TY - JOUR
T1 - Elevation of glucosylceramide in multidrug-resistant cancer cells and accumulation in cytoplasmic droplets
AU - Morjani, Hamid
AU - Aouali, Nasséra
AU - Belhoussine, Rajae
AU - Veldman, Robert Jan
AU - Levade, Thierry
AU - Manfait, Michel
PY - 2001/10/15
Y1 - 2001/10/15
N2 - Multidrug-resistant (MDR) cancer cells have been shown to have an accumulation of glucosylceramide (GlcCer). In this study, we aim at localizing, at subcellular level, where these lipids accumulate. Neutral lipids and phospholipid containing organelles have been identified using confocal fluorescence microscopy and microspectrofluorometry by monitoring the emission of the fluorescent probe Nile-red. Data from confocal fluorescence microscopy analysis shows accumulation of neutral lipids in cytoplasmic droplets of MDR human carcinoma MCF7R cells. Microspectrofluorometric measurements show an increase of the gold-yellow emission intensity in MCF7R cells, corresponding to neutral lipids. Similar observations were made in human MDR vincristine-HL60 and doxorubicin-KB selected cells. Total cellular glucosylceramide (GlcCer) measurements using [3H]-palmitic acid and thin layer chromatography show a significant increase of GlcCer in MCF7R cells. Moreover, MCF7R cells treated with fluorescent GlcCer-bodipy exhibit an accumulation of this lipid in cytoplasmic droplets. Treatment of MCF7R cells with 1-phenyl-2-palmitoylamino-3-morpholino-1-propanolol (PPMP), a potent inhibitor of GlcCer synthase, attenuates the Nile-red fluorescence emission emanating from these structures and reverses MDR. Moreover, Golgi compartments stained with fluorescent PPMP-bodipy, show an increase in the Golgi compartments density. Treatment of MCF7R cells with cyclosporine A (CSA), tamoxifen (TMX) and 3′-azido-3′deoxythymidine (AZT) leads to the same effect observed in the presence of PPMP. Treatment of MCF7 and MCF7R with the β-glucosidase inhibitor conduritol β-epoxide (CBE) significantly increases resistance to daunorubicin only in MCF7R cells. These data demonstrate also that: (i) CSA, an inhibitor of MDR, has an additional target in addition to P-glycoprotein; and (ii) TMX (used in breast cancer treatment and prevention) and AZT (used in the treatment of HIV) could have side effects by disturbing lipid metabolism and inhibiting many cellular functions required in normal cells.
AB - Multidrug-resistant (MDR) cancer cells have been shown to have an accumulation of glucosylceramide (GlcCer). In this study, we aim at localizing, at subcellular level, where these lipids accumulate. Neutral lipids and phospholipid containing organelles have been identified using confocal fluorescence microscopy and microspectrofluorometry by monitoring the emission of the fluorescent probe Nile-red. Data from confocal fluorescence microscopy analysis shows accumulation of neutral lipids in cytoplasmic droplets of MDR human carcinoma MCF7R cells. Microspectrofluorometric measurements show an increase of the gold-yellow emission intensity in MCF7R cells, corresponding to neutral lipids. Similar observations were made in human MDR vincristine-HL60 and doxorubicin-KB selected cells. Total cellular glucosylceramide (GlcCer) measurements using [3H]-palmitic acid and thin layer chromatography show a significant increase of GlcCer in MCF7R cells. Moreover, MCF7R cells treated with fluorescent GlcCer-bodipy exhibit an accumulation of this lipid in cytoplasmic droplets. Treatment of MCF7R cells with 1-phenyl-2-palmitoylamino-3-morpholino-1-propanolol (PPMP), a potent inhibitor of GlcCer synthase, attenuates the Nile-red fluorescence emission emanating from these structures and reverses MDR. Moreover, Golgi compartments stained with fluorescent PPMP-bodipy, show an increase in the Golgi compartments density. Treatment of MCF7R cells with cyclosporine A (CSA), tamoxifen (TMX) and 3′-azido-3′deoxythymidine (AZT) leads to the same effect observed in the presence of PPMP. Treatment of MCF7 and MCF7R with the β-glucosidase inhibitor conduritol β-epoxide (CBE) significantly increases resistance to daunorubicin only in MCF7R cells. These data demonstrate also that: (i) CSA, an inhibitor of MDR, has an additional target in addition to P-glycoprotein; and (ii) TMX (used in breast cancer treatment and prevention) and AZT (used in the treatment of HIV) could have side effects by disturbing lipid metabolism and inhibiting many cellular functions required in normal cells.
UR - http://www.scopus.com/inward/record.url?scp=0035888295&partnerID=8YFLogxK
U2 - 10.1002/ijc.1449
DO - 10.1002/ijc.1449
M3 - Article
C2 - 11668492
AN - SCOPUS:0035888295
SN - 0020-7136
VL - 94
SP - 157
EP - 165
JO - International Journal of Cancer
JF - International Journal of Cancer
IS - 2
ER -