Detection of P-glycoprotein in the nuclear envelope of multidrug resistant cells.
P-glycoprotein is a plasma membrane efflux pump which is responsible for multidrug resistance of many cancer cell lines. A number of studies have demonstrated the presence of P-glycoprotein molecules, besides on the plasma membrane, also in intracellular sites, such as the Golgi apparatus and the nucleus. In this study, the presence and function of P-glycoprotein in the nuclear membranes of human breast cancer cells (MCF-7 WT) and their multidrug resistant variants (MCF-7 DX) were investigated. Electron and confocal microscopy immunolabelling experiments demonstrated the presence of P-glycoprotein molecules in the nuclear membranes of MCF-7 DX cells. Moreover, the labelling pattern was strongly dependent on pH values of the incubation buffer. At physiological pH (7.2), a strong labelling was detected in the cytoplasm and the nuclear matrix in both sensitive and resistant MCF-7 cells. By raising the pH to 8.0, the P-glycoprotein molecules were easily detected in the cytoplasm (transport vesicles and Golgi apparatus), plasma and nuclear membranes exclusively in MCF-7 DX cells. Furthermore, drug uptake and efflux studies, performed by flow cytometry on isolated nuclei in the presence of the P-glycoprotein inhibitor cyclosporin A, suggested the presence of a functional P-glycoprotein in the nuclear membrane, but not in the nuclear matrix, of drug resistant cells. Therefore, P-glycoprotein in the nuclear envelope seems to represent a further defense mechanism developed by resistant cells against antineoplastic agents.
Calcabrini
A
A
Laboratorio di Ultrastrutture, Istituto Superiore di Sanità, Rome, Italy.
Meschini
S
S
Stringaro
A
A
Cianfriglia
M
M
Arancia
G
G
Molinari
A
A
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000032:analysis*; D000970:Antineoplastic Agents / Q000493:pharmacokinetics / Q000494:pharmacology; D004273:DNA, Neoplasm / Q000032:analysis; D004317:Doxorubicin / Q000493:pharmacokinetics / Q000494:pharmacology; D018432:Drug Resistance, Multiple; D005260:Female; D005434:Flow Cytometry; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D018613:Microscopy, Confocal; D016253:Microscopy, Immunoelectron; D009685:Nuclear Envelope / Q000737:chemistry* / Q000648:ultrastructure; D014407:Tumor Cells, Cultured
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000970:Antineoplastic Agents; D004273:DNA, Neoplasm; D004317:Doxorubicin