Contribution of Cholesterol and Phospholipids to Inhibitory Effect of Dimethyl-β-Cyclodextrin on Efflux Function of P-Glycoprotein and Multidrug Resistance-Associated Protein 2 in Vinblastine-Resistant Caco-2 Cell Monolayers

Publisher Website
Google Scholar
Abstract
Purpose. The purpose of this study is to reveal the contribution of membrane components to the inhibitory effect of 2,6-di-O-methyl-β-cyclodextrin (DM-β-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-carboxy- ethyl)-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 Westerrn 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 β-cyclodextrin derivatives (β-CyDs), DM-β-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-β-CyD lowered cholesterol content and P-gp level in caveolar membranes. In addition, DM-β-CyD released not only cholesterol and phospholipids but also proteins including P-gp and MRP2 from apical membranes of the monolayers. Conclusions. DM-β-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-β-CyD may require the extraction of not only cholesterol but also phospholipids from the monolayers.