title: Contribution of cholesterol and phospholipids to inhibitory effect of dimethyl-beta-cyclodextrin on efflux function of P-glycoprotein and multidrug resistance-associated protein 2 in vinblastine-resistant Caco-2 cell monolayers.
abstract: OBJECTIVE
  The purpose of this study is to reveal the contribution of membrane components to the inhibitory effect of 2,6-di-O-methyl-beta-cyclodextrin (DM-beta-CyD) on P-glycoprotein (P-gp) and multidrug resistance-associated protein 2 (MRP2) function in vinblastine-resistant Caco-2 (Caco-2R) cell monolayers.
  
  
  METHODS
  The transport of rhodamine-123 and 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF) was studied in Caco-2R cell monolayers. P-gp and MRP2 residing in the monolayers and releasing in cell supernatants were detected by Western blotting. The mRNA levels of MDR1 and MRP2 were detected by reverse transcription-polymerase chain reaction (RT-PCR) method. Cholesterol, phospholipids, and proteins were mainly determined by each assay kit.
  
  
  RESULTS
  Of various beta-cyclodextrin derivatives (beta-CyDs), DM-beta-CyD most significantly impaired the efflux function of P-gp and MRP2 without changing cell viability and membrane integrity. The treatment with CyDs did not change the mRNA levels of MDR1 and MRP2. DM-beta-CyD lowered cholesterol content and P-gp level in caveolar membranes. In addition, DM-beta-CyD released not only cholesterol and phospholipids but also proteins including P-gp and MRP2 from apical membranes of the monolayers.
  
  
  CONCLUSIONS
  DM-beta-CyD may impair P-gp and MRP2 function in Caco-2R cell monolayers, probably, at least in part, through the release of these transporters from the apical membranes of monolayers, and the exertion of the inhibitory effect of DM-beta-CyD may require the extraction of not only cholesterol but also phospholipids from the monolayers.
authors.affiliation: Graduate School of Pharmaceutical Sciences, Kumamoto University, 5-1 Oe-honmachi, Kumamoto 862-0973, Japan.
authors.forename: Hidetoshi
authors.initials: H
authors.lastname: Arima
authors.forename: Kiyokazu
authors.initials: K
authors.lastname: Yunomae
authors.forename: Tadatoshi
authors.initials: T
authors.lastname: Morikawa
authors.forename: Fumitoshi
authors.initials: F
authors.lastname: Hirayama
authors.forename: Kaneto
authors.initials: K
authors.lastname: Uekama
mesh_terms: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000037:antagonists & inhibitors* / Q000378:metabolism; D018938:Caco-2 Cells; D002784:Cholesterol / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D019008:Drug Resistance, Neoplasm / Q000187:drug effects* / Q000502:physiology; D006801:Humans; D049990:Membrane Transport Modulators; D026901:Membrane Transport Proteins / Q000037:antagonists & inhibitors*; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000037:antagonists & inhibitors*; D010743:Phospholipids / Q000378:metabolism; D014747:Vinblastine / Q000494:pharmacology*; D047392:beta-Cyclodextrins / Q000494:pharmacology*
chemical_list: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D002784:Cholesterol; D049990:Membrane Transport Modulators; D026901:Membrane Transport Proteins; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D010743:Phospholipids; D014747:Vinblastine; D047392:beta-Cyclodextrins; C000715087:ABCC2 protein, human; C038119:heptakis(2,6-O-dimethyl)beta-cyclodextrin