Abstract
BackgroundMost cancer cells are characterized by mobile lipids visible on proton NMR (1H-NMR), these being comprised mainly of methyl and methylene signals from lipid acyl chains. Erythroleukemia K562 cells show narrow signals at 1.3 and 0.9 ppm, corresponding to mobile lipids (methylene and methyl, respectively), which are reduced when K562 cells are multidrug resistant (MDR). While the significance of the mobile lipids is unknown, their subcellular localization is still a matter of debate and may lie in the membrane or the cytoplasm. In this study, we investigate the role of cholesterol in the generation of mobile lipid signals.ResultsThe proportion of esterified cholesterol was found to be higher in K562-sensitive cells than in resistant cells, while the total cholesterol content was identical in both cell lines. Cholesterol extraction in the K562 wild type (K562wt) cell line and its MDR counterpart (K562adr), using methyl-β-cyclodextrin, was accompanied by a rise of mobile lipids in K562wt cells only. The absence of caveolae was checked by searching for the caveolin-1 protein in K562wt and K562adr cells. However, cholesterol was enriched in another membrane microdomain designated as "detergent-insoluble glycosphingomyelin complexes" or rafts. These microdomains were studied after extraction with triton X-100, a mild non-ionic detergent, revealing mobile lipid signals preserved only in the K562wt spectra. Moreover, following perturbation/disruption of these microdomains using sphingomyelinase, mobile lipids increased only in K562wt cells.ConclusionThese results suggest that cholesterol and sphingomyelin are involved in mobile lipid generation via microdomains of detergent-insoluble glycosphingomyelin complexes such as rafts. Increasing our knowledge of membrane microdomains in sensitive and resistant cell lines may open up new possibilities in resistance reversion.
Highlights
Most cancer cells are characterized by mobile lipids visible on proton NMR (1HNMR), these being comprised mainly of methyl and methylene signals from lipid acyl chains
When studied by NMR proton spectroscopy, most cancer cells are characterized by increased narrow signals at 0.9 and 1.3 ppm corresponding, respectively, to methyl and methylene resonances that belong to lipid acyl chains moving isotropically
Data points are the percentages of the cellular concentration normalized to cellular concentration at T = 0 hours expressed as means, with vertical bars representing standard deviation (SD)
Summary
Most cancer cells are characterized by mobile lipids visible on proton NMR (1HNMR), these being comprised mainly of methyl and methylene signals from lipid acyl chains. When studied by NMR proton spectroscopy, most cancer cells are characterized by increased narrow signals at 0.9 and 1.3 ppm corresponding, respectively, to methyl and methylene resonances that belong to lipid acyl chains moving isotropically. This so-called "mobile lipid signal" (or ML signal) has been studied for several decades (for review see[1]). Research has failed to elucidate the molecular origin of mobile lipids, their subcellular localization or their physiological significance Regarding their molecular origin, acyl chains can form part of triglycerides or esterified cholesterol. These studies (op cit.) tend to show that cytosolic lipid droplets generate mobile lipid signals in NMR spectra, without excluding the possibility that such signals could arise independently of the presence of cytosolic lipid bodies
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