Submitted on November 29, 2005
Revised on March 16, 2006
Accepted on March 17, 2006
Gangliosides do not affect ABC transporter function in human neuroblastoma cells
Anne-Jan Dijkhuis, Karin Klappe, Willem Kamps, Hannie Sietsma, and Jan Willem Kok
Cell Biology, section Membrane Cell Biology, University Medical Center Groningen, Groningen 9713 AV
Corresponding Author: j.w.kok{at}med.umcg.nl
Previous studies have indicated a role for glucosylceramide synthase (GCS) in multidrug resistance (MDR), either related to turnover of ceramide (Cer) or generation of gangliosides, which modulate apoptosis and/or activity of ATP binding cassette (ABC) transporters. This study challenges the hypothesis that gangliosides modulate the activity of ABC transporters and was performed in two human neuroblastoma cell lines, expressing either functional P-glycoprotein (Pgp) or multidrug resistance related protein 1 (MRP1). Two inhibitors of GCS, D,L-threo-1-phenyl-2-hexadecanoylamino-3-pyrrolidino-1-propanol (t-PPPP) or N-butyldeoxynojirimycin (NB-dNJ), very efficiently depleted ganglioside content in two human neuroblastoma cell lines. This was established by three different assays, i.e. equilibrium radiolabeling, cholera toxin binding and mass analysis. FACS analysis showed that ganglioside depletion only slightly and in opposite direction affected Pgp- and MRP1-mediated efflux activity. Moreover, both effects were marginal compared to well established inhibitors of either MRP1, i.e. MK571 or Pgp, i.e. GF120918. t-PPPP slightly enhanced cellular sensitivity to vincristine, as determined by MTT analysis, in both neuroblastoma cell lines, while NB-dNJ was without effect. MRP1 expression and its localization in detergent-resistant membranes (DRMs) were not affected by ganglioside depletion. Altogether, these results show that gangliosides are not relevant to ABC transporter-mediated MDR in neuroblastoma cells.
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