

Section: Propidium Iodidementioning

confidence: 60%

N-hexanoyl-sphingomyelin potentiates in vitro doxorubicin cytotoxicity by enhancing its cellular influx

et al. 2004

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Anticancer drugs generally have intracellular targets, implicating transport over the plasma membrane. For amphiphilic agents, such as the anthracycline doxorubicin, this occurs by passive diffusion. We investigated whether exogenous membrane-permeable lipid analogues improve this drug influx. Combinations of drugs and lipid analogues were coadministered to cultured endothelial cells and various tumour cell lines, and subsequent drug accumulation in cells was quantified. We identified N-hexanoyl-sphingomyelin (SM) as a potent enhancer of drug uptake. Low micromolar amounts of this short-chain sphingolipid, being not toxic itself, enhanced the uptake of doxorubicin up to 300% and decreased its EC 50 toxicity values seven-to 14-fold. N-hexanoyl SM acts at the level of the plasma membrane, but was found not incorporated in (isolated) lipid rafts, and artificial disruption or elimination of raft constituents did not affect its drug uptake-enhancing effect. Further, any mechanistic role of the endocytic machinery, membrane leakage or ABCtransporter-mediated efflux could be excluded. Finally, a correlation was established between the degree of drug lipophilicity, as defined by partitioning in a two-phase octanol -water system, and the susceptibility of the drug towards the uptake-enhancing effect of the sphingolipid. A clear optimum was found for amphiphilic drugs, such as doxorubicin, epirubicin and topotecan, indicating that N-hexanoyl-SM might act by modulating the average degree of plasma membrane lipophilicity, in turn facilitating transbilayer drug diffusion. The concept of short-chain sphingolipids as amphiphilic drug potentiators provides novel opportunities for improving drug delivery technologies.

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Section: Propidium Iodidementioning

confidence: 60%

N-hexanoyl-sphingomyelin potentiates in vitro doxorubicin cytotoxicity by enhancing its cellular influx

et al. 2004

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“…These results indicated that the reduction of cellular SM to ϳ15% of the wild-type level has no effect on the steady-state level of cellular cholesterol and its distribution to the plasma membrane. Enrichment of SM in the plasma membrane has been hypothesized to force cholesterol to be also enriched in the plasma membrane, based on the observations that addition of SM to cultured human skin fibroblasts results in an increase in cellular cholesterol content according to the increase in cellular SM level (35), and that cholesterol interacts with SM more strongly than with glycerophospholipids in model membranes (6,36,37). However, our analyses with CHO cells suggest that neither SM, glycosphingolipids, nor ceramide is essential for the preferential distribution of cholesterol to the plasma membrane.…”

Section: Discussionmentioning

confidence: 99%

Reduction of Sphingomyelin Level without Accumulation of Ceramide in Chinese Hamster Ovary Cells Affects Detergent-resistant Membrane Domains and Enhances Cellular Cholesterol Efflux to Methyl-β-cyclodextrin

Fukasawa¹,

Nishijima²,

Itabe³

et al. 2000

Journal of Biological Chemistry

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We examined the effects of reduction of sphingomyelin level on cholesterol behavior in cells using 2 types of Chinese hamster ovary cell mutants deficient in sphingomyelin synthesis: LY-A strain defective in intracellular trafficking of ceramide for sphingomyelin synthesis, and LY-B strain defective in the enzyme catalyzing the initial step of sphingolipid biosynthesis. Although the sphingomyelin content in LY-A and LY-B cells was ϳ40 and ϳ15%, respectively, of the wild-type level without accumulation of ceramide, these mutant cells were almost identical in cholesterol content and also in plasma membrane cholesterol level to the wild-type cells. However, density gradient fractionation analysis of Triton X-100-treated lysates of cells prelabeled with [ 3 H]cholesterol showed that the [ 3 H]cholesterol level in the lowdensity floating fraction was lower in sphingomyelindeficient cells than in wild-type cells. When cells were exposed to methyl-␤-cyclodextrin, cholesterol was more efficiently fluxed from sphingomyelin-deficient cells than wild-type cells. These results suggest that the steady state level of cholesterol at the plasma membrane is little affected by the sphingomyelin levels in Chinese hamster ovary cells, but that sphingomyelin levels play an important role in the retention of cholesterol in the plasma membrane against efflux to extracellular cholesterol-acceptors, due to interaction between sphingomyelin and cholesterol in detergent-resistant membrane domains. Both cholesterol and sphingomyelin (SM)1 are preferentially distributed in the plasma membrane of cells (1). Recent developments in membrane biology have demonstrated various lines of evidence that the plasma membrane has microdomains, termed detergent-resistant membrane (DRM) domains or lipid rafts, which are involved in various cellular events, including signal transduction and membrane trafficking (2, 3). DRM domains are highly enriched in cholesterol and sphingolipids, and probably exist as liquid-ordered phase, characterized by a conformationally ordered state of the acyl chains of phospholipids that are laterally diffusible (3). Formation of the liquidordered phase reflects the fact that cholesterol interacts favorably with phospholipid acyl chains in an extended conformation. SM and saturated glycerophospholipids readily form the extended acyl chain conformation, while unsaturated phospholipids having cis-configuration of the double bond do not. Many studies with model membranes have indicated that cholesterol interacts with SM more strongly than with unsaturated glycerophospholipids, the predominant forms of natural glycerophospholipids (4 -7). In addition, previous studies with model membrane vesicles consisting of pure lipids have shown that cholesterol enhances detergent insolubility of SM, and that sphingolipids or saturated glycerophospholipids tending to form the lipid-ordered phase are also important for detergent insolubility of cholesterol (8,9).Only a few reports, however, have addressed the issue of the participation of SM in ...

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“…It is well established that lipid structural order in membranes is largely determined by cholesterol and sphingomyelin content and by the degree of saturation of the phospholipid fatty acyl chains (7,8). Most of the techniques used to study cellular membrane lipid involve destruction or chemical perturbation of the membrane under study.…”

Section: Introductionmentioning

confidence: 99%

1H-NMR detectable fatty acyl chain unsaturation in excised leiomyosarcoma correlate with grade and mitotic activity.

Singer¹,

Sivaraja²,

Souza³

et al. 1996

J. Clin. Invest.

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Quantitative contributions of cholesterol and the individual classes of phospholipids and their degree of fatty acyl (un)saturation to membrane fluidity measured by fluorescence polarization

Cited by 200 publications

References 42 publications

N-hexanoyl-sphingomyelin potentiates in vitro doxorubicin cytotoxicity by enhancing its cellular influx

N-hexanoyl-sphingomyelin potentiates in vitro doxorubicin cytotoxicity by enhancing its cellular influx

Reduction of Sphingomyelin Level without Accumulation of Ceramide in Chinese Hamster Ovary Cells Affects Detergent-resistant Membrane Domains and Enhances Cellular Cholesterol Efflux to Methyl-β-cyclodextrin

1H-NMR detectable fatty acyl chain unsaturation in excised leiomyosarcoma correlate with grade and mitotic activity.

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