Calcabrini A, Meschini S, Stringaro A, Cianfriglia M, Arancia G, Molinari A
Laboratorio di Ultrastrutture, Istituto Superiore di Sanità, Rome, Italy.
Histochem J. 2000 Oct;32(10):599-606. doi: 10.1023/a:1026732405381.
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.
P-糖蛋白是一种质膜外排泵，它导致许多癌细胞系产生多药耐药性。大量研究表明，P-糖蛋白分子除了存在于质膜上，还存在于细胞内位点，如高尔基体和细胞核。在本研究中，对人乳腺癌细胞（MCF-7 WT）及其多药耐药变体（MCF-7 DX）核膜中P-糖蛋白的存在及功能进行了研究。电子显微镜和共聚焦显微镜免疫标记实验证明了MCF-7 DX细胞核膜中存在P-糖蛋白分子。此外，标记模式强烈依赖于孵育缓冲液的pH值。在生理pH（7.2）时，在敏感和耐药的MCF-7细胞的细胞质和核基质中均检测到强标记。将pH提高到8.0时，仅在MCF-7 DX细胞的细胞质（运输小泡和高尔基体）、质膜和核膜中容易检测到P-糖蛋白分子。此外，在P-糖蛋白抑制剂环孢素A存在的情况下，通过流式细胞术对分离的细胞核进行药物摄取和外排研究，表明耐药细胞核膜中存在功能性P-糖蛋白，但核基质中不存在。因此，核被膜中的P-糖蛋白似乎代表了耐药细胞对抗肿瘤药物形成的另一种防御机制。
