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Molecular mechanism of steroid transporter P-glycoprotein

类固醇转运蛋白P-糖蛋白的分子机制

基本信息

批准号：
04660085
负责人：
UEDA Kazumitsu
金额：
$ 1.34万
依托单位：
KYOTO UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1992
资助国家：
日本
起止时间：
1992 至 1993
项目状态：
已结题

项目摘要

P-glycoprotein acts as an energy-dependent efflux pump that exports anticancer agents out of the cell, lowering their intracellular concentration to sublethal levels, and is considered to be important in multidrug resistance (MDR) of human tumors. P-glycoprotein is expressed in several normal human tissues, but physiological substrates of P-glycoprotein have not been fully understood.To analyze the mechanism of drug transport and physiological substrates, we established a transepithelial transport system. We expressed MDR1 cDNA isolated from human normal adrenal in LLC-PK1 cells, which are derived from porcine kidney proximal tubule. One transformant (LLC-GA5-COL300) showed 54-, 150-, and 167-fold resistance to colchicine, vinblastine, and adriamycin, respectively, compared to LLC-PK1 cells. LLC-PK_1 and the P-glycoprotein-expressing transfectant cells form a highly polarized epithelium, and electron microscopic immunocytochemistry using a monoclonal antibody MRK16 showed that human P- … More glycoprotein is specifically expressed on the apical surface of the transfectant cells.By using this transepithelial transport system, we measured transport of ^3H-labeled aldosterone, corticosterone, cortisol, deoxycorticosterone, dexamethasone, estradiol, estriol, progesterone, testesterone. In the transfectant cells, basal to apical transport of aldosterone, cortisol, estriol and dexamethasone increased and their apical to basal transport decreased compared to those in the host cells. But transport of corticosterone, deoxycorticosterone, estradiol, progesterone, testesterone was indistinguishable in both cell lines. Transepithelial transport of aldosterone, cortisol, estriol and dexamethasone was inhibited by verapamil. These results indicate that human P-glycoprotein transports aldosterone, cortisol, estriol and dexamethasone.Cortisol, a glucocorticoid, is the main steroid produced in the human adrenal. Aldosterone is the main mineralocorticoid produced in the human adrenal. Estriol is the main estrogen produced during pregnancy in the fetoplacental unit. P-glycoprotein is expressed in the human adrenal cortex and the pregnant uterus and placenta. These results suggest that human P-glycoprotein functions as a steroid transporter in the adrenal cortex and in the fetoplacental unit during pregnancy. Less

P-糖蛋白作为一种能量依赖性外排泵，将抗癌药物输出到细胞外，将其细胞内浓度降低至亚致死水平，并被认为在人类肿瘤的多药耐药（MDR）中很重要。P-糖蛋白在人体多种正常组织中均有表达，但其生理底物尚未完全了解，为了分析药物转运机制和生理底物，我们建立了一个跨上皮转运系统。我们从人正常肾上腺中分离MDR 1 cDNA，在来源于猪肾近曲小管的LLC-PK 1细胞中表达。其中一株LLC-GA 5-COL 300细胞对秋水仙碱、长春碱和阿霉素的耐药性分别是LLC-PK 1细胞的54倍、150倍和167倍。LLC-PK_1和表达P-糖蛋白的转染细胞形成高度极化的上皮细胞，用单克隆抗体MRK_（16）进行的电镜免疫细胞化学显示，人P-糖蛋白的表达与LLC-PK_1的表达有关。 ...更多信息利用这种跨上皮转运系统，我们测定了^3 H标记的醛固酮、皮质酮、皮质醇、脱氧皮质酮、地塞米松、雌二醇、雌三醇、孕酮和睾酮的转运。与宿主细胞相比，转染细胞中醛固酮、皮质醇、雌三醇和地塞米松的基底向顶端转运增加，而它们的顶端向基底转运减少。但皮质酮、去氧皮质酮、雌二醇、孕酮、睾酮的转运在两种细胞系中没有区别。维拉帕米抑制醛固酮、皮质醇、雌三醇和地塞米松的跨上皮转运。这些结果表明，人P-糖蛋白转运醛固酮、皮质醇、雌三醇和地塞米松，皮质醇是人肾上腺产生的主要类固醇激素。醛固酮是人体肾上腺中产生的主要盐皮质激素。雌三醇是妊娠期间胎儿胎盘单位产生的主要雌激素。P-糖蛋白在人肾上腺皮质、妊娠子宫和胎盘中表达。这些结果表明，人类P-糖蛋白的功能作为类固醇转运蛋白在肾上腺皮质和胎儿胎盘单位在怀孕期间。少